Effects of non-steroidal anti-inflammatory drugs on proliferation, differentiation and migration in equine mesenchymal stem cells

In equine medicine, stem cell therapies for orthopaedic diseases are routinely accompanied by application of NSAIDs (non-steroidal anti-inflammatory drugs). Thus, it has to be analysed how NSAIDs actually affect the growth and differentiation potential of MSCs (mesenchymal stem cells) in vitro in order to predict the influence of NSAIDs such as phenylbutazone, meloxicam, celecoxib and flunixin on MSCs after grafting in vivo. The effects of NSAIDs were evaluated regarding cell viability and proliferation. Additionally, the multilineage differentiation capacity and cell migration was analysed. NSAIDs at lower concentrations (0.1-1 μM for celecoxib and meloxicam and 10-50 μM for flunixin) exert a positive effect on cell proliferation and migration, while at higher concentrations (10-200 μM for celecoxib and meloxicam and 100-1000 μM for flunixin and phenylbutazone), there is rather a negative influence. While there is hardly any influence on the adipogenic as well as on the chondrogenic MSC differentiation, the osteogenic differentiation potential, as demonstrated with the von Kossa staining, is significantly disturbed. Thus, it can be concluded that the effects of NSAIDs on MSCs are largely dependent on the concentrations used. Additionally, for some differentiation lineages, also the choice of NSAID is critical.     

NSAIDs and scavenging birds: potential impacts beyond Asia's critically endangered vultures

Veterinary treatment of livestock with diclofenac, a non-steroidal anti-inflammatory drug (NSAID), has caused catastrophic declines of Gyps vultures in Asia. This has highlighted a lack of knowledge on the potential impacts of NSAIDs on scavenging birds. Surveys of veterinarians and zoos document the outcomes of the treatment of over 870 scavenging birds from 79 species. As well as diclofenac, carprofen and flunixin were associated with mortality, with deaths observed in 13 and 30% of cases, respectively. Mortality was also found following treatment with ibuprofen and phenylbutazone. NSAID toxicity was reported for raptors, storks, cranes and owls, suggesting that the potential conservation impact of NSAIDs may extend beyond Gyps vultures and could be significant for New World vultures. In contrast, there were no reported mortalities for the NSAID meloxicam, which was administered to over 700 birds from 60 species. The relative safety of meloxicam supports other studies indicating the suitability of this NSAID to replace diclofenac in Asia.     

The Pied Crow (Corvus albus) is insensitive to diclofenac at concentrations present in carrion

Diclofenac, a nonsteroidal anti-inflammatory drug (NSAID), kills vultures (Gyps spp.) that consume tainted carcasses. As a result, vulture populations in India, Nepal, and Pakistan have been devastated. Studies on meloxicam and ketoprofen demonstrated that the toxicity of the NSAIDs is unpredictable, thereby necessitating individual testing of all available NSAIDs. Because it is no longer practical to use vultures for toxicity testing, we evaluated the Pied Crow (Corvus albus) as a model. Pied Crows (n=6) were exposed to a dose of 0.8 and 10 mg/kg of diclofenac, with no signs of toxicity, and a rapid half-life of elimination. Using primary renal cell and hepatocyte cultures, a high tolerance was demonstrated at the cellular level. Meta-analysis of pharmacokinetic data for the Domestic Chicken (Gallus gallus) and the African White-backed (Gyps africanus), Cape Griffon (Gyps coprotheres), and Turkey Vultures (Cathartes aura) showed a trend toward toxicity when the half-life of elimination increased. We conclude that the crow is not susceptible to diclofenac and, more important, that toxicity in the Gyps species is probably related to zero-order metabolism.     

Effects of non-steroid anti-inflammatory drugs in membrane bilayers

The thermal effects of non-steroidal anti-inflammatory drugs (NSAIDs) meloxicam, tenoxicam, piroxicam and lornoxicam have been studied in dipalmitoylphosphatidylcholine (DPPC) membrane bilayers using neutral and acidic environments (pH 2.5). The strength of the perturbing effect of the drugs is summarized to a lowering of the main phase transition temperature and a broadening of the phase transition temperature as well as broadening or abolishment of the pretransition of DPPC bilayers. The thermal profiles in the two environments were very similar. Among the NSAIDs studied meloxicam showed the least perturbing effect. The differential scanning calorimetry results (DSC) in combination with molecular modeling studies point out that NSAIDs are characterized by amphoteric interactions and are extended between the polar and hydrophobic segments of lipid bilayers. The effects of NSAIDs in membrane bilayers were also investigated using Raman spectroscopy. Meloxicam showed a gauche:trans profile similar to DPPC bilayers while the other NSAIDs increased significantly the gauche:trans ratio. In conclusion, both techniques show that in spite of the close structural similarity of the NSAIDs studied, meloxicam appears to have the lowest membrane perturbing effects probably attributed to its highest lipophilicity.     

Comparative plasma pharmacokinetics of meloxicam in sheep and goats following intravenous administration

Meloxicam, a novel cyclooxygenase-2 selective nonsteroidal anti-inflammatory drug (NSAID), has been used extensively in humans and recently in some domestic animal species. Although it is an attractive NSAID for use in small ruminants, meloxicam pharmacokinetics have not been investigated in sheep and goats and this information is essential for rational therapeutic use of the drug in these species. In this investigation, comparative pharmacokinetic properties of meloxicam were studied in sheep and goats after a single intravenous dose of 0.5 mg kg(-1) body mass. Blood samples were collected via jugular venepuncture into heparinised tubes at predetermined times after drug administration. Plasma concentrations of meloxicam were determined by reversed-phase high performance liquid chromatography. The plasma concentrations of meloxicam were detectable in sheep and goats up to 72 and 48 h, respectively. The plasma concentration versus time data of meloxicam in both sheep and goats were adequately described by a two-compartment open model. The values obtained for sheep and goats for distribution half-life, volume of distribution at steady state and volume of the central compartment were almost similar in sheep and goats. The elimination half-life (t(1/2beta)), area under the plasma concentration-time curve (AUC), mean residence time (MRT) and total systemic clearance (Cl(B)) in sheep were significantly different from those of goats. The mean+/-S.E. values of t(1/2beta), MRT, AUC and Cl(B) in sheep were 10.85+/-1.21 h, 15.13+/-1.67 h, 31.88+/-2.97 microg h mL(-1) and 0.016+/-0.002 L h(-1) kg(-1), respectively whereas the respective values in goats were 6.73+/-0.58 h, 9.37+/-0.83 h, 19.23+/-2.23 microg h mL(-1) and 0.03+/-0.01 L h(-1) kg(-1). The results indicate that elimination kinetics of meloxicam differ significantly between sheep and goats and the elimination of the drug tends to be faster in goats compared to sheep.     

Thermal Nociceptive Threshold Testing Detects Altered Sensory Processing in Broiler Chickens with Spontaneous Lameness

Lameness is common in commercially reared broiler chickens but relationships between lameness and pain (and thus bird welfare) have proved complex, partly because lameness is often partially confounded with factors such as bodyweight, sex and pathology. Thermal nociceptive threshold (TNT) testing explores the neural processing of noxious stimuli, and so can contribute to our understanding of pain. Using an acute model of experimentally induced articular pain, we recently demonstrated that TNT was reduced in lame broiler chickens, and was subsequently attenuated by administration of Non-Steroidal Anti-Inflammatory Drugs (NSAIDs). This study extended these findings to a large sample of commercial broilers. It examined factors affecting thermal threshold (Part 1) and the effect of an NSAID drug (meloxicam, 5 mg/kg) and of an opioid (butorphanol; 4 mg/kg) (Part 2). Spontaneously lame and matched non-lame birds (n = 167) from commercial farms were exposed to ramped thermal stimulations via a probe attached to the lateral aspect of the tarsometatarsus. Baseline skin temperature and temperature at which a behavioural avoidance response occurred (threshold) were recorded. In Part 1 bird characteristics influencing threshold were modelled; In Part 2 the effect of subcutaneous administration of meloxicam or butorphanol was investigated. Unexpectedly, after accounting for other influences, lameness increased threshold significantly (Part 1). In Part 2, meloxicam affected threshold differentially: it increased further in lame birds and decreased in non-lame birds. No effect of butorphanol was detected. Baseline skin temperature was also consistently a significant predictor of threshold. Overall, lameness significantly influenced threshold after other bird characteristics were taken into account. This, and a differential effect of meloxicam on lame birds, suggests that nociceptive processing may be altered in lame birds, though mechanisms for this require further investigation.     

Modulation of NSAID-induced antinociceptive and anti-inflammatory effects by alpha2-adrenoceptor agonists with gastroprotective effects

Tizanidine, an alpha2-adrenergic receptor agonist with myospasmolytic action, is indicated for the treatment of back pain either as monotherapy or in combination with nonsteridal anti-inflammatory drugs (NSAIDs). Tizanidine (0.25-1.0 mg/kg) significantly produced analgesic and anti-inflammatory effect in acetic acid induced writhing in mice and carrageenan-induced paw edema in rats, respectively. The effects were comparable with clonidine (0.25 and 0.50 mg/kg), another alpha2-agonist. Yohimbine (1 mg/kg), alpha2-adrenergic antagonist reversed the effect of tizanidine. Tizanidine (0.25 mg/kg) and clonidine (0.25 mg/kg) significantly potentiated the antinociceptive and anti-inflammatory effect of NSAIDs (nimesulide, meloxicam and naproxen). Tizanidine (1 mg/kg) did not alter basal pH, acidity (free and total) of gastric content and did not produce any mucosal injury in fasted rats. Tizanidine (1 mg/kg) significantly reduced meloxicam (UD50 3.21 mg/kg), nimesulide (UD50 24.52 mg/kg) and naproxen (UD50 14.10 mg/kg)-induced ulcerogenic effect (ulcer index, pH and free/total acidity). It is expected that tizanidine exerted gastrotprotection through stimulation of gastric and central alpha2-adrenergic receptors. Present investigation suggested that tizanidine not only enhance the analgesic and anti-inflammatory effect of NSAIDs but also improved gatstrointestinal tolerability of NSAIDs through modulation of central alpha-2-receptors. From this study, it can be speculated that tizanidine and NSAID combination therapy would prove to be a novel approach to treat nociceptive/inflammatory conditions with improved gastric tolerability of NSAIDs.     

Proteomic analysis in NSAIDs-treated primary cardiomyocytes

NSAIDs (non-steroidal anti-inflammatory drugs) are widely used for the treatment of a variety of inflammatory diseases, but many of them were withdrawn from the market due to their cardiovascular toxicity. In this study, we tried to identify proteins responding to the cellular toxicity in NSAIDs-treated primarily cultured cardiomyocytes using 2-D proteomic analysis. We used seven different NSAIDs (celecoxib, rofecoxib, valdecoxib, diclofenac, naproxen, ibuprofen, and meloxicam) possessing each different degree of cardiovascular risk. Overall protein spots were similar in all NSAIDs-treated cells although numbers of decreased proteins were about 2-fold higher in celecoxib or rofecoxib-treated cells than in cells incubated with other NSAIDs. Many stress-related proteins, cardiac muscle movement proteins and proteins involved in membrane organization have been isolated. Among them, Septin-8, a filament scaffolding protein, showed its specific expression pattern depending on the extent of drug toxicity. Its expression level was low in cells treated by relatively high toxic drugs such as celecoxib, diclofenac, valdecoxib, and rofecoxib. On the contrary, Septin-8 was similarly expressed in control cells in the presence of less toxic drugs such ibuprofen, naproxen, and meloxicam. This data suggests that Septin-8 differentially responds to each NSAID.     

Oxicams Bind in a Novel Mode to the Cyclooxygenase Active Site via a Two-water-mediated H-bonding Network

Oxicams are widely used nonsteroidal anti-inflammatory drugs (NSAIDs), but little is known about the molecular basis of the interaction with their target enzymes, the cyclooxygenases (COX). Isoxicam is a nonselective inhibitor of COX-1 and COX-2 whereas meloxicam displays some selectivity for COX-2. Here we report crystal complexes of COX-2 with isoxicam and meloxicam at 2.0 and 2.45 angstroms, respectively, and a crystal complex of COX-1 with meloxicam at 2.4 angstroms. These structures reveal that the oxicams bind to the active site of COX-2 using a binding pose not seen with other NSAIDs through two highly coordinated water molecules. The 4-hydroxyl group on the thiazine ring partners with Ser-530 via hydrogen bonding, and the heteroatom of the carboxamide ring of the oxicam scaffold interacts with Tyr-385 and Ser-530 through a highly coordinated water molecule. The nitrogen atom of the thiazine and the oxygen atom of the carboxamide bind to Arg-120 and Tyr-355 via another highly ordered water molecule. The rotation of Leu-531 in the structure opens a novel binding pocket, which is not utilized for the binding of other NSAIDs. In addition, a detailed study of meloxicam·COX-2 interactions revealed that mutation of Val-434 to Ile significantly reduces inhibition by meloxicam due to subtle changes around Phe-518, giving rise to the preferential inhibition of COX-2 over COX-1.     

Pharmacokinetic interactions of flunixin meglumine and enrofloxacin in ICR mice.

We examined the pharmacokinetic interactions of enrofloxacin and flunixin in male ICR mice that were subcutaneously (SC) administered with both or either one of the drugs. The experiments were performed on the following three groups: flunixin alone (2 mg/kg, SC), combination of flunixin (2 mg/kg, SC) and enrofloxacin (10 mg/kg, SC), and enrofloxacin alone (10 mg/kg, SC). Blood samples were collected at 5, 15 and 30 min, and 1, 2, 3, 4, 5 and 6 h after the drug administration, and the pharmacokinetic parameters of flunixin and enrofloxacin were evaluated from the plasma drug concentrations. Significant changes were detected in the pharmacokinetics of flunixin following its coadministration with enrofloxacin. Coadministration of flunixin and enrofloxacin resulted in a 41% increase of the area under the curve (AUC) and a 53% extension of the terminal half-life of flunixin; moreover, flunixin attained the maximum plasma drug concentration 2.75 times faster than when administered alone. The terminal rate constant and the maximum plasma drug concentration showed significant decreases of 34% and 33%, respectively, following the coadministration of enrofloxacin and flunixin as compared to those following the administration of flunixin alone. In contrast, no significant difference in the pharmacokinetics of enrofloxacin was detected following its coadministration with flunixin, as compared to those following the administration of enrofloxacin alone. Following the administration of enrofloxacin alone or its coadministration with flunixin, the plasma level of ciprofloxacin, the metabolite of enrofloxacin, was very low or undetectable. In conclusion, the pharmacokinetics of flunixin in ICR mice are altered by the coadministration of flunixin and enrofloxacin.     

Pharmacokinetics of Repeated Sodium Salicylate Administration to Laying Hens: Evidence for Time Dependent Increase in Drug Elimination from Plasma and Eggs

Salicylates were the first non-steroid anti-inflammatory drugs (NSAIDs) to be used in any species and are still widely used in humans and livestock. However, the data on their pharmacokinetics in animals is limited, especially after repeated administration. Evidence exist that in chickens (Gallus gallus) salicylate (SA) may induce its own elimination. The aim of this study was to investigate salicylate pharmacokinetics and egg residues during repeated administration of sodium salicylate (SS) to laying hens. Pharmacokinetics of SA was assessed during 14 d oral administration of SS at daily doses of 50 mg/kg and 200 mg/kg body weight to laying hens. On the 1^st, 7^th and 14^th d a 24 h-long pharmacokinetic study was carried out, whereas eggs were collected daily. Salicylate concentrations in plasma and eggs were determined using high-performance liquid chromatography with ultraviolet detection and pharmacokinetic variables were calculated using a non-compartmental model. Mean residence time (MRT), minimal plasma concentration (C_min, C_16h) and elimination half-life (T_1/2el) of SA showed gradual decrease in layers administered with a lower dose. Total body clearance (Cl_B) increased. Layers administered with the higher dose showed a decrease only in the T_1/2el. In the low dose group, SA was found only in the egg white and was low throughout the experiment. Egg whites from the higher dose group showed initially high SA levels which significantly decreased during the experiment. Yolk SA levels were lower and showed longer periods of accumulation and elimination. Repeated administration of SS induces SA elimination, although this effect may differ depending on the dose and production type of a chicken. Decreased plasma drug concentration may have clinical implications during prolonged SS treatment.     

Interaction of oxicam NSAIDs with DMPC vesicles: differential partitioning of drugs

Small unilamellar vesicles (SUVs) formed by the dimyristoylphosphatidylcholine (DMPC), a phospholipid; serve as a membrane mimetic system that can be used to study the effect of absence of net surface charges on drug-membrane interaction. The targets of non-steroidal anti-inflammatory drugs (NSAIDs) are cyclooxygenases, which are membrane active enzymes. Hence, to approach their targets NSAIDs have to pass different bio-membranes. Different membrane parameters are expected to guide the first level of interaction of these drugs before they are presented to their targets. Our earlier studies have demonstrated the crucial role of surface charges of membrane mimetic systems like micelles and mixed micelles on the interaction of oxicam NSAIDs. In order to see whether net surface charges of membranes are essential for the interaction of oxicam NSAIDs, we have studied the incorporation of two oxicam NSAIDs, viz., piroxicam and meloxicam in DMPC vesicles using the intrinsic fluorescence properties of the drugs. To see whether different prototropic forms of the drugs can interact with DMPC vesicles, studies were carried out under different pH conditions. Transmission electron microscopy (TEM) was used to characterize the SUVs those were formed at different pH values. Steady state fluorescence anisotropy measurements show that both forms of the two drugs, viz., global neutral and anion can be incorporated into the DMPC vesicles. Partition coefficient (KP) between DMPC and the aqueous buffer used has been calculated in all cases from fluorescent intensity measurements. The KP values for the neutral and anionic forms of piroxicam are 219.0 and 25.8, respectively, and that for meloxicam are 896.7 and 110.2, respectively. From the KP values it is evident that irrespective of the nature of the prototropic forms, meloxicam has a higher KP value than piroxicam. This correlates with the previously calculated log KP values between n-octanol and aqueous phase, which demonstrates that in absence of net surface charges of DMPC vesicles the hydrophobic interaction is the principal driving force for incorporation. Our results imply that for bio-membranes having no net surface charges hydrophobic effect plays a principal role to guide these NSAIDs to their targets.     

Incorporation of NSAIDs in micelles: implication of structural switchover in drug-membrane interaction

Non-steroidal anti-inflammatory drugs (NSAIDs) of oxicam group are not only effective as anti-inflammatory agents but also show diverse functions. Their principal targets are cyclooxygenases, which are membrane-associated enzymes. To bind with the targets these drugs have to pass through the membrane and hence their interactions with biomembranes should play a major role in guiding their interactions with cyclooxygenases. Here we have studied the interactions of three NSAIDs of oxicam group viz. piroxicam, meloxicam and tenoxicam with micelles having different headgroup charges, as simple membrane mimetic systems. Spectroscopic methods have been used to understand the interaction of these drugs with Cetyl N,N,N-trimethyl ammonium bromide (cationic), Sodium dodecyl sulphonate (anionic) and Triton X-100 (neutral) micelles. Our results demonstrate that the environment of the drugs i.e. the nature of the micelles plays a decisive role in choosing a specific prototropic form of the drugs for incorporation. Additionally it induces a switch over or change between different prototropic forms of piroxicam, which is correlated with the change in their reactivities in presence of different surface charges, given by the change in pK(a) values. These results together, indicate that in vivo, the diverse nature of biomembranes might play a significant role in choosing the particular form of oxicam NSAIDs that would be presented to their targets.     

In vitro scavenging activity for reactive oxygen and nitrogen species by nonsteroidal anti-inflammatory indole, pyrrole, and oxazole derivative drugs

This study was undertaken to evaluate the scavenging activity for reactive oxygen species (ROS) and reactive nitrogen species (RNS) by several nonsteroidal anti-inflammatory drugs (NSAIDs), namely indole derivatives (indomethacin, acemetacin, etodolac), pyrrole derivatives (tolmetin and ketorolac), and an oxazole derivative (oxaprozin). The inhibition of prostaglandin synthesis constitutes the primary mechanism of the anti-inflammatory action of these drugs. Nevertheless, it has been suggested that the anti-inflammatory activity of NSAIDs may be also partly due to their ability to scavenge ROS and RNS and to inhibit the respiratory burst of neutrophils triggered by various activator agents. Thus, the scavenging activity of these NSAIDs was evaluated against an array of ROS (O(2)(-), HO, HOCl, and ROO) and RNS (NO and ONOO(-)) using noncellular in vitro systems. The results obtained demonstrated that tolmetin, ketorolac, and oxaprozin were not active against O(2)(-), while acemetacin, indomethacin, and etodolac exhibited concentration-dependent effects. Oxaprozin was also the least active scavenger for HO, among all the tested NSAIDs shown to be active. The scavenging effect for HOCl was not observed for any of the tested NSAIDs. The ROO was effectively scavenged by etodolac, with the other tested NSAIDs being much less active. NO and ONOO(-) were scavenged by all the tested NSAIDs. These effects may strongly contribute to the anti-inflammatory therapy benefits that may be attained with some of the studied NSAIDs.     

Successful infection of pigeons and chickens with Histoplasma capsulatum

Summary The first repeatedly successful infection withHistoplasma capsulatium of pigeons and chickens is reported.From 44 intravenously infected pigeonsHistoplasma capsulatum could be recovered by culture in 20 cases as long as 45 days after injection; from 10 chickens 5 positive cultures were obtained.This research was in part supported by grant E-576 of the National Institutes of Health.     

Removing the threat of diclofenac to critically endangered Asian vultures

Veterinary use of the nonsteroidal anti-inflammatory (NSAID) drug diclofenac in South Asia has resulted in the collapse of populations of three vulture species of the genusGyps to the most severe category of global extinction risk. Vultures are exposed to diclofenac when scavenging on livestock treated with the drug shortly before death. Diclofenac causes kidney damage, increased serum uric acid concentrations, visceral gout, and death. Concern about this issue led the Indian Government to announce its intention to ban the veterinary use of diclofenac by September 2005. Implementation of a ban is still in progress late in 2005, and to facilitate this we sought potential alternative NSAIDs by obtaining information from captive bird collections worldwide. We found that the NSAID meloxicam had been administered to 35 captiveGyps vultures with no apparent ill effects. We then undertook a phased programme of safety testing of meloxicam on the African white-backed vultureGyps africanus, which we had previously established to be as susceptible to diclofenac poisoning as the endangered AsianGyps vultures. We estimated the likely maximum level of exposure (MLE) of wild vultures and dosed birds by gavage (oral administration) with increasing quantities of the drug until the likely MLE was exceeded in a sample of 40G. africanus. Subsequently, sixG. africanus were fed tissues from cattle which had been treated with a higher than standard veterinary course of meloxicam prior to death. In the final phase, ten Asian vultures of two of the endangered species(Gyps bengalensis,Gyps indicus) were dosed with meloxicam by gavage; five of them at more than the likely MLE dosage. All meloxicam-treated birds survived all treatments, and none suffered any obvious clinical effects. Serum uric acid concentrations remained within the normal limits throughout, and were significantly lower than those from birds treated with diclofenac in other studies. We conclude that meloxicam is of low toxicity toGyps vultures and that its use in place of diclofenac would reduce vulture mortality substantially in the Indian subcontinent. Meloxicam is already available for veterinary use in India.     

Association of Individual Non-Steroidal Anti-Inflammatory Drugs and Chronic Kidney Disease: A Population-Based Case Control Study

Background

Non-steroidal anti-inflammatory agents (NSAIDs) are known to be associated with renal damage. No clear evidence exists regarding differential risk of chronic kidney disease (CKD), specifically, across various NSAIDs.

Aim

The aim of this population-based case-control study was to evaluate the association between use of individual NSAIDs and risk of CKD in a general population of Southern Italy.

Methods

A nested case-control study was carried out using the general practice Arianna database, identifying incident CKD patients as cases and matched controls from 2006 to 2011. The date of first CKD diagnosis was defined as the index date (ID). Conditional logistic regressions were performed to estimate the risk of CKD associated with NSAIDs by class and individual drugs as compared to non-use during different time windows (within one year, six or three months prior to ID), with the latter being defined as current users. Among current users, the effect of cumulative exposure to these drugs was evaluated.

Results

Overall, 1,989 CKD cases and 7,906 matched controls were identified. A statistically significant increase in the risk of CKD was found for current users of oxicams (adjusted OR: 1.68; 95% CI: 1.15-2.44) and concerning individual compounds, for ketorolac (adj. OR: 2.54; 95% CI: 1.45-4.44), meloxicam (adj. OR: 1.98; 95% CI: 1.01-3.87) and piroxicam (adj. OR: 1.95; 95% CI: 1.19-3.21).

Conclusions

The risk of CKD varies across individual NSAIDs. Increased risk has been found for ketorolac, which may precipitate subclinical CKD through acute renal damage, and long-term exposure to oxicams, especially meloxicam and piroxicam.     

Carbachol interactions with nonsteroidal anti-inflammatory drugs

The inhibition of cyclooxygenase enzymes by nonsteroidal anti-inflammatory drugs (NSAIDs) does not completely explain the antinociceptive efficacy of these agents. It is known that cholinergic agonists are antinociceptive, and this study evaluates the interactions between carbachol and some NSAIDs. Antinociceptive activity was evaluated in mice by the acetic acid writhing test. Dose-response curves were constructed for NSAIDs and carbachol, administered either intraperitoneally (i.p.) or intrathecally (i.t.). The interactions of carbachol with NSAIDs were evaluated by isobolographic analysis after the simultaneous administration of fixed proportions of carbachol with each NSAID. All of the drugs were more potent after spinal than after systemic administration. The combinations of NSAIDs and carbachol administered i.p. were supra-additive; however, the i.t. combinations were only additive. Isobolographic analysis of the coadministration of NSAIDs and carbachol and the fact that atropine antagonized the synergistic effect suggest that carbachol may strongly modulate the antinociceptive activity of NSAIDs; thus, central cholinergic modulation would be an additional mechanism for the antinociceptive action of NSAIDs, unrelated to prostaglandin biosynthesis inhibition.     

Investigation of shiga toxin-producing Escherichia coli in avian species in India

AIMS: To investigate the presence or absence of shiga toxin-producing Escherichia coli (STEC) in avian species in India. METHODS AND RESULTS: Faecal samples originating from 500 chicken and 25 free flying pigeons were screened for the presence of E. coli. A total of 426 (chicken, 401; pigeons, 25) E. coli strains were isolated. Of 426 E. coli strains, 387 were grouped into 77 serogroups, while 70 and 59 strains were untypable and rough, respectively. All isolates were subjected to multiplex polymerase chain reaction (m-PCR) for the detection of stx(1), stx(2), eaeA, hlyA and saa genes. None of the E. coli strains studied showed the presence of stx(1), stx(2) or their variants and saa genes. Overall 11 (2.74%) and seven (1.74%) strains from chickens possessed eaeA and hlyA genes, respectively, while as only six (1.49%) strains from chickens possessed both eaeA and hlyA genes. O9, O8, O60 and O25 serogroups were most predominant of which there were 24 (5.63%), 23 (5.39%), 23 (5.39%) and 20 (4.69%) strains, respectively. None of the isolates from pigeons showed the presence of any of the virulence genes studied. CONCLUSIONS: STEC are absent in chickens and pigeons. However, further studies are required in this direction to confirm or contradict our findings. E. coli strains originating from birds are carrying a low percentage eaeA or hlyA genes. SIGNIFICANCE AND IMPACT OF THE STUDY: The present study is the first attempt to investigate STEC in chickens and free flying pigeons in India. The chickens and pigeons cannot be considered as important carrier of STEC in India.     

The Safety and Pharmacokinetics of Carprofen,Flunixin and Phenylbutazone in the Cape Vulture (Gyps coprotheres) following Oral Exposure

The following study evaluates the overt toxic potential of carprofen (CRP), flunixin (FXN) and phenylbutazone (PBZ) in Old world vultures in relation to historic toxicity data for diclofenac and ketoprofen, with the Cape vulture (Gyps coprotheres) being the indicator species. The toxic potential of a single oral dose of CRP (11.5 mg/kg), FXN (1 mg/kg),PBZ (1.7 mg/kg) or water was evaluated by means of a four-way parallel study (n = 2), as means of ascertaining if these drugs were as toxic as diclofenac in the vulture. No unscheduled deaths or pathological lesions were noted following exposure. Clinical signs of lethargy and depression were, however, noted in one CRP, two FXN and one PBZ treated birds. Mild reversible inhibition of UA excretion was evident in all three groups, although UA remained within the population reference interval in contrast to the effects previously described for diclofenac and ketoprofen. All treatment groups had a drug concentration responsive increase in alanine transferase activity. CRP, FXN and PBZ were characterised by a maximum plasma concentration (Cmax) of 1051.8 ± 620.7 ng/ml, 335.9 ± 36.3 ng/ml and 11150 ± 2474.9 ng/ml at 4 ± 4.3, 0.45 ± 0.02 and 5.3 ± 5.2 hours (Tmax) respectively and a half-life of elimination of 13.3 ±5, 1.8±1 and 18.7 ±11.4 hours respectively. While we could not demonstrate a lethal effect of the tested substances, the presence of toxic clinical signs, clinical pathological changes and/or long half-lives of elimination suggests that all three drugs have a potential for toxicity in a larger population or on repeat administration. In conclusion while the studied substances were not as overtly toxic as diclofenac, they are of safety concern.