Anomalous upper critical field in CeCoIn5/YbCoIn5 superlattices with a Rashba-type heavy Fermion interface.

We report a highly unusual angular variation of the upper critical field (H(c2)) in epitaxial superlattices CeCoIn(5)(n)/YbCoIn(5)(5), formed by alternating layers of n and a 5 unit-cell thick heavy-fermion superconductor CeCoIn(5) with a strong Pauli effect and normal metal YbCoIn(5), respectively. For the n=3 superlattice, H(c2)(θ) changes smoothly as a function of the field angle θ. However, close to the superconducting transition temperature, H(c2)(θ) exhibits a cusp near the parallel field (θ=0°). This cusp behavior disappears for n=4 and 5 superlattices. This sudden disappearance suggests the relative dominance of the orbital depairing effect in the n=3 superlattice, which may be due to the suppression of the Pauli effect in a system with local inversion symmetry breaking. Taking into account the temperature dependence of H(c2)(θ) as well, our results suggest that some exotic superconducting states, including a helical superconducting state, might be realized at high magnetic fields.

CDK4 and cyclin D1 allow human myogenic cells to recapture growth property without compromising differentiation potential.

In vitro culture systems of human myogenic cells contribute greatly to elucidation of the molecular mechanisms underlying terminal myogenic differentiation and symptoms of neuromuscular diseases. However, human myogenic cells have limited ability to proliferate in culture. We have established an improved immortalization protocol for human myogenic cells derived from healthy and diseased muscles; constitutive expression of mutated cyclin-dependent kinase 4, cyclin D1 and telomerase immortalized human myogenic cells. Normal diploid chromosomes were preserved after immortalization. The immortalized human myogenic cells divided as rapidly as primary human myogenic cells during the early passages, and underwent myogenic, osteogenic and adipogenic differentiation under appropriate culture conditions. The immortalized cells contributed to muscle differentiation upon xenotransplantation to immunodeficient mice under conditions of regeneration following muscle injury. We also succeeded in immortalizing cryopreserved human myogenic cells derived from Leigh disease patients following primary culture. Forced expression of the three genes shortened their cell cycle to < 30 h, which is similar to the doubling time of primary cultured human myogenic cells during early passages. The immortalization protocol described here allowed human myogenic cells to recapture high proliferation activity without compromising their differentiation potential and normal diploidy.

A clinical study of patients with pemphigus vulgaris and pemphigus foliaceous: an 11-year retrospective study (1996-2006).

Only a few reports have been published of detailed clinical studies of pemphigus in Japan. The aim of this study was to determine the clinical characteristics of patients with pemphigus vulgaris (PV) and pemphigus foliaceous (PF), who were newly diagnosed in the dermatology department of Kurume University Hospital, Japan, over the past 11 years. The primary site of involvement was the oral mucosa in 21 patients (75%) with PV. At the initial visit, most of the patients with PV had moderate to severe disease. With regard to management, systemic corticosteroids were the mainstay of treatment for patients with PV, and plasmapheresis was the most frequently used adjuvant therapy. Dapsone was the mainstay of treatment for the patients with PF. The patients were investigated for any association with an underlying malignancy; in patients with PV, lung, stomach and uterine cancers (one patient each) were seen.

Hormonal regulation of the transformation phenotype in simian virus 40-transformed rat embryonic preadipose cell lines.

The regulation of transformed phenotypes was studied in newly isolated preadipose cell lines which were established after infection with simian virus 40 tsA58 dl2009. The clonal cell lines isolated exhibited most of the characteristics typical of transformed cells. The transformants, however, were able to differentiate into adipocytes in the presence of low calf serum (0.5%) and a combination of several hormones, including hydrocortisone and insulin. Treatment with insulin alone stimulated the growth of these cells but did not induce lipid accumulation without added hydrocortisone. The effect of hydrocortisone was accompanied by a restoration of growth control in the transformants after they reached high cell density. The blot hybridization analysis of cellular DNAs digested by restriction enzymes revealed that simian virus 40 genomes were integrated at multiple separate sites at which a head-to-tail oligomeric insertion took place. Large T antigen was synthesized in growing cells but was regulated at high cell density when cells were committed to differentiate by glucocorticoids. These results suggest that the glucocorticoid hydrocortisone is capable of restoring growth regulation at high cell densities to simian virus 40-transformed preadipose cell lines.

Differential early viral gene expression in two stages of human papillomavirus type 16 DNA-induced malignant transformation.

Human papillomavirus (HPV) type 16 DNA induces progressive transformation in NIH 3T3 cells. Two types of cell lines, PM3T3G0 and PM3T3Fo, were isolated by G418 or focus selection, respectively, after transfection of cells by a recombinant HPV 16 DNA carrying the neo gene. These cell lines exhibited distinct phenotypes compared with controls. Saturation densities of PM3T3G0 and PM3T3Fo lines were two- to three- and five- to sevenfold greater than that of control NIH 3T3 cells, respectively. Neither cell type required high serum for growth, in contrast to NIH 3T3 cells. PM3T3G0 lines were premalignant, whereas PM3T3Fo lines manifested tumorigenicity within 2 weeks. Subpopulations of three PM3T3G0 lines underwent progressive transformation as reflected by focus formation. Analysis of HPV 16-specific mRNA species demonstrated that high levels of early and late gene expression were detected in premalignant PM3T3G0 lines, whereas relatively low quantities of selected gene messages were expressed in malignant transformants. Thus, high levels of viral gene expression are not crucial for malignant transformation.

Calyculin A, an inhibitor of protein phosphatases, a potent tumor promoter on CD-1 mouse skin.

Calyculin A, isolated from a marine sponge, has a novel spiro ketal skeleton. Structurally unrelated to okadaic acid, calyculin A bound to the okadaic acid receptors in particulate and soluble fractions of mouse skin. The biochemical and tumor-promoting activities of calyculin A were studied with those of okadaic acid. Calyculin A inhibited the activity of protein phosphatases, which serve as the okadaic acid receptors. The effective dose of calyculin A for 50% inhibition was 0.3 nM, similar to that of okadaic acid. Like okadaic acid, calyculin A induced ornithine decarboxylase in mouse skin and hyperphosphorylation of a Mr 60,000 protein in human papilloma virus type 16-transformed human keratinocytes. A two-stage carcinogenesis experiment on mouse skin, initiated by 100 micrograms (390 nmol) of 7,12-dimethylbenz(a)anthracene and followed by 1 microgram (1.0 nmol) of calyculin A, revealed that calyculin A is an additional member of the okadaic acid class of tumor promoters. The percentages of tumor-bearing mice in the groups treated with DMBA plus calyculin A, and with DMBA followed by 1 microgram (1.2 nmol) of okadaic acid were 86.7 and 80.0%, respectively, in week 30. The mechanisms of action of calyculin A and okadaic acid, in addition to dinophysistoxin-1 (35-methylokadaic acid), are discussed. Calyculin A is the first tumor promoter to be screened by the okadaic acid receptor binding test.

Telomerase activity in a specific cell subset co-expressing integrinbeta1/EGFR but not p75NGFR/bcl2/integrin beta4 in normal human epithelial cells.

Telomerase activity is correlated with the immortality of various cultured cells and cancer cells. The activity, however, is also demonstrated in various normal regenerating cells which normally have a finite life span in vivo and in vitro, though its biological implication remains unclear. Using cultured normal human epithelial cells, we show that telomerase activity is associated with epithelial cell subsets which actively proliferate in vitro. Unlike in most cancer cell lines, telomerase activity was evidently up-regulated when the cells entered into S phase in primary human epithelial cells. To characterize the cells which have telomerase activity, the primary human epithelial cell population of uterine cervix was dissociated into several distinctive cellular subsets by means of immunocytochemical cell fractionation. Telomerase activity was found to be closely associated with the subset which expressed predominantly integrin beta1 and epidermal growth factor receptor. We further identified the telomerase-negative subpopulation which contained a small subset that strongly coexpresses p75NGFR low affinity nerve growth factor receptor, integrin beta4 and bcl-2 protein. The location of the p75NGFR-expressing cells contrasts to that of the Ki-67 positive cells in vivo and is distinctive of telomerase positive cycling cells, indicating that these cells remain at the G0 phase. The present study supports the notion that telomerase activity is linked to cell cycle regulation, implying that telomerase is activated upon cell proliferation in regenerating normal human epithelial cells.

Telomerase activity in normal human epithelial cells.

Telomerase activity is found in most cancer tissues and many immortalized cell lines as well as in germ line cells but it is generally undetected in normal human somatic tissues. There is weak telomerase activity in some cell types of hematopoietic lineage in which a stem cell-like subpopulation may exist. Likewise, physiologically regenerating somatic tissues and organs such as skin, small intestine, and most other epithelia of the human body are supposed to contain similar cell lineages to maintain their renewal throughout the life span of individuals. It is therefore of interest whether telomerase activity is present in physiologically regenerating epithelial cells. Telomerase activity was detected, though very weakly, in cultured normal epidermal keratinocytes and at higher levels in a subpopulation that adhere rapidly on collagen IV-coated culture dishes. No telomerase activity was detected in a subpopulation that was less adherent on the coated dishes. The rapidly adherent subpopulation of keratinocytes was enriched in small proliferating cells with macrocolony forming potential. It was also passaged through more generations in culture, and expressed integrin beta 1 at higher levels than the less adherent subpopulation. Telomerase activity was similarly found in ectocervical keratinocytes as well as in simple endocervical epithelial cells. These findings provide the evidence of a telomerase-positive population among physiologically regenerating normal human epithelial cells. The identity of the telomerase-positive cells remains to be defined.

Ultraviolet-B irradiation alters cytokine production by immune lymphocytes in herpes simplex virus-infected mice.

Previous studies from our laboratories have shown that UV-B irradiation at the site of an intradermal infection of herpes simplex virus (HSV) resulted in a higher incidence of zosteriform lesions and suppressed cellular immune responses to HSV in mice. In order to determine whether the production of T-cell-derived cytokine (IFN-gamma, IL-2 and IL-4) by immune cells from irradiated mice is also suppressed, we examined the production of cytokines by lymph node cells and spleen cells taken from UV-B irradiated, HSV type 1 (HSV-1)-infected mice. UV-B irradiation (120 mJ/cm2) prior to HSV-1 infection was found to markedly suppress IFN-gamma production compared to that of the non-irradiated control. IL-4 production was enhanced compared to IL-2 in the UV-B irradiated mice. These results suggest that alteration(s) in the cytokine production profile may therefore be involved in the development of severe skin lesions caused by HSV infection in UV-B irradiated mice.

The proliferative properties of tumor cells differentially correlate with the host immune responses in anogenital Bowen's disease.

Bowen's disease is a squamous cell carcinoma in situ that rarely invades into the underlying dermis. In order to evaluate the relationship between the cytological properties of the tumor cells and the host immune response, we have examined the expression of p53 and proliferating cell nuclear antigen (PCNA), and the number of mitotic cells, clumping cells, koilocytes, Langerhans cells (LCs) and dermal lymphoid cell infiltration in 18 cases of anogenital Bowen's disease. When compared with normal anogenital skins (n = 10), a statistically significant number of p53-positive cells, PCNA-positive cells, mitotic cells, clumping cells, koilocytes and dermal lymphoid cells was observed in the cases of Bowen's disease. Importantly, there existed a very strong correlation between the number of PCNA-positive tumor cells and the number of infiltrated dermal lymphoid cells. Moreover, the number of mitotic cells significantly correlated with the number of intratumoral LCs. The in situ hybridization technique for human papilloma virus (HPV) demonstrated that the HPV-infected Bowen's disease showed a similar histological and immunohistological pattern as the HPV-non-infected counterparts, except for increased koilocyte formation and decreased p53 positivity. The present data suggest that the proliferative activity of Bowen's disease significantly correlates with the host immune reaction, and that the host immune system may differentially recognize the different cytological properties of tumor cells in the Bowen's disease.

Differentiation of the rat myelomonocytic leukemia cell line c-WRT-7 by in vitro culture with the rat bone marrow preadipocyte cell line REC A16.

Differentiation of the rat myelomonocytic leukemia cell line (c-WRT-7) was investigated, by co-culture with a rat embryonic bone marrow preadipose cell line (REC A16). Co-cultivation with REC A16, or with conditioned medium from REC A16 cultures (REC-CM), induced differentiation of c-WRT-7 cells to macrophages. A soluble factor(s) produced by REC A16 appeared to be responsible for the differentiation of c-WRT-7. Because REC-CM was associated with colony-stimulating activity on murine marrow progenitors, c-WRT-7 cells were cultured with various colony-stimulating factors (CSF) and it was found that macrophage CSF (M-CSF) significantly induced differentiation of c-WRT-7. We further demonstrated that both the colony-stimulating and differentiation-inducing activities of REC-CM were significantly blocked by anti-M-CSF antiserum. These results suggest that the differentiation of c-WRT-7 is due to M-CSF produced by REC A16. Co-culture of these two cell lines should provide a useful model to study the mechanisms of interaction between leukemia cells and marrow stroma.

A case of hemiplegic migraine in childhood: transient unilateral hyperperfusion revealed by perfusion MR imaging and MR angiography.

We report on an 8-year-old girl with a typical attack of hemiplegic migraine, in whom MR angiography and perfusion MR imaging revealed unilateral dilation of branches of both the middle and posterior cerebral arteries and hyperperfusion of the ipsilateral hemisphere, respectively. The findings resolved spontaneously after the attack. These imaging techniques should be indicated for patients with migraine attacks and may play a role in assessing the vascular events in migraine headache.

Growth suppression and astrocytic differentiation of glioma cells by interleukin-1.

The effect of recombinant human interleukin-1 (rHuIL-1) derivatives on human glioma cell lines was examined in vitro. Five glioma cell lines, U-251 MG, U-373 MG, U-87 MG, A-172, and T98G, were incubated in medium containing 1% fetal calf serum and various concentrations of different type of rHuIL-1: OCT-43 (rHuIL-1 beta), OCT-7000 (rHuIL-1 alpha), and OCT-8000 (rHuIL-1 alpha). The high-affinity IL-1 receptors were expressed in the U-251 MG and U-373 MG cell lines, and rHuIL-1 was found to suppress cell growth and to induce morphological differentiation of these cell lines. Growth inhibition occurred in a dose-dependent manner in concentrations or rHuIL-1 ranging between 1 and 100 ng/ml. Interestingly, rHuIL-1 induced a transient growth of glioma cells shortly after administration, then suppressed cell growth with accompanying elongation of cytoplasmic processes. This unique process of transient growth stimulation followed by growth suppression was parallel to the efficacy of bromodeoxyuridine uptake in the rHuIL-1-treated cells. Concomitantly, accumulation of glial fibrillary acidic protein and cyclic adenosine monophosphate contents was observed in four glioma cell lines. Continuous rHuIL-1 treatment for longer than 30 days elicited irreversible astrocytic terminal differentiation. These results indicate that IL-1 is an effector on the growth regulation of glioma cells, resulting in astrocytic differentiation in vitro.

Gene expression of telomerase related proteins in human normal oral and ectocervical epithelial cells.

We analyzed telomerase activities and gene expressions of telomerase components: hTERT, hTR, hTEP1, telomeric repeat binding factors: TRF1, TRF2, and c-myc, Max and Mad in human normal oral and ectocervical epithelial keratinocytes, comparing with those of squamous carcinoma cells and HPV16- or SV40-immortalized cells. Significant telomerase activity and hTERT expression were detected in primary keratinocytes. However, both were dramatically down-regulated during serial passages. The down-regulation of hTERT mRNA was associated with augmented expression of TRF1. Expression of c-myc was slightly decreased, whereas Mad was expressed in parallel with that of hTERT during passages. We also detected an alternate splicing of hTERT transcript in two of four cancer cells and normal aged epithelial cells. These results suggest that the senescence of normal oral and ectocervical keratinocytes is accompanied with up-regulation of TRF1 and down-regulation of telomerase activity due to transcriptional suppression of active form of hTERT in vitro.

Japanese monozygotic twins with Rett syndrome.

We present a study of 28-year-old Japanese monozygotic female twins with Rett syndrome (RS). To our knowledge, this is the first report of monozygotic twins with RS from Japanese family. There are some differences between twins about seizures, scoliosis and stereotypical hand movements during adolescence. Monozygosity was confirmed by both blood typing and HLA titers.

The effectiveness of clonazepam on the Rolandic discharges.

Rolandic discharge (RD), noted in the electroencephalography (EEG) of patients with benign epilepsy in childhood with centrotemporal spikes (BECCT) has several unique features. One feature is that the amount or frequency of RDs does not correlate well with the incidence of seizures in BECCT although it is a key finding in the diagnosis of this epileptic syndrome. In this study, we examined the efficacy of antiepileptic drugs focusing on the disappearance of RDs in relationship with seizure control. Forty patients with BECCT who were not medically treated prior to this study were randomly sorted into three groups. Twenty patients were assigned for clonazepam (CZP) treatment, 10 patients for valproate (VPA) and the remaining 10 patients for carbamazepine (CBZ). Each drug was administered for 4 consecutive weeks. EEGs were recorded twice during the study, before and 4 weeks after the medication trial. The effects of each treatment on RDs were assessed. RDs disappeared in 15 of the 20 cases treated with CZP (75%) within 4 weeks while the same was observed in only one of the 10 cases treated with VPA (10%). CBZ failed to demonstrate any effect on RD. In the group treated with CZP, there were no differences in seizure incidence, seizure type and blood concentration of CZP between the patients whose RDs disappeared and those whose RDs remained.