Stimulation of osteoclastic bone resorption by hydrogen peroxide.

The molecular mechanisms underlying the pathophysiology of bone destruction still remain poorly understood. We have found that hydrogen peroxide (H2O2), a reactive oxygen species (ROS), is a potent stimulator of osteoclastic bone resorption and cell motility. A marked enhancement of bone resorption was noted when rat osteoclasts, cultured on devitalised bovine cortical bone, were exposed to 10 nM [H2O2]. Apart from exposing osteoclasts to a low extracellular pH, which is known to enhance osteoclastic bone resorption, we provide first evidence for a molecule that stimulates osteoclastic bone resorption in osteoclast cultures that do not respond to parathyroid hormone and 1, 25 dihydroxyvitamin D3. We envisage that both basic biological and practical clinical implications may eventually follow from these studies.     

Molecular species of cholesteryl esters formed in abetalipoproteinemia: effect of apoprotein B-containing lipoproteins

In order to study the effects of very low density (VLDL) and low density (LDL) lipoproteins on the activity and specificity of lecithin:cholesterol acyltransferase (LCAT), we determined the molecular species of cholesteryl esters (CE) synthesized in the plasma from three abetalipoproteinemic (ABL) patients, before and after supplementation with normal VLDL or LDL. The patients' plasma had significantly lower concentration of 18:2 CE and higher concentrations of 16:0 CE and 18:1 CE compared to normal plasma. Incubation of ABL plasma with [4-14C]cholesterol at 37 degrees C and the subsequent analysis of labeled CE formed by high performance liquid chromatography revealed that the major species formed was 16:0 CE (34% of total label), whereas similar incubation of the d greater than 1.063 g/ml fraction of normal plasma resulted in the formation of predominantly 18:2 CE (45% of total label). Addition of normal VLDL or LDL to ABL plasma stimulated the total LCAT activity by 30-80% and normalized the CE species synthesized. The LCAT activity of a normal d greater than 1.063 g/ml fraction also was stimulated by the normal VLDL or LDL, but there was no alteration in the species of CE formed. Most of the CE synthesized was found in the added VLDL or LDL with both ABL and normal plasma, indicating that the CE transfer (CET) activity was not affected in ABL plasma. These results suggest that while the VLDL and LDL are required for the maximal activity of LCAT, the species of CE formed are primarily determined by the molecular species composition of phosphatidylcholine in the plasma.     

Lipid A from endotoxin: antigenic activities of purified fractions in liposomes.

Isolation of lipid A by acid hydrolysis of Shigella flexneri lipopolysaccharide resulted in a product that consisted of a heterogeneous mixture of bands when visualized by thin layer chromatography. Differential extraction with ethyl acetate and chloroform, or extraction with EDTA, followed by chloroform-methanol-water (Bligh-Dyer extraction), or a combination of both extraction schemes, resulted in partial purification of immunologically active lipid A. Eight fractions were purified further by preparative thin layer chromatography, and each of the fractions had phosphate, carbohydrate, and esterified fatty acids. Upon incorporation into liposomes, five of the eight purified fractions reacted with anti-lipid A serum, but the three fractions with the most number of esterified fatty acids failed to react with anti-lipid A serum. At least one fraction that originally was unreactive with anti-lipid A serum became reactive as a hapten inhibitor upon removal of esterified fatty acids by alkaline hydrolysis. Alkali-treated fractions from "unreactive" and "reactive" lipid A had similar activities as hapten inhibitors. Our data suggest that lipid A can exist in multiple forms that differ by the number and placement, and possibly by the type, of fatty acids linked to the carbohydrate of lipid A. Highly acylated forms of lipid A do not react with antiserum against the unpurified lipid A mixture, but removal of fatty acids does expose immunoreactive groups.     

Partial inhibition of the decay of Hill activity in isolated chloroplasts by kinetin.

Kinetin when included in the grinding medium inhibits degeneration of isolated spinach chloroplasts. This is indicated by the relatively greater retention of Hill activity by chloroplasts isolated in the presence of Kinetin. It appears that the Kinetin effect is probably on the integrity of the chloroplasts membrane. The technique seems to be of great practical value in photosynthetic studies.     

Ester-to-amide rearrangement of ethanolamine-derived prodrugs of sobetirome with increased blood-brain barrier penetration

Current therapeutic options for treating demyelinating disorders such as multiple sclerosis (MS) do not stimulate myelin repair, thus creating a clinical need for therapeutic agents that address axonal remyelination. Thyroid hormone is known to play an important role in promoting developmental myelination and repair, and CNS permeable thyromimetic agents could offer an increased therapeutic index compared to endogenous thyroid hormone. Sobetirome is a clinical stage thyromimetic that has been shown to have promising activity in preclinical models related to MS and X-linked adrenoleukodystrophy (X-ALD), a genetic disease that involves demyelination. Here we report a new series of sobetirome prodrugs containing ethanolamine-based promoieties that were found to undergo an intramolecular O,N acyl migration to form the pharmacologically relevant amide species. Several of these systemically administered prodrugs deliver more sobetirome to the brain compared to unmodified sobetirome. Pharmacokinetic properties of the parent drug sobetirome and amidoalcohol prodrug 3 are described and prodrug 3 was found to be more potent than sobetirome in target engagement in the brain from systemic dosing.     

Neo-tanshinlactone D-ring modified novel analogues induce apoptosis in human breast cancer cell via DNA damage

Neo-tanshinlactone (NTL) a natural product is known for its specificity and selectivity towards the breast cancer cells. By NTL D-ring modification approach, 13 new analogues were synthesized (1A–1M). Among them 1J showed the best anticancer activity in MCF-7 (ER+, PR+/?, HER2?), SKBR3 (ER?, PR?, HER2+) and MDA-MB-231 (ER?, PR?, HER2?) cells lines with IC₅₀ value 11.98 nM, 23.71 nM, and 62.91 nM respectively. 1J showed minor grove binding interaction with DNA at AT-rich region and induced DNA double strand breaks (DDSBs). This had triggered several key molecular events involving, activation of ATM, Chk2 and p53, reduction in mitochondrial potential (Δψ_m) leading to caspase-3 and PARP cleavage mediated apoptosis. These results along with other biochemical studies strongly suggest that novel NTL analogue 1J caused DNA cleavage mediated apoptosis in the breast cancer cells and this may serve as potential lead for future breast cancer treatment.     

Modelling of Lipase Catalysed Transesterification Reactions

Transesterification of substituted ethanols with ethyl acetate is studied in the presence of two different lipases. The steric bulk of the substituent group plays a more important role than the electronic factors in the reaction. The reversible reaction is modelled mathematically, and the rate parameters in the model estimated by a transient parameter estimation procedure. The effect of lipase quantity, reaction temperature and substrate concentration on the reaction is studied independently. Product quantity exhibits a maxima for increasing substrate concentration which is modelled successfully with a substrate product inhibition model, and compared with the experimental data