Ginseng increases intestinal elimination of albendazole sulfoxide in the rat

Herbal products show potential drug interactions, some of them with adverse effects. The main aim of this work was to study the effect of Panax ginseng on the intestinal elimination of the benzimidazole derivative albendazole sulfoxide (ABZSO). An upper small intestine segment was isolated and perfused in situ with saline, while ABZSO solution (10 mg/kg i.v.) was administered intravenously. Blood samples and intestinal secretion were collected over 60 min and analysed by HPLC. The intestinal clearance of ABZSO was 0.106+/-0.010 ml/min. Systemic co-administration of ginseng (10 mg/kg i.v.) increased significantly (P<0.05) the clearance of ABZSO (0.132+/-0.005 ml/min). The increase in ABZSO elimination could be the result of the effect of ginseng on metabolic pathways. These results highlight the interactions between herbal products (sometimes dietary constituents) and drugs such as benzimidazoles, since ginseng modifies the luminal clearance of this anthelminthic drug and could potentially interfere with drugs that undergo the same intestinal processes.     

Disruption of the endocytic protein HIP1 results in neurological deficits and decreased AMPA receptor trafficking

Huntingtin interacting protein 1 (HIP1) is a recently identified component of clathrin-coated vesicles that plays a role in clathrin-mediated endocytosis. To explore the normal function of HIP1 in vivo, we created mice with targeted mutation in the HIP1 gene (HIP1(-/-)). HIP1(-/-) mice develop a neurological phenotype by 3 months of age manifest with a failure to thrive, tremor and a gait ataxia secondary to a rigid thoracolumbar kyphosis accompanied by decreased assembly of endocytic protein complexes on liposomal membranes. In primary hippocampal neurons, HIP1 colocalizes with GluR1-containing AMPA receptors and becomes concentrated in cell bodies following AMPA stimulation. Moreover, a profound dose-dependent defect in clathrin-mediated internalization of GluR1-containing AMPA receptors was observed in neurons from HIP1(-/-) mice. Together, these data provide strong evidence that HIP1 regulates AMPA receptor trafficking in the central nervous system through its function in clathrin-mediated endocytosis.     

How rational should bioethics be? The value of empirical approaches

Rational justification of claims with empirical content calls for empirical and not only normative philosophical investigation. Empirical approaches to bioethics are epistemically valuable, i.e., such methods may be necessary in providing and verifying basic knowledge about cultural values and norms. Our assumptions in moral reasoning can be verified or corrected using these methods. Moral arguments can be initiated or adjudicated by data drawn from empirical investigation. One may argue that individualistic informed consent, for example, is not compatible with the Asian communitarian orientation. But this normative claim uses an empirical assumption that may be contrary to the fact that some Asians do value and argue for informed consent. Is it necessary and factual to neatly characterize some cultures as individualistic and some as communitarian? Empirical investigation can provide a reasonable way to inform such generalizations. In a multi-cultural context, such as in the Philippines, there is a need to investigate the nature of the local ethos before making any appeal to authenticity. Otherwise we may succumb to the same ethical imperialism we are trying hard to resist. Normative claims that involve empirical premises cannot be reasonable verified or evaluated without utilizing empirical methods along with philosophical reflection. The integration of empirical methods to the standard normative approach to moral reasoning should be reasonably guided by the epistemic demands of claims arising from cross-cultural discourse in bioethics.     

Association of insulin receptor substrate 1 (IRS-1) y895 with Grb-2 mediates the insulin signaling involved in IRS-1-deficient brown adipocyte mitogenesis

We have recently generated immortalized fetal brown adipocyte cell lines from insulin receptor substrate 1 (IRS-1) knockout mice and demonstrated an impairment in insulin-induced lipid synthesis as compared to wild-type cell lines. In this study, we investigated the consequences of IRS-1 deficiency on mitogenesis in response to insulin. The lack of IRS-1 resulted in the inability of insulin-stimulated IRS-1-deficient brown adipocytes to increase DNA synthesis and enter into S/G2/M phases of the cell cycle. These cells showed a severe impairment in activating mitogen-activated protein kinase kinase (MEK1/2) and p42-p44 mitogen-activated protein kinase (MAPK) upon insulin stimulation. IRS-1-deficient cells also lacked tyrosine phosphorylation of SHC and showed no SHC-Grb-2 association in response to insulin. The mitogenic response to insulin could be partially restored by enhancing IRS-2 tyrosine phosphorylation and its association with Grb-2 by inhibition of phosphatidylinositol 3-kinase activity through a feedback mechanism. Reconstitution of IRS-1-deficient brown adipocytes with wild-type IRS-1 restored insulin-induced IRS-1 and SHC tyrosine phosphorylation and IRS-1-Grb-2, IRS-1-SHC, and SHC-Grb-2 associations, leading to the activation of MAPK and enhancement of DNA synthesis. Reconstitution of IRS-1-deficient brown adipocytes with the IRS-1 mutant Tyr895Phe, which lacks IRS-1-Grb-2 binding, restored SHC-IRS-1 association and SHC-Grb-2 association. However, the lack of IRS-1-Grb-2 association impaired MAPK activation and DNA synthesis in insulin-stimulated mutant cells. These data provide strong evidence for an essential role of IRS-1 and its direct association with Grb-2 in the insulin signaling pathway leading to MAPK activation and mitogenesis in brown adipocytes.     

Environmental reprogramming of the expression of protein kinase CK2β subunit in fish

Interleukin-8 (IL-8) is considered as the major polymorphonuclear neutrophils (PMNs) chemoattractant cytokine in lung diseases such as asthma and adult respiratory distress syndrome (ARDS). However, controversial results were obtained regarding the involvement of IL-8 in the pathogenesis of pneumonia. This study examines the role of IL-8 in the recruitment and activation of PMNs in the lung of pneumonia patients. The interesting aspect of this study is that it is a site- specific analysis of the infected and uninfected lungs of the same patient. The level of IL-8 mRNA, protein and myeloperoxidase present in the cells of the bronchioalveolar lavages (BALs) taken from the areas of known pneumonic consolidations on chest X-ray (infected lung) are compared with the BALs obtained from areas of no obvious infiltrate (non-infected lung). The results obtained from the infected and non-infected lungs of pneumonic patients were further compared with that of a control group of non-smoking patients. The level of IL-8 mRNA and protein were determined by RT-PCR and ELISA respectively. There was a significant increase in the level of IL-8 mRNA in the infected lung as compared to its level in the non-infected lung (p < 0.001). In correlation with the increase in mRNA, IL-8 protein concentrations in BAL fluids from the infected lung were 6 fold higher than those taken from the non-infected lung (p < 0.0001). This pattern was also consistent with MPO activity in the BALs (4.5 fold more MPO activity in the infected lung as compared to that of the non-infected lung), indicating that IL-8 is directly implicated in neutrophil accumulation that follows acute respiratory infection. The results of the present study, therefore, indicate the involvement of IL-8 in the pathogenesis of pneumonia.     

The membrane transport factor p115 recycles only between homologous compartments in intact heterokaryons

Cytosolic proteins that participate in membrane traffic are assumed to be recruited from the cytosol onto specific membrane sites where they perform their function, and then released into cytosol before rebinding to catalyze another round of transport. To examine whether the ER to Golgi transport factor p115 recycles through release into a cytosolic pool, we formed heterokaryons between rat NRK and simian COS-7 cells and examined the dynamics of rat p115 transfer from the rat to the simian portion of the heterokaryon. The heterokaryons shared a common cytosolic pool, as shown by the efficient relocation of a cytosolic green fluorescent protein (GFP) from the COS-7 to the NRK part of the heterokaryon. Unexpectedly, even 24 h after cell fusion, rat p115 did not redistribute to the COS-7 part of the heterokaryon. This was not due to the inability of the rat p115 to associate with simian membranes since rat p115 expressed in COS-7 cells was efficiently targeted to and associated with simian Golgi complex. Furthermore, rat p115 associated with heterologous simian membranes after the NRK and COS-7 Golgi fused into a single chimeric structure. Our results indicate that p115 is not freely diffusible in intact cells and might remain tethered to membranes throughout its life cycle. These findings suggest that p115, and perhaps other cytosolic proteins involved in membrane traffic, recycle not by being released into cytosol, but in association with recycling membranes.     

The crystal structure of tetrameric methionine adenosyltransferase from rat liver reveals the methionine-binding site

Most of the transmethylation reactions use the same methyl donor, S-adenosylmethionine (SAM), that is synthesised from methionine and ATP by methionine adenosyltransferase (MAT). In mammals, two MAT enzymes have been detected, one ubiquitous and another liver specific. The liver enzyme exists in two oligomeric forms, a tetramer (MAT I) and a dimer (MAT III), MAT I being the one that shows a higher level of affinity for methionine but a lower SAM synthesis capacity. We have solved the crystal structure of rat liver MAT I at 2.7 A resolution, complexed with a methionine analogue: l-2-amino-4-methoxy-cis-but-3-enoic acid (l-cisAMB). The enzyme consists of four identical subunits arranged in two tight dimers that are related by crystallographic 2-fold symmetry. The crystal structure shows the positions of the relevant cysteine residues in the chain, and that Cys35 and Cys61 are perfectly oriented for forming a disulphide link. This result leads us to propose a hypothesis to explain the control of MAT I/III exchange and hence, the effects observed on activity. We have identified the methionine-binding site into the active-site cavity, for the first time. The l-cisAMB inhibitor is stacked against Phe251 aromatic ring in a rather planar conformation, and its carboxylate group coordinates a Mg(2+), which, in turn, is linked to Asp180. The essential role of the involved residues in MAT activity has been confirmed by site-directed mutagenesis. Phe251 is exposed to solvent and is located in the beginning of the flexible loop Phe251-Ala260 that is connecting the N-terminal domain to the central domain. We postulate that a conformational change may take place during the enzymatic reaction and this is possibly the reason of the unusual two-step mechanism involving tripolyphosphate hydrolysis. Other important mechanistic implications are discussed on the light of the results. Moreover, the critical role that certain residues identified in this study may have in methionine recognition opens further possibilities for rational drug design.     

Calpain-3 gene expression is decreased during experimental cancer cachexia

The Yoshida AH-130 rat ascites hepatoma is a model system for studying the mechanisms involved in the protein hypercatabolism associated with cancer cachexia. The present study was aimed at investigating if the calpain-3 gene expression in skeletal muscle was affected by tumor growth. The results presented clearly show that calpain-3 gene expression is considerably reduced in experimental cancer cachexia, while there is a reciprocal change in the expression of the ubiquitin-dependent proteolytic system and in the ubiquitous m-calpain. The results, observed during cancer cachexia, suggest a potential counterregulatory role of calpain-3 in muscle proteolysis.     

Mitochondrial DNA haplotyping of Testudo graeca on both continental sides of the Straits of Gibraltar

Testudo graeca is an endangered species of tortoise that inhabits Mediterranean areas of Africa, Asia, and Europe. Western populations are found on both sides of the Straits of Gibraltar. The effects of geographical isolation on genetic divergence were assessed by the sequence analysis of two mitochondrial DNA regions of the 12S rRNA and cytochrome b genes. Four different haplotypes were identified. A single haplotype was shared by all Spanish and some east Moroccan specimens. Two haplotypes were unique to the west Moroccan T. graeca populations and allowed the clear discrimination between individual specimens found west of the Moulouya River. Phylogenetic analysis based on the estimation of nucleotide sequence distances of the haplotypes suggests an African origin for the Spanish populations and a subspecies status for the west Moroccan pool.     

Effect of trehalose and EDTA on cryoprotective action of ram semen diluents

To obtain better ram semen extenders for artificial insemination (AI), we developed effective trehalose-containing hypertonic diluents. The cryoprotective action of trehalose has been explained by its dehydrating activity and interaction with cell membranes. Accordingly, we tested the cryopreserving capacity of different combinations of a Salamon's modified plus trehalose extender with EDTA. Evaluations were based on the percentage of motile spermatozoa and acrosome integrity, measured after thawing and after a 4-h post-thaw resistance test at 37 degrees C. We conclude that the combination of trehalose plus EDTA confers the highest cryopreserving activity tested, not only for freeze-thawing but also for post-thawing resistance, possibly by removing calcium from the medium thereby preventing cation competition with trehalose for membrane-binding sites.