Multifarious Pharmacological Applications of Green Routed Eco-Friendly Iron Nanoparticles Synthesized by Streptomyces Sp. (SRT12)

Biological Trace Element Research - A simple, eco-friendly, green routine co-precipitation method was experimented to synthesize iron nanoparticles (Fe-NPs) using the cell-free supernatant of...     

Omeprazole attenuates hyperoxic injury in H441 cells via the aryl hydrocarbon receptor

Hyperoxia contributes to the development of bronchopulmonary dysplasia in premature infants. Earlier we observed that aryl hydrocarbon receptor (AhR)-deficient mice are more susceptible to hyperoxic lung injury than AhR-sufficient mice, and this phenomenon was associated with a lack of expression of cytochrome P450 1A enzymes. Omeprazole, a proton pump inhibitor used in humans with gastric acid-related disorders, activates AhR in hepatocytes in vitro. However, the effects of omeprazole on AhR activation in the lungs and its impact on hyperoxia-induced reactive oxygen species (ROS) generation and inflammation are unknown. In this study, we tested the hypothesis that omeprazole attenuates hyperoxia-induced cytotoxicity, ROS generation, and expression of monocyte chemoattractant protein-1 (MCP-1) in human lung-derived H441 cells via AhR activation. Experimental groups included cells transfected with AhR small interfering RNA (siRNA). Hyperoxia resulted in significant increases in cytotoxicity, ROS generation, and MCP-1 production, which were significantly attenuated with the functional activation of AhR by omeprazole. The protective effects of omeprazole on cytotoxicity, ROS production, and MCP-1 production were lost in H441 cells whose AhR gene was silenced by AhR siRNA. These findings support the hypothesis that omeprazole protects against hyperoxic injury in vitro via AhR activation that is associated with decreased ROS generation and expression of MCP-1.     

Synthesis of 2-substituted 17β-hydroxy/17-methylene estratrienes and their in vitro cytotoxicity in human cancer cell cultures

Synthesis of various types of 2-(alkylaminomethyl) and 2-(aroyl) 17β-estradiol analogs are reported. The synthesis of similar types of 2-substituted 17-methylene estratriene analogs was also achieved. Synthesis of chalcone derivatives of 17β-estradiol and 17-methylene estratriene were also realized. All these 2-substituted estratrienes were tested for their antiproliferative activity by using four different cell lines from colon, lung, glioma and breast cancers. Among the various 2-substituted estratrienes, the compounds 10d, 14a-h and 17e were found to have in vitro antiproliferative activity comparable to that of parent analogs 1-4. Comparison of the SAR pattern of these 2-susbtituted estratriene derivatives confirmed that relatively, 17-methylene estratrienes are more active than that of 17β-estradiol analogs.     

Discovery of [7-(2,6-dichlorophenyl)-5-methylbenzo [1,2,4]triazin-3-yl]-[4-(2-pyrrolidin-1-ylethoxy)phenyl]amine--a potent, orally active Src kinase inhibitor with anti-tumor activity in preclinical assays

We describe the identification of [7-(2,6-dichlorophenyl)-5-methylbenzo [1,2,4]triazin-3-yl]-[4-(2-pyrrolidin-1-ylethoxy)phenyl]amine (3), a potent, orally active Src inhibitor with desirable PK properties, demonstrated activity in human tumor cell lines and in animal models of tumor growth.     

Active Site Coupling in PDE:PKA Complexes Promotes Resetting of Mammalian cAMP Signaling

Cyclic 3′5′ adenosine monophosphate (cAMP)-dependent-protein kinase (PKA) signaling is a fundamental regulatory pathway for mediating cellular responses to hormonal stimuli. The pathway is activated by high-affinity association of cAMP with the regulatory subunit of PKA and signal termination is achieved upon cAMP dissociation from PKA. Although steps in the activation phase are well understood, little is known on how signal termination/resetting occurs. Due to the high affinity of cAMP to PKA (K_D ∼ low nM), bound cAMP does not readily dissociate from PKA, thus begging the question of how tightly bound cAMP is released from PKA to reset its signaling state to respond to subsequent stimuli. It has been recently shown that phosphodiesterases (PDEs) can catalyze dissociation of bound cAMP and thereby play an active role in cAMP signal desensitization/termination. This is achieved through direct interactions with the regulatory subunit of PKA, thereby facilitating cAMP dissociation and hydrolysis. In this study, we have mapped direct interactions between a specific cyclic nucleotide phosphodiesterase (PDE8A) and a PKA regulatory subunit (RIα isoform) in mammalian cAMP signaling, by a combination of amide hydrogen/deuterium exchange mass spectrometry, peptide array, and computational docking. The interaction interface of the PDE8A:RIα complex, probed by peptide array and hydrogen/deuterium exchange mass spectrometry, brings together regions spanning the phosphodiesterase active site and cAMP-binding sites of RIα. Computational docking combined with amide hydrogen/deuterium exchange mass spectrometry provided a model for parallel dissociation of bound cAMP from the two tandem cAMP-binding domains of RIα. Active site coupling suggests a role for substrate channeling in the PDE-dependent dissociation and hydrolysis of cAMP bound to PKA. This is the first instance, to our knowledge, of PDEs directly interacting with a cAMP-receptor protein in a mammalian system, and highlights an entirely new class of binding partners for RIα. This study also highlights applications of structural mass spectrometry combined with computational docking for mapping dynamics in transient signaling protein complexes. Together, these results present a novel and critical role for phosphodiesterases in moderating local concentrations of cAMP in microdomains and signal resetting.     

Effects of stress hormone cortisol on the mRNA expression of myogenenin,MyoD, Myf5, PAX3 and PAX7

The present investigation was carried out to evaluate the effect of stress hormone cortisol on the myogenic markers in the C2C12 cells co-cultured with 3T3-L1 preadipocytes. Co-culturing was achieved by transwell inserts with a 0.4 μm porous membrane. C2C12 and 3T3-L1 cells were grown independently on the transwell plates. After differentiation, inserts containing 3T3-L1 cells were transferred to C2C12 plates for co-culturing. 10 μg/μl of cortisol was added to the medium. After 72 h of treatment, C2C12 cells which were in the lower well were harvested for analysis. RT-PCR analysis of myogenic markers such as of myogenin, MyoD, Myf5, PAX3 and PAX7 showed a significant reduction in the mRNA expression of these myogenic markers. In addition, cortisol increased calpain activity, which led to accelerated protein degradation, which in turn reduced the myogenic rate. In conclusion, cortisol treatment reduced mRNA expression of myogenic markers in the co-cultured C2C12 cells, which is quite distinct from one dimensional mono-cultured C2C12 cells.     

IGF2BP1: a novel binding protein of p38 MAPK

Maneb (MB) and paraquat (PQ) provoke oxidative stress-mediated cell damage. Role of xanthine oxidase (XO) in oxidative stress and its association with nitric oxide (NO)/NO synthase (NOS) have been widely reported. While inducible NOS (iNOS) is implicated in MB+PQ-induced toxicity in rat polymorphonuclear leukocytes (PMNs), role of XO and its alliance with iNOS have not yet been established. The study investigated the role of XO in MB+PQ-induced oxidative stress in rat PMNs and its regulation by iNOS and inflammatory cytokines. MB+PQ-augmented reactive oxygen species (ROS), superoxide, nitro-tyrosine, lipid peroxidation (LPO), and nitrite levels along with the catalytic activity of iNOS, superoxide dismutase (SOD), and XO. XO inhibitor, allopurinol (AP), alleviated MB+PQ-induced changes except nitrite content and iNOS activity. Conversely, an iNOS inhibitor, aminoguanidine, mitigated MB+PQ-induced LPO, nitrite, iNOS, and nitro-tyrosine levels; however, no change was observed in ROS, SOD, and XO. Nuclear factor-κB inhibitor, pyrrolidine dithiocarbamate (PDTC), tumor necrosis factor-alpha (TNF-α) inhibitor, pentoxyfylline, and an anti-inflammatory agent, dexamethasone, attenuated MB+PQ-induced increase in XO, superoxide, and ROS with parallel reduction in the expression of interferon-gamma (IFN-γ), TNF-α, and interleukin-1β (IL-1β) in rat PMNs. Exogenous IFN-γ, TNF-α, and IL-1β enhanced superoxide, ROS, and XO in the PMNs of control and MB+PQ-treated rats; however, IFN- γ was found to be the most potent inducer. Moreover, AP ameliorated cytokine-induced free radical generation and restored XO activity towards normalcy. The results thus demonstrate that XO mediates oxidative stress in MB+PQ-treated rat PMNs via iNOS-independent but cytokine (predominantly IFN-γ)-dependent mechanism.     

All three domains of the Epstein-Barr virus-encoded latent membrane protein LMP-1 are required for transformation of rat-1 fibroblasts.

LMP-1, the Epstein-Barr virus latent membrane protein 1, is the only protein encoded by the virus that has been shown to have the properties of a transforming oncogene in rodent fibroblasts such as Rat-1 cells. LMP-1 is phosphorylated and proteolytically cleaved in Rat-1 cells in a manner similar to that seen in human lymphocytes. In this study, we demonstrate that all three major domains of LMP-1 (N-terminal, transmembrane, and C-terminal domains) are required for the ability to transform Rat-1 cells in culture, as assayed by loss of contact inhibition. This study is the first demonstration of a functional role for the C-terminal domain of LMP-1. Our analysis suggests that there are at least three distinct regions of the C terminus involved in signalling. Amino acids 306 to 334, which generate a toxic signal in the absence of amino acids 334 to 364, and the last 23 amino acids, 364 to 386, are essential for transformation. Biochemical analysis of the LMP-1 mutants with the three domains deleted indicate that the mutant N-terminal with the domain deleted is phosphorylated normally but is inefficiently cleaved compared with the wild-type LMP-1. The mutant with the transmembrane domain deleted is also phosphorylated but is not cleaved, showing that phosphorylation of LMP-1 does not require membrane association. The nontransforming mutant with the C-terminal domain deleted that lacks the last 23 amino acids is phosphorylated and cleaved. Therefore, these processing events alone are insufficient to generate a transforming signal.