Effects of ultraviolet B irradiation on cell–cell interaction; implication of morphological changes and actin filaments in irradiated cells

We studied the effects of ultraviolet B (UV-B) irradiation on cell–cell interactions using mouse lymphoma RMA cells and T cell hybridoma HTB-176.10 cells. RMA cells act as stimulators by presenting H-2K^b surface antigens to HTB-176.10 cells, inducing IL-2 production in HTB-176.10 cells. Irradiating RMA cells with 1000 J/m² UV-B suppressed cell cluster formation between RMA and HTB-176.10 cells and reduced the level of IL-2 production in HTB-176.10 cells, although H-2K^b surface antigens of RMA cells were still expressed. Electron microscopic observations of irradiated RMA cells revealed that UV-B irradiation damaged cell structures, resulting in the disappearance of microvilli on the cell surface, destruction of mitochondria, vacuolation of cytoplasm and swelling of the perinuclear cisterna space. We found that these alterations were accompanied by polymerization of filamentous actin quantified by flow cytometry after NBD-phallacidin staining. Our results suggest that a target of UV-B-induced alterations is actin filaments, which support the cell morphology as the cytoskeleton, and that modification of filamentous actin inhibits interaction between RMA and HTB-176.10 cells. This underlying mechanism may account for the impaired interaction between antigen-presenting cells and T cells after transfusion with UV-B-irradiated allogeneic blood components.     

Oral Tolerance Induced by Enterobacteria Altered the Process of Lymphocyte Recruitment to Intestinal Microvessels: Roles of Endothelial Cell Adhesion Molecules,TGF‐beta and Negative Regulators of TLR Signaling

Objective: Although enterobacteria are implicated in intestinal immune response, there has been no report on how intraluminal pathogens affect lymphocyte recruitment. The aim of this study was to determine how the presence of intestinal flora affects lymphocyte migration to intestine under physiological and lipopolysaccharide (LPS)‐induced inflammatory conditions. Methods: Interaction of T‐cells with ileal microvessels was monitored by using an intravital microscope in mice under germ‐free (GF) and specific pathogen‐free (SPF) conditions. LPS was administered into either the peritoneal cavity or duodenum before lymphocyte injection. Results: Adherence of T‐cells was greater in SPF than in GF mice, indicating that the presence of enterobacteria upregulated migration under physiological conditions. Intraperitoneally administered LPS significantly increased the adherence of T‐cells in both GF and SPF mice accompanied by the expression of adhesion molecules and proinflammatory cytokines. However, intraluminally administered LPS did not enhance the adherence of T‐cells in SPF mice. A significant induction of increase in mRNA expression of IRAK‐M, a negative regulator of TLR4 signaling, and transforming growth factor beta (TGF‐beta), a regulatory cytokine, was observed in SPF mice after luminal LPS treatment. Conclusions: Tolerance to intraluminally administered LPS in the lymphocyte recruitment process was induced by enterobacteria, possibly via the induction of IRAK‐M and TGF‐beta.     

Pharmacokinetics of antibiotics in children

In designing the dosage regimen of antibiotics including β-lactams in the field of pediatrics, the choice of adequate drugs, as well as the dosage and the number of administrations have not been performed exactly. This is possibly because of the extended spectrum of antibacterial activity in recent drugs. Even the timing of discontinuation has not been seriously considered. The present report mainly concentrates on several cephalosporins and demonstrates the pharmacokinetic characteristics of the drugs, and further, describes the difference in pharmacokinetic parameters between age groups. On the other hand, it has been suggested that based on the minimum inhibitory concentration for 80% (MIC₈₀) of antibiotics against subject micro-organisms, the effective area under the concentration-time curve, time above MIC₈₀ and C_max/MIC₈₀ should be considered and the suitability of drug choice, the dosage of suitable drugs, and the number of administrations should be decided. The dosage regimen in the present study was not designed to take into account the effects of sub-MIC and post-antibiotic effect. However, for monotherapy, the dosage regimen was designed to maintain the concentration of at least MIC, which is sufficient from the viewpoint of saving life.     

H n.m.r. spectrum from the flexible N-terminal segment of Streptomyces subtilisin inhibitor

The ¹H n.m.r. spectrum of Streptomyces subtilisin inhibitor shows a limited number of unusually sharp signals at room temperature. Some of these signals are assigned uniquely to protons of the side chains of the N-terminal segment, Aspl-Ala2-Pro3-Ser4-Ala5-Leu6-Tyr7-based on experiments of spin decoupling, pH titration, and enzymatic cleavage of the protein. Quantitative examination of these signals indicates that the N-terminal end of this protein is heterogeneous in that the protein contains a considerable fraction whose sequence starts with Ala2 rather than with Asp1. The pK_a values for the amino groups of Asp1 and Ala2 exposed at the N-terminus are determined to be 8.9 ± 0.4 and 9.0 ± 0.1, respectively. Furthermore, examination of the line-widths of the methyl proton resonances of Ala2 and Ala5 residues indicates that the N-terminal peptide segment is free and undergoes rapid segmental motions in the order of 10^?9s.     

Detection of apoptosis in keloids and a comparative study on apoptosis between keloids, hypertrophic scars, normal healed flat scars, and dermatofibroma

Recent studies have suggested that the regulation of apoptosis during wound healing is important in scar establishment and the development of pathological scarring. In this study, we demonstrate that keloid fibroblasts can be identified as apoptotic cells because of their highly condensed chromatin and discrete nuclear fragments. To further reveal the phenomenon of apoptosis, we quantified the number of terminal deoxynucleotide transferase-mediated dUTP nick-end labeling (TUNEL)-positive cells in surgically resected tissues of keloids (N = 10), hypertrophic scars (N = 10), normal healed flat scars (N = 10), and dermatofibroma (N = 10). The number of TUNEL-positive cells was relatively low, but was significantly higher for the keloid group compared with the normally healed flat scar group (p = 0.004), suggesting reduced cell survival and increased apoptotic cell death in a subpopulation of keloid fibroblasts. Furthermore, the number of TUNEL-positive cells was significantly higher for the keloid group compared with the dermatofibroma group (p = 0.044), suggesting that a subpopulation of keloid fibroblasts may suppress tumorgenicity at a greater rate than dermatofibroma by undergoing cell death. Hypertrophic scars had significantly higher levels of apoptosis than normally healed flat scars (p = 0.033). Therefore, these results suggest that selected fibroblasts in keloids and hypertrophic scars undergo apoptosis, which may play a role in the process of pathological scarring.     

Sodium dodecyl sulfate-bovine plasma albumin complex

The acid-induced isomerization (the N-F transition) and expansion of sodium dodecyl sulfate-bovine plasma albumin complex (AD_m;m, molar ratio of added sodium dodecyl sulfate to bovine plasma albumin; 0 ± m ± 12) were studied by measuring CD-resolved secondary structure, fluorescence polarization and lifetime of tryptophyl fluorophors, acid-titration with the electrostatic correction for the surface potential, ¹H-n.m.r. spectra and cross relaxation time between irradiated and observed protons. The immobilization of tryptophyl fluorophors observed in the F-form of AD₀ was suppressed in the F-form of AD₁₀. The acid-titration analysis of AD₁₂ showed non salt-bonding between carboxylate groups and cationic side chains in the F-form, as in the case of AD₀, indicating charged side chains being presumably mobile. ¹H-n.m.r. spectra and cross relaxation times between irradiated and observed protons in the F-form of AD₁₀ indicated the increase in the local motion. On the other hand, AD₁₀ and AD₁₂ did not show any significant change in the CD-resolved secondary structure in the N-F transition region. The F-form of AD₁₀ or AD₁₂ may therefore be the molten-globule state which has secondary structure similar to the N-form of the complexes with fluctuating tertiary structure (side chains).