Isolation, characterization, and disruption of dnr1, the areA/nit-2-like nitrogen regulatory gene of the zoophilic dermatophyte, Microsporum canis.

A homolog of the major nitrogen regulatory genes areA from Aspergillus nidulans and nit-2 from Neurospora crassa was isolated from the zoophilic dermatophyte, Microsporum canis. This gene, dnr1, encodes a polypeptide of 761 amino acid residues containing a single zinc-finger DNA-binding domain, which is almost identical in amino acid sequence to the zinc-finger domains of AREA and NIT-2. The functional equivalence of dnr1 to areA was demonstrated by complementation of an areA loss-of-function mutant of A. nidulans with dnr1 cDNA. To further characterize this gene, dnr1 was disrupted by gene replacement based on homologous recombination. Of 100 transformants analyzed, two showed the results expected for replacement of dnr1. The growth properties of the two dnr1(-) mutant strains on various nitrogen sources were examined. Unlike the A. nidulansareA(-) mutant, these dnr1(-) mutants showed significantly reduced growth on ammonia, a preferred nitrogen source for fungi. These mutant strains were also able to utilize various amino acids for growth. In comparison with wild-type M. canis, the two dnr1(-) mutants showed reduced growth on medium containing keratin as the sole nitrogen source. This is the first report describing successful production of targeted gene-disrupted mutants by homologous recombination and their phenotypic analysis in dermatophytes.     

A Comparative Analysis of the Murine Thymic Microenvironment in Normal,Autoimmune, and Immunodeficiency States

It is widely accepted that the thymic microenvironment regulates normal thymopoiesis through a highly coordinated and complex series of cellular and cytokine interactions. A direct corollary of this is that abnormalities within the microenvironment could be of etiologic significance in T-cell-based diseases. Our laboratory has developed a large panel of monoclonal antibodies (mAbs) that react specifically with epithelial or nonepithelial markers in the thymus. We have taken advantage of these reagents to characterize the thymic microenvironment of several genetic strains of mice, including BALB/cJ, C57BL/6J, NZB/BlnJ, SM/J, NOD/Ltz, NOD/Ltz-scid/sz, C57BL/6J-Hcph m^e/Hcph m^e, and ALY/NscJcl-aly/aly mice, and littermate control animals. We report herein that control mice, including strains of several backgrounds, have a very consistent phenotypic profile with this panel of monoclonal antibodies, including reactivity with thymic epithelial cells in the cortex, the medulla and the corticomedullary junction, and the extracellular matrix. In contrast, the disease-prone strains studied have unique, abnormal staining of thymic cortex and medulla at both the structural and cellular levels. These phenotypic data suggest that abnormalities in interactions between developing thymocytes and stromal cells characterize disease-prone mice.     

Molecular identification of a novel human rotavirus in relation to subgroup and electropherotype of genomic RNA

A total of 41 stool rotavirus specimens collected from children with acute diarrhea at four different locations in Akita Prefecture, Japan, during the peak of the winter diarrhea epidemic in 1988 were analyzed by polyacrylamide gel electrophoresis of viral RNA in conjunction with subgrouping assay. We found that a single strain predominated, with cocirculating strains with less common electropherotypes at a given location, and that two different strains could predominate at geographically close but different locations even during a very limited time of the epidemic season. Furthermore, we isolated a human rotavirus strain (AU125) that was similar to the AU-1 strain in that it possessed a long RNA pattern yet belonged to subgroup I. Genetic analysis by RNA-RNA hybridization assay indicated that the AU125 strain was distinct from two previously identified human rotavirus gene groups (genogroups) represented by the Wa strain (subgroup II with long RNA electropherotype) and the DS-1 strain (subgroup I with short RNA electropherotype), but was very closely related to the AU-1 strain. These data suggest that the genetic diversity of human rotaviruses may be more extensive than was previously thought.     

A case of a placental site trophoblastic tumor

A rare placental site trophoblastic tumor (PSTT) in a 39-year-old female was studied. This tumor, protruding into the uterine cavity, was histologically similar to tumors in previously reported cases of PSTT. Ultrastructurally, the characteristic finding was the presence of perinuclear filaments. Also, the tumor cells were strongly positive for hPL by immunohistochemical method. These findings suggest that this was a tumor caused by neoplastic proliferation of the extravillous intermediate trophoblast.     

Functional analysis of the effect of forced activation of STAT3 on M1 mouse leukemia cells

M1 mouse myeloid leukemia cells exhibit growth arrest and differentiation to monocytes/macrophages in response to leukemia inhibitory factor (LIF) stimulation. Although recent studies have demonstrated that STAT3 plays a central role in this process, it is unknown whether STAT3 activation alone is sufficient. To address this issue, we have established M1/STAT3ER cells, where STAT3 is selectively activated by 4-hydroxytamoxifen (4HT). 4HT stimulation did not have any effect on growth and morphology of M1/ STAT3ER cells, and did not induce the down-regulation of mRNA of c-myc and c-myb, which is necessary for M1 cell differentiation. On the other hand, mRNA of jun-B, IRF1 and p19 was increased by 4HT. DNA precipitation assay indicated that both stimulation of LIF and 4HT similarly activated STAT3ER. Introduction of a constitutive active MAP kinase kinase (MEK1) into M1/STAT3ER cells did not induce differentiation either. Together, our present data suggest that signaling other than the activation of STAT3 and MEK1 may be necessary for M1 cell-growth arrest and differentiation, while a set of early genes of LIF are induced by only STAT3 activation.     

A "floral" variant of nodal marginal zone lymphoma

We describe 6 cases of a specific variant of nodal marginal zone lymphoma with "floral" lymph follicles in patients ranging in age from 18 to 66 years. All 6 patients had lymphadenopathy, either local (n = 5) or systemic (n = 1), and good performance status (0), and none had fever, weight loss, or night sweating. They all underwent excisional biopsy. Histologically, all lesions had a distinctive morphology, with proliferation of medium-sized atypical lymphoid cells in the marginal zone, hyperplastic lymph follicles with enlarged germinal centers, and a thickened mantle zone. In places, folliculolysis was observed. On immunohistochemical staining, the atypical lymphoid cells showed a B-cell phenotype (CD20 +), IgM positivity in 2 of 5 cases, and negativity for CD5, CD10, CD23, CD43, bcl-6, and IgD. Polymerase chain reaction examination for immunoglobulin heavy chain in 5 cases showed monoclonality in all. Five patients did not receive adjuvant chemotherapy and had no recurrences. The patient with systemic lymphadenopathy received chemotherapy and had a complete response without relapse. This variant should be differentiated from the usual nodal marginal zone lymphoma because of its specific clinical and pathological features.     

Effects of body position on snoring in apneic and nonapneic snorers

STUDY OBJECTIVES: The positional dependency of obstructive sleep apnea (OSA) is well known, but objective evidence for the positional effect on snoring is lacking. The aim of this study is to elucidate the effect of body position on snoring, and that of sleep stage as well. DESIGN: Retrospective analysis of the effects of body position and sleep stage on snoring in nonapneic snorers (snorer group) and OSA patients (apneic group). SETTING: A sleep laboratory in a national hospital in Japan. PATIENTS: Seventy-two patients who complained of habitual snoring and underwent overnight polysomnography. INTERVENTIONS: N/A MEASUREMENTS AND RESULTS: In the lateral position, most subjects in the snorer group showed decreased snoring both in time (p = 0.0004) and intensity (p = 0.0003), but subjects in the apneic group showed variable changes. In the apneic group, the positional dependency of snoring (the ratio of lateral value to supine value) was correlated with supine apnea-hypopnea index (AHI), that is, OSA patients with higher supine AHI tended to show increased snoring in the lateral position. AS to the effect of sleep stage, snoring was increased in deeper non-rapid eye movement sleep and decreased in rapid eye movement sleep in a given position. CONCLUSIONS: This study demonstrated that the positional dependency is different between nonapneic snorers and OSA patients. Most of the nonapneic snorers snore less in the lateral position than in the supine position in contrast to OSA patients who often fail to decrease snoring even in the lateral position.     

A model for the embryological instruction of the branchial arteries

It is difficult to teach students about the embryological transformation of the branchial arteries. In mammals, six pairs of branchial arteries develop, but all are not present at the same embryological stage. The first, second, fifth and a part of right sixth branchial artery disappear early in embryological development. The remaining third, fourth and sixth branchial arteries mainly constitute the arterial system of the breast. The aorta and pulmonary trunk are derived from the truncus arteriosus. Because of the complexity of this developmental process, we have devised a user-friendly model in order to assist with educational presentations. In this model, the shrinkage of a vessel has been represented by inserting a wire inside of the hose representing the artery. Degenerated or disappearing parts of the vessel are removable by using hooks and Velcro tape. Branchial arteries, truncus arteriosus, aortic sac and dorsal arteries are represented by different colors. The descent of the heart is represented by the relational change between larynx and heart. Additionally we represented the vagus nerve and recurrent laryngeal nerve by using strings. The right vagus nerve can move dorsally and the left ventrally by rotating the digestive tract. The right recurrent laryngeal nerve can move superiorly to hook around the right subclavian artery, and left recurrent laryngeal nerve can hook around the ductus arteriosus formed by the left sixth branchial artery.