Intracellular transport of endocytosed chylomicron [3H]retinyl ester in rat liver parenchymal cells. Evidence for translocation of a [3H]retinoid from endosomes to endoplasmic reticulum

The intracellular transport of chylomicron remnants labeled with [3H]retinyl ester was studied in rat liver parenchymal cells by means of subcellular fractionation in Nycodenz and sucrose density gradients. The data presented indicate that endocytosed chylomicron remnant [3H]retinyl ester initially is located in low density endosomes. Radioactivity is subsequently transferred to a denser vesicle. Equilibrium as well as rate zonal centrifugation suggest that this denser [3H] retinoid-containing vesicle may represent endoplasmic reticulum. We have compared the intracellular transport of chylomicron remnant [3H]retinyl ester and 125I-asialofetuin. The receptor-mediated endocytosis of asialoglycoproteins in rat liver parenchymal cells is a thoroughly studied system. Our results suggest that the [3H] retinoid and 125I-asialofetuin follow the same path initially to the endosomes. After transit in endosomes, the intracellular transport differs. While asialofetuin is transported to the lysosomes, the retinoid is probably transferred to the endoplasmic reticulum.     

Seasonal and annual variability in the phytoplankton community of the Raunefjord,west coast of Norway from 2001–2006

Changes in phytoplankton community composition potentially affect the entire marine food web. Because of seasonal cycles and inter-annual variations in species composition, long-term monitoring, covering many sequential years, is required to establish a baseline study and to reveal long-term trends. The current study describes the phytoplankton biomass variations and species composition in relation to hydrographic and meteorological conditions in the Raunefjord, western Norway, over a 6-year period from 2001 to 2006. The extent of inflow or upwelling in the fjord varied from year to year and resulted in pronounced differences in water column stability. The annual phytoplankton community succession showed some repeated seasonal patterns, but also high variability between years. Two to four diatom blooms were observed per year, and the spring blooms occurring before water column stratification in March were dominated by Skeletonema marinoi and Chaetoceros socialis, and other Chaetoceros and Thalassiosira spp. Blooms of the haptophytes Phaeocystis pouchetii and Emiliania huxleyi were irregular and in some years totally absent. Although E. huxleyi was present all year round it appeared in bloom concentrations only in 2003, when the summer was warm and the water column characterized by high surface temperatures and pronounced stratification. The annual average abundance of both diatoms and flagellates increased during the six years. Despite the high variation from year to year, our investigation provides valuable knowledge about annual phytoplankton community patterns in the region, and can be used as a reference to detect possible future changes.     

Downregulation of c-myc RNA is not a prerequisite for reduced cell proliferation, but is associated with G1 arrest in B-lymphoid cell lines

We related the effects of c-myc expression on the ability of growth inhibitors to block the cells in the G0/G1 phase of the cell cycle. In two different B-cell lines, there was an association between the accumulation of cells in the middle to late G1 phase of the cell cycle and a rapid transient downregulation of c-myc mRNA levels. The phorbol ester TPA and the adenylate cyclase activator forskolin reduced the c-myc RNA, levels and after 3 days of treatment a proportion of the cells accumulated in G1. In contrast, neither interferon-gamma, tumor necrosis factor-alpha nor the monoclonal antibody 33-1 against DQ major histocompatibility antigens changed the cell-cycle distribution or regulated the c-myc RNA levels. Yet, all five growth inhibitors reduced the proliferation to approximately the same extent. The growth reduction was not accompanied by definite differentiation, as judged by the absence of the B-cell differentiation marker B1 (CD20).     

Intracellular transport and degradation of chylomicron remnants in rat liver cells after in vivo endocytosis

The intracellular transport and degradation of in vivo endocytosed chylomicron remnants labelled with 125I in the protein moiety was studied in rat liver cells by means of subcellular fractionation in Nycodenz and sucrose density gradients. Initially, the radioactivity was located in low-density endosomes and was sequentially transferred to light and dense lysosomes. Data from gel filtration of the light and dense lysosomal fractions showed radioactive material with a molecular weight of about 1000-2000, representing short peptide fragments or amino acids which remain attached to iodinated tyramine cellobiose. In addition, undegraded apoproteins accumulated in both types of lysosome. Our data suggest that endocytosed chylomicron remnant apoproteins are first located in low-density endosomes and are sequentially transferred to light and dense lysosomes. Furthermore, the degradation process starts in the light lysosomes.     

Molecular cloning of a pair of human pepsinogen A genes which differ by a Glu→Lys mutation in the activation peptide

Summary Three human cosmid clones containing pepsinogen A (PGA) encoding sequences were isolated from a genomic bank derived from a single individual. One cosmid contains two PGA genes in tandem in a head-to-tail orientation, while the other two cosmids each contain a single PGA gene. The three cosmids were characterized by restriction mapping and sequence analysis (exons 1 and 2 and flanking regions). As judged from these data, three of the four PGA genes isolated appear to be nearly identical, but one of the tandem genes is clearly different from the other genes. The first exon of all four genes codes for the same amino acid sequence. However, in the second exon of one of the tandem genes we found a nucleotide substitution giving rise to a GluLys substitution of the 43rd amino acid residue of the activation peptide, leading to a charge difference of the corresponding isozymogens. The presence of two distinct PGA genes in the isolated gene pair conclusively proves the multigene structure of the PGA system. These genes might be responsible for at least part of the electrophoretic polymorphism at the protein level.     

Retinol and retinyl esters in parenchymal and nonparenchymal rat liver cell fractions after long-term administration of ethanol

Chronic ethanol consumption reduces the liver retinoid store in man and rat. We have studied the effect of ethanol on some aspects of retinoid metabolism in parenchymal and nonparenchymal liver cells. Rats fed 36% of total energy intake as ethanol for 5-6 weeks had the liver retinoid concentration reduced to about one-third, as compared to pair-fed controls. The reduction in liver retinoid affected both the parenchymal and the nonparenchymal cell fractions. Plasma retinol level was normal. Liver uptake of injected chylomicron [3H]retinyl ester was similar in the experimental and control group. The transport of retinoid from the parenchymal to the nonparenchymal cells was not found to be significantly retarded in the ethanol-fed rats. Despite the reduction in total retinoid level in liver, the concentrations of unesterified retinol and retinyl oleate were increased in the ethanol fed rats. Hepatic retinol esterification was not significantly affected in the ethanol-fed rats. Since our study has demonstrated that liver uptake of chylomicron retinyl ester is not impaired in the ethanol-fed rat, we suggest that liver retinoid metabolism may be increased.     

Effect of dextran and dextran modifications on the thermal and proteolytic stability of conjugated bovine testis beta-galactosidase and human serum albumin

In order to study carbohydrate-induced protein stabilization bovine testis beta-galactosidase and human serum albumin were conjugated with dextran, partially acetylated dextran and partially methylated dextran. The conjugates and the free proteins were compared with respect to thermal stability at 50 degrees C and resistance to proteolytic digestion by subtilopeptidase A. Both beta-galactosidase and serum albumin were stabilized by conjugation with polysaccharide. However, higher stability was achieved by conjugating the proteins with the hydrophilic polysaccharides, dextran and acetylated dextran, than by conjugation with the hydrophobic polysaccharide, methylated dextran. The results are discussed in relation to possible explanations of carbohydrate-induced protein stabilization.     

Compounds in urban air compete with 2,3,7,8-tetrachlorodibenzo-p-dioxin for binding to the receptor protein

Acetone extracts of filter-collected urban airborne particulate matter contain compounds which can competitively inhibit 2,3,7,8-[1,6-3H]tetrachlorodibenzo-p-dioxin (TCDD) binding to the rat liver TCDD-receptor protein. The concentration of conventional polycyclic aromatic hydrocarbons (PAHs) or chlorinated dioxins and dibenzofurans cannot account for more than 1-30% of the observed competition for [3H]TCDD binding to the receptor protein. The difference in potency between samples collected in urban areas during different periods of the year and a background sample is 25-400-fold. Collecting samples in the presence of increased concentrations of nitrogen dioxide, nitrous acid, nitric acid or ozone did not increase the amount of compounds with receptor affinity. However, with nitrogen dioxide and especially with nitric acid, a substantial increase of the mutagenic effects in the Ames Salmonella assay in the absence of mammalian activation as well as a degradation of several PAHs were noted. Affinity for the TCDD-receptor protein, mutagenicity in the absence of mammalian metabolic activation in the Ames Salmonella assay and PAH-content are characteristics of urban particulate matter showing the presence of compounds, that represent potential health risks. The compounds with affinity for the receptor may constitute a group of substances different from both conventional PAHs and direct-acting mutagens.     

Uptake of guanine by synchronizedChlorella fusca

1. The transport of guanine in autospores of light-dark synchronizedChlorella fusca was studied using radioactive guanine in the concentration range of 4 nM to 50 μM.
2. The transport system was constitutive, it had high specificity for the permeant, and theQ ₁₀ value was in the range of 1.5 to 2.2. At concentrations lower than 0.2 μM the half saturating constant, S_0.5 was 1 μM both for cells kept in dark and cells kept in light. At higher concentrations the S_0.5 of darkened cells was about 0.23 μM, while that of illuminated cells was unchanged. Only above 0.2 μM guanine did illumination of the cells or addition of glucose increase the transport rate.
3. Guanine which had accumulated did not leak out at temperatures below 45°C or by treatment with 10 μM dinitrophenol, which completely inhibited transport. Furthermore, the accumulated guanine did not exchange with exogenous guanine.
4. The guanine accumulated, more than 10⁵-fold over the external concentration, showing that the transport, was active.
5. The initial transport rate per cell revealed annual fluctuations.