Five New Phenolic Glycosides from Viburnum luzonicum

Viburnum luzonicum is widely distributed in China. Its branch extracts showed potential α-amylase and α-glucosidase inhibitory activities. In order to discover new bioactive constituents, five undescribed phenolic glycosides, viburozosides A−E ( 1 – 5 ), were obtained by bioassay-guided isolation coupled with HPLC-QTOF-MS/MS analysis. Their structures were elucidated by spectroscopic analyses, including 1D NMR, 2D NMR, ECD, and ORD. All compounds were tested for their α-amylase and α-glucosidase inhibitory potency. Compound 1 showed significantly competitive inhibition against α-amylase (IC₅₀=17.5 μM) and α-glucosidase (IC₅₀=13.6 μM).     

Phytoconstituents of Selaginella effusa Alston and Their α-Glucosidase as well as Urease Inhibitory Activities

Three new compounds ( 1 – 2 , 14 ), as well as 22 known compounds ( 3 – 13 , 15 – 25 ), were extracted for the first time from the Selaginella effusa Alston (S. effusa). For the unknown compounds, the planar configurations were determined via NMR and by high-resolution mass spectrometry, while their absolute configurations were determined by calculated electronic circular dichroism (ECD), and the configuration of the stereogenic center of biflavones 4 – 5 were established for the first time. The pure compounds ( 1 – 25 ) were tested in vitro to determine the inhibitory activity of the enzyme-catalyzed reactions. Compounds 1 – 9 inhibited α-glucosidase with IC₅₀ values ranging from 0.30±0.02 to 4.65±0.04 μM and kinetic analysis of enzyme inhibition indicated that biflavones 1 – 3 were mixed-type α-glucosidase inhibitors. Compounds 12 – 13 showed excellent inhibitory activity against urease, with compound 12 (IC₅₀=4.38±0.31 μM) showing better inhibitory activity than the positive control drug AHA (IC₅₀13.52±0.61 μM). In addition, molecular docking techniques were used to simulate inhibitor-enzyme binding and to estimate the binding posture of the α-glucosidase and urease catalytic sites.     

CX3CL1 induces cell migration and invasion through ICAM-1 expression in oral squamous cell carcinoma cells

Human oral squamous cell carcinoma (OSCC) has been associated with a relatively low survival rate over the years and is characterized by a poor prognosis. C-X3-C motif chemokine ligand 1 (CX3CL1) has been involved in advanced migratory cells. Overexpressed CX3CL1 promotes several cellular responses related to cancer metastasis, including cell movement, migration and invasion in tumour cells. However, CX3CL1 controls the migration ability, and its molecular mechanism in OSCC remains unknown. The present study confirmed that CX3CL1 increased cell movement, migration and invasion. The CX3CL1-induced cell motility is upregulated through intercellular adhesion molecule-1 (ICAM-1) expression in OSCC cells. These effects were significantly suppressed when OSCC cells were pre-treated with CX3CR1 monoclonal antibody (mAb) and small-interfering RNA (siRNA). The CX3CL1-CX3CR1 axis activates promoted PLCβ/PKCα/c-Src phosphorylation. Furthermore, CX3CL1 enhanced activator protein-1 (AP-1) activity. The CX3CR1 mAb and PLCβ, PKCα, c-Src inhibitors reduced CX3CL1-induced c-Jun phosphorylation, c-Jun translocation into the nucleus and c-Jun binding to the ICAM-1 promoter. The present results reveal that CX3CL1 induces the migration of OSCC cells by promoting ICAM-1 expression through the CX3CR1 and the PLCβ/PKCα/c-Src signal pathway, suggesting that CX3CL1-CX3CR1-mediated signalling is correlated with tumour motility and appealed to be a precursor for prognosis in human OSCC.     

Identification and partial characterization of α2-macroglobulin from the tuatara (Sphenodon punctatus)

α₂-Macroglobulin (α₂-M), a large molecular mass proteinase-binding protein, was identified in plasma from tuatara (Sphenodon), a rare reptile endemic to New Zealand. In this genus, α₂-M constitutes 11–13% of total plasma protein (∼2.2–3.9 mg/ml). Analysis of blood samples collected at approximately monthly intervals from individual tuatara indicated that the plasma level of α₂-M remains fairly constant. The subunits of tuatara α₂-M have an apparent molecular mass of ∼160 kDa as determined by SDS-polyacrylamide gel electrophoresis and the intact protein is an oligomer that contains inter-chain disulfide bonds. N-terminal sequence analyses of tuatara α₂-M revealed a distinct similarity to α-macroglobulins of other vertebrates and that at least two types of α₂-M subunits are present in plasma of tuatara.     

PLD activation in Chinese hamster ovary (CHO) cells transfected with PGF2α receptor cDNA

Pertussis toxin-insensitive GTP-binding protein was observed to be involved in prostaglandin F2α(PGF2α)-induced phosphoinositide metabolism in Chinese hamster ovary (CHO) cells transfected with PGF2α receptor cDNA (CHO-PGF2α·R cells) (Ito, S. et al. Biochem. Biophys. Res. Commun. 200: 756, 1994). In the present study, we investigated PGF2α-induced PLD activation in CHO-PGF2α·R cells. PLD activation was examined by measuring the production of [³H]phosphatidylbutanol ([³H]PBut), a specific product of the PLD-catalyzed transphosphatidylation reaction. PGF2α-induced [³H]PBut formation was concentration-dependent with the maximal level obtained at 1 μM PGF2α. The maximal [³H]PBut formation was observed at 2 min after addition of PGF2α. Depletion of extracellular Ca²⁺ with EGTA suppressed PGF2α-induced PLD activation by 50%. PKC inhibitors Ro31–8425 and calphostin C inhibited PGF2α-induced [³H]PBut formation by 50%. PTK inhibitors genistein and herbimycin A failed to inhibit PGF2α-induced PLD activation. A combination of maximal effective concentrations of PGF2α (1 μM) and PMA (100 nM) enhanced PLD activation in an additive manner. Pretreatment of the cells with PMA for 2 h down-regulated PKCα and decreased PGF2α-induced PLD activation. These results suggest that PLD activation by PGF2α is mediated by both PKC-dependent and -independent pathways and that PKCα is involved in the former pathway.     

Biosynthesis, processing and release of pro-opiomelanocortin related peptides in the intermediate lobe of the pituitary gland of the frog (Rana ridibunda)

The biosynthesis of pro-opiomelanocortin (POMC) and related peptides by the intermediate lobe of the pituitary gland was studied in the frog Rana ridibunda using the pulse-chase technique. Analysis of radioactive proteins by dodecyl sulfate polyacrylamide gel electrophoresis showed that during pulse incubations a 36,000 dalton (36K) glycosylated prohormone was synthesized. It disappeared slowly during chase incubations, giving rise to another glycosylated protein (Mr 18K), identified as the N-terminal fragment of POMC. This latter protein was secreted to the incubation medium. High performance liquid chromatography analysis of peptides synthesized during chase incubations revealed the biosynthesis of two peptides related to gamma-MSH, three peptides related to alpha-MSH, one endorphin-related and one CLIP-related peptides. These newly synthesized peptides were slowly secreted to the incubation medium. Among the alpha-MSH related peptides, only the des-N alpha-acetyl alpha-MSH form of the peptide was found to be present within the cells, in contrast to the incubation medium where the presence of des-N alpha-acetyl alpha-MSH and a modified alpha-MSH was demonstrated.     

The distribution of alpha-melanocyte stimulating hormone (alpha-MSH) in the central nervous system of the rat: an immunohistochemical study. II. Lower brain stem

The distribution of immunoreactive alpha-melanocyte stimulating hormone (alpha-MSHI) in the rat lower brain stem was examined by indirect immunofluorescence or peroxidase- anti-peroxidase immunohistochemical method using an antiserum against synthetic alpha-MSH. The results confirmed the presence of alpha-MSHI fibers in the midbrain central gray matter and parabrachial area, and demonstrated a much more extensive distribution of these fibers in various parts of the lower brain stem areas previously thought not contain alpha-MSHI fibers. In addition, the commissural nucleus was identified as a new alpha-MSHI neurons-containing site. No alpha-MSHI neurons were seen in other regions of the rat lower brain stem.     

Quantification of the permeability of the blood-CSF barrier to alpha-MSH in the rat

The ability of alpha-MSH to cross the blood-CSF barrier of the rat was assessed by measurement of the rate of appearance of immunoreactive alpha-MSH in a cerebrospinal fluid (CSF) perfusate following intravenous injection of peptide. Comparisons were made with the rate of appearance of a simultaneously administered dose of 14C-inulin which is poorly permeable at the blood-CSF barrier. Concentrations of drugs measured in plasma were fitted to two-compartment pharmacokinetic models, and those measured in the CSF perfusate to one-compartment open systems receiving an input from the plasma compartment. The rate constant for entry of alpha-MSH into CSF was 0.00087 min-1, which was not significantly different from that for inulin of 0.00055 min-1. As alpha-MSH penetrated into CSF at a rate comparable to inulin, it was concluded that the limited entry of peptide was by aqueous diffusion along with other water-soluble macromolecules.     

The effect of chloroplast coupling factor ATP synthetase (CF1 · CF0) reconstitution on fluidity properties of isolated thylakoid lipid vesicles

The reconstitution of chloroplast coupling factor ATP synthetase (CF₁ · CF₀) with thylakoid lipids by cholate dialysis produced vesicles that displayed higher steady-state anisotropy (r_s) values for both 1,6-diphenyl-1,3,5-hexatriene (DPH) and trimethylammonium-diphenyl hexatriene fluorescence than the pure lipid alone. This is interpreted as meaning that the insertion of protein into the lipid bilayer brings about an increase in the ordering of acyl chains. This ordering effect became more obvious as the protein-to-lipid ratio was increased. Time-resolved decay analyses of DPH fluorescence anisotropy confirmed the conclusion drawn from the steady-state measurements, but further indicated that the dynamic motion of the probe was also slightly restricted after CF₁ · CF₀ incorporation. The restriction of DPH motion and the change in the half-angle for its cone of rotation was observed at relatively low protein-to-lipid ratios as compared with other reconstituted or biological membranes, suggesting that perhaps lipid-protein interactions occur with the inserted CF₁ · CF₀ complex.     

A 250-kilodalton cellular protein is induced by progestins in two human breast cancer cell lines MCF7 and T47D

We have studied the effect of R5020, a synthetic progestin, on the biosynthesis of cellular proteins extracted from the MCF7 and T47D human breast cancer cells, using gel electrophoresis. R5020 stimulates the synthesis, as measured after [35S]-methionine labelling, and the accumulation, as shown by silver staining, of a protein of molecular weight approximately equal to 250,000. The increase of the labelled 250-kilodalton protein was rapid (3 hours) and after 3 days this protein represented approximately equal to 6% of the total cellular proteins (approximately equal to 1 microgram/150,000 cells). The induction of the 250-kilodalton protein was obtained by physiologically active concentrations of several progestins and high concentrations of 5 alpha-dihydrotestosterone but not by estradiol or dexamethasone. It was inhibited by R486 , a progestin antagonist, but not by flutamide, an androgen antagonist. These results indicate a mediation by the progesterone receptor. The 250-kilodalton protein appears to be an excellent probe to study in cell culture the mechanism of action of progestin on human cells.