Lactic dehydrogenase virus infection reduces the expulsion of the nematode <Emphasis Type="Italic">Nippostrongylus brasiliensis</Emphasis>

The interleukin (IL)-13-mediated goblet cell response is the major host effector system involved in the expulsion of Nippostrongylus brasiliensis. Lactic dehydrogenase virus (LDV) induced higher levels of N. brasiliensis egg production compared with controls, but the effect of LDV infection on worm expulsion of, and goblet cell and IL-13 responses to, N. brasiliensis have not been studied. In this study, the effects of LDV infection on these host responses against N. brasiliensis were examined. Mice with chronic LDV infection showed significantly lower worm expulsion rates than non-LDV-infected mice after N. brasiliensis infection, and there were no significant differences in the ratio of female versus male adult worms between control and LDV-infected mice. The number of goblet cells in LDV-infected mice was significantly lower than that in controls. In addition, the levels of IL-13 gene expression in lymph nodes were significantly lower in LDV-infected mice compared with controls. These results suggest that LDV infection reduces the protective immune responses against N. brasiliensis infection by the suppression of IL-13 production.     

Immunohistochemical study of skin lesions in herpes zoster

Summary Thirty-seven biopsy skin tissues of herpes zoster taken from 27 patients were analysed immunohistochemically using two monoclonal antibodies detecting either nucleocapsid or glycoproteins of varicella-zoster virus (VZV) on paraffin sections of formalin fixed tissues. Skin lesions of herpes zoster were divided clinically into four stages: erythematous, vesicular, pustular and ulcerative. In the erythematous stage, VZV antigens, if detected, were found only within ballooning cells in the lower epidermis or follicular epithelium. In the vesicular stage, antigens were detected in the cells around and within the intraepidermal vesicles and in histiocytes or fibrocytes of the dermis in all cases and in the endothelial or perineural cells in 10 of 14 cases. In the pustular stage, the antigens were observed in degenerated or necrotic keratinocytes and multinucleated giant cells within pustules and some necrotic cells in the dermis. In the ulcerative stage, the viral antigens were detected only at the ulcer margin and around the hair shaft in 2 of 7 cases. These results suggest that VZV initially involves the epidermis in the erythematous stage, subsequently invades the dermis in the vesicular stage, and disappears in the early ulcerative stage.     

CD27/CD70 interaction directly drives B cell IgG and IgM synthesis

CD27 is a T cell activation antigen expressed on a majority of peripheral blood T cells. CD27 is also expressed on a subpopulation of human B cells, and it is reported that CD27⁺ B cells secrete both IgG and IgM. CD70, a ligand for CD27, is expressed on activated T and B cells, suggesting an interaction between T and B cells via CD27/CD70 ligation. Here, we analyze B cell immunoglobulin synthesis using a CD70 transfectant and present functional data showing that B cells secrete large amounts of IgG and IgM as a result of the CD27/CD70 interaction. A flow cytometric analysis showed that CD27 expression was increased and CD70 was expressed on tonsillar and peripheral blood B cells after activation with Staphylococcus aureus Cowan strain (SAC) plus interleukin (IL-2). In addition, the proliferation of B cells was enhanced mildly by the addition of CD70 transfectant, and its proliferation was blocked by anti-CD70 mAb. More importantly, the CD70 transfectant enhanced IgG and IgM production by purified B cells greatly in the presence of SAC plus IL-2. The enhancement was completely blocked by the addition of either anti-CD70 mAb or anti-CD27 mAb. Strongly suggesting that the interaction of CD27 with its ligand, CD70, on B cells plays an important role in B cell growth and differentiation to produce IgG and IgM.     

Genetic dissection of the complex pathological manifestations of collagen disease in MRL/lpr mice

An MRL strain of mice bearing a Fas-deletion mutant gene, lpr, MRL/MpJ-lpr/lpr (MRL/lpr) develops collagen disease involving vasculitis, glomerulonephritis, arthritis and sialoadenitis, each of which has been studied as a model for polyarteritis, lupus nephritis, rheumatoid arthritis and Sjögren’s syndrome, respectively. Development of such lesions seems dependent on host genetic background since the congenic C3H/HeJ-lpr/lpr (C3H/lpr) mice rarely develop them. To identify the gene loci affecting each lesion, a genetic dissection of these complex pathological manifestations was carried out. First, histopathological features in MRL/lpr, C3H/lpr, (MRL/lpr × C3H/lpr) F1 intercross, and MRL/lpr × (MRL/lpr × C3H/lpr) F1 backcross mice were analyzed. Genomic DNA of the backcross mice were subjected to association studies by Chi-squared analysis for determining which polymorphic microsatellite locus occurs at higher frequency among affected compared to unaffected individuals for each lesion. As a result, gene loci recessively associated with each lesion were mapped on different chromosomal positions. We concluded that each of these lesions in MRL/lpr mice is under the control of a different set of genes, suggesting that the complex pathological manifestations of collagen disease result from polygenic inheritance.     

DNA typing of HLA in the patients with moyamoya disease

Summary Moyamoya disease is a clinical entity demonstrating a chronic occlusion of the cerebrovascular system. Although some possible etiological factors have been postulated, the etiology of this disease is still unknown. So far, some investigations have suggested the association between moyamoya disease and HLA in the serological typing. However, DNA typing of HLA have not been performed yet. Thus, we performed DNA-typing of HLA in the unrelated Japanese patients with definite moyamoya disease, using the polymerase chain reaction-sequence specific oligonucleotide probe (PCR-SSOP) technique. In the total patients,DQB1*0502 had a positive association with the disease. On the other hand,DRB1*0405 andDQB1*0401 showed a negative association. In comparing the early-onset and late-onset groups, two groups did not share the same disease associated alleles at all. Thus, the etiology of moyamoya disease seem to have a genetic background. Furthermore, different genetic factors might also be involved in the difference between the early-onset and late-onset groups.     

Sequential compound exocytosis of large dense-core vesicles in PC12 cells studied with TEPIQ (two-photon extracellular polar-tracer imaging-based quantification) analysis

We investigated exocytosis of PC12 cells using two-photon excitation imaging and extracellular polar tracers (TEP imaging) at the basal region of PC12 cells adjacent to the glass cover slip. TEPIQ (two-photon extracellular polar-tracer imaging-based quantification) analysis revealed that most exocytosis was mediated by large dense-core vesicles (LVs) with a mean diameter of 220 nm, and that exocytosis of LVs occurred slowly with a mean latency of ∼7 s even though exocytosis was induced with large increases in cytosolic Ca²⁺ concentration by uncaging of a caged-Ca²⁺ compound. We also found that 97% of exocytic LVs remained poised at the plasma membrane, 72% maintained their fusion pores in an open conformation for more than 30 s, and 76% triggered sequential compound exocytosis of vesicles that were located deeper in the cytosol. Sequential compound exocytosis by PC12 cells was confirmed by electron microscopic investigation with photoconversion of diaminobenzidine by FM1-43 (a polar membrane tracer). Our data suggest that pre-stimulus docking of LVs to the plasma membrane does not necessarily hasten the fusion reaction, while docking and resulting stability of exocytic LVs facilitates sequential compound exocytosis, and thereby allowing mobilization of deep vesicles.     

The morphologic feature of mucus leakage appearing in low papillary carcinoma of the breast.

This report summarizes the clinicopathologic findings in 11 cases of low papillary carcinoma of the breast accompanied by the morphologic feature of mucus leakage into the mammary stroma. These cases were characterized by two morphologic findings. First, abundant mucus produced by the tumor cells filled the intraductal spaces where neoplastic epithelium formed very low papillary projections, ie, a feature of mucinous-producing low papillary carcinoma in situ. Second, there was expansive leakage of mucus into the mammary stroma occasionally accompanied by a few epithelial cells. All the cases showed a high level of mucus production and contained no elements of invasive ductal carcinoma or ordinary invasive mucinous carcinoma. These cases have no evidence of direct invasion of the mammary stroma by malignant cells. The average age of the 11 patients was 41 years. Foci of microcalcification were seen in some tumors (seven cases; 64%). There were no cases with lymph node metastases. All the patients underwent mastectomy with no adjuvant therapy, and they are currently alive and well.     

Use of sulfonated probes for in situ detection of amylase mRNA in formalin-fixed paraffin sections of human pancreas and submaxillary gland

A sulfonated probe and its applicability to in situ hybridization is described and discussed. The DNA probes were modified by introducing an antigenic sulfone group into the cytidine residues of the denatured DNA (Budowsky EI, Sverdlov ED, Monastyrkaya GS: New method of selective and rapid modification of the cytidine residues. FEBS Lett 25:201, 1972). Hybridization of the sulfonated DNA with the target nucleic acid sequences was confirmed by an avidin-biotin peroxidase complex method using a monoclonal antibody specific to sulfonated DNA. The detection limit of this system was estimated to be about 1.25 pg of actual target sequences by dot blot hybridization analysis. When the sulfonated probes of human amylase cDNA were applied to in situ hybridization immunohistochemistry on formalin-fixed paraffin sections of the human pancreas and submaxillary gland, hybridization signals were clearly localized in the cytoplasm of the acinar cells of the pancreas, and in the serous cells of the submaxillary gland. Suitable probe lengths for in situ hybridization immunohistochemistry were between 100 and 800 bases. The in situ hybridization technique utilizing a sulfonated DNA probe is sensitive, simple, and easy to perform and applicable to studies of cell biology.     

Quantitative determination of dihydroetorphine in rat plasma and brain by liquid chromatography-tandem mass spectrometry

The extraordinarily strong analgesic dihydroetorphine (DHE) was registered as one of the most strictly controlled narcotic drugs by the United Nations in 1999. However, an effective detection method for DHE in biological samples has not yet been established. We developed a quantitative method for assay of DHE in rat plasma and brain by liquid chromatography-tandem mass spectrometry equipped with an ionspray interface. A 0.5-ml volume of plasma and brain homogenate spiked with buprenorphine (internal standard) was purified by the solid-phase extraction column Bond Elute Certify. DHE produced numerous weak fragment ions by collision induced dissociation. Therefore, collision energy was utilized to decompose the interferences, and the protonated molecular ion was used for both precursor and product ion monitoring. As a result of the method validation, the dynamic concentration range was determined as 0.05-10 ng/ml. DHE in these samples was stable for 2 months at -4 degrees C and for 24 h at ambient temperatures. Using the present method, DHE was detected in rat plasma and brain tissue after intravenous injection (0.5 microg/kg).