Microwell bag culture for large-scale production of homogeneous islet-like clusters.

Pluripotent stem-cell derived cells can be used for type I diabetes treatment, but we require at least 10-10 islet-like clusters per patient. Although thousands of uniform cell clusters can be produced using a conventional microwell plate, numerous obstacles need to be overcome for its clinical use. In this study, we aimed to develop a novel bag culture method for the production of uniform cell clusters on a large scale (10-10 clusters).

Lateral access mechanism of LPA receptor probed by molecular dynamics simulation.

G-protein-coupled receptors (GPCR) are a family of membrane receptors that play important roles in the regulation of various physiological phenomena. LPA receptors (LPA1-6) are members of the class A GPCRs, which transduce a lysophosphatidic acid (LPA) signal across the cell membrane and evoke various responses, including cellular survival, proliferation, differentiation, and migration. The crystal structure of LPA6 revealed a gap between its transmembrane helices (TMs), which is opened toward the membrane side.

Cost-effectiveness of a "treat-all" strategy using Direct-Acting Antivirals (DAAs) for Japanese patients with chronic hepatitis C genotype 1 at different fibrosis stages.

Aim: To evaluate the cost-effectiveness of therapeutic strategies initiated at different stages of liver fibrosis using three direct-acting antivirals (DAAs), sofosbuvir-ledipasvir (SL), glecaprevir-pibrentasvir (GP), and elbasvir plus grazoprevir (E/G), for Japanese patients with chronic hepatitis C (CHC) genotype 1.

Methods: We created an analytical decision model reflecting the progression of liver fibrosis stages to evaluate the cost-effectiveness of alternative therapeutic strategies applied at different fibrosis stages. We compared six treatment strategies: treating all patients regardless of fibrosis stage (TA), treating individual patients with one of four treatments starting at four respective stages of liver fibrosis progression (F1S: withholding treatment at stage F0 and starting treatment from stage F1 or higher, and three successive options, F2S, F3S, and F4S), and administering no antiviral treatment (NoRx).

Bilateral lesions of the mesencephalic trigeminal sensory nucleus stimulate hippocampal neurogenesis but lead to severe deficits in spatial memory resetting.

The mesencephalic trigeminal sensory nucleus (Me5), which receives signals originating from oral proprioceptors, becomes active at weaning and contributes to the acquisition of active exploratory behavior [Ishii, T., Furuoka, H., Kitamura, N.

Central L-arginine reduced stress responses are mediated by L-ornithine in neonatal chicks.

Recently, we observed that central administration of L-arginine attenuated stress responses in neonatal chicks, but the contribution of nitric oxide (NO) to this response was minimal. The sedative and hypnotic effects of L-arginine may be due to L-arginine itself and/or its metabolites, excluding NO. To clarify the mechanism, the effect of intracerebroventricular (i.

Intracerebroventricular injection of L-arginine induces sedative and hypnotic effects under an acute stress in neonatal chicks.

L-arginine participates in many important and diverse biochemical reactions associated with the normal physiology of the organism. In the present study, we investigated the effect of central administration of L-arginine on the stress response and its mechanism in neonatal chicks. Intracerebroventricular (i.

Relationships between the sedative and hypnotic effects of intracerebroventricular administration of L-serine and its metabolites, pyruvate and the derivative amino acids contents in the neonatal chicks under acute stressful conditions.

Intracerebroventricular (i.c.v.

Intracerebroventricular injection of glutathione and its derivative induces sedative and hypnotic effects under an acute stress in neonatal chicks.

Glutathione-related enzymes glyoxalase 1 and glutathione reductase 1 regulates anxiety in mice. To clarify the central function of glutathione as a neurotransmitter in the stress reaction, the effect of intracerebroventricular (i.c.

Behavioral analyses of wind-evoked escape of the cricket, Gryllodes sigillatus.

The wind-evoked escape behavior of the cricket Gryllodes sigillatus was investigated using an air puff stimulus. A high velocity air puff elicited the escape behavior in many crickets. The crickets tended to escape away from the stimulus source, but the direction was not accurately oriented 180 degrees from the stimulus.

In vitro-activated human lupus T cells express normal estrogen receptor proteins which bind to the estrogen response element.

We have shown that estrogen receptor (ERalpha, ERbeta) transcripts are expressed in SLE and normal T cells. In this study, T cell nuclear extracts from female lupus patients and normal donors were tested for biologically active ER proteins capable of binding to the human estrogen response element (hERE) by electrophoretic mobility shift assays. When peripheral blood T cells were stimulated with 17beta-estradiol (E2), PMA and ionomycin, two major retarded bands in T cell nuclear extracts exhibited a migration pattern similar to slow migrating protein-ERE complexes in human breast cancer cell extracts.

Gender differences in autoimmune diseases: estrogen increases calcineurin expression in systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) predominantly affects women (9:1 compared to men) of childbearing age and often decreases its intensity in postmenopausal women, suggesting that sex hormones play a role in its pathogenesis. Comparison of steady-state levels of calcineurin mRNA using RNase protection assays revealed increased calcineurin expression in response to estradiol in cultured T cells from nine female lupus patients. Calcineurin mRNA levels did not increase significantly in T cells from eight age-matched normal control female volunteers.

Isolation of anti-nucleosome antibodies from the plasma of lupus nephritis patients.

Anti-nucleosome antibodies, which recognise conformational epitopes consisting of histone and DNA in chromatin, have been described in autoimmune diseases. In this study, an attempt was made to isolate anti-nucleosome antibodies from the anti-DNA-depleted plasma IgG of two lupus patients either with or without nephritis by nucleohistone affinity chromatography. The purified nucleohistone-binding antibodies bound to nucleohistone in a specific manner and contained enriched anti-histone antibodies.

Peripheral blood T cells and monocytes and B cell lines derived from patients with lupus express estrogen receptor transcripts similar to those of normal cells.

Objective: To identify and characterize estrogen receptor (ER) transcripts expressed in immune cells of patients with systemic lupus erythematosus (SLE) and healthy donors.

Methods: Peripheral blood monocytes and T cells were prepared from patients with SLE (n = 6) and healthy donors (n = 8). T cells were separated into CD4 and CD8.

V gene sequences of lupus-derived human IgM anti-ssDNA antibody: implication for the importance of the location of DNA-binding amino acids.

Binding and structural characteristics of human IgMk anti-ssDNA antibody 7B3 were determined. 7B3 was derived from Epstein-Barr virus-transformed peripheral blood B cells of a lupus nephritis patient. Purified 7B3 bound ssDNA from various species, but not dsDNA or structurally unrelated antigens.

Binding affinity and quantity of estrogen receptor in peripheral blood monocytes of patients with systemic lupus erythematosus.

Binding affinity and quantity of the estrogen receptor in monocytes of patients with systemic lupus erythematosus (SLE) were studied. The tritiated-estradiol binding assay was performed using peripheral blood adherent cells (> 95% monocytes) derived from six lupus patients (SLEDAI score: 2-30) and five age-comparable normal women during the mid-follicular phase of the menstrual cycle. Dissociation constant (Kd) and number of binding sites (Ro) were estimated by Scatchard analysis.

Anti-(DNA-histone) antibodies in active lupus nephritis.

Objective: To detect and characterize anti-(DNA-histone) antibodies in patients with active lupus nephritis.

Methods: Calf thymus double stranded DNA was reassociated with histone in vitro. Polynucleosomes were prepared from chicken erythrocyte nuclei, calf thymus nucleohistone, and human peripheral blood mononuclear cells.

V gene sequences of human anti-ssDNA antibodies secreted by lupus-derived CD5-negative B cell hybridomas.

V gene sequences encoding two lupus-derived human monoclonal IgMk anti-ssDNA antibodies (2F7 and 1A6) and CD5 mRNA expression by the corresponding hybridomas were investigated. Both antibodies displayed V gene sequences nearly in germline configuration compared with their putative germline counterparts. It appeared that 2F7 used hv3019b9/HUD-3/JH6 and 12La/Jk2, while 1A6 utilized HHG19/D31-HUD-3/JH2 and Humkv328h5/Jk1.

Heparan sulphate-ELISA gives false positive results for anti-DNA-DNA/histone immune complexes in sera of patients with SLE.

Heparan sulphate-reactive antibodies in lupus sera have been suggested to be anti-DNA-DNA/histone immune complexes and to be associated with lupus nephritis. In this study, 23 anti-DNA-positive lupus sera including 13 active nephritis sera were tested for the presence of circulating anti-DNA-DNA/histone immune complexes by solid phase heparan sulphate-ELISA. Because of high background binding to protamine chloride-linked heparan sulphate plates, poly-L-lysine (PLL) was used as a linker and the remaining active sites of PLL were blocked with poly-L-glutamic acid.

Cationic and high affinity serum IgG anti-dsDNA antibodies in active lupus nephritis.

To investigate differences between cationic anti-dsDNA antibodies during active and inactive nephritis, low- and high-affinity IgG anti-dsDNA antibodies were prepared from sera of a lupus patient and compared for their binding affinity, spectrotype, and idiotype expression. The ratio of high-affinity to low-affinity anti-DNA antibodies and the relative avidity of the high-affinity anti-DNA antibodies decreased when active nephritis became inactive. Isoelectric focusing showed that cationic anti-dsDNA populations were present predominantly in the high-affinity fraction during active nephritis and in the low-affinity fraction during inactive nephritis.

Lupus-derived human monoclonal IgM anti-DNA antibody displays monospecificity, high affinity and private idiotype specificity.

A human monoclonal IgM k anti-DNA antibody, designated 2F7, was prepared by somatic hybridization of peripheral blood lymphocytes from a lupus patient with a human-mouse heterohybridoma cell line, K6H6/B5. 2F7 was tested for its antigen binding and idiotypic specificity by direct binding and inhibition enzyme-linked immunosorbent assays. 2F7 had a high binding activity to single-stranded DNA (ssDNA) but not to double-stranded DNA.

Expression of inactive stage anti-dsDNA idiotypes on anti-ssDNA antibodies in a lupus patient during active stage of lupus cerebritis.

The possibility that idiotypes (Ids) defined on anti-double stranded DNA (dsDNA) antibodies during active and inactive stages of lupus (1/84 Id and 4/90 Id, respectively) were expressed on anti-DNA antibodies during a subsequent active period (9/90) of the disease was investigated in a lupus patient with lupus cerebritis. Using rabbit (R)-anti-Ids specific to 1/84 Id and 4/90 Id in inhibition assays, the 4/90 Id was shown to be expressed on the framework regions of anti-single stranded DNA (ssDNA) but poorly on co-existing anti-dsDNA antibodies of active (9/90) stage. The 1/84 Id was poorly expressed on both types of 9/90 anti-DNA antibodies.

Shift of private and not of cross-reactive anti-DNA idiotypes in systemic lupus erythematosus.

The shift of private idiotype (Id) and cross-reactive Id (CRI) on anti-DNA antibodies in a lupus patient KE was investigated during a 7-year period. Anti-private Id and anti-CRI activities were separated by affinity chromatography from rabbit (R)-anti-Ids raised against KE anti-DNA antibodies during active (1/84) and inactive (4/90) stages of the disease. Anti-CRI isolated from the 84 R-anti-Id appeared to recognize binding site-related Ids that are shared with KE non-anti-DNA antibodies, unrelated lupus patients' sera, and certain normal sera.

Idiotypic and immunochemical differences of anti-DNA antibodies of a lupus patient during active and inactive disease.

IgG anti-DNA antibodies of a lupus patient during active and inactive stages of her disease were studied. There were no significant differences in the amounts, in double-stranded DNA-binding activity, or in complement-fixing ability between purified IgG anti-DNA antibodies of both stages. However, their idiotype (Id) expressions were different as revealed by binding to rabbit anti-Ids raised against each of the anti-DNA antibodies.

Detection and purification of antiidiotypic antibody against anti-DNA in intravenous immune globulin.

Pooled normal human IgG for therapeutic use, following depletion of anti-DNA, anti-Fc, and anti-F(ab')2 of normal IgG, expressed antiidiotypic activity against anti-DNA derived from lupus sera. The antiidiotype enriched by elution from anti-DNA affinity columns bound directly to anti-DNA IgG and inhibited the binding of lupus sera to DNA but did not bind to normal IgG or inhibit the binding of anti-tetanus toxoid to tetanus toxoid. Antiidiotypes in pooled normal sera may have a role in the clinical improvement seen in patients with autoimmune diseases receiving intravenous immune globulin.

Lack of correlation between HLA types and anti-idiotypic production in family members of a lupus patient.

Correlations of anti-single-stranded (ss) DNA, anti-F(ab')2, and anti-idiotypes to HLA types of 16 healthy family members of a lupus patient were studied. High levels of anti-ss DNA (63%) and anti-F(ab')2 (69%) were detected. Of the 12 family members who expressed HLA-DR2 antigen, 8 had anti-ss DNA and anti-F(ab')2 antibodies.