BIOCHEMICAL PHARMACOLOGY

Resveratrol is a natural molecule with antioxidant action. Moreover, resveratrol is also considered to be a molecule with anti-inflammatory action, an effect attributed to suppression of prostaglandin (PG) biosynthesis. The aim of the present study was to investigate the effects of resveratrol, a polyphenol present in most red wines, on reactive oxygen species formation as well as on arachidonic acid (AA) release, cyclooxygenase expression, and PG synthesis in murine resident peritoneal macrophages. Results show that resveratrol exerted a strong inhibitory effect on superoxide radical (O2−) and hydrogen peroxide (H2O2) produced by macrophages stimulated by lipopolysaccharides (LPS) or phorbol esters (PMA). Resveratrol also significantly decreased [3H]AA release induced by LPS and PMA or by exposure to O2− or H2O2. Resveratrol treatment caused a significant impairment of cyclooxygenase-2 (COX-2) induction stimulated by LPS and PMA or by O2− or H2O2 exposure. These effects of resveratrol on [3H]AA release and COX-2 overexpression were correlated with a marked reduction of PG synthesis. Our results indicate that the antioxidant action of resveratrol affects AA mobilization and COX-2 induction.

The metallothioneins (MT) are small, cysteine-rich heavy metal-binding proteins which participate in an array of protective stress responses. Although a single essential function of MT has not been demonstrated, MT of higher eukaryotes evolved as a mechanism to regulate zinc levels and distribution within cells and organisms. These proteins can also protect against some toxic metals and oxidative stress-inducing agents. In mice, among the four known MT genes, the MT-I and -II genes are most widely expressed. Transcription of these genes is rapidly and dramatically up-regulated in response to zinc and cadmium, as well as in response to agents which cause oxidative stress and/or inflammation. The six zinc-finger metal-responsive transcription factor MTF-1 plays a central role in transcriptional activation of the MT-I gene in response to metals and oxidative stress. Mutation of the MTF-1 gene abolishes these responses, and MTF-1 is induced to bind to the metal response elements in proximal MT promoter in cells treated with zinc or during oxidative stress. The exact molecular mechanisms of action of MTF-1 are not fully understood. Our studies suggest that the DNA-binding activity of MTF-1 in vivo and in vitro is reversibly activated by zinc interactions with the zinc-finger domain. This reflects heterogeneity in the structure and function of the six zinc fingers. We hypothesize that MTF-1 functions as a sensor of free zinc pools in the cell. Changes in free zinc may occur in response to chemically diverse inducers. MTF-1 also exerts effects on MT-I gene transcription which are independent of a large increase in MTF-1 DNA-binding activity. For example, cadmium, which has little effect on the DNA-binding activity of MTF-1 in vivo or in vitro, is a more potent inducer of MT gene expression than is zinc. The basic helix-loop-helix-leucine zipper protein, USF (upstream stimulatory factor family), also plays a role in regulating transcription of the mouse MT-I gene in response to cadmium or H 2 O 2 . Expression of dominant negative USF-1 or deletion of its binding site from the proximal promoter attenuates induction of the mouse MT-I gene. USF apparently functions in this context by interacting with as yet unidentified proteins which bind to an antioxidant response element which overlaps the USF-binding site (USF/ARE). Interestingly, this composite element does not participate in the induction of MT-I gene transcription by zinc or redox-cycling quinones. Thus, regulation of the mouse MT-I gene by metals and oxidative stress involves multiple signaling pathways which depend on the species of metal ion and the nature of the oxidative stress. BIOCHEM PHARMACOL 59;1:95-104, 2000.

␣-Difluoromethylornithine (DFMO) is commonly used as a specific ornithine decarboxylase (ODC, EC4.1.1.17) irreversible inhibitor. ODC is the enzyme responsible for polyamine biosynthesis, which has been shown to be strictly necessary for cell proliferation. In HT-29 Glc Ϫ/ϩ cells, L-arginine is the major precursor of these molecules through the sequential actions of arginase, which leads to L-ornithine generation and ODC. L-ornithine, a substrate for ODC, retroinhibits arginase. Since DFMO is an ornithine analogue, we searched for a direct effect of this agent upon arginase. The flux of L-arginine through arginase in intact cells was inhibited by 51 Ϯ 11% by 10 mM of DFMO whereas 10 mM of L-valine, a known potent arginase inhibitor, inhibited this flux by 73 Ϯ 6%. DFMO equilibrated between extracellular and intercellular spaces and, when used at 10-mM concentration, was without effect on L-arginine net uptake. Measurement of arginase activity in HT-29 cell homogenates with increasing concentrations of DFMO and L-arginine led to an inhibition with a calculated K i (inhibitory constant) equal to 3.9 Ϯ 1.0 mM. L-ornithine was less effective than DFMO in inhibiting arginase activity. Bovine liver arginase, used as another source of the enzyme, was also severely inhibited by DFMO. The inhibitory effect of DFMO upon arginase, one step upstream of the ODC reaction in the metabolic conversion of L-arginine to polyamines, is of potential physiological importance, since it could alter the production of ornithine and thus its metabolism in pathways other than the ODC pathway.

Tissue morphogenesis during development is regulated by growth factors and cytokines, and is characterized by constant remodeling of extracellular matrix in response to signaling molecules. MEK kinase 1 (MEKK1) is a mitogen-activated protein kinase (MAPK) kinase kinase originally identified as an upstream activator for several MAPK pathways. During mouse embryogenesis, MEKK1 controls cell shape changes and formation of actin stress fibers that are required for sealing epidermis in the embryos in a process known as eyelid closure. MEKK1-null mice display eye-open at birth (EOB), a phenotype found also in mice impaired in activin, a subgroup of the transforming growth factor b (TGFb) family, or in epidermal growth factor receptor (EGFR) or its ligand TGFa, or in transcription factor c-Jun. Molecular analyses have revealed at least two signaling mechanisms in the control of eyelid closure. One is originated from the activins and is transduced through MEKK1, leading to transcription-independent actin stress fiber formation and transcription-dependent keratinocyte migration. Another is the TGFa/EGFR signal that is transduced through a MEKK1-independent pathway to the activation of the ERK MAPK, which also leads to keratinocyte migration. c-Jun might serve as a connection between the two pathways. As embryonic eyelid closure is a specific morphogenetic process that is easily detectable, genetic mutant mice with EOB will be ideal models to understand the signaling mechanisms in the control of epithelial cell migration and the morphogenetic process of epithelial sheet movement.

with two sulfonate groups on adjacent phenyl rings; AlPcS 2a , aluminium phthalocyanine with two sulfonate groups on adjacent phenyl rings; E-64, trans-epoxysuccinyl-L-leucylamido(4-guanidino)butane; MTT, 3-[4,5-dimethylthiazol-2-y]-2,5 diphenyltetrazolium bromide a v a i l a b l e a t w w w . s c i e n c e d i r e c t . c o m j o u r n a l h o m e p a g e : w w w . e l s e v i e r . c o m / l o c a t e / b i o c h e m p h a r m

Generation of free radicals is thought to mediate the cytotoxic action of alloxan on the pancreatic ␤-cell. In this investigation, the early effects of alloxan on cell function were studied. When INS-1D insulinoma cells were exposed to alloxan (1 mM) for 45 min followed by a 3-hr recovery period, the drug increased basal insulin release while abolishing the effect of glucose in static incubations. This was associated with impaired stimulation of cellular metabolism by glucose and reduced viability, both monitored colorimetrically with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). These alterations were largely counteracted by the antioxidant butylated hydroxyanisol (BHA). Similar changes occurred when glucose was added directly after 5 min of alloxan treatment, whereas KCl-induced secretion was only partially inhibited. In perifusion, alloxan caused transient insulin secretion to 50% of the rates obtained with glucose 30 min later. Under these conditions, epinephrine abolished the stimulation due to both agents. Membrane potential and cytosolic calcium concentrations ([Ca 2ϩ ] i ) were recorded to clarify the action of alloxan. Alloxan-induced insulin release correlated with depolarization of INS-1D cells and a rise in [Ca 2ϩ ] i . Alloxan did not augment [Ca 2ϩ ] i in the presence of BHA or the absence of extracellular calcium. Nickel chloride blocked the effect of alloxan on [Ca 2ϩ ] i , whereas verapamil was ineffective. This suggests that alloxan promotes Ca 2ϩ influx through channels distinct from L-type channels, perhaps through non-selective cation channels. Thus, alloxan causes changes in INS-1D cells prevented by antioxidant treatment, suggesting that free radicals may modulate the ionic permeability leading to functional activation.

Mouse kidney thiol transmethylase and S-adenosylmethionine were incubated with the radioactive antithyroid drugs. I-thiouracil (TU). 6.propyl-2-thiouracil (PTU). methima7ole (MMI). 6. methyl-I-thiouracil (6-methyl TU) or thiourea. Radioactive metaholites were produced with TU. PTU and h-methyl TU and, in each case. were identified as the corresponding S-methyl derivatives. No measurable metabolism of MM1 or thiourea was observed. Kinetic studies with the partially purified enzyme demonstrated K, values for TU, PTU and h-methyl TU of 1 x lo-' M. 2.5 x IF3 M and 1.54 x IF3

We reported previously that treatment with antibody to transforming growth factor-p (TGF-B) caused a marked attenuation of bleomycin (BL)-induced lung fibrosis (LF) in mice. Decorin (DC), a proteoglycan, binds TGF-B and thereby down-regulates all of its biological activities. In the present study, we evaluated the antifibrotic potential of DC in a three-dose BL-hamster mode1 of lung fibrosis. Hamsters were placed in the following groups: (1) saline (SA) + phosphate-buffered saline (PBS) (SA + PBS); (2) SA + DC; (3) BL + PBS; and (4) BL + DC. Under pentobarbital anesthesia, SA (4 mL/kg) or BL was instilled intratracheally in three consecutive doses (2.5, 2.0, 1.5 units/kg/4 mL) at weekly intervals. DC (1 mg/mL) or PBS was instilled intratracheally in 0.4 mL/hamster on days 3 and 5 foll owing instillation of each dose of SA or BL. In week 4, hamsters received three doses of either DC or PBS every other day. The hamsters were killed at 30 days following the first instillation, and their lungs were appropriately processed. Lung hydroxyproline levels in SA + PBS, SA + DC, BL + PBS, and BL + DC groups were 965, 829, 1854, and 1387 p,g/lung, respectively. Prolyl hydroxylase activities were 103, 289, and 193% of SA + PBS control in SA + DC, BL + PBS, and BL + DC groups, respectively. The myeloperoxidase activities in the corresponding groups were 222, 890, and 274% of control (0.525 units/lung). Intratracheal instillation of BL caused significant increases in these biochemical markers, and instillation of DC diminished these increases in the BL + DC group. DC treatment also caused a significant reduction in the infiltration of neutrophils in the bronchoalveolar lavage fluid (BALF) of hamsters in the BL + DC group. However, DC treatment had little effect on BL-induced increases in lung superoxide dismutase activity and lipid peroxidation and leakage of plasma proteins in the BALF of the BL + DC group. Hamsters in the BL + PBS group showed severe multifocal fibrosis and accumulation of mononuclear inflammatory cells and granulocytes. In contrast, hamsters in the BL + DC group showed mild multifocal septal thickening with aggregations of mononuclear inflammatory cells. Hamsters in both control groups (SA + PBS

Ammonia is a major neurotoxin implicated in hepatic encephalopathy (HE). Here we discuss evidence that many aspects of ammonia toxicity in HE-affected brain are mediated by glutamine (Gln), synthesized in excess from ammonia and glutamate by glutamine synthetase (GS), an astrocytic enzyme. The degree to which Gln is increased in brains of patients with HE was found to positively correlate with the grade of HE. In animals with HE, a GS inhibitor, methionine sulfoximine (MSO), reversed a spectrum of manifestations of ammonia toxicity, including brain edema and increased intracranial pressure, even though MSO itself increased brain ammonia levels. MSO inhibited, while incubation with Gln reproduced the oxidative stress and cell swelling observed in ammonia-exposed cultured astrocytes. Recent studies have shown that astrocytes swell subsequent to Gln transport into mitochondria and its degradation back to ammonia, which then generates reactive oxygen species and the mitochondrial permeability transition. This sequence of events led to the formulation of the "Trojan Horse" hypothesis. Further verification of the role of Gln in the pathogenesis of HE will have to account for: 1) modification of the effects of Gln by interaction of astrocytes with other CNS cells; and 2) direct effects of Gln on these cells. Recent studies have demonstrated a "Trojan Horse"-like effect of Gln in microglia, as well as an interference by Gln with the activation of the NMDA/NO/cGMP pathway by ammonia as measured in whole brain, a process that likely also involves neurons.

Polymyalgia rheumatica (PMR) is an inflammatory disease characterized by bilateral pain involving predominantly the shoulders and proximal aspects of the arms and less commonly the neck and the pelvic girdle. This review discusses briefly the main epidemiological data and clinical features of this condition. Especial attention is paid in the management of the disease. For this reason, both the classic management and the impact of new therapies are discussed in depth. In general, patients with PMR experience a rapid response to 12.5-25 mg/prednisone/day in less than a week. Patients with poor response to glucocorticoids or with relapsing disease require other therapies aimed mainly to spare glucocorticoids. Among them, methotrexate is the most commonly used. Nevertheless, different studies indicate that this agent yields only a modest effect. Biologic therapies against the main cytokines involved in the pathogenesis of the disease have been used in refractory patients. However, randomized controlled trials do not support the use of anti-tumor necrosis factor agents in PMR. In contrast, several case series and retrospective studies highlight the efficacy of the antiinterleukin-6 receptor tocilizumab in PMR. Nonetheless, controlled trials are needed to fully establish the beneficial effect of this agent. The potential favorable effect of the Janus-kinase inhibitors and new anti-interleukin-6 antagonists remains to be determined.

Antiangiogenic therapy has become a mainstay of cancer therapeutics, but clinical responses are generally short-term owing to the development of secondary resistance. Tumor starvation by antiangiogenic drugs is largely attributed to increased hypoxia and impaired nutrients supply, suggesting that angiogenesis inhibition causes remarkable metabolic perturbations in the tumor microenvironment. We review here recent acquisitions concerning metabolic effects of angiogenesis blockade in tumors and discuss the possibility that some metabolic features of tumor cells -i.e. their dependency from glucose as primary energy substrate -might affect tumor responses to anti-vascular endothelial growth factor treatment. Moreover, we discuss the hypothesis that anti-angiogenic therapy might foster metabolic evolution of tumors. The therapeutic implications of this hypothesis will be discussed further here. ß

Oncolytic virotherapy is a growing treatment modality that uses replicating viruses as selective antineoplastic agents. Safety and efficacy considerations dictate that an ideal oncolytic agent would discriminate between normal and cancer cells on the basis of common genetic abnormalities in human cancers. Here, we identify a naturally occurring alphavirus (M1) as a novel selective killer targeting zinc-finger antiviral protein (ZAP)-deficient cancer cells. In vitro, in vivo, and ex vivo studies showed potent oncolytic efficacy and high tumor tropism of M1. We showed that the selectivity depends on ZAP deficiency by systematic identification. A largescale multicenter pathology study using tissue microarrays reveals that ZAP is commonly deficient in human cancers, suggesting extensive application prospects for M1. Additionally, M1 killed cancer cells by inducing endoplasmic reticulum stress-mediated apoptosis. Our report provides novel insights into potentially personalized cancer therapy using oncolytic viruses.

Polyamines are ubiquitous molecules, which, like iron, are essential for cell growth. All eukaryotic cells are equipped with a specific polyamine transport system (PTS). Polyamines have primary and secondary amino groups which chelate bivalent metal cations such as Fe and Cu. In the present study, we investigated the potential contribution of naturally occurring polyamines and their active transport system to iron uptake. In presence of subtoxic Fe(III) (10 mM), treatment of CHO cells with spermine, and to a lesser extent with spermidine (10-100 mM), resulted in a marked cytotoxic effect. This cytotoxicity was prevented by the addition of an iron-chelator, deferioxamine, and was not observed in CHO-MG cells, a mutant cell line devoid of polyamine transport activity. Experiments using 14 C-polyamines and 55 Fe(III) revealed that these toxic effects were related to polyamine-modulation of iron uptake, and were dependent on the presence of the active PTS. These results demonstrated active uptake of polyamine-iron complexes via the PTS. The number of amino groups affected the efficacy of the studied natural polyamines to transport iron via the PTS. Spermine, a tetramine, was more efficient than the triamine spermidine. Co-transport of iron by the diamine putrescine was not observed. These results demonstrate that the cell polyamine transport system is a potential cell entry pathway for iron. The studied polyamines, spermine and spermidine, may be components of the pool of transferrin-independent iron-chelating vectors, which have recently attracted the attention of many investigators.

Screen, marked differences were observed. Evaluation of the activity profile using the COMPARE program revealed a different pattern for both agents: the cisplatin activity profile was similar to those of other diamine-platinum compounds, alkylating agents including melphalan, and camptothecin analogs, whereas the activity profile of oxaliplatin resembled those of other "dach" {diaminocyclohexane) platinum compounds and of acridine derivatives. The sensitivity profiles are influenced by the target(s)/mechanism(s) of action and the mechanism(s) of resistance of a drug. The dissimilarity in profiles suggests that these two platinum compounds have a different target(s)/mechanism(s) of action, a different mechanism(s) of resistance, or most likely both. Studies evaluating combinations of cisplatin/oxaliplatin suggest that the activities of these two agents are at least additive and possibly synergistic. Oxaliplatin has a different spectrum of activity and low cross-resistance to cisplatin and should be valuable in cisplatin refractory patients or in combination with cisplatin.

A~t~ct-Affinity, specificity and kinetics for [3H]-DHA binding to human red cell ghost were determined by ultra-filtration. At 2" an apparent dissociation constant of 0.96 nM was found with maximum specific binding of 29 fmoles per mg protein. The low dissociation constant was confirmed by kinetic studies with a value of 0.86 nM. Propranolol and isoproterenol inhibited t3H]-DHA binding stereospecifically. Agonist potency (IPR > EPI > NE) indicated that human erythrocytes had an adtenergic receptor of be&z-2 subtype. Isoproterenol in the presence of theophylline resulted in a concentrationdependent increase of intracellular CAMP levels in intact cells. Basal and maximal levels were 2.3 and 7.5 pmoles/lOa cells respectively after 2.5 min stimulation. Et& for isoproterenol was 0.27 PM. Propranolol shifted the isoproterenol concentration response curve to the right. The present results show that human erythrocytes possess recognition sites for beta-adrenergic ligands with binding characteristics similar to that of adrenergic receptors of beta-2 subtype. At least a small number of these binding sites are functionally coupled to adenylate cyclase.

It has been shown previously that 4-anilino quinazolines compete with the ability of ATP to bind the epidermal growth factor receptor (EGF-R), inhibit EGF-stimulated autophosphorylation of tyrosine residues in EGF-R, and block EGF-mediated growth. Since millimolar concentrations of ATP in cells could reduce the efficacy of 4-anilino quinazolines in cells and the activity of these compounds would not be sustained once they were removed from the body, we reasoned that irreversible inhibitors of EGF-R might improve the activity of this series of compounds in animals. Molecular modeling of the EGF-R kinase domain was used to design irreversible inhibitors. We herein describe one such inhibitor: N-[4-[(3-bromophenyl)amino]-6-quinazolinyl]2-butynamide, known as CL-387,785. This compound covalently bound to EGF-R. It also specifically inhibited kinase activity of the protein (IC50 = 370+/-120 pM), blocked EGF-stimulated autophosphorylation of the receptor in cells (ic50 approximately 5 nM), inhibited cell proliferation (IC50 = 31-125 nM) primarily in a cytostatic manner in cell lines that overexpress EGF-R or c-erbB-2, and profoundly blocked the growth of a tumor that overexpresses EGF-R in nude mice (when given orally at 80 mg/kg/day for 10 days, daily). We conclude that CL-387,785 is useful for studying the interaction of small molecules with EGF-R and may have clinical utility.

The plant-derived phenolic compounds gossypol, quercetin and myricetin are powerful inhibitors of iron-induced lipid peroxidation in rat liver microsomes, under all five experimental conditions tested and at low micromolar concentrations (ICjo < 1.5 yM). However, they greatly accelerate the generation of hydroxyl radicals (*OH) from H,O, in the presence of Fe'+-EDTA at pH7.4, as measured by the deoxyribose assay. At 100 PM, the three phenolic compounds enhanced *OH formation up to eight-fold. The hydroxyi radical generation was inhibited by catalase and superoxide dismutase, suggesting a mechanism in which the phenols oxidize to produce superoxide radical, which then assists *OH generation from HzO, in the presence of Fe '+-EDTA. At concentrations up to 75 PM, quercetin and myricetin also accelerate bleomycin-dependent DNA damage in the presence of Fe3+, possibly by reducing the Fe 3+-bleomycin-DNA complex to the Fe 2t form. Hence these naturally-occurring substances can have nro-oxidant effects under some reaction conditions and cannot be classified simplistically as "antioxidants".

Intraperitoneal injection of adenosine into Swiss albino mice, RB strain, sensitive to audiogenic convulsions, rapidly produces, in proportion to the dose administered: sedation, modification of the EEG, lowering of arterial pressure and protection against audiogenic seizures. ...

The relationship between methylene blue (MB) and Alzheimer's disease (AD) has recently attracted increasing scientific attention since it has been suggested that MB may slow down the progression of this disease. In fact, MB, in addition to its well characterized inhibitory actions on the cGMP pathway, affects numerous cellular and molecular events closely related to the progression of AD. Currently, MB has been shown to attenuate the formations of amyloid plaques and neurofibrillary tangles, and to partially repair impairments in mitochondrial function and cellular metabolism. Furthermore, various neurotransmitter systems (cholinergic, serotonergic and glutamatergic), believed to play important roles in the pathogenesis of AD and other cognitive disorders, are also influenced by MB. Recent studies suggest that the combination of diverse actions of MB on these cellular functions is likely to mediate potential beneficial effects of MB. This has lead to attempts to develop novel MB-based treatment modalities for AD. In this review article, actions of MB on neurotransmitter systems and multiple cellular and molecular targets are summarized with regard to their relevance to AD. ß

In the present study, we have examined the effect of dietary fat on paracetamol-induced liver injury in an in vitro rat liver slice model. Rats were fed, for 7-10 days, diets containing either butter or polyunsaturated vegetable margarine, two fat sources commonly consumed in the human diet. Liver slices were then exposed to paracetamol for 2 hr and further incubated for 4 hr without paracetamol. Cell damage in the slices was quantified at 6 hr by measuring leakage of lactate dehydrogenase, increase in water content and potassjum loss. Covalent binding of radioactive paracetamol to liver and the membrane fatty acid composition of the liver were also measured. Liver slices from rats fed butter diets were significantly more sensitive to the toxic effects of paracetamol than those from margarine fed rats. The membrane lipid composition of the livers also reflected the differing fatty acid content of the two diets.

b i o c h e m i c a l p h a r m a c o l o g y 7 4 ( 2 0 0 7 ) 8 3 1 -8 4 0 GTPase Cell division a b s t r a c t Cinnamaldehyde is a natural product from spices that inhibits cell separation in Bacillus cereus. Cell division is regulated by FtsZ, a prokaryotic homolog of tubulin. FtsZ assembles into the Z-ring at the site of cell division. Here, we report the effect of cinnamaldehyde on FtsZ and hence on the cell division apparatus. Cinnamaldehyde decreases the in vitro assembly reaction and bundling of FtsZ. It is found that cinnamaldehyde perturbs the Zring morphology in vivo and reduces the frequency of the Z ring per unit cell length of Escherichia coli. In addition, GTP dependent FtsZ polymerization is inhibited by cinnamaldehyde. Cinnamaldehyde inhibits the rate of GTP hydrolysis and binds FtsZ with an affinity constant of 1.0 AE 0.2 mM À1 . Isothermal titration calorimetry reveals that binding of cinnamaldehyde to FtsZ is driven by favorable enthalpic interactions. Further, we map the cinnamaldehyde binding region of FtsZ, using the saturation transfer difference-nuclear magnetic resonance and an in silico docking model. Both predict the cinnamaldehyde binding pocket at the C terminal region involving the T7 loop of FtsZ. Our results show that cinnamaldehyde binds FtsZ, perturbs the cytokinetic Z-ring formation and inhibits its assembly dynamics. This suggests that cinnamaldehyde, a small molecule of plant origin, is a potential lead compound that can be developed as an anti-FtsZ agent towards drug design. a v a i l a b l e a t w w w . s c i e n c e d i r e c t . c o m j o u r n a l h o m e p a g e : w w w . e l s e v i e r . c o m / l o c a t e / b i o c h e m p h a r m

Objective structured practical examination (OSPE) a new
assessment tool has been implemented in a few medical colleges.
This method of assessment was planned to be implemented in our
medical college, before which the authors planned to compare the
scores obtained by the students by conventional method of
examination and OSPE. The marks obtained by the two methods
of examination were compared and a feedback was collected from
students to study their perspective towards OSPE. There was
statistically significant difference in the mean of the scores
obtained by conventional method and that of OSPE. The students’
perception about OSPE was good. This study enabled the authors
to standardize the pattern of OSPE and implement it.

Drug addiction is a chronic, relapsing disorder, characterized by an uncontrollable motivation to seek and use drugs. Converging clinical and preclinical observations implicate pathologies within the corticolimbic glutamate system in the genetic predisposition to, and the development of, an addicted phenotype. Such observations pose cellular factors regulating glutamate transmission as likely molecular candidates in the etiology of addiction. Members of the Homer family of proteins regulate signal transduction through, and the trafficking of, glutamate receptors, as well as maintain and regulate extracellular glutamate levels in corticolimbic brain regions. This review summarizes the existing data implicating the Homer family of protein in acute behavioral and neurochemical sensitivity to drugs of abuse, the development of drug-induced neuroplasticity, as well as other behavioral and cognitive pathologies associated with an addicted state. Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers

The monoamine hypothesis has been the prevailing hypothesis of depression over the last several decades. It states that depression is associated with reduced monoamine function. Hence efforts to increase monoamine transmission by inhibiting serotonin (5-HT) and norepinephrine (NE) transporters has been a central theme in depression research since the 1960s. The selective 5-HT reuptake inhibitors (SSRIs) and 5-HT and NE reuptake inhibitors (SNRIs) that have emerged from this line of research are currently first line treatment options for major depressive disorder (MDD). One of the recent trends in antidepressant research has been to refine monoaminergic mechanisms by targeting monoaminergic receptors and additional transporters (e.g. with multimodal drugs and triple re-uptake inhibitors) or by adding atypical antipsychotics to SSRI or SNRI treatment. In addition, several other hypotheses of depression have been brought forward in pre-clinical and clinical research based on biological...

Cancer Chemofher Pharmacol24: 65566, 1989. 3. Marshall ME, Butler K and Hermansen D, Treatment of hormone-refractory stage D carcinoma of prostate with coumarin (1 ,Zbenzopyrone) and cimetidine: a pilot study. Prostate 17: 95-99, 1990. 4. Dexeus FH, Logothetis CJ, Sella A, Fitz K, Amato R, Reuben JM and Dozier N. Phase II studv of coumarin and cimetidine in patients with metastatic renal cell carcinoma. J Clin &co1 8: 325-329, 1990. 5. Cohen AJ, Critical review of the toxicoiogy of coumarin with special reference to interspecies differences in metabolism and hepatotoxic response and their significance to man. Food Cosmet Toxic01 17: 277-289, 1979. 6. Shilling WH, Crampton RF and Longland RC, Metabolism of coumarin in man. Nature 221: 664-665, 1969.

The rate-limiting, hormone-regulated, enzymatic step in steroidogenesis is the conversion of cholesterol to pregnenolone by the cholesterol side-chain cleavage enzyme system (CSCC), which is located on the matrix side of the inner mitochondrial membrane. However, it has long been observed that hydrophilic cholesterol-like substrates capable of traversing the mitochondrial membranes are cleaved to pregnenolone by the CSCC in the absence of any hormone stimulation. Therefore, the true regulated step in the acute response of steroidogenic cells to hormone stimulation is the delivery of cholesterol to the inner mitochondrial membrane and the CSCC. It has been known for greater than three decades that transfer of cholesterol requires de novo protein synthesis; however, prior to this time the regulatory protein(s) had yet to be identified conclusively. It is the purpose of this commentary to briefly review a number of the candidates that have been proposed as the acute regulatory protein. As such, we have summarized the available information that describes the roles of transcription, translation, and phosphorylation in this regulation, and have also reviewed the supporting cases that have been made for several of the proteins put forth as the acute regulator. We close with a comprehensive description of the Steroidogenic Acute Regulatory protein (StAR) that we and others have identified and characterized as a family of proteins that are synthesized and imported into the mitochondria in response to hormone stimulation, and for which strong evidence exists indicating that it is the long sought acute regulatory protein.

2,3-Dihydroxybenzoic acid is present in blood plasma and urine of healthy human volunteers after aspirin ingestion. Its identity has been confirmed by mass spectrometry and by electrochemical analysis. Methods for its identification and measurement are described. The concentration of 2,3dihydroxybenzoic acid is much lower than that of 2,5-dihydroxybenzoic acid, salicylic acid or salicyluric acid.

The presence of a residual amount of unstable proteins in wines is a concern for winemakers because these polymers can precipitate from solution during storage causing appearance of sediments and hazes. Such precipitates are commonly the result of denaturation and subsequent aggregation of heat-unstable wine proteins. It has been demonstrated that the vast majority of the wine proteins derive from grapes and that proteins responsible for haze formation are Pathogenesis-Related (PR) proteins, in particular thaumatin-like (TL) proteins and chitinases. Moreover, these proteins are highly resistant to acidic pHs, proteolysis and fermentation conditions. However, they can became insoluble during wine storage and thus originate perceptible turbidity in the bottles. Despite the efforts made in the recent past, the white wine protein instability is still a main problem during white wine making and bentonite treatments are even now indispensable to stabilise white wines.

a-Tocopherol is the most important lipophilic antioxidant of the chromanol type protecting biomembranes from lipid peroxidation (LPO). Therefore, a-tocopherol and its derivatives are frequently used in the therapy or prevention of oxygen radical-derived diseases. In the present study, novel chromanol-type antioxidants (twin-chromanol, cis-and trans-oxachromanol) as well as the well-known shortchain analogue of a-tocopherol, pentamethyl-chromanol, were tested for their antioxidative potency in rat heart mitochondria (RHM). Our experiments revealed that the bioenergetic parameters of mitochondria were not deteriorated in the presence of chromanols (up to 50 nmol/mg protein). Exposure of RHM to cumene hydroperoxide and Fe 2+ (final concentrations 50 mM each), inducing LPO, significantly affected their bioenergetic parameters which were determined in the presence of glutamate and malate (substrates of mitochondrial complex I). Alterations of the bioenergetic parameters were partially prevented in a concentration-dependent manner by preincubating RHM with antioxidants before adding the radical-generating system. In the lower concentration range, twin-chromanol turned out to be more efficient than pentamethyl-chromanol, both being far more protective than cis-and trans-oxachromanol. Measurement of protein-bound SH groups and thiobarbituric acid-reactive substances revealed that this protective effect was due to their antioxidative action. Furthermore, HPLC measurements of a-tocopherol and a-tocopheryl quinone in rat liver mitochondria demonstrated an a-tocopherol-sparing effect of twin-chromanol. In conclusion, new chromanol-type antioxidants, especially twinchromanol, were able to improve bioenergetic and biochemical parameters of mitochondria exposed to oxidative stress. #

Hydroxytyrosol (HT) (also known as dihydroxyphenylethanol (DPE)) is a polyphenol extracted from virgin olive oil. HT is known to exert an antioxidant effect but the mechanism of action and the identity of the reactive oxygen molecule(s) targeted are not known. In this study, we show that HT inhibits luminol-amplified chemiluminescence of human neutrophils stimulated with N-formyl-methionyl-leucylphenylalanine (fMLP), phorbol myristate acetate (PMA) and opsonized zymosan. This effect was dose-dependent and occurred immediately after the addition of HT. However, HT had no effect on lucigenin-amplified chemiluminescence, suggesting that it does not inhibit NADPH oxidase activation or scavenge superoxide anions. Furthermore, HT inhibited H 2 O 2 -dependent-dichlorofuoroscein (DCFH) fluorescence of activated neutrophils, as measured by flow cytometry. Finally, HT inhibited luminol-amplified chemiluminescence in a cell-free system consisting of H 2 O 2 and HRPO. These results suggest that HT, a polyphenol derived from olive oil, could exert its antioxidant effect by scavenging hydrogen peroxide but not superoxide anion released during the respiratory burst. #