Effect of Asparagus racemosus Extract on Transdermal Delivery of Carvedilol: A Mechanistic Study

This study was designed for investigating the effect of Asparagus racemosus (AR) extract and chitosan (CTN) in facilitating the permeation of carvedilol (CDL) across rat epidermis. Transdermal flux of carvedilol through heat-separated rat epidermis was investigated in vitro using vertical Keshary–Chien diffusion cells. Biophysical and microscopic manifestations of epidermis treated with AR extract, CTN, and AR extract–CTN mixture were investigated by using differential scanning calorimetry, transepidermal water loss, scanning electron microscopy (SEM), and transmission electron microscopy (TEM). Biochemical estimations of cholesterol, sphingosine, and triglycerides were carried out for treated excised as well as viable rat epidermis. The antihypertensive activity of the patches in comparison with that of oral carvedilol was studied in deoxycorticosterone acetate-induced hypertensive rats. The permeation of carvedilol across excised rat epidermis was significantly higher (p < 0.05) when AR extract, CTN, or AR extract–CTN mixture was used as donor vehicle as compared to propylene glycol/ethanol (7:3) mixture. Epidermis obtained after 12 h treatment of viable rat skin with AR extract–CTN mixture showed significantly higher (p < 0.05) permeability to CDL as compared to that after treatment with AR extract or CTN alone. Further, the application of patches containing AR extract–CTN mixture resulted in sustained release of CDL which was able to control the hypertension in deoxycorticosterone acetate-induced hypertensive rats through 36 h. Estimation of micro constituents in rat epidermis revealed maximum extraction of cholesterol, sphingosine, and triglycerides after treatment with AR extract–CTN mixture. This was manifested in altered lipid and protein-specific thermotropic transitions. Further, increase in intercellular space, disordered lipid structure, and corneocyte detachment as observed in SEM and TEM suggested great potential of AR extract for use as percutaneous permeation enhancer. The developed transdermal patches of CDL containing AR extract–CTN mixture exhibited better performance as compared to oral administration in controlling hypertension in rats.     

Smooth muscle contractility and protein tyrosine phosphorylation

During the last 5 years several studies have documented an involvement of protein tyrosine kinases (PTKs) in smooth muscle contraction and Ca²⁺mobilization. Most of these studies have utilized highly selective inhibitors of PTKs, genistein and tyrphostin and have shown that these inhibitors attenuated smooth muscle contraction induced by growth factors - epidermal growth factor (EGF) and platelet derived growth factor (PDGF) and several vasoactive peptides. It has also been demonstrated that inhibitors of protein tyrosine phosphatases (PTPases) such as vanadate and pervanadate mimic growth factors and vasoactive peptides in causing the contraction of smooth muscle. In this brief review, we have summarized some of the recent observations suggesting a possible link between protein tyrosine phosphorylation pathway and smooth muscle contraction.     

Primary structure of an invertebrate dihydrolipoamide dehydrogenase with phylogenetic relationship to vertebrate and bacterial disulfide oxidoreductases

Dihydrolipoamide dehydrogenase (E₃) is a flavoprotein component of multi-enzyme complexes catalyzing oxidative decarboxylation of -ketoacids in the Krebs' cycle. We have cloned a 2.4-kb E₃ cDNA from an arthropod, Manduca sexta, that codes for 497 amino acids and translates to a 51-kDa protein in vitro. Sequences at and around the dinucleotide binding domains, disulfide active site and the C-terminal interface domain involved in substrate binding are highly conserved in Manduca E₃. Phylogenetic analysis of protein sequences from the flavoprotein class of disulfide oxidoreductases family of enzymes suggests that in spite of the homologous nature of E₃ and glutathione reductase (goR) in sequence and structure, E₃ shares a common ancestor with mercuric reductase (merA), whereas goR is more related to trypanothione reductase (tryR) than to other members. All members, except goRs, seemed to be monophyletic. Plant goRs seemed to have arisen differently and are more closely related to tryRs than to bacterial and vertebrate goRs. Earlier speculation on the nature of origin of E₃ in Pseudomonas is not supported by phylogenetic data. A possible structural relationship of Manduca E₃ to other pyridine-binding proteins, such as the neurotransmitter transporters and channels, is proposed.     

Revised structures of daturalactone and 12-oxowithanolide

Recently Dhar et al. reported the isolation of two new steroidal lactones from the leaves of Datura quercifolia HBK and formulated them as 5α,12α,17α-trihydroxy-1-oxo-6α, 7α-epoxy-22 S-witha-2,24-dienolide and 5α,17α-dihydroxy-1, 12-dioxo-6α,7α-epoxy-22 S-witha-2,24-dienolide on the basis of UV, IR, NMR and MS studies. Further detailed chemical and spectral studies have led to revised structures, namely 5α, 12α-dihydroxy-1-oxo-6α,7α: 24α, 25α-diepoxy-20 S, 22 R-with-2-enolide and 5α-hydroxy-1, 12-dioxo-6α, 7α: 24α, 25α-diepoxy-20 S, 22 R-with-2-enolide, respectively, for the above two compounds.     

Photophysical behaviour of 1-(4-N,N-dimethylaminophenylethynyl)pyrene (DMAPEPy) in homogeneous media.

The photophysical behaviour of a new pyrene derivative, 1-(4-N,N-dimethylaminophenylethynyl)pyrene (DMAPEPy), in various solvents has been studied. Due to the presence of an ethynyl link with a cylindrical pi cloud between the donor (N,N-dimethyl group) and the acceptor (pyrene), the molecule shows efficient intramolecular charge transfer, with a high extinction coefficient in all the solvents. There is significant solvatochromism in the fluorescence with a large increase in the Stokes' shift of around 125 nm between n-hexane and acetonitrile. The solvent-dependent spectral data show a good correlation with the Kamlet-Taft solvent polarity parameter (pi*). The plots of Stokes' shifts with E(T)(30) are linear for non-protic solvents and for protic solvents but with different slopes. The fluorescence quantum yields are high for non-polar solvents and decrease as the solvent polarity increases. Unlike the parent molecule pyrene, DMAPEPy shows a short lifetime, which is fairly insensitive to oxygen-induced quenching and is dependent on solvent polarity. The molecule shows high steady-state fluorescence anisotropy, which is very sensitive to the viscosity change of the medium.     

Reduced LINE-1 methylation is associated with arsenic-induced genotoxic stress in children

Early life exposure to arsenic has profound effect towards development of arsenic induced toxic outcomes. Some districts in the state of West Bengal, India are highly affected by arsenic, mainly through ground water. In children, not much of the toxic outcomes like dermatological lesions are observed but it is thought that the exposure leads to transient alteration in their biological processes that leads to various deleterious health effects later on. We evaluated the global methylation status by analyzing the LINE-1 methylation profile in children from arsenic exposed region between the age group 5–15 years along with the cytogenetic stress induced by arsenic as measured by lymphocyte micronucleus (MN) frequency. A total of 52 arsenic exposed and 32 unexposed children were analyzed. Whole blood DNA was used to measure the LINE-1 methylation by qRT-MSP. We found a significant association of MN-frequency in exposed individuals with highly depleted LINE-1 methylation compared to the exposed individuals with near baseline (which was comparable to unexposed control) methylation index as well as with those with the hypermethylated LINE-1 promoters. From our results, we interpret that LINE-1 methylation index may serve as a potent global epigenetic mark to detect the degree of arsenic genotoxicity at a very early age. We propose that this may be utilized to determine the extent of toxic influence exerted by arsenic, from a very early age.     

Phenotypic consequences of transforming growth factor β1 gene ablation in murine embryonic fibroblasts: Autocrine control of cell proliferation and extracellular matrix biosynthesis

The profound effects of transforming growth factor β1 (TGF-β1) on the immune system, cardiogenesis, in yolk sac hematopoeisis and in differentiation of endothelium have been demonstrated by detailed analyses of TGF-β1 knockout mice during embryogenesis. We have systematically examined the autocrine and paracrine roles of TGF-β1 in cell proliferation and in its ability to modulate the gene expression of selected components of extracellular matrix (ECM) using embryonic fibroblasts from TGF-β1 null mice (TGF-β1^−/−). The rates of cell proliferation of embryonic fibroblasts from normal mice (TGF-β1^+/+) and TGF-β1 null mice were compared by cell counting, by ³H thymidine incorporation, and by measuring the fraction of cells in the G1, S, and G2/M phases of the cell cycle by fluorescent activated cell sorting (FACS). Concurrently, the expression of pro-α1(I) collagen, fibronectin, and plasminogen activator inhibitor-1 (PAI-1) was also quantified by hybridization of total mRNA from TGF-β1^+/+ and TGF-β1^−/− embryonic fibroblasts. We report that TGF-β1^−/− cells proliferated at about twice the rate of TGF-β1^+/+ cells. Further, TGF-β1 null fibroblasts accumulated and synthesized lower constitutive levels of pro-α1(I) collagen, fibronectin, and PAI-1 mRNA. The quantitative differences in the rates of cell proliferation and ECM gene expression between TGF-β1^+/+ and TGF-β1^−/− cells could be eliminated by treatment of TGF-β1^+/+ cells with a neutralizing antibody of TGF-β1. Thus, our results are consistent with the hypothesis that TGF-β1 acts as a negative autocrine regulator of growth and a positive autocrine regulator of ECM biosynthesis in embryonic fibroblasts. 176:67–75, 1998. Published 1998 Wiley-Liss, Inc.

1 This article was prepared by a group of United States government employees and non-United States government employees, and as such is subject to 17 U.S.C. Sec. 105.