Selective elimination of double-positive immature thymocytes by a thymic epithelial cell line

A cloned epithelial cell line, TEL-2, has been established from the stroma tissues of normal mouse thymus. Incubation of mouse thymocytes on TEL-2 cells resulted in the selective elimination of double-positive (CD4+CD8+) cells from the culture, whereas single-positive (CD4+CD8- or CD4-CD8+) thymocytes remaining in the culture were concentrated in non-integrated cell population. The CD3- or CD3 low-positive thymocytes were also eliminated by the TEL-2 cells from the culture, followed by the concentration of CD3 high-positive cells in the culture. Only intact viable thymocytes were integrated into TEL-2 cells. Electron microscopic examination showed that the integrated cells into TEL-2 cytoplasm were gradually degenerated. Mature single-positive T cells, mature B cells or double-negative thymocytes were not integrated into TEL-2 cells. The TEL-2 cell may provide information on the mechanism of selective disappearance of double-positive immature cells from the thymus.     

An investigation of Japanese subjects maps susceptibility to type 1 (insulin-dependent) diabetes mellitus close to the DQA1 gene.

Insulin-dependent diabetic and control subjects of Japanese origin were HLA-DRB1, -DQB1, and -DQA1 typed using restriction fragment length polymorphism analysis and sequence-specific oligonucleotide gene probing. The DQA1 allele DQA1*0301 was positively associated with the disease [48/52 (92%) diabetic patients versus 44/64 (69%) control subjects, Pc less than 0.03, RR = 4.97]. Alleles of the DRB1 and DQB1 genes showed no significant association with the disease. The frequency of DQB1 genotypes encoding the amino acid aspartic acid at position 57 of the DQ beta chain did not differ significantly between subjects with insulin-dependent diabetes mellitus (IDDM) and controls. These findings suggest that a susceptibility allele for IDDM in the Japanese is more closely associated with the DQA1 gene than the DQB1 gene.     

EXFOLIATIVE CYTOPATHOLOGY OF ALVEOLAR RHABDOMYOSARCOMA

Exfoliative cytopathology of alveolar rhabdomyosarcoma is presented. A 34-year-old male was found to have alveolar rhabdomyosarcoma of the retroperitoneum at autopsy. Ascites was recognized in the course of this disease. Cytological features in the ascites were characterized by relatively small-sized ovoid cells observed singly or in loose clusters. The neoplastic cells possessed small amount of rather vesicular cytoplasm and an eccentric nucleus. The nucleus characteristically revealed multiple deep indentations in the nuclear membrane, a few prominent eosinophilic nucleoli and finely granular or evenly distributed chromatin pattern. These cytological findings, especially the nuclear features, might be useful to differentiate alveolar rhabdomyosarcoma from other neoplasms which consist of relatively small-sized ovoid cells even if the striated muscle origin cannot be confirmed by immunoperoxidase staining.     

Methods for culture ofBombyx mori wing discs

Summary A culture method forBombyx wing discs is described. Grace's insect medium,Bombyx hemolymph, and moderate doses of ecdysterone (0.02 to 0.1 µg/ml) were necessary for complete imaginal differentiation ofBombyx wing discs. This procedure is useful for understanding ecdysteroid action on insect imaginal differentiation and metamorphosis.     

The effect of exhaustive exercise on the antioxidant enzyme system in skeletal muscle from calcium-deficient rats

 The aim of the current study was to elucidate the synergism of dietary calcium restriction and exhaustive exercise in the antioxidant enzyme system of rat soleus muscle, and to investigate the involvement of neutrophils in exercise-induced muscle damage. Forty-eight male Wistar rats were assigned to the following groups: control (C) or calcium-restricted [1 month (1 M) or 3 months (3 M)]. Each group was subdivided into acutely exercised or non-exercised groups. Soleus muscle from each rat was analysed to determine the levels of antioxidant enzymes [Mn-superoxide dismutase (SOD), Cu,Zn-SOD, glutathione peroxidase (GPX), and catalase (CAT)]. Dietary calcium restriction resulted in calcium deficiency and upregulated the antioxidant enzymes examined except GPX. Conversely, exhaustive exercise significantly decreased GPX and CAT, but not SODs activities in the calcium-restricted (1 M and/or 3 M) rats. Contents of immunoreactive Mn-SOD and Cu,Zn-SOD were only increased in the 3 M rats. During calcium restriction, the mRNA expression of both forms of SOD showed initial upregulation, followed by downregulation. Exhaustive exercise significantly increased the mRNA expressions only in the 3 M rats. Moreover, exhaustive exercise markedly increased myeloperoxidase activity in soleus muscles from the 1 M and 3 M rats compared with the C rats, and significantly enhanced the ability of neutrophils to generate superoxide in the 3 M rats. The results demonstrate that dietary calcium restriction upregulates certain antioxidant enzyme activities in rat soleus muscle, indicating an enhanced resistance to potential increases in intracellular reactive oxygen species. The results also suggest that exhaustive exercise may cause oxidative damage in soleus muscle of calcium-deficient rats through the activation of neutrophils. Received: 4 August 1997 / Received after revision: 29 September 1997 / Accepted: 26 November 1997     

Neuropsin is essential for early processes of memory acquisition and Schaffer collateral long-term potentiation in adult mouse hippocampus in vivo

Long-term potentiation (LTP) is thought to be particularly important in the acquisition of hippocampus-associated memory, in part because it develops quickly and persists for indefinite periods. Extracellular proteolysis has been hypothesized to contribute to LTP by modifying adhesive relations of synapses and thus the morphology of excitatory synapses. Here we report that neuropsin (NP), an extracellular serine protease, is critically involved in the formation of both the potentiation effect and hippocampus-dependent forms of memory. NP-knockout mice were significantly impaired in the Morris water maze and Y-mazes and failed to exhibit early phase LTP induced by a single tetanus. Potentiation was also impaired or completely blocked by in vivo application of a specific inhibitor or a neutralizing monoclonal antibody for NP. Intriguingly, recombinant (r-) NP alone, without tetanic stimulation, elicited either long-lasting potentiation or depression, depending on the applied dose. The r-NP-elicited potentiation was occluded by prior induction of LTP, while theta-burst-elicited LTP was occluded by application of r-NP alone, suggesting that the two forms of plasticity have a common signalling pathway. r-NP-elicited potentiation and depression increased phosphorylation at different sites on the GluR1 subunit of the AMPA receptor that had previously been associated with LTP or long-term depression. Thus, we conclude that NP is necessary for establishment of LTP and has a significant role in memory acquisition.     

The numerical aberrations of chromosome 7 detected by fluorescence in situ hybridization in human breast cancers

The relationship between the numerical aberrations of chromosome 7 in interphase cells and the clinicopathological behavior of breast tumors was investigated in 51 touch imprinted preparations of breast tumors. Using fluorescence in situ hybridization with a chromosome 7-specific DNA probe, the fluoresceinisothiocyanate (FITC) spots mean and the representative copy number of each breast tumor were examined. The FITC spots mean (2.34) of 40 breast cancers increased compared with that of 11 benign lesions (1.98) (P < 0.02). The FITC spots mean tended to increase with the advancing stage and tumor size of the breast cancer. The FITC spots mean in the case with metastasis was also of a higher value than that without metastasis (P < 0.01). Furthermore, the existence of trisomy or over-trisomy of the copy number was related to the advancing stage and tumor size (P < 0.05 and P < 0.01, respectively). These findings suggest that the FITC spots mean and polysomy of the number of chromosome 7 may be highly predictive for breast tumor aggressiveness.     

Redistribution of fibroblasts and macrophages as micrometastases develop into established liver metastases

Fibroblastic tissue is an important component of malignant tumors, involved in the establishment of metastatic foci from micrometastases, and thought to prevent invasion of metastatic tumor cells into surrounding tissue. However, experimental models of fibrosis during the growth of micrometastasis into established metastases were not previously available. In the present paper, we performed immunohistochemical studies on experimental hepatic metastasis with colon 38 mouse colon carcinoma cells injected into syngeneic C57BL/6 mice. Early and late stages of metastatic nodules were examined for the distribution of endothelial cells, fibroblasts, and macrophages by the use of markers of these cells. One week after intrasplenic injection of colon 38 cells, micrometastases mainly appeared in the region of sinusoids accompanied with invasion of F4/80-positive Kupffer cells. Transitional metastases can be defined based on the histological appearance and intensive infiltration of both macrophages and fibroblasts. These transitional metastases were connected by protrusions of fibroblast-rich tissues co-localized with collagen-rich matrix and CD31-positive cells. This protrusion preceded fibrosis formation characteristics to established metastases associated with angiogenesis and segregation of tumor cells from host cells. Three stages can thus be classified during the development of hepatic metastasis in this syngeneic experimental system: micrometastasis, transitional metastasis, and established metastasis. This revised version was published online in July 2006 with corrections to the Cover Date.