The estradiol-stimulated lipoprotein receptor of rat liver. A binding site that membrane mediates the uptake of rat lipoproteins containing apoproteins B and E

Hepatic catabolism of lipoproteins containing apolipoproteins B or E is enhanced in rats treated with pharmacologic doses of 17 alpha-ethinyl estradiol. Liver membranes prepared from these rats exhibit an increased number of receptor sites that bind 125I-labeled human low density lipoproteins (LDL) in vitro. In the present studies, this estradiol-stimulated hepatic receptor was shown to recognize the following rat lipoproteins: LDL, very low density lipoproteins obtained from liver perfusates (hepatic VLDL), and VLDL-remnants prepared by intravenous injection of hepatic VLDL into functionally eviscerated rats. The receptor also recognized synthetic lamellar complexes of lecithin and rat apoprotein E as well as canine high density lipoproteins containing apoprotein E (apo E-HDLc). It did not recognize human HDL or rat HDL deficient in apoprotein E. Much smaller amounts of this high affinity binding site were also found on liver membranes from untreated rats, the number of such sites increasing more than 10-fold after the animals were treated with estradiol. Each of the rat lipoproteins recognized by this receptor was taken up more rapidly by perfused livers from estrogen-treated rats. In addition, enrichment of hepatic VLDL with C-apoproteins lowered the ability of these lipoproteins to bind to the estradiol-stimulated receptor and diminished their rate of uptake by the perfused liver of estrogen-treated rats, just as it did in normal rats. The current data indicate that under the influence of pharmacologic doses of estradiol the liver of the rat contains increased amounts of a functional lipoprotein receptor that binds lipoproteins containing apoproteins B and E. This hepatic lipoprotein receptor appears to mediate the uptake and degradation of lipoproteins by the normal liver as well as the liver of estradiol-treated rats. The hepatic receptor bears a close functional resemblance to the LDL receptor previously characterized on extrahepatic cells.     

Ionophore-induced disassembly of blood platelet microtubules: effect of cyclic AMP and indomethacin

Cytoplasmic calcium levels are believed to be important in blood platelet activation. Upon activation, the discrete marginal microtubule band, which maintains the discoid shape of non-activated platelets, becomes disrupted. Present studies demonstrate that the extent of assembly of the marginal microtubule band is related to cytoplasmic calcium levels. The divalent cationophore, A23187, causes platelet aggregation, secretion, and contraction by promoting calcium transport from intraplatelet storage sites into the cytoplasm. A23187 caused disassembly of platelet microtubules. Quantitation of electron micrographs revealed that numbers of microtubules were reduced by approximately 80% after A23187 treatment. Secondly, assembled microtubules in homogenates of platelets, in which microtubules were stabilized prior to homogenization, were decreased in favor of free tubulin in A23187-treated platelets. Thirdly, A23187 increased 14C-colchicine binding by intact platelets; this also indicated a shift in the microtubule subunit equilibrium to favor free, colchicine-binding tubulin subunits. In control experiments, A23187 did not affect the stability of platelet tubulin, the colchicine binding reaction, or the total tubulin content of platelets. Stimulation of colchicine binding depended on A23187 concentration (0.05-0.5 microM) and did not require extracellular calcium. A23187-stimulation of colchicine binding was blocked by dibutyryl cyclic AMP (0.80 mM) and/or 3-isobutyl-1-methylxanthine (50 microM) and by indomethacin (10 microM). Cyclic AMP or indomethacin also interferes with A23187-induced platelet activation, but indomethacin is not likely to completely inhibit the perturbation of intraplatelet calcium gradients by A23187. It is suggested that A23187-induced microtubule disassembly may be an indirect effect of calcium on microtubules.     

Linkage of the HMP pathway to ATP generation by the proline cycle

The reactions catalyzed by proline oxidase and pyrroline-5-carboxylate reductase form a catalytic cycle linking the hexose-monophosphate pentose (HMP) pathway to mitochondrial ATP generation. The cycling of proline and pyrroline-5-carboxylate couples glucose oxidation to ATP generation by a mechanism independent of the Embden-Meyerhof pathway and the tricarboxylic acid cycle.     

DNA sequence directs placement of histone cores on restriction fragments during nucleosome formation.

Restriction fragments, 203 and 144 base pairs in length, bearing the Escherichia coli lac control region have been reconstituted with the core histones from calf thymus to form nucleosomes. By several criteria the reconstituted nucleosomes are similar to native nucleosomes obtained by micrococcal nuclease digestion of calf thymus nuclei. However, sensitive nuclease digestion studies reveal subtle and important differences between native monosomes and the lac reconstitutes. Each reconstitute consists mainly of nucleosomes containing histone cores placed nonrandomly with respect to the DNA sequence. The shorter reconstitute forms asymmetric nucleosomes as evidenced by the DNase I digestion pattern. Exonuclease III digestion followed by 5'-end analysis of the larger reconstitute suggests that, of the many possible arrangements of histone core with DNA sequence, only two are highly favored.     

13C- and 1H-nmr studies of cis-trans conformers of oligoprolines

In aqueous solutions, ¹³C- and ¹H-nmr studies show that the percentage of trans conformation of proline oligomers ⁺H₂H Pro-(Pro)_n-CO increases substantially from n = 1 (65% trans) to n = 2 (90% trans). The relatively low percentage of trans structure for the dimer (n = 1) very likely is caused by the extra stability acquired by the end-to-end intramolecular H-bonding of the cis dimer. As n increase from 2 to 3 (or 5) in ⁺H₂N-Pro-(Pro)_n-CO, the percentage of trans conformation stays more or less constant (～0.9). A high salt concentration (4M CaCl₂) causes a conformation randomization, so that the short-chain oligomer (n = 1, 3, 5) and the long-chain poly (L -proline) all show about the same frantion of trans conformation (0.7-0.8).     

Ultrastructural alterations in skeletal muscle fibers of streptozotocin-diabetic rats

Summary The ultrastructure of fast-twitch-oxidative-glycolytic (FOG), fasttwitch-glycolytic (FG) and slow-twitch-oxidative (SO) fibers in plantaris and soleus muscles of normal and streptozotocin-diabetic rats was studied. In the diabetic animals, the mitochondria of FOG and SO fibers showed a loss of cristae and an increase in electron-dense granules. There was also an increased number of lipid droplets in close proximity to the mitochondria and the nuclei, and a separation of individual muscle nuclei to form satellite cells. Higher incidences of surface projections and sarcoplasmic splittings at the nuclear region were noticed in SO fibers. The FG fibers showed some disorientation of the T-tubular system. It is concluded that streptozotocin-diabetes has differential effects on the fine structure of the three fiber types of rat skeletal muscle.Supported by USPHS Grant AM 18280-04, Boston University Grant GRS-405-BI, and a grant-in-aid award from Sigma Xi Society     

Circular RNA AFF4 modulates osteogenic differentiation in BM-MSCs by activating SMAD1/5 pathway through miR-135a-5p/FNDC5/Irisin axis

Bone marrow-derived mesenchymal stem cells (BM-MSCs), the common progenitor cells of adipocytes and osteoblasts, have been recognized as the key mediator during bone formation. Herein, our study aim to investigate molecular mechanisms underlying circular RNA (circRNA) AFF4 (circ_AFF4)-regulated BM-MSCs osteogenesis. BM-MSCs were characterized by FACS, ARS, and ALP staining. Expression patterns of circ_AFF4, miR-135a-5p, FNDC5/Irisin, SMAD1/5, and osteogenesis markers, including ALP, BMP4, RUNX2, Spp1, and Colla1 were detected by qRT-PCR, western blot, or immunofluorescence staining, respectively. Interactions between circ_AFF4 and miR-135a-5p, FNDC5, and miR-135a-5p were analyzed using web tools including TargetScan, miRanda, and miRDB, and further confirmed by luciferase reporter assay and RNA pull-down. Complex formation between Irisin and Integrin αV was verified by Co-immunoprecipitation. To further verify the functional role of circ_AFF4 in vivo during bone formation, we conducted animal experiments harboring circ_AFF4 knockdown, and born samples were evaluated by immunohistochemistry, hematoxylin and eosin, and Masson staining. Circ_AFF4 was upregulated upon osteogenic differentiation induction in BM-MSCs, and miR-135a-5p expression declined as differentiation proceeds. Circ_AFF4 knockdown significantly inhibited osteogenesis potential in BM-MSCs. Circ_AFF4 stimulated FNDC5/Irisin expression through complementary binding to its downstream target molecule miR-135a-5p. Irisin formed an intermolecular complex with Integrin αV and activated the SMAD1/5 pathway during osteogenic differentiation. Our work revealed that circ_AFF4, acting as a sponge of miR-135a-5p, triggers the promotion of FNDC5/Irisin via activating the SMAD1/5 pathway to induce osteogenic differentiation in BM-MSCs. These findings gained a deeper insight into the circRNA-miRNA regulatory system in the bone marrow microenvironment and may improve our understanding of bone formation-related diseases at physiological and pathological levels.Subject terms: Stem cells, Diseases     

Multiple species of wild tree peonies gave rise to the ‘king of flowers’, Paeonia suffruticosa Andrews

The origin of cultivated tree peonies, known as the ‘king of flowers'' in China for more than 1000 years, has attracted considerable interest, but remained unsolved. Here, we conducted phylogenetic analyses of explicitly sampled traditional cultivars of tree peonies and all wild species from the shrubby section Moutan of the genus Paeonia based on sequences of 14 fast-evolved chloroplast regions and 25 presumably single-copy nuclear markers identified from RNA-seq data. The phylogeny of the wild species inferred from the nuclear markers was fully resolved and largely congruent with morphology and classification. The incongruence between the nuclear and chloroplast trees suggested that there had been gene flow between the wild species. The comparison of nuclear and chloroplast phylogenies including cultivars showed that the cultivated tree peonies originated from homoploid hybridization among five wild species. Since the origin, thousands of cultivated varieties have spread worldwide, whereas four parental species are currently endangered or on the verge of extinction. The documentation of extensive homoploid hybridization involved in tree peony domestication provides new insights into the mechanisms underlying the origins of garden ornamentals and the way of preserving natural genetic resources through domestication.