Differential regulation of IgA production by TGF-beta and IL-5: TGF-beta induces surface IgA-positive cells bearing IL-5 receptor, whereas IL-5 promotes their survival and maturation into IgA-secreting cells.

Transforming growth factor beta (TGF-beta) and IL-5 have been shown to augment IgA production by LPS-stimulated murine B cells. We investigated the effect of TGF-beta on the expression of surface Ig-isotype and IL-5 receptor on LPS-stimulated B cells. TGF-beta increased the proportion of both surface IgA-positive (sIgA+) B cells and sIgG2b+ B cells and enhanced IgA and IgG2b production by LPS-stimulated B cells. TGF-beta synergized with IL-5 only for IgA production of the seven Ig-isotypes and in combination with IL-5 caused a significant increase in the proportion of sIgA+ B cells up to 17.4%. In contrast, IL-5 decreased the proportion of sIgG2b+ B cells and sIgG3+ B cells and inhibited the production of IgG2b and IgG3 by LPS-stimulated B cells. About 50% of sIgA+ cells induced by TGF-beta expressed IL-5 receptor. They secreted peak levels of IgA and seemed to maintain long viability in the presence of IL-5; whereas TGF-beta had the opposite effects on sIgA+ B cells and down-regulated the IL-5 receptor expression. These results indicate that TGF-beta increases the number of sIgA(+)- and IL-5 receptor-positive B cells which respond to IL-5 giving rise to IgA-secreting cells and also support the notions that TGF-beta preferentially induces switching to sIgA+ B cells and IL-5 induces the maturation of postswitch sIgA+ B cells into IgA-secreting cells in a stepwise fashion.     

Redistribution and fate of colchicine-induced alkaline phosphatase in rat hepatocytes: possible formation of autophagosomes whose membrane is derived from excess plasma membrane

The redistribution and fate of colchicine-induced alkaline phosphatase (ALPase) in rat hepatocytes were investigated by electron microscopic enzyme cytochemistry and biochemistry. ALPase activity markedly increased in rat hepatocytes after colchicine treatment (2.0 mg/kg body weight, intraperitoneal injection). At 20–24 h after colchicine treatment, the liver showed the highest activity of ALPase. Thereafter, ALPase activity decreased and returned to normal levels at 48 h. In normal hepatocytes from control rats, ALPase activity was seen only on the bile canalicular membrane. However, at 20–24 h after colchicine treatment, colchicine-induced ALPase was redistributed in the sinusoidal and lateral (basolateral) membranes as well as in the bile canalicular membrane. At 30–36 h after colchicine treatment, ALPase activity on the basolateral membrane gradually decreased. In contrast, ALPase in the bile canalicular membrane increased along with the enlargement of bile canaliculi, suggesting that ALPase in the basolateral membrane had been transported to the bile canalicular membrane. Furthermore, ALPase-positive vesicles, cisternae and autophagosome-like structures were frequently seen in the cytoplasm. ALPase was also positive in some lysosomal membranes. ALPase in hepatocytes at 48 h after colchicine treatment returned to almost the same location as in control hepatocytes. Altogether, it is suggested that excessively induced ALPase is at least partially retrieved by invagination of the bile canalicular membrane and then transported to lysosomes for degradation. In addition, this study indicates that excess plasma membrane might be a possible origin of autophagosomal membrane.     

Immunocytochemical study on photoreceptor cell differentiation in the cultured retina of the chick

Photoreceptor cell differentiation was investigated in a dissociated monolayer culture of chick embryonic retinas with electron microscopic immunohistochemistry. The antibody was raised against bovine rhodopsin purified on SDS-polyacrylamide gel electrophoresis. In the developing retina, immunoreactivity was first recognized on the 14th day of incubation and was localized on the plasma membrane of the growing inner segment. On the 16th day, immunoreactivity was observed on some differentiating outer segments but not on inner segments. In the culture from 6 1/2-day-old embryonic retinas, immunoreactivity was found on the 7th day of culturing on the plasma membrane of large-sized neurons. Electron microscopic observations confirmed that such stained cells showed reaction product on the plasma membrane, and that they displayed fine structures characteristic of intact photoreceptor cells. They had a number of microvillous processes and often one thick process, both of which were intensely stained. Outer segment formation, however, was not observed in the present monolayer culture. These results indicate that opsin synthesis and its transport to the plasma membrane begins prior to and probably independently of outer segment formation.     

Accumulation of O-methyl-L-homoserine by methanol-utilizing bacteria

Summary Several classes of mutants derived from facultative methylotrophs, Microcyclus eburneus and Protaminobacter ruber, accumulate 0.5–3.8 mg/ml of O-methyl-L-homoserine (OMH) in a medium containing methanol as sole carbon source. The wild-type parent strains also accumulate OMH when the medium is supplemented with L-homoserine; the accumulation of OMH by the mutants is ascribed to an increased L-homoserine pool.     

Genetic polymorphism of the complement C2 in Japanese

Summary Genetic polymorphism of the second component of human complement (C2) was investigated in 521 unrelated healthy adult Japanese using isoelectric focusing in polyacrylamide gel followed by a specific hemolytic overlay method. Besides the phenotypes reported previously (C, AC and BC), a relatively infrequent double-banded phenotype (tentatively named A'C) was observed. Moreover, a homozygous variant (A) and a heterozygous double variant (AB) were observed. The estimated frequencies for the common allele. C2 ² (=C2 ¹ ), and the variant alleles, C2 ^A , C2 ^B (=C2 ² ) and C2 ^A were 0.939, 0.034, 0.022, and 0.006, respectively.The results of further typing for HLA-A,-B,-C specificities indicated the presence of significant associations of C2 ^A with HLA-B15 and with A26, and of C2 ^B with HLA-Bw61. These findings support our previous observation that in Japanese there are allelic combinations showing linkage disequilibrium between C2 and HLA loci which are different from those in Caucasians, and that the C2 structural locus is more closely linked to HLA-B than to HLA-A.C2 hemolytic activities of each phenotypes were assayed. The mean activity of type AC sera was significantly higher than that of type C or type BC, while there were no differences in the activities among the types C, BC or A'C.Also presented are two pedigrees demonstrating the segregation of C2 with HLA alleles in which a homozygous C2A or C2B individual was observed.     

Immunochemical approach to characterize advanced glycation end products of the Maillard reaction. Evidence for the presence of a common structure

Reaction of protein amino groups with glucose (the Maillard reaction) leads from early stage products such as Schiff base and Amadori products to advanced glycation end products (AGE), structures implicated in diabetic complications and the aging process. We have prepared the polyclonal anti-AGE antibody and the monoclonal anti-AGE antibody against AGE-bovine serum albumin and made an immunochemical approach to characterize AGE structures. Both polyclonal and monoclonal antibodies reacted with AGE-proteins such as AGE-bovine serum albumin, AGE-human serum albumin, and AGE-hemoglobin but not with unmodified counterparts. Treatments of these AGE-proteins with borohydride had no effect on the immunoreactivity. Moreover, fructosyl-epsilon-caproic acid, a synthetic Amadori compound, did not serve as an antigen, indicating that these antibodies were specific for AGE products but not for early stage products of the Maillard reaction. In addition, these antibodies were also able to recognize AGE products prepared either from alpha-tosyl-1-lysine, alpha-tosyl-1-lysine methyl ester, monoaminocarboxylic acid such as epsilon-aminocaproic acid, gamma-amino-n-butyric acid, and beta-alanine. Thus, these results strongly suggest the presence of a common structure in AGE preparations, regardless of whether AGE products are generated from proteins, amino acids, or monoaminocarboxylic acids.     

Structure of triphosphonoglycosphingolipid containing N-acetylgalactosamine 6-O-2-aminoethylphosphonate in the nervous system of Aplysia kurodai

A phosphonoglycosphingolipid, named F-21, was found in the nervous system of Aplysia kurodai by two-dimensional thin-layer chromatography (Abe, S., Araki, S., and Satake, M. (1986) Biomed. Res. (Tokyo) 7, 47-51). F-21 was isolated from the nervous tissue of Aplysia in this study, and its chemical structure was characterized as follows, where 2-AEP is 2-aminoethylphosphonate. (Formula; see text) The major aliphatic components of the ceramide portion were palmitic acid (75%), stearic acid (22%), octadeca-4-sphingenine (43%), and anteisononadeca-4-sphingenine (54%). Some information on the steric interactions in the sugar moiety was obtained by NMR spectroscopy. The ring protons of the internal galactose, H1, H3, and H4 and the H3 of the side chain galactose were shifted, as compared to the corresponding protons of dephosphonylated F-21. This may indicate the interactions between the 2-AEP residue of N-acetylgalactosamine and the internal galactose and between the N-acetyl group of N-acetylgalactosamine and the side chain galactose, implying a sterically restricted and unique structure that may relate to some biological functions of F-21.