Potential of immunomodulatory agents as adjunct host-directed therapies for multidrug-resistant tuberculosis

Treatment of multidrug-resistant tuberculosis (MDR-TB) is extremely challenging due to the virulence of the etiologic strains of Mycobacterium tuberculosis (M. tb), the aberrant host immune responses and the diminishing treatment options with TB drugs. New treatment regimens incorporating therapeutics targeting both M. tb and host factors are urgently needed to improve the clinical management outcomes of MDR-TB. Host-directed therapies (HDT) could avert destructive tuberculous lung pathology, facilitate eradication of M. tb, improve survival and prevent long-term functional disability. In this review we (1) discuss the use of HDT for cancer and other infections, drawing parallels and the precedent they set for MDR-TB treatment, (2) highlight preclinical studies of pharmacological agents commonly used in clinical practice which have HDT potential, and (3) outline developments in cellular therapy to promote clinically beneficial immunomodulation to improve treatment outcomes in patients with pulmonary MDR-TB. The use of HDTs as adjuncts to MDR-TB therapy requires urgent evaluation

Neurochemistry

Neurochemistry is a flourishing academic field that contributes to our understanding of molecular, cellular and medical neurobiology. As a scientific discipline, neurochemistry studies the role of chemicals that build the nervous system, it explores the function of neurons and glial cells in health and disease, it discovers aspects of cell metabolism and neurotransmission, and it reveals how degenerative processes are at work in the nervous system. Accordingly, this book contains chapters from a variety of topics that fall into the following broad sections: I. Neural Membranes and Intracellular Signaling, II. Neural Processing and Intercellular Signaling, III. Growth, Development and Differentiation, and IV. Neurodegenerative Diseases. The book presents comprehensive reviews in these different areas written by experts in their respective fields. Neurodegeneration and neuronal diseases are featured prominently and are a recurring theme throughout most chapters. This book will be a most valuable resource for neurochemists and other scientists alike. In addition, it will contribute to the training of current and future neurochemists and, hopefully, will lead us on the path to curing some of the biggest challenges in human health

Studies on Cell Adhesion and Activation

The role of plasmalemmal lipids in the adhesion of Baby Hamster Kidney (BHK) fibroblasts to polystyrene tissue culture grade dishes in serum and serum-free medium was investigated using various fatty acids that differ in their hydrocarbon chain length and saturation. A number of biologically active agents which include arachidonic acid metabolites, as well as some of the factors which affect the calcium/protein kinase C pathway were also considered in the present study. Fatty acids were the first to be tested. The saturated fatty acid derivative stearoyl-CoA (18:0) had two different effects on BHK cell adhesion in serum-free Ham's F-10. Concentrations, ranged between 1.75 and 7 muM, enhanced the adhesion in almost all cases, while higher concentrations such as 14 and 28 uM reduced the adhesion of BHK cells to polystyrene tissue culture dishes by more than 50% of the control values. BHK cells in the presence of long chain unsaturated fatty acids, (oleoyl-CoA (18:1), linoleoyl-CoA (18:2), linolenoyl-CoA (18:3), and arachidonoyl-CoA (18:4), responded in different ways depending on the type of the fatty acid added. When oleoyl-CoA was used at concentrations lower than 7 muM the adhesion was maintained at the control level or slightly enhanced. However,the use of concentrations higher than that reduced the attachment sharply. Linoleoyl-CoA, linolenoyl-CoA and arachidonoyl-CoA generally maintained the adhesion at the control level, though linoleoyl-CoA enhanced the adhesion at fairly low concentrations (lower than 14 muM) . Both, stearoyl-CoA and oleoyl-CoA (1.75 muM and higher) decreased the spreading area of BHK cells by 30-50% of control samples in serum-free medium. The above fatty acyl-CoAs did not however alter either the adhesion or the spreading of BHK fibroblasts in serum-containing medium. In contrast to that, linoleic and arachidonic acid used as free acid form (unbound to the coenzyme A) at concentrations ranged between 2 and 20 microg/ml in combination with 1.25x10e-5 M ATP and 5x10 6 M Coenzyme A, decreased significantly (P< 0.001) the adhesion in both 3%serum and serum-free Ham's F-10 medium. A noticeable loss of viability was found at concentrations higher than 5microg/ml. The effect on cell viability was less pronounced in serum-containing medium. Incorporation studies in serum-free conditions revealed that approximately 40% of the total labeled oleoyl-CoA was taken up by the cells in a period of 20 mins incubation. 21% of the total incorporated is present in the plasma membrane. This results suggest that the observed effect on cell adhesion might be due to changes in plasmalemmal lipids. Hence, the effect on the adhesion and spreading could be explained either in terms of the action of membrane electrodynamic forces or membrane fluidity. It should be noted however, that the total oleoyl-CoA incorporated was reduced by 43% when the experiment was carried out in serum-containing medium. This may explain the lack of effect of acyl-Coenzyme A on BHK cell adhesion as well as other functions in serum-containing medium. The fatty acid metabolites, Prostacyclin (PGI2), prostaglandin E2, E1 and leukotrienes B4 (LTB4) exerted different effects. At the time where 1.25 and 0.125 muM LTB4 increased BHK and slightly endothelial cell-cell attachment and PGE1 (1-50muM) enhanced BHK cell-polystyrene adhesion, prostacyclin and prostaglandin E2 at concentrations up to 5 mug/ml did not have any clear effect on BHK cell-substratum adhesion. Inhibitors of arachidonic acid release, Bromophenacyl Bromide and mepacrine (quinacrine), significantly reduced BHK fibroblast adhesion to polystyrene surfaces. There was an irreversible inhibition of adhesion. Viability tests revealed that approximately 95% of the cells were in a viable state. Tests were also made on calcium and protein kinase C modulators. A diacylglycerol kinase inhibitor (R59022), reduced BHK fibroblast as well as endothelial cell adhesion to polystyrene tissue culture dishes in both serum and serum-free conditions. Endothelial cell adhesion in 3% serum Ham's F-10 was reduced by approximately 90% of the control samples when a concentration of 18.4 mug/ml was used. In contrast, a concentration of 30 mug/ml was needed to get the same reduction with BHK cells under the same conditions. The above concentration (30microg/ml) however, decreased BHK cell adhesion by 40% of the control samples in serum-free conditions

DUAL LOX/COX INHIBITION: A NOVEL STRATEGY TO PREVENT NEUROVASCULAR LEAKAGE IN EPILEPSY

Epilepsy affects 3.4 million patients in the USA and is characterized by recurring seizures. The blood-brain barrier is leaky in epilepsy and may contribute to seizure progression but the mechanisms which cause this leakage are not fully understood. We hypothesized that seizures trigger LOX- and COX-mediated blood-brain barrier leakage and that dual LOX/COX inhibition prevents barrier leakage in vivo. To test this hypothesis, we administered either the dual LOX/COX inhibitor licofelone or a combination of the 5-LOX inhibitor zileuton and the COX-2 inhibitor celecoxib to rats that experienced status epilepticus (SE). Serum and brain capillaries were isolated 48 hours after SE and serum S100β levels were measured and Texas Red™ leakage rates were determined. Dual inhibition of 5-LOX and COX prevented serum S100β elevations observed in SE rats in a dose-dependent manner with licofelone. Inhibition of 5-LOX and COX-2 with zileuton and celecoxib completely prevented serum S100β elevation. Texas Red™ leakage rates for SE rats were also reduced in a dose-depended manner with licofelone and reduced to control rates with zileuton and celecoxib. These data support our hypothesis that seizure-induced blood-brain barrier leakage is mediated by LOX and COX, and inhibition of these enzymes prevents barrier leakage

Multiple P2Y receptors couple to calcium-dependent, chloride channels in smooth muscle cells of the rat pulmonary artery

BACKGROUND: Uridine 5'-triphosphate (UTP) and uridine 5'-diphosphate (UDP) act via P2Y receptors to evoke contraction of rat pulmonary arteries, whilst adenosine 5'-triphosphate (ATP) acts via P2X and P2Y receptors. Pharmacological characterisation of these receptors in intact arteries is complicated by release and extracellular metabolism of nucleotides, so the aim of this study was to characterise the P2Y receptors under conditions that minimise these problems. METHODS: The perforated-patch clamp technique was used to record the Ca(2+)-dependent, Cl(- )current (I(Cl,Ca)) activated by P2Y receptor agonists in acutely dissociated smooth muscle cells of rat small (SPA) and large (LPA) intrapulmonary arteries, held at -50 mV. Contractions to ATP were measured in isolated muscle rings. Data were compared by Student's t test or one way ANOVA. RESULTS: ATP, UTP and UDP (10(-4)M) evoked oscillating, inward currents (peak = 13–727 pA) in 71–93% of cells. The first current was usually the largest and in the SPA the response to ATP was significantly greater than those to UTP or UDP (P < 0.05). Subsequent currents tended to decrease in amplitude, with a variable time-course, to a level that was significantly smaller for ATP (P < 0.05), UTP (P < 0.001) and UDP (P < 0.05) in the SPA. The frequency of oscillations was similar for each agonist (mean≈6–11.min(-1)) and changed little during agonist application. The non-selective P2 receptor antagonist suramin (10(-4)M) abolished currents evoked by ATP in SPA (n = 4) and LPA (n = 4), but pyridoxalphosphate-6-azophenyl-2',4'-disulphonic acid (PPADS) (10(-4)M), also a non-selective P2 antagonist, had no effect (n = 4, 5 respectively). Currents elicited by UTP (n = 37) or UDP (n = 14) were unaffected by either antagonist. Contractions of SPA evoked by ATP were partially inhibited by PPADS (n = 4) and abolished by suramin (n = 5). Both antagonists abolished the contractions in LPA. CONCLUSION: At least two P2Y subtypes couple to I(Cl,Ca )in smooth muscle cells of rat SPA and LPA, with no apparent regional variation in their distribution. The suramin-sensitive, PPADS-resistant site activated by ATP most resembles the P2Y(11 )receptor. However, the suramin- and PPADS-insensitive receptor activated by UTP and UDP does not correspond to any of the known P2Y subtypes. These receptors likely play a significant role in nucleotide-induced vasoconstriction

Studies on the pathology of bacterial kidney disease (Renibacterium salmoninarum) in coho (Oncorhynchus kisutch) and Atlantic salmon (Salmo salar)

With the intensification of the aquaculture industry along the west coast of Canada, there has been a concomitant increase in the prevalance of bacterial kidney disease (BKD) (Renibacterium salmoninarum)(Rs) in production and government hatchery facilities. Due to the paucity of information on the pathogenesis and epizootiology of Rs, treatment and control measures have been confounded and emergance of BKD exacerbated. Initially, to enhance specificity over conventional histochemistry for demonstration of Rs in histological sections an avidin biotin conjugated immunoperoxidase technique was developed for us to monitor the histopathological manifestation of BKD. To ascertain the temporal and spatial hlstogenesis of BKD. coho (Oncorhvnchus kisutch) and Atlantic (Salmo salar) salmon ware challenged experimentally by intraperitoneal injection and naturally by cohabitation with Rs-innoculated fish than serially sampled. Histopathology revealed lesions consistent with past case reports and investigations, as well as previously undescribed manifestations including, inflammatory call kinetics in the renal perivascular compartment, cresentic glomerulonephritis, extrarenal dissemination (via septic emboli or direct extension), and ovarian follicular cell accumulation of Rs. Pseudocyst formation, renal interstitial hyperplasia and meningoencephalitis were also characterized. The granulomatous response in coho salmon was predominantly histiocytic, whereas, in Atlantic salmon a tuberculoid response was more apparent. In either species a profound call mediated immunity was adduced. To further resolve the nature of the inflammatory response, coho and Atlantic salmon were immunosuppressed by administration of suprapharmacologic doses of glucocorticosteroids. On challange with Rs both species incurred an earlier onset and greater rate of mortality than immunocompetent cohorts. The granulomatous response appeared irregular and expansive with exuberent intra- and extracellular Rs growth. These observations may be attributed to inhibition of prostaglandin and leukotrienne synthesis, as well as an inability to immunologically prima phagocytes. To evaluate the inflammogenic potential o£ somatic, cell- wall associated, and soluble fractions of Rs, coho and Atlantic salmon were injected and serially sampled for histopathology. Past studies on Rs virulance determinants and pathogenic mechanisms have focused almost exclusively on a soluble protein, designated p57. In this investigation no histological alterations were appreciated in fish challenged with cell-wall associated or soluble (p57) fractions, however, mild, multifocal pyogranulomata were noted in the renal interstitium of coho and Atlantic salmon challenged with the somatic or peptidoglycan fraction of Rs. Peptidoglycans of a number of mammalian pathogens are strongly inflammogenic, poorly biodegradable, and persist in host tissue for protracted periods. Chemical resolution and in vivo evaluatlon of subcellular components of Rs is warranted to further resolve the pathogenesis of BKD in salmonid species

The Economic Impact of Payer Policies after the Rx-to-OTC Switch of Second-Generation Antihistamines* *Preliminary results of this analysis were presented at the 9th annual HMO Research Network Conference April 1-2, 2003

AbstractObjectiveAs a result of the over-the-counter (OTC) introduction of loratadine, health plans have been struggling to determine the best policy to incorporate this change within their existing drug benefit structure for second-generation antihistamines (SGA). The objective of this study was to examine the economic impact of payer policies in response to the Rx-to-OTC switch of loratadine.Study DesignDecision analysis was used to model the budgetary impact and cost-effectiveness of four policies for SGA benefits for the managed care organization (MCO), employer, and Medicaid perspectives separately.Patients and MethodsOutcomes included direct medical costs and lost productivity (employers only), discounted, quality-adjusted life-years (QALYs) saved because of amelioration of allergic rhinitis symptoms and avoidance of unintentional injuries associated with the use of first-generation antihistamines (FGA). Bayesian probabilistic sensitivity analysis was conducted using second-order Monte Carlo simulation.ResultsProviding limited OTC and second-tier prescription benefits would cost approximately $0.13 and$0.30 compared to third-tier prescription benefits for employers and MCOs, respectively, and would save Medicaid $.02 per member per month (PMPM). Providing limited coverage for OTC loratadine while retaining second-tier prescription benefits for SGA was the optimal policy for a willingness to pay below$26,200 per QALY for all payers.ConclusionsOffering second-tier prescription and limited OTC benefits provides greater effectiveness and is not significantly more expensive PMPM than discontinuation. Some of the drug savings from limiting coverage of prescription SGA may be attenuated by the cost of lost productivity and direct medical expenditures due to unintentional injuries associated with increased FGA use in addition to the increased cost of therapeutic substitutes