Bis(η-allyl) (η-pentalene) zirconium: preparation, structure and structural dynamics

The preparation of a novel mononuclear complex of zirconium having an η⁸-bonded pentalene ligand and two η³-allyl groups is described. Its structure has been determined by ¹H and ¹³C NMR spectroscopy. At room temperature some of the NMR signals are broadened, revealing that the compound is structurally dynamic. It is shown that the compound has C₂ symmetry with the enantiomeric forms undergoing racemisation.     

Cardiovascular actions of prostaglandins F2α; 15-me-F2α; F1α; F2β and F1β in the cat

Prostaglandin F_2α (5μg/kg, i.v.) causes an increase in pulmonary arterial pressure, decrease in systemic arterial pressure, and reflex bradycardia in the anesthetized cat. The same dose of the 15-methyl analogue of PGF_2α produces the same triad of effects but of greater magnitude and duration. Although prostaglandins F_1α, F_2β and F_1β also cause the same cardiovascular effects as F_2α, there is a decrease in potency for all parameters measured, with PGF_2α>PGF_1α>PGF_2β>PGF_1β. When compared to the actions of PGF_2α in producing an increase in pulmonary arterial pressure, PGs F_1α, F_2β and F_1β were less potent by approximately 10, 100, and 1000 fold respectively.     

Antinoceptive effect of triterpenoid α,β-amyrin in rats on orofacial pain induced by formalin and capsaicin

The effects of α,β-amyrin, a pentacyclic triterpene isolated from Protium heptaphylum was investigated on rat model of orofacial pain induced by formalin or capsaicin. Rats were pretreated with α,β-amyrin (10, 30, and 100 mg/kg, i.p.), morphine (5 mg/kg, s.c.) or vehicle (3% Tween 80), before formalin (20 μl, 1.5%) or capsaicin (20 μl, 1.5 μg) injection into the right vibrissa. In vehicle-treated controls, formalin induced a biphasic nociceptive face-rubbing behavioral response with an early first phase (0–5 min) and a late second phase (10–20 min) appearance, whereas capsaicin produced an immediate face-rubbing (grooming) behavior that was maximal at 10–20 min. Treatment with α,β-amyrin or morphine significantly inhibited the face-rubbing response in both test models. While morphine produced significant antinociception in both phases of formalin test, α,β-amyrin inhibited only the second phase response, more prominently at 30 mg/kg, in a naloxone-sensitive manner. In contrast, α,β-amyrin produced much greater antinociceptive effect at 100 mg/kg in the capsaicin test, which was also naloxone-sensitive. These results provide first time evidence to show that α,β-amyrin attenuates orofacial pain atleast, in part, through a peripheral opioid mechanism but warrants further detailed study for its utility in painful orofacial pathologies.     

Phosphorylations of αA- and αB-crystallin

In addition to being refractive proteins in the vertebrate lens, the two α-crystallin polypeptides (αA and αB) are also molecular chaperones that can protect proteins from thermal aggregation. The αB-crystallin polypeptide, a functional member of the small heat shock family, is expressed in many tissues in a developmentally regulated fashion, is stress-inducible, and is overexpressed in many degenerative diseases and some tumors indicating that it plays multiple roles. One possible clue to α-crystallin functions is the fact that both polypeptides are phosphorylated on serine residues by cAMP-dependent and cAMP-independent mechanisms. The cAMP-independent pathway is an autophosphorylation that has been demonstrated in vitro, depends on magnesium and requires cleavage of ATP. Disaggregation of αA-, but not αB-crystallin into tetramers results in an appreciable increase in autophosphorylation activity, reminiscent of other heat shock proteins, and suggests the possibility that changes in the aggregation state of αA-crystallin are involved in yet undiscovered signal transduction pathways. The α-crystallin polypeptides differ with respect to their abilities to undergo cAMP-dependent phosphorylation, with preference given to the αB-crystallin chain. These differences and complexities in α-crystallin phosphorylations, coupled with the differences in expression patterns of the two α-crystallin polypeptides, are consistent with the idea that each polypeptide has distinctive structural and metabolic roles.     

η-Allyl-cobalt (II) complexes

(η³-Cyclooctenyl)Co(bisphosphine) compounds react with HBF₄ in the presence of alkenes with oxidation of the metal to give the novel, paramagnetic organocobalt(II) species [(η³-cyclooctenyl)Co(bisphosphine)]⁺BF₄⁻, (η³-2-RC₃H₄)Co(bisphosphine) complexes react similarly. The Co(II) compounds form adducts with CO and NO (the latter being diamagnetic) and undergo facile chemical and electrochemical reduction.     

α3β1 integrin promotes cell survival via multiple interactions between 14-3-3 isoforms and proapoptotic proteins

Laminin-5 and α3β1 integrin promote keratinocyte survival; however, the downstream signaling pathways for laminin-5/α3β1 integrin-mediated cell survival had not been fully established. We report the unexpected finding of multiple interactions between 14-3-3 isoforms and proapoptotic proteins in the survival signaling pathway. Ln5-P4 motif within human laminin-5 α3 chain promotes cell survival and anti-apoptosis by inactivating Bad and YAP. This effect is achieved through the formation of 14-3-3ζ/p-Bad and 14-3-3σ/p-YAP complexes, which is initiated by α3β1 integrin and FAK/PI3K/Akt signaling. These complexes result in cytoplasmic sequestration of Bad and YAP and their subsequent inactivation. An increase in Akt1 activity in cells induces 14-3-3ζ and σ, p-Bad, and p-YAP, promoting cell survival, whereas decreasing Akt activity suppresses the same proteins and inhibits cell survival. Suppression of 14-3-3ζ with RNA-interference inhibits cell viability and promotes apoptosis. These results reveal a new mechanism of cell survival whereby the formation of 14-3-3ζ/p-Bad and 14-3-3σ/p-YAP complexes is initiated by laminin-5 stimulation via the α3β1 integrin and FAK/PI3K/Akt signaling pathways, thereby resulting in cell survival and anti-apoptosis.     

ILSMRs (intensifier of β-lactam-susceptibility in methicillin-resistant Staphylococcus aureus) from Tara [Caesalpinia spinosa (Molina) Kuntze]

Four quinic acid gallates were isolated from the dried pods of Tara [Caesalpinia spinosa (Molina) Kuntze]. These compounds intensified the susceptibility of methicillin-resistant Staphylococcus aureus (MRSA) to oxacillin. 3,4,5-Tri-O-galloylquinic acid methyl ester (2) was the most effective compound of them.     

γ -Ray-Induced DNA Damage and Repair in Methanosarcina barkeri

The capacity of the mesophilic archaeon, Methanosarcina barkeri (DSM 804) for DNA double strand break repair following⁶⁰Co- γ irradiation was investigated. The genome (1.9 Mb) of Methanosarcina barkeri was largely fragmented and was found to be repaired on incubation in medium under anaerobic conditions at 37°C for 4 h. To get an insight into its repair process a set of inhibitors were used. The methanogenesis inhibitor, bromoethanesulfonate showed partial inhibition of repair in Methanosarcina barkeri but not in Escherichia coli or human peripheral blood mononuclear cells. The Methanosarcina barkeri cells could also partially repair the DNA damage in a non-nutrient medium. Arabinosine-CTP, a nucleoside analogue and a polymerase inhibitor, completely inhibited repair in this archaeon. Arabinosine-CTP did not affect DSB (double-strand break) repair in human peripheral blood mononuclear cells but completely inhibited repair in Escherichia coli (a bacterium). The involvement of polymerase indicates recombination to be the underlying mechanism in DSB repair of Methanosarcina barkeri. 3-Aminobenzamide, a poly (ADP-ribose) polymerase inhibitor, completely inhibited repair in this archaeon as well as in eukarya but not in Escherichia coli showing the involvement of poly (ADP-ribose) polymerase in the DSB repair of Methanosarcina barkeri.     

β-secretase (BACE1)-inhibiting stilbenoids from Smilax Rhizoma

In the course of searching for BACE1 (beta-secretase) inhibitors from natural products, the ethyl acetate soluble fraction of Smilax Rhizoma (the dried rhizomes of Smilax china L.) showed potent inhibitory activity. The active compounds were identified as a trans/cis-resveratrol mixture, oxyresveratrol, veraphenol, and cis-scirpusin A. They were shown to non-competitively inhibit BACE1 with the Ki values of 5.4 x 10(-6), 5.4 x 10(-6), 3.4 x 10(-6), and 5.4 x 10(-6)M and IC(50) values of 1.5 x 10(-5), 7.6 x 10(-6), 4.2 x 10(-6), and 1.0 x 10(-5)M, respectively. The active compounds were less inhibitory to alpha-secretase (TACE) and other serine proteases such as chymotrypsin, trypsin, and elastase, suggesting that they were relatively specific inhibitors of BACE1.     

Differential steric effects in the axial ligation of pyridine and imidazole to α- and β-alkylcobinamides

One subclass of B₁₂-requiring enzymes is now known to bind their B₁₂ coenzymes “base-off,” with a histidine residue from the protein supplying an imidazole ligand to the cobalt center. Recent results from Sirovatka and Finke (J.M. Sirovatka and R.G. Finke, J.Am. Chem. Soc. 119, (1997) 3057) show that imidazole has an extraordinary trans effect on the mode of carbon–cobalt bond cleavage in coenzyme B₁₂ analogs, compared to pyridine or the natural 5,6-dimethylbenzimidazole ligand, and it was suggested that a differential steric effect could, in part, account for the uniqueness of the imidazole ligand. Such a differential steric effect for imidazole and pyridine is now demonstrated by studies of the thermodynamics of ligation of these ligands to the α and β diastereomers of two alkylcobinamides (RCbi⁺s, derivatives of cobalamins which lack the normal axial nucleotide) based on the known differences in steric crowding of the α (“lower”) and β (“upper”) axial ligand positions of cobalt corrinoids. Imidazole binds more tightly than pyridine to both diastereomers of NCCH₂Cbi⁺ and CF₃Cbi⁺, in all cases due to a more favorable entropy change, which is the result of lowered steric interference with corrin side chain thermal motions.     

Bioactive and osteoblast cell attachment studies of novel α- and β-chitin membranes for tissue-engineering applications

Chitin is a novel biopolymer and has excellent biological properties such as biodegradation in the human body and biocompatible, bioabsorable, antibacterial and wound healing activities. In this work, α- and β-chitin membranes were prepared using α- and β-chitin hydrogel. The bioactivity studies were carried out using these chitin membranes with the simulated body fluid solution (SBF) for 7, 14 and 21 days. After 7, 14 and 21 days the membranes were characterized using SEM, EDS and FT-IR. The SEM, EDS and FT-IR studies confirmed the formation of calcium phosphate layer on the surface of the both chitin membranes. These results indicate that the prepared chitin membranes were bioactive. Cell adhesion studies were also carried out using MG-63 osteoblast-like cells. The cells were adhered and spread over the membrane after 24 h of incubation. These results indicated that the chitin membranes could be used for tissue-engineering applications.     

Organization, sequence and nuclease hypersensitivity of repetitive elements flanking the chicken apoVLDLII gene: extended sequence similarity to elements flanking the chicken vitellogenin gene.

Analysis of nuclease hypersensitivity of regions flanking the estrogen-dependent, chicken apoVLDLII gene has revealed an hepatic, DNaseI hypersensitive site whose sensitivity is influenced by both the developmental stage and sex of the bird. The site is located 3.0kb upstream from the gene, in a block of middle repetitive elements. Contact hybridization studies indicate that the block consists of contiguous copies of two elements with reiteration frequencies of 500-1000 and 10,000-30,000 copies per haploid genome. Sequencing of 1.8kb spanning the repeats has revealed that the higher frequency element is a member of the CR1 family. The adjacent lower frequency repeat can also be found next to another member of the CR1 family located in the 3' flanking region of the vitellogenin gene. The hypersensitive site has been mapped to one of the two most highly conserved regions of the CR1 element. This region displays homology with a silencer sequence recently identified in a CR1 element flanking the chicken lysozyme gene.     

Antibacterial activity of α-mangostin against vancomycin resistant Enterococci (VRE) and synergism with antibiotics

alpha-Mangostin, isolated from the stem bark of Garcinia mangostana L., was found to be active against vancomycin resistant Enterococci (VRE) and methicillin resistant Staphylococcus aureus (MRSA), with MIC values of 6.25 and 6.25 to 12.5 microg/ml, respectively. Our studies showed synergism between alpha-mangostin and gentamicin (GM) against VRE, and alpha-mangostin and vancomycin hydrochloride (VCM) against MRSA. Further studies showed partial synergism between alpha-mangostin and commercially available antibiotics such as ampicillin and minocycline. These findings suggested that alpha-mangostin alone or in combination with GM against VRE and in combination with VCM against MRSA might be useful in controlling VRE and MRSA infections.     

Separation and concentration of Δ-6-keto-PGF1α using monoclonal antibody to ω3-olefin structure of trienoic prostanoids

A monoclonal antibody against cis-3-hexen-1-ol was prepared and used to separate and/or concentrate Δ¹⁷-6-keto-prostaglandin F_1α (PGF_1α) in the human sera. cis-3-Hexen-1-ol was conjugated with the human serum albumin (HSA) according to the N-succinimidylester method and hyperimmunized to BALB/c mouse. The monoclonal afntibodies were obtained from hybridoma clones established by a fusion between SP2/0-Ag14-k13 mouse myeloma cells and splenocytes of a mouse. A monoclonal antibody, named 4G9-12B, recognized the epitope characteristic for ω3-olefin structure. The 4G9-12B antibody became more specific for Δ¹⁷-6-keto-PGF_1α than 6-keto-PGF_1α by applying inhibition ELISA using amino-residue coating plates. Using the prepared immunoaffinity columns of this antibody, Δ¹⁷-6-keto-PGF_1α was clearly detected in 6 pg/ml of the human blood sera by GC/MS analysis. These results suggest that the monoclonal antibody to the partial structure of trienoic prostanoid, ω3-olefin unit, and that its immunoaffinity columns are useful in separating and concentrating Δ¹⁷-6-keto-PGF_1α in the human blood or urine.     

Monitoring detrital input and resuspension effects on sediment trap material using mineralogy and stable isotopes (δO and δC): the case of Lake Neuchâtel (Switzerland)

Isotopic analyses on bulk carbonates are considered a useful tool for palaeoclimatic reconstruction assuming calcite precipitation occurring at oxygen isotope equilibrium with local water and detrital carbonate input being absent or insignificant. We present results from Lake Neuchâtel (western Switzerland) that demonstrate equilibrium precipitation of calcite, except during high productivity periods, and the presence of detrital and resuspended calcite. Mineralogy, geochemistry and stable isotope values of Lake Neuchâtel trap sediments and adjacent rivers suspension were studied. Mineralogy of suspended matter in the major inflowing rivers documents an important contribution of detrital carbonates, predominantly calcite with minor amounts of dolomite and ankerite. Using mineralogical data, the quantity of allochthonous calcite can be estimated by comparing the ratio ankerite + dolomite/calcite + ankerite + dolomite in the inflowing rivers and in the traps. Material taken from sediment traps shows an evolution from practically pure endogenic calcite in summer (10–20% detrital material) to higher percentages of detrital material in winter (up to 20–40%). Reflecting these mineralogical variations, δ¹³C and δ¹⁸O values of calcite from sediment traps are more negative in summer than in winter times. Since no significant variations in isotopic composition of lake water were detected over one year, factors controlling oxygen isotopic composition of calcite in sediment traps are the precipitation temperature, and the percentage of resuspended and detrital calcite. Samples taken close to the river inflow generally have higher δ values than the others, confirming detrital influence. SEM and isotopic studies on different size fractions (<2, 2–6, 6–20, 20–60, >60 μm) of winter and summer samples allowed the recognition of resuspension and to separate new endogenic calcite from detrital calcite. Fractions>60 and <2 μm have the highest percentage of detritus. Fractions 2–6 and 6–20 μm are typical for the new endogenic calcite in summer, as given by calculations assuming isotopic equilibrium with local water. In winter such fractions show similar values than in summer, indicating resuspension. Using the isotopic composition of sediment traps material and of different size fractions, as well as the isotopic composition of lake water, the water temperature measurements and mineralogy, we re-evaluated the bulk carbonate potential for palaeoclimatic reconstruction in the presence of detrital and re-suspended calcite. This re-evaluation leads to the following conclusion: (1) the endogenic signal can be amplified by applying a particle-size separation, once the size of endogenic calcite is known from SEM study; (2) resuspended calcite does not alter the endogenic signal, but it lowers the time resolution; (3) detrital input decreases at increasing distances from the source, and it modifies the isotopic signal only when very abundant; (4) influence of detrital calcite on bulk sediment isotopic composition can be calculated.     

Transformation of 3-hydroxy-steroids by Fusarium moniliforme 7α-hydroxylase

Transformation of physiologically important 3-hydroxy-steroids by the DHEA-induced 7α-hydroxylase of F. moniliforme was investigated. Whereas DHEA was almost totally 7α-hydroxylated, PREG, EPIA and ESTR were only partially converted into their 7α-hydroxylated derivatives because hydroxylation at other undetermined positions as well as reduction of ketone at C17 or C20 into hydroxyl also occurred. Cholesterol was not transformed by the enzyme. Kinetic parameters of the 7α-hydroxylation for these substrates were determined and confirmed that DHEA was the best substrate of the 7α-hydroxylase. Inhibition studies of DHEA 7α-hydroxylation by the other 3-hydroxy-steroids were also carried out and proved that DHEA, PREG, EPIA and ESTR shared the same active site of the enzyme. Induction effects of these steroids were compared, and DHEA appeared to be the best inducer of the 7α-hydroxylase of F. moniliforme.     

Integrins and the activation of latent transforming growth factor β1 – An intimate relationship

Integrins are crucial for the ability of cells to sense mechanical perturbations and to transmit intracellular stress to their environment. We here review the more recently discovered role of integrins in activating the pleiotrophic cytokine transforming growth factor beta 1 (TGF-β1). TGF-β1 controls tissue homeostasis in embryonic and normal adult tissues and contributes to the development of fibrosis, cancer, autoimmune and vascular diseases when being mis-regulated. In most of these conditions, active TGF-β1 is generated by dissociation from a large latent protein complex that sequesters latent TGF-β1 in the extracellular matrix (ECM). Two main models are proposed how integrins contribute to latent TGF-β1 activation: (1) In a protease-dependent mechanism, integrins αvβ8 and αvβ3 are suggested to simultaneously bind the latent TGF-β1 complex and proteinases. This close vicinity at the cell surface improves enzymatic cleavage of the latent complex to release active TGF-β1. (2) Integrins αvβ3, αvβ5, αvβ6, and αvβ8 appear to change the conformation of the latent TGF-β1 complex by transmitting cell traction forces. This action requires association of the latent complex with a mechanically resistant ECM and is independent from proteolysis. Understanding that different integrins use different mechanisms to activate latent TGF-β1 opens new possibilities to develop cell-specific therapeutic strategies for TGF-β1-induced pathologies.     

A synthetic combination of mutations, including fs(1)pyr? Su(b) , r? Su(b) and b , causes female sterility and reduces embryonic viability in Drosophila melanogaster

A Drosophila melanogaster mutant, fs(1)pyr ^Su(b) , carrying a mutation that maps to the tip of the X chromosome, has been isolated. The mutation, when present alone, does not confer a detectable phenotype. However, this mutation causes female sterility and reduces embryonic viability when combined with mutations which deregulate the pyrimidine and β-alanine pools. Embryos that are homozygous for the mutations fs(1)pyr ^Su(b) , r ^Su(b) [previously designated as Su(b)] and b, and originate from a female parent homozygous for the three mutations show severely reduced viability. Newly laid eggs begin development normally, but the majority of the embryos die just before the eggs are due to hatch. Received: 15 May 1998 / Accepted: 18 January 1999     

Neuro-steroids: 3β-hydroxy-Δ5-derivatives in the rodent brain

Pregnenolone, dehydroepiandrosterone and their sulfate esters have been characterized in the rat brain. Their formation or accumulation depend on in situ mechanisms unrelated to the peripheral endocrine glands. Although their functions are still poorly understood, they may affect the brain by metabolism to sex steroid hormones and they may be functionally related to sexual behavior, possibly through direct modulations of the firing rates of neurons.