Increased UDP-GlcNAc:Gal beta 1-3GaLNAc-R (GlcNAc to GaLNAc) beta-1, 6-N-acetylglucosaminyltransferase activity in metastatic murine tumor cell lines. Control of polylactosamine synthesis

Malignant transformation of rodent cell lines by polyoma virus and by activated ras genes is associated with increased UDP-GlcNAc:Man alpha-R beta-1,6-N-acetylglucosaminyltransferase V (GlcNAc-transferase V) activity and it product -GlcNAc beta 1-6Man alpha 1-6Man beta 1-branched Asn-linked oligosaccharides. In this report, we have compared beta 1-6GlcNAc branching of core O- and N-linked oligosaccharides in three experimental models of malignancy, namely (a) rat2 fibroblasts and their malignant T24H-ras-transfected counterpart; (b) benign SP1 mammary carcinoma cells and two metastic sublines of SP1; and (c) the metastatic MDAY-D2 lymphoma cell line and its poorly metastatic glycosylation mutant KBL-1. In addition to the previously reported increase in GlcNAc-transferase V activity, UDP-GlcNAc:Gal beta 1-3GalNAc alpha-R (GlcNAc to GalNAc) beta-1,6-N-acetylglucosaminyltransferase (core 2 GlcNAc-transferase, EC 2.4.1.102) activity was found to be elevated by 70% in the malignant rat2 and SP1 cell lines while several other glycosyltransferase activities were not significantly different. The action of core 2 GlcNAc-transferase followed by beta 1-4Gal-transferase provides an N-acetyllactosamine antenna that can be extended with polylactosamine (i.e. repeating Gal beta 1-4GlcNAc beta 1-3) provided UDP-GlcNAc:Gal beta-R beta 1-3GlcNAc-transferase (GlcNAc-transferase) (i)) activity is present. Polylactosamine content in microsomal membrane glycoproteins was quantitated by labeling the GlcNAc termini resulting from the action of Escherichia freundii endo-beta-galactosidase with bovine galactosyltransferase/UDP-[3H] Gal. Glycopeptidase F- sensitive and -insensitive fractions were measured to assess the N- and O-linked components. In the SP1 tumor model, the metastatic sublines showed increased core 2 GlcNAc-transferase and GlcNAc-transferase V activities but no change in GlcNAc-transferase (i) activity, yet polylactosamine was increased in both O- and N-linked oligosaccharides. In rat2 cells, down-regulation of GlcNAc-transferase (i) following transformation was associated with decreased polyactosamine even though core 2 GlcNAc-transferase and GlcNAc-transferase V were elevated in the cells. Finally, a 3-fold decrease in GlcNAc-transferase V in KBL-1, the glycosylation mutant of MDAY-D2 cells, resulted in complete loss of polylactosamine in N-linked but no change in O-linked polylactosamine content. These results suggest that, provided GlcNAc-transferase (i) is not limiting, the beta 1-6-branching enzymes core 2 GlcNAc-transferase and GlcNAc-transferase V regulate the levels of polyactosamine in O- and N-linked oligosaccharides, respectively.(ABSTRACT TRUNCATED AT 400 WORDS)     

Characterization of insulin-like growth factor I (IGF-I) receptors of human breast cancer cells

Studies of binding of IGF-I to a plasma-membrane-enriched subcellular fraction prepared from MCF-7 human breast cancer cells reveal the presence of 0.2 pmols specific binding sites for this mitogen per mg membrane protein, with an equilibrium affinity constant of 1.45 nM-1. Competition studies with insulin, IGF-II, and an anti-IGF-I receptor antibody are consistent with the presence of specific IGF-I receptors, and SDS-PAGE showed binding to a 130 kDa subunit identical to that of receptors from human placenta. In addition, we show that IGF-I is more potent than estradiol and comparable to EGF in stimulating in vitro proliferation of MCF-7 cells, and that IGF-I-stimulated proliferation of these cells is inhibited by a blocking monoclonal antibody against the IGF-I receptor. These results demonstrate that IGF-I is an important mitogen for MCF-7 cells and that the mitogenic effect is mediated by specific IGF-I receptors.     

Tn antigen and UDP-Gal:GalNAc alpha-R beta 1-3Galactosyltransferase expression in human breast carcinoma.

The serine/threonine O-linked carbohydrates GalNAc alpha and Gal beta 1-3GalNAc alpha, referred to as Tn and T antigens, respectively, appear to be more prevalent in some human carcinomas than in surrounding tissues. Tn/T antigens may represent incomplete synthesis of O-linked oligosaccharides, due to decreased activity of specific glycosyltransferases, or alternatively, increased glycosidases activity in tumors which may expose these internal O-linked oligosaccharide sequences. To explore these possibilities, we measured UDP-Gal:GalNAc alpha-R beta 1-3 galactosyltransferase (beta 3Gal-T) and Gal beta 1-3GalNAc alpha-R beta 1-3 galactosidase in a series of human breast tumors. In addition, glycoproteins extracted from the tumors were separated by SDS-PAGE and stained with the lectins HPA (GalNAc alpha-R reactive) and PNA (Gal beta-3GalNAc alpha-R reactive). The relative levels of HPA- to PNA-reactive glycoproteins in the carcinomas correlated inversely with beta 3Gal-T activities. The results suggest that Tn antigen expression in human breast carcinoma is due in part to low beta 3Gal-T activity, a situation similar to that observed previously in haematopoietic cells of individuals with a condition called Tn syndrome.     

The effect of somatostatin on the production of human interferons by mononuclear cells.

Somatostatin (SMS) is a tetradecapeptide which can inhibit the secretion of a number of peptides produced by the endocrine or nervous systems. SMS 201-995 (octreotide) is a somatostatin analogue with very potent somatostatin activities. We have been investigating the effects of both SMS and octreotide on the production of human interferon (IFN). We obtained human peripheral blood mononuclear cells from normal donors and induced them to produce IFN in the presence or absence of a number of peptides possessing somatostatin activities. SMS and octreotide were shown to inhibit the secretion of INF-gamma but not IFN-alpha. Concentrations of 10(-6) M were shown to decrease yields when Concanavalin A or phytohemagglutin were used as the inducer. Higher concentrations had a progressively greater effect. No effects were observed on IFN-gamma production if interleukin 2, ionomycin, or various natural antigens were used to induce the cells. The 28-amino acid form of somatostatin had some effects on gamma IFN yields but the first 14-amino acid fragment of this peptide moiety did not. No effect of any of these compounds was observed on IFN bioactivity. These studies indicate SMS may have some regulatory action on the secretion of immunomodulators in vitro but the concentrations required are well above those encountered under physiologic circumstances, suggesting SMS may not play an important regulatory role governing such secretion in vivo.     

Comparison of 5.8S and ITS2 rDNA RFLP Patterns Among Isolates of Acremonium obclavatum, A. kiliense, and A. strictum from Diverse Sources

Polymerase chain reaction (PCR) products of nuclear 5.8S and internal transcribed spacer regions (ITS2) of rDNA from reference cultures of Acremonium obclavatum (a rarely recognized species first reported from India) were compared with cultures of Acremonium spp. isolated from Georgia, USA. Digestion of amplicons sequentially with Hinfl and Sau3AI divided the isolates into four restriction fragment length polymorphism (RFLP) groups. A representative isolate of primary colonizers of insulation facings from a building in Georgia appeared identical to the type culture of A. obclavatum, whereas other cultures from Indian soils showed variation in the ITS2 region that divided them into further subgroups. Reference cultures of A. kiliense (ATCC 14489) and A. strictum (ATCC 10141) and two additional isolates from metropolitan Atlanta, assigned to this latter species complex on a morphological basis, represented two additional RFLP groups both of which were distinct from the RFLP groups in A. obclavatum. A. kiliense and A. strictum could be placed into similar subgroups on the basis of morphological differences and distinct RFLP patterns. Received: 6 June 1997 / Accepted: 22 July 1997     

Investigation of intermolecular hydrogen bond interactions in crystalline l-Cysteine by DFT calculations of the oxygen-17, nitrogen-14, and hydrogen-2 EFG tensors and AIM analysis

A systematic computational study is carried out to investigate hydrogen bond (HB) interactions in the real crystalline structures of l-Cysteine at 30 and 298 K by density functional theory (DFT) calculations of electric field gradient (EFG) tensors at the sites of O-17, N-14, and H-2 nuclei. One-molecule (monomer) and nine-molecule (cluster) models of l-Cysteine are created by available crystal coordinates at both temperatures and the EFG tensors are calculated for both models to indicate the effect of HB interactions on the tensors. The calculated EFG tensors at the level of B3LYP and B3PW91 DFT methods and 6-311++G?? and cc-pVTZ basis sets are converted to those experimentally measurable nuclear quadrupole resonance (NQR) parameters i.e. quadrupole coupling constants (qcc) and asymmetry parameters (η_Q). The evaluated NQR parameters reveal that the EFG tensors of ¹⁷O, ¹⁴N, and ²H are influenced and show particular trends from monomer to the target molecule in the cluster due to the contribution of target molecule to classic N–H…O, and non-classic S–H…O and S–H…S types of HB interactions. On the other hand, atoms in molecules (AIM) analyses confirm the presence of HB interactions and rationalize the observed EFG trends. The results indicate different contribution of various nuclei to HB interactions in the cluster where O2 and N1 have major contributions. The EFG tensors as well as AIM analysis at the H6 site show that the N1-H6…O2 HB undergoes a significant change from 30 to 298 K where changes in other N–H…O interactions are almost negligible. There is a good agreement between the calculated ¹⁴N NQR parameters and reported experimental data.     

Levels of formaldehyde in residential indoor air of Gonabad,Iran

Abstract

Human health has been identified to be affected more significantly by indoor air quality. Among numerous pollutants present in indoor air, formaldehyde (FA) is of great concern because of its highly hazardous nature. The concentrations of FA were determined from 20 newly decorated homes in the city of Gonabad, Iran during 2015. It was found that the indoor air levels of FA in all the sampled houses were exceptionally high in the range of 21 to 360 µg/m³ (mean of 149.3 µg/m³). If the 24-h average concentrations of FA measured from those sites were concerned, nearly 40% of them were seen to exceed the WHO guideline values (i.e., 100 µg/m³). One of the important reasons for the high concentrations could be low air exchange rates in those houses (e.g., from 0.18 to 0.37?h^?1), high levels of humidity in the newly decorating houses and stronger sources in the indoor environment. Furthermore, its pollution in homes with natural ventilation was seen to be much higher than those of mechanical ventilation. Due to high levels of indoor FA, more effective control procedures should be developed and employed to reduce the risk associated with formaldehyde exposure.