Muscarinic acetylcholine receptors in rat gastric mucosa

Summary The muscarinic cholinergic innervation of the rat gastric mucosa was investigated by localizing the muscarinic receptors using a tritiated muscarinic antagonist, pirenzepine. Radioautography was performed by freeze drying stomach tissue, which was then embedded in Epon and wet sectioned with ethylene glycol, and dry mounting on emulsion film by the wire-loop method to prevent loss of the labelled substance during fixation and the radioautographic procedure. Light and electron microscopy showed that the specific pirenzepine-binding sites were localized predominantly on parietal cells, chief cells and perivascular plexuses. Analysis of the grain distribution on parietal cells revealed that the silver grains corresponding to the pirenzepine-binding sites were mainly on the basolateral plasma membrane. On the other hand, the surface mucous or mucous neck cells had few pirenzepine-binding sites.     

Genetic polymorphism of human factor I (C3b inactivator)

Summary Genetic polymorphism of human factor I (C3b inactivator) has been described using polyacrylamide gel isoelectric focusing electrophoresis of neuraminidase-treated EDTA plasma samples followed by electrophoretic blotting technique. In 435 individuals three different common patterns were observed, and these were controlled by two common alleles at a single locus. The results of typing family material confirmed autosomal codominant Mendelian inheritance. Two common alleles were designated FI^*B and FI^*A, and gene frequencies were estimated to be 0.8931 and 0.1069 for FI^*B and FI^*A, respectively. The distribution of phenotypes fitted the Hardy-Weinberg equilibrium. Linkage studies failed to show close linkage between factor I and the major histocompatibility complex.     

False positive reaction for carcinoembryonic antigen in paget cells

Virchows Archiv B Cell Pathology - Paget cells from cases of mammary and extramammary Paget’s disease were examined for carcinoembryonic antigen (CEA) and CEA-related antigens by the...     

Restriction fragment length polymorphism detected by human salivary amylase cDNA

Summary The plasmid clone which contains human salivary amylase cDNA was used to detect restriction fragment length polymorphisms (RFLPs). After double digestion with Pst 1 and Bam H1, a polymorphism with two alleles was observed. In Japanese, frequencies of these alleles, tentatively called 5.7kb and 6.5kb fragment alleles, are 0.55 and 0.45, respectively.     

Arylsulfatase a from normal human lung and lung tumors showed different patterns of microheterogeneity

Arylsulfatase A was purified from human lung to apparent homogeneity as determined by electrophoresis in the presence of sodium dodecyl sulfate. The enzyme from normal lung as well as that from lung adenocarcinoma showed considerable microheterogeneity when examined by isoelectric focussing, with an isoelectric point (pI) ranging from 5.1 to 4.6. The tumor enzyme was more heterogeneous and contained more acidic components than the normal lung enzyme. The cause of the charge heterogeneity was examined by treatment with exogenous hydrolases. Upon treatment with sialidase, phosphatase or endo-beta-N-acetylglucosaminidase H (endoglycosidase H), the acidic enzyme forms shifted to an alkaline region on isoelectric focussing gels. Combined treatment of the arylsulfatase A with endoglycosidase H and sialidase resulted in complete loss of the most acidic components to give the less acidic components with pI 5.1, 5.0, and 4.9. These results strongly suggest that the charge heterogeneity of arylsulfatase A is due not only to sialylation but also to phosphorylation at the carbohydrate moiety of the enzyme, and the extent of substitution by acidic groups is markedly increased in the tumor enzyme.     

Characterization of the (Ca2+-Mg2+)ATPase purified by calmodulin-affinity chromatography from bovine aortic smooth muscle

A calmodulin inhibitor, trifluoperazine, suppresses ATP-dependent Ca2+ uptake into microsomes prepared from bovine aortic smooth muscle. From this microsomal preparation which we expected to contain calmodulin-dependent Ca2+-transport ATPase [EC 3.6.1.3], we purified (Ca2+-Mg2+)ATPase by calmodulin affinity chromatography. The protein peak eluted by EDTA had calmodulin-dependent (Ca2+-Mg2+)ATPase activity. The major band (135,000 daltons) obtained after sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) accounted for about 80% of the total protein eluted. This major band was phosphorylated by [gamma-32P]ATP in a Ca2+-dependent manner. All the 32P incorporated into the major band was released by hydroxylaminolysis. The ATPase reconstituted in soybean phospholipid liposomes showed ATP, calmodulin-dependent Ca2+ uptake. The affinity of the ATPase for Ca2+, Km, was 7 microM and the maximum ATPase activity was 1.4 mumol/mg/min. These values were changed to 0.17 microM and 3.5 mumol/mg/min, respectively by the addition of calmodulin. The activity of the purified (Ca2+-Mg2+)ATPase was inhibited by orthovanadate, and the concentration required for half-maximal inhibition was about 1.8 microM which is close to that of plasma membrane ATPases. Judging from the effect of orthovanadate and the molecular weight, the purified (Ca2+-Mg2+)ATPase was considered to have originated from the plasma membrane not from the sarcoplasmic reticulum.     

DNA-DNA hybridization analysis of nylon oligomer-degradative plasmid pOAD2: identification of the DNA region analogous to the nylon oligomer degradation gene.

Fine structure of the gene of 6-aminohexanoic acid cyclic dimer hydrolase, one of the enzymes responsible for the degradation of the nylon oligomer (6-aminohexanoic acid cyclic dimer), on the plasmid pOAD2 harbored in Flavobacterium sp. KI72 was determined by constructing miniplasmids from plasmid pNDH5 (a hybrid plasmid consisting of pBR322 and a 9.1-kilobase-pair HindIII fragment of pOAD2 ). The 6-aminohexanoic acid cyclic dimer hydrolase produced by cells of Escherichia coli C600 harboring pNDH5 or its miniplasmid was examined immunologically and electrophoretically and was found to be identical to that of Flavobacterium sp. KI72 . A fragment of pOAD2 (17.2- to 19.1-kilobase-pair region on pOAD2 ) was detected as hybridized fragment by Southern blotting experiments, indicating the presence of the DNA region analogous to the 6-aminohexanoic acid cyclic dimer hydrolase gene on the plasmid.     

Cell-mediated cytotoxicity against allogeneic mutant H-2 antigens: restricted and nonrestricted responses

Two "gain and loss" type mutations of the H-2D region, the H- 2bm13 and H- 2bm14 , resulted in the expression of noncross-reactive CML determinants that are unique to each mutation, the Dbm13 gains and Dbm14 gains, respectively. According to the results of direct cytolytic and competitive inhibition assays of in vitro induced primary cytotoxic T lymphocytes, allogeneic responses specific for Dbm13 gains are generated by responders bearing the H-2b ( KbIbDb ) haplotype, but not by responders bearing the H- 2bm14 ( KbIbDbm14 ), KbIbDd , KbIbDk , or KbIb / qDq haplotype. Responses by the non-H-2b responders against Dbm13 are limited to those determinants shared by the Dbm13 and Db molecules. Because congenic mice differing only at the H-2D region are either responsive or nonresponsive to Dbm13 gains, the responsiveness is controlled by gene(s) in the H-2D region. F1 hybrid offspring of responsive (H-2b) and nonresponsive (non-H-2b) parents are invariably responsive, indicating genetic dominance of the responsiveness. In contrast to the response against Dbm13 gains, cytotoxicity specific for Dbm14 gains is generated by responders bearing the H-2b, H- 2bm13 , KbIbDd , KbIbDk , or KbIb / qDq haplotype. These data indicate the existence of two types of allogeneic MHC determinants; one, represented by Dbm14 gains, is the classic type capable of eliciting CML responses in mice of a wide range of H-2 haplotypes, whereas the other, exemplified by Dbm13 gains, elicits CTL responses only in mice of a few related haplotypes. It is proposed that recognition of Dbm13 gains is restricted by structures shared by Db and Dbm13 but missing from other D (or L, R, etc.) molecules, such as Dbm14 , Dd, Dk, and Dq. Availability of various restricting structures in self MHC molecules may thus influence the alloreactive CTL repertoire.     

Conversion to adipocytes of a clonal bone marrow preadipocyte line (H-1/A) and fatty acid composition of the resultant adipocytes

Conversion to adipocytes and fatty acid composition were investigated in a clonal bone marrow preadipocyte line (H-1/A). The growing cells exhibited a fibroblastic appearance. After the cessation of growth, triacylglyceride (TG) synthesis in the cells increased as they incorporated precursor from the growth medium and became adipocytes. Hydrocortisone and insulin accelerated the TG synthesis in H-1/A cells in a dose-dependent manner when they were cultured in the growth medium containing 10% horse serum. The rate of conversion to adipocytes was reduced as the concentration of horse serum was decreased, and this reduction was not influenced by the addition of insulin and/or hydrocortisone. These results suggest that conversion to adipocytes of H-1/A cells is primarily dependent on some component(s) of the serum. Conversion to adipocytes of the cells may involve a process of differentiation since the conversion was completely inhibited when the cells were cultured in the presence of bromodeoxyuridine. Fatty acid composition was significantly different between adipose H-1/A cells and adipocytes derived from other marrow preadipocyte line MC3T3-G2/PA6 cells. Unsaturated fatty acids accounted for 76% of the fatty acid composition of adipose H-1/A cells; in contrast, saturated fatty acids constituted 65% of the fatty acid composition of the adipose MC3T3-G2/PA6 cells. These results suggest that there is a heterogeneity of preadipocytes in bone marrow. These two preadipocyte lines thus provide a useful tool for the study of marrow adipocytes and can also be used to analyze the hematopoietic microenvironment through studies of the effect of these cells on hematopoietic cell proliferation.     

Sensitivity of mice to systemic hyperthermia: effect of the transplantation of ascites tumor cells modified by preheating, and by an injection of CDDP or interferon

Effects of preheating and injection of cis-DDP (CDDP) or interferon on tumor-induced sensitization to systemic hyperthermia (SH) was investigated in mice. LD50 of SH at 42.0 +/- 0.2 degrees C (core body temperature) was 43 min in normal mice and 8 min in mice which were i.p. transplanted with FMA3 cells at a dose of 10(5) one day before. In mice which had received the SH for 10 min one hour before, one hour after or one day before the transplantation, LD50 of the SH one day after the transplantation was 41, 35 and 22 min, respectively. An injection of CDDP given i.p. at a dose of 4 mg/kg one day after the transplantation, which was effective to kill about 99% of the tumor cells, did not change the course of thermosensitization after the transplantation. An i.p. injection of mouse interferon did not change the thermosensitivity of normal mice, but greatly suppressed the thermosensitizing effect of tumor cells when it was given one day before the transplantation.