Horses are companion animals that are reared for multipurpose such as riding, racing, plowing, pulling carts, etc.Therefore, the population of horses is increasing in Punjab.Horses are very sensitive to internal parasites and may frequently be prone to infection caused by small strongyles.Nematodes are responsible for poor health, reduced growth, weight loss, anorexia, diarrhea, gestational lesions, liver damage, and a high mortality rate.The management strategies practiced by the farm owners have an impact on the prevalence of parasitic infections in animals.Farmers adopt different management practices for the well-being of horses.Anthelmintic drugs are the primary control method for their prevention.The drugs used for parasite control give varying efficacy.Therefore, a study was planned to conduct a questionnaire-based survey in Punjab to estimate the stud farmer's practices for parasite control and to check the efficacy of 6 brands of drugs commercially available in Punjab.The data of the questionnaire was recorded on Google Forms and fecal samples were also collected from various stud farms for egg hatch assay.Based on the survey, Systamax ® was the most widely accepted brand of anthelmintic by farmers for the treatment of helminth infections in horses (82.98%).In the egg hatch assay, it is estimated that Parazole ® had the lowest dose (0.2 mg/mL) at which 50% inhibition of cyathostomins egg hatching occurred.While Oxafax ® showed the highest dose (2.4 mg/mL) at which 50% inhibition of egg hatching occurred.In vitro, the efficacy of Oxafax ® was less as compared to other brands.However, the dose-response of all six brands had a non-significant difference.It is concluded that the different strategies adopted by horse owners had a role in the prevalence of parasitic infections in horses and all six brands had varying efficacy against small nematodes.

Although early reperfusion after stroke salvages the still-viable ischemic tissue, peri-infarct selective neuronal loss (SNL) can cause sensorimotor deficits (SMD). We designed a longitudinal protocol to assess the effects of cytoprotectants on SMD, microglial activation (MA) and SNL, and specifically tested whether the KCa3.1-blocker TRAM-34 would prevent SNL. Spontaneously hypertensive rats underwent 15 min middle-cerebral artery occlusion and were randomized into control or treatment group, which received TRAM-34 intraperitoneally for 4 weeks starting 12 h after reperfusion. SMD was assessed longitudinally using the sticky-label test. MA was quantified at day 14 using in vivo [11C]-PK111195 positron emission tomography (PET), and again across the same regions-of-interest template by immunofluorescence together with SNL at day 28. SMD recovered significantly faster in the treated group (p = 0.004). On PET, MA was present in 5/6 rats in each group, with no significant between-group difference. On immunofluorescence, both SNL and MA were present in 5/6 control rats and 4/6 TRAM-34 rats, with a non-significantly lower degree of MA but a significantly (p = 0.009) lower degree of SNL in the treated group. These findings document the utility of our longitudinal protocol and suggest that TRAM-34 reduces SNL and hastens behavioural recovery without marked MA blocking at the assessed time-points.

Introduction: We have previously shown that 15min distal MCAo in the spontaneously hypertensive rat (SHR) consistently induces isolated selective neuronal loss (SNL) and topographically congruent microglial activation (MA), together with slowly recovering sensori-motor deficit (SMD) (Ejaz et al, Stroke , 2015). Whether MA exacerbates SNL in the reperfused penumbra is unclear. We tested whether reducing MA by means of the KCa3.1 blocker TRAM-34 also impacts SNL and SMD in this model. Methods: 15min microclip distal MCAo was performed in 12 isoflurane-anesthetized adult male SHRs, randomized in two groups of n=6 rats, a control group and a treated group that received TRAM-34 40mg/kg i.p. bd for 1 week starting 12hrs post-MCAo, then od for 3 weeks. To quantify SMD, the sticky label test was administered 3 times a week for 28 days. NeuN and Iba4 immunoflorescence was then performed. Manual delineation of areas of SNL and MA was carried out by two independent assessors on randomly presented IF sections, and volumes of SNL and MA were determined. Both assessments were done blinded to subject’s group. Results: Although less so than in controls, SMD was significant in the TRAM-34 group (p<.001 and .01, respectively), with a significant Group x Time interaction (p=0.004, repeated-measures ANOVA) and a trend for faster recovery from day 7 in the latter. On IF, the volume (mm 3 ) of MA was lower in the TRAM-34 relative to control group (1.33 ± 1.7 vs 0.27 ± 0.46), which also applied to SNL (1.41 ± 1.79 vs 0.28 ± 0.41) (p = 0.018 and 0.015, respectively; two-sample t-test). Conclusion: In this pilot study, the expected marked reduction in MA induced by TRAM34 was associated with concomitant reductions in SNL and faster SMD recovery. Thus, the early administration of an MA blocker following treatment-induced reperfusion in the clinical setting might reduce cell damage in the salvaged penumbra and in turn enhance neurological recovery.

Background Mapping the hypoxic brain in acute ischemic stroke has considerable potential for both diagnosis and treatment monitoring. PET using 18 F-fluoro-misonidazole (FMISO) is the reference method; however, it lacks clinical accessibility and involves radiation exposure. MR-based T2′ mapping may identify tissue hypoxia and holds clinical potential. However, its validation against FMISO imaging is lacking. Here we implemented back-to-back FMISO-PET and T2′ MR in rodents subjected to acute middle cerebral artery occlusion. For direct clinical relevance, regions of interest delineating reduced T2′ signal areas were manually drawn. Methods Wistar rats were subjected to filament middle cerebral artery occlusion, immediately followed by intravenous FMISO injection. Multi-echo T2 and T2* sequences were acquired twice during FMISO brain uptake, interleaved with diffusion-weighted imaging. Perfusion-weighted MR was also acquired whenever feasible. Immediately following MR, PET data reflecting the history of FMISO brain uptake during MR acquisition were acquired. T2′ maps were generated voxel-wise from T2 and T2*. Two raters independently drew T2′ lesion regions of interest. FMISO uptake and perfusion data were obtained within T2′ consensus regions of interest, and their overlap with the automatically generated FMISO lesion and apparent diffusion coefficient lesion regions of interest was computed. Results As predicted, consensus T2′ lesion regions of interest exhibited high FMISO uptake as well as substantial overlap with the FMISO lesion and significant hypoperfusion, but only small overlap with the apparent diffusion coefficient lesion. Overlap of the T2′ lesion regions of interest between the two raters was ∼50%. Conclusions This study provides formal validation of T2′ to map non-core hypoxic tissue in acute stroke. T2′ lesion delineation reproducibility was suboptimal, reflecting unclear lesion borders.

Purpose Mapping brain hypoxia is a major goal for stroke diagnosis, pathophysiology and treatment monitoring. 18F-fluoro-misonidazole (FMISO) positron emission tomography (PET) is the gold standard hypoxia imaging method. Normobaric hyperoxia (NBO) is a promising therapy in acute stroke. In this pilot study, we tested the straightforward hypothesis that NBO would markedly reduce FMISO uptake in ischemic brain in Wistar and spontaneously hypertensive rats (SHRs), two rat strains with distinct vulnerability to brain ischemia, mimicking clinical heterogeneity. Methods Thirteen adult male rats were randomized to distal middle cerebral artery occlusion under either 30% O2 or 100% O2. FMISO was administered intravenously and PET data acquired dynamically for 3hrs, after which magnetic resonance imaging (MRI) and tetrazolium chloride (TTC) staining were carried out to map the ischemic lesion. Both FMISO tissue uptake at 2-3hrs and FMISO kinetic rate constants, determined based on previously published kinetic modelling, were obtained for the hypoxic area. In a separate group (n = 9), tissue oxygen partial pressure (PtO2) was measured in the ischemic tissue during both control and NBO conditions. Results As expected, the FMISO PET, MRI and TTC lesion volumes were much larger in SHRs than Wistar rats in both the control and NBO conditions. NBO did not appear to substantially reduce FMISO lesion size, nor affect the FMISO kinetic rate constants in either strain. Likewise, MRI and TTC lesion volumes were unaffected. The parallel study showed the expected increases in ischemic cortex PtO2 under NBO, although these were small in some SHRs with very low baseline PtO2. Conclusions Despite small samples, the apparent lack of marked effects of NBO on FMISO uptake suggests that in permanent ischemia the cellular mechanisms underlying FMISO trapping in hypoxic cells may be disjointed from PtO2. Better understanding of FMISO trapping processes will be important for future applications of FMISO imaging.

'True' transient ischaemic attacks are characterized not only clinically, but also radiologically by a lack of corresponding changes on magnetic resonance imaging. During a transient ischaemic attack it is assumed that the affected tissue is penumbral but rescued by early spontaneous reperfusion. There is, however, evidence from rodent studies that even brief focal ischaemia not resulting in tissue infarction can cause extensive selective neuronal loss associated with long-lasting sensorimotor impairment but normal magnetic resonance imaging. Selective neuronal loss might therefore contribute to the increasingly recognized cognitive impairment occurring in patients with transient ischaemic attacks. It is therefore relevant to consider treatments to reduce brain damage occurring with transient ischaemic attacks. As penumbral neurons are threatened by markedly constrained oxygen delivery, improving the latter by increasing arterial O2 content would seem logical. Despite only small increases in arterial O2 content, normobaric oxygen therapy experimentally induces significant increases in penumbral O2 pressure and by such may maintain the penumbra alive until reperfusion. Nevertheless, the effects of normobaric oxygen therapy on infarct volume in rodent models have been conflicting, although duration of occlusion appeared an important factor. Likewise, in the single randomized trial published to date, early-administered normobaric oxygen therapy had no significant effect on clinical outcome despite reduced diffusion-weighted imaging lesion growth during therapy. Here we tested the hypothesis that normobaric oxygen therapy prevents both selective neuronal loss and sensorimotor deficits in a rodent model mimicking true transient ischaemic attack. Normobaric oxygen therapy was applied from the onset and until completion of 15 min distal middle cerebral artery occlusion in spontaneously hypertensive rats, a strain representative of the transient ischaemic attack-prone population. Whereas normoxic controls showed normal magnetic resonance imaging but extensive cortical selective neuronal loss associated with microglial activation (present both at Day 14 in vivo and at Day 28 post-mortem) and marked and long-lasting sensorimotor deficits, normobaric oxygen therapy completely prevented sensorimotor deficit (P < 0.02) and near-completely Day 28 selective neuronal loss (P < 0.005). Microglial activation was substantially reduced at Day 14 and completely prevented at Day 28 (P = 0.002). Our findings document that normobaric oxygen therapy administered during ischaemia nearly completely prevents the neuronal death, microglial inflammation and sensorimotor impairment that characterize this rodent true transient ischaemic attack model. Taken together with the available literature, normobaric oxygen therapy appears a promising therapy for short-lasting ischaemia, and is attractive clinically as it could be started at home in at-risk patients or in the ambulance in subjects suspected of transient ischaemic attack/early stroke. It may also be a straightforward adjunct to reperfusion therapies, and help prevent subtle brain damage potentially contributing to long-term cognitive and sensorimotor impairment in at-risk populations.

INTRODUCTION AND OBJECTIVES:Selective neuronal loss (SNL) in the reperfused penumbra may impact clinical recovery and is thus important to investigate. Brief proximal middle cerebral artery occlusion (MCAo) results in predominantly striatal SNL, yet cortical damage is more relevant given its behavioral implications and that thrombolytic therapy mainly rescues the cortex. Distal temporary MCAo (tMCAo) does target the cortex, but the optimal occlusion duration that results in isolated SNL has not been determined. In the present study, we assessed different distal tMCAo durations looking for consistently pure SNL.METHODS:Microclip distal tMCAo (md-tMCAo) was performed in ~6-month old male spontaneously hypertensive rats (SHRs). We previously reported that 45 min md-tMCAo in SHRs results in pan-necrosis in the majority of subjects. Accordingly, three shorter MCAo durations were investigated here in decremental succession, namely 30, 22, and 15 min (n = 3, 3, and 7 subjects, respectively). Recanalization was confirmed by MR angiography just prior to brain collection at 28 days and T2-weighted MRI was obtained for characterization of ischemic lesions. NeuN, OX42, and GFAP immunohistochemistry appraised changes in neurons, microglia, and astrocytes, respectively. Ischemic lesions were categorized into three main types: (1) pan-necrosis; (2) partial infarction; and (3) SNL.RESULTS:Pan-necrosis or partial infarction was present in all 30 min and 22 min subjects, but not in the 15 min group (p < 0.001), in which isolated cortical SNL was consistently present. MRI revealed characteristic hyperintense abnormalities in all rats with pan-necrosis or partial infarction, but no change in any 15 min subject.CONCLUSION:We found that 15 min distal MCAo consistently resulted in pure cortical SNL, whereas durations equal or longer than 22 min consistently resulted in infarcts. This model may be of use to study the pathophysiology of cortical SNL and its prevention by appropriate interventions.

Background and Purpose— New-definition transient ischemic attacks (TIAs) are frequent but difficult to diagnose because magnetic resonance imaging (MRI)-negative by definition. However, hidden underlying cell damage might be present and account for the reported long-lasting cognitive impairment after TIAs. Most prior rodent models of true TIA targeted the striatum or have not been fully characterized. Here we present the MRI, behavioral, and quantitative cell changes characterizing a new rodent model of true TIA targeting the more behaviorally relevant cerebral cortex. Methods— Fifteen-minute distal middle cerebral artery occlusion was performed in 29 spontaneously hypertensive rats allowed to survive for 7 to 60 days. Behavior was assessed serially using both global neurological and fine sensorimotor tests. Diffusion- and T2-weighted MRI was obtained 20 min postreperfusion and again 7 to 60 days later, and then changes in neurons and microglia were quantified across the middle cerebral artery territory using immunohistochemistry. Results— No MRI changes or pan-necrosis were observed at any time point, but patchy cortical selective neuronal loss affected 28/29 rats, regardless of survival interval, together with topographically congruent microglial activation that gradually declined over time. The Neuroscore was unchanged, but there was marked contralateral sensorimotor impairment, still recovering by day 28. Conclusions— Our new rodent model mimicking true cortical TIA is characterized by normal MRI, but consistent cortical selective neuronal loss and microglial activation and long-lasting sensorimotor deficits. By causing selective neuronal loss, TIAs and silent microemboli might affect neuronal reserve, thereby increasing long-term cognitive impairment risk. Selective neuronal loss and microglial activation might represent novel therapeutic targets that could be detectable in vivo after TIAs using appropriate imaging tracers.

Histopathologic assessment in transient middle cerebral artery occlusion (MCAo) rodent models generally lacks comprehensiveness and exposes to interobserver bias. Here we compared a novel quantitative assessment of regional infarction, selective neuronal loss (SNL) and microglial activation (MA) across the MCA territory to a previously published semiquantitative visual protocol. NeuN and OX42 immunohistochemistry was applied after either 15 or 45 minutes distal MCAo to maximize SNL and infarction, respectively. Survival times varied from 28 to 60 days to cover potential biases such as delayed tissue shrinkage. Damage was assessed using a template of 44 cytoarchitectonic regions of interest (ROIs) mapped onto a subset of digitized coronal sections spanning the MCA territory. For each ROI were obtained a semiquantitative visually determined index of histopathologic changes (method 1), and lpsilateral/contralesional ratios of remaining neurons and activated microglia cell counts (method 2). There was excellent agreement between the two methods for 28-day survival for both MCAo durations, whereas method 2 more sensitively detected subtle SNL and MA at 45 days and 60 days after 15-minute MCAo. Thus the visual method is accurate for usual degrees of ischemic damage, but absolute cell quantification is superior to detect subtle changes and should therefore be preferred in brief MCAo models, although requires optimal staining quality.

Angiogenesis is the formation of new blood vessels from the existing blood vessels and is involved in both physiological and pathological events. Pathological angiogenesis provokes in several important diseases as in inflammatory diseases, ischemic conditions, diabetic neuropathy and cancer. The discovery of new drugs from phyto-chemicals has a long history and anti-angiogenic agents from plant sources provide a platform for the development of phyto-medicines to treat/control pathological angiogenesis. The current project was designed to determine the efficacy of different concentrations of aqueous extract from Opuntia dillenii (OD) on angiogenesis by employing chicken chorioallantoic membrane (CAM) assay. Computer based 3D image probing was utilized to quantify anti-angiogenic effects of OD extract. Extremely aggravated dose dependent anti-angiogenic response was recorded in all groups treated with OD extract with significant reduction (P<0.01) in total area, diameter of the primary, secondary, tertiary blood vessels and capillary plexuses. Analysis of 3D parameters of OD treated CAMs revealed deteriorated angular spectrum and reduction in the height of Abbott curves. In addition, histological data revealed thinning of ectodermal layer and damaged extracellular matrix. The results spotlight that OD extract possess considerable anti-angiogenic potential and further characterization/isolation of diverse phyto-chemicals would turn up with the discovery of novel phyto-agents to control/mitigate pathological angiogenesis.

Infectious bursal disease (IBD) caused by non-enveloped double stranded RNA virus is an acute and contagious poultry disease. Outbreak of IBD could result in 10–75% mortality of the birds; hence it has gained socio-economic importance worldwide. Medicinal plants have shown broad spectrum anti-viral activities against RNA and DNA viruses. Moringa oleifera Lam (MOL), Phyllanthus emblicus Linn (PEL), Glycyrrhiza glabra Linn (GGL), and Eugenia jambolana Lam (EJL) are commonly available medicinal plants of the sub-continent and exhibited anti-viral potential against different viruses. Ethanolic extracts of the leaves of MOL and EJL, roots of GGL and dried fruit of PEL were investigated for their cytotoxic and anti-viral potential against IBD virus using MTT colorimetric assay and anti-viral assay. Significant anti-viral potential (P<0.001) was demonstrated at concentrations 12.5, 25, 50 and 100 µg ml−1 of GGL, PEL, MOL and EJL, respectively, with no cytotoxicity. Data also spotlighted that all tested plant extracts possess significant anti-viral potential and this trend was higher in GGL followed by PEL, MOL, and EJL. The data undoubtedly conclude that these medicinal plants contain several health beneficial phyto-chemicals which got significant anti-viral potential and effectively be utilized against IBD virus. Moreover, the outcomes of this study provide a platform on the way to discover novel anti-viral agents against IBD virus and other viruses from plant origin.

Environmental toxicants invariably affect all biological organisms resulting to sufferings ranging from subclinical to debilitating clinical conditions. This novel research aimed to determine the toxic burdens of increased environmental elements in some vital organs/tissues of the wild animals (starling, owl, crow and pigeon), exposed to air polluted environment were assessed using particle induced X-ray emission and histopathological approaches. The presence of significantly elevated amounts of elemental toxicants namely: Aluminum (Al), Chlorine (Cl), Iron (Fe), Potassium (K), Magnesium (Mg), Manganese (Mn), Silicon (Si) and Vanadium (V) from the skin, muscle, lungs, liver and kidney of sampled animals were in concurrence with the observed histopathological changes. The skin of sampled starling, owl, pigeon and crow spotlighted highly significant increase (P < 0.001) in Al, Cl, Mg and Si. Muscle samples with myodegenerative lesions and mineral depositions highlighted substantial augmentation (P < 0.001) in the amount of Al, Fe, Mn, Si and V. The lungs of starling, owl, and pigeon were severely intoxicated (P < 0.001) with increased amount of Al, Fe, K, Mn and Si producing pulmonary lesions of congestion, edema, pneumonitis and mineral debris depositions. Liver samples revealed that the sampled animals were laden with Cl, Fe, Mg, Mn and V with histopathological profound degenerative changes and hepatic necrosis. Kidney sections presented severe tubular degenerative and necrotic changes that may be attributed to increased amounts of Cl and Fe. These current findings implied that the environmental/elemental toxicants and the accompanying lesions that were discerned in the organs/tissues of sampled birds may as well be afflicting people living within the polluted area. Further assessment to more conclusively demonstrate correlations of current findings to those of the populace within the area is encouraged.

Excessive use of pesticides poses increased risks to non target species including humans. In the developing countries, lack of proper awareness about the toxic potential of pesticides makes the farmer more vulnerable to pesticide linked toxicities, which could lead to diverse pathological conditions. The toxic potential of a pesticide could be determined by their ability to induce genetic mutations and cytotoxicity. Hence, determination of genetic mutation and cytotoxicity of each pesticide is unavoidable to legislate health and safety appraisal about pesticides. The objective of current investigation was to determine the genotoxic and cytotoxic potential of Endosulfan (EN) and Lambda-cyhalothrin (LC); individually and in combination. 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) assay was utilized to determine cytotoxicity, while two mutant histidine dependent Salmonella strains (TA98, TA100) were used to determine the mutagenicity of EN and LC. Moreover, mutagenicity assay was conducted with and without S9 to evaluate the effects of metabolic activation on mutagenicity. Even though a dose dependent increase in the number of revertant colonies was detected with EN against both bacterial strains, a highly significant (p<0.05) increase in the mutagenicity was detected in TA98 with S9. In comparison, data obtained from LC revealed less mutagenic potential than EN. Surprisingly, the non-mutagenic individual-concentrations of EN and LC showed dose dependent mutagenicity when combined. Combination of EN and LC synergistically induced mutagenicity both in TA98 and TA100. MTT assay spotlighted comparable dose dependent cytotoxicity effects of both pesticides. Interestingly, the combination of EN and LC produced increased reversion and cytotoxicity at lower doses as compared to each pesticide, concluding that pesticide exposure even at sub-lethal doses can produce cytotoxicity and genetic mutations, which could lead to carcinogenicity.

Tuberculosis (TB) is one of the leading infectious causes of death due to single infectious agent after HIV/AIDS. Rifampicin (RIF), Isoniazid (INH), Ethambutol (EMB), Pyrazinamide (PZA) and/or their combinations are extensively prescribed to treat TB. Despite several therapeutic implications, these drugs also produce several toxic effects at cellular level. MTT assay and Ames test were adopted in this study for the determination of cytotoxic and mutagenic potential of these anti-TB drugs. Among all tested drugs, cytotoxic potential of RIF was strongest with highly significant decline (p<0.001) in cell numbers at the concentration of 250μg/ml with LC50 at 325μg/ml, while significant decline (p<0.01) in cell count was observed in INH treated group at the concentration 500μg/ml with LC50 at 1000μg/ml. Moreover, combination RIPE demonstrated significant reduction (p<0.01) in cell number at the concentration of 25-500-500-500μg/ml with LC50 at 60-1200-1200-1200μg/ml. It is apparent from the data that almost all drugs represented identical mutagenic pattern i.e., more significant results were achieved in TA100 with metabolic activation (+S9). RIF proved to be highly mutagenic of all tested drugs with significant mutagenicity (p<0.01) at 0.0525μg/plate against TA98 strain with S9. The combination RIPE exhibited highly significant mutagenic activity (p<0.01) at concentration 0.125-3-3-3μg/plate without S9, while addition of S9 resulted in similar activity at lower doses, i.e., 0.0525-1-1-1μg/plate. It was concluded from the data that all anti-TB drugs possess significant cytotoxic and mutagenic potential, especially in combination, making TB patient more vulnerable to cytotoxic and mutagenic effects of anti-TB drugs, which could produce further health complications in TB patients.

Major transformations in the environmental composition are principally attributable to the combustion of fuels by automobiles. Motorized gasoline-powered two-stroke auto-rickshaws (TSA) and compressed natural gas (CNG)-powered four-stroke auto-rickshaws (FSA) are potential source of air pollution in south Asia and produce toxic amount of particulate matter (PM) to the environment. In this study, we attempted to characterize elemental pollutants from the PM of TSA and FSA using proton-induced X-ray emission (PIXE) analysis. The observations of the existing investigation recognized significant increase in Al (P < 0.05), P (P < 0.01), and Zn (P < 0.01) from the PM samples of FSA. In addition, the concentrations of Cu, Fe, K, Mg, Na and S were also observed exceeding the recommended National Institute for Environmental Studies limits. On the contrary, increased concentration of Sr and V were observed in the PM samples from TSA. It is generally believed that FSA generates smaller amount of PM but data obtained from FSA are clearly describing that emissions from FSA comprised potentially more toxic substances than TSA. The current research is specific to metropolitan population and has evidently revealed an inconsistent burden of exposure to air pollutants engendered by FSA in urban communities, which could lead to the disruption of several biological activities and may cause severe damage to entire ecological system.

Introduction: Severe brain hypoxia in the territory of the occluded artery is a key feature of ischemic stroke. This region can be imaged using positron emission tomography (PET) and the standard hypoxia radiotracer F-18-fluoromisonidazole (F-18-FMISO). However, the utility of F-18-FMISO is limited by its slow accumulation in the lesion. Therefore, this study investigated three hypoxia-sensitive radiotracers, namely the nitroimidazole F-18-fluoroazomycin arabinoside (F-18-FAZA) and two Cu-64 bis(thiosemicarbazone) complexes (Cu-64-ATSM and Cu-64-ATSE), expected to have improved pharmacokinetic profiles relative to F-18-FMISO, in a rodent model of ischemic stroke.Methods: In anaesthetised Wistar rats, the distal middle cerebral artery was permanently occluded by electrocoagulation, the radiotracers administered intravenously and animals PET scanned for up to 3 hours, followed by T2-weighted magnetic resonance imaging to map the infarct.Results: As expected, late and prominent F-18-FMISO retention was observed despite lower tracer delivery into the affected region. Time-activity curves revealed that both Cu-64-ATSM and Cu-64-ATSE showed rapid entry and efflux from the brain, but did not show significant accumulation in the lesion. F-18-FAZA showed limited brain penetration, and accumulation in the lesion was inconsistent, low and as slow as F-18-FMISO.Conclusions: This study suggests further development of these radiotracers as hypoxia markers for ischemic stroke may not be warranted. Crown Copyright (C) 2013 Published by Elsevier Inc. All rights reserved.

Background and purpose: Current models dictate that, depending on occurrence of early reperfusion, the ischemic penumbra either undergoes or escapes infarction (i.e., “pan-necrosis”). However, tissue outcome following temporary middle-cerebral artery occlusion (tMCAo) in rodents can also include selective neuronal loss (SNL), which even if subtle may impede functional recovery. In order to explore the pathophysiology of ischemic stroke, determine potential therapeutic targets and monitor effects of therapy, in vivo imaging surrogates of these varied histopathological outcomes applicable in the clinical setting would be useful. Although hyperintense signal on T2-weighted MRI in the chronic post-stroke stage is considered a reliable surrogate of tissue infarction, SNL is not associated with T2W abnormal signal. In the clinical setting, the neuron-specific PET ligand 11C-flumazenil (FMZ) has been used to identify both pan-necrosis and peri-infarct SNL, but this inference has not been histopathological confirmed so far. Here we investigated the late tissue sequelae of tMCAo in the rodent using in vivo T2W MRI and FMZ-PET against post mortem immunohistochemistry as gold standard. Methods: Adult spontaneously hypertensive rats (SHRs) underwent 45 min distal-clip middle-cerebral artery occlusion and, 28 days later, FMZ-PET and T2W-MRI, immediately followed by immunohistochemistry for neuronal loss (NeuN), activated microglia and astrocytosis. Based on standard histopathological definitions, ischemic lesions were classified into pan-necrosis, partial infarction or SNL. NeuN changes and FMZ binding across the whole hemisphere were quantified in the same set of 44 regions-of-interest according to previously validated protocols; linear regressions between these two measures were carried out both within and across subjects. Results: Both cortical pan-necrosis/partial infarction and SNL were present in all rats except one, where SNL was isolated and extensive. Infarction/partial infarction, but not SNL, was associated with T2W hyperintense signals and cortical atrophy. In contrast, FMZ binding was decreased in all types of lesions including SNL, in proportion with NeuN staining intensity both within (p<0.05 to <0.001) and across (p<0.001) subjects, including the subject that showed pure SNL (p=0.01). Conclusion: This novel study revealed three main facts: i) long-term histopathological cortical changes following 45 min tMCAo in SHRs included all three of SNL, partial infarction and frank infarction; ii) T2W MRI showed conspicuous high signal lesions for complete or partial infarction, but no changes for SNL; and iii) FMZ-PET was sensitive to all three types of tMCAo-induced histopathological changes, including isolated SNL, suggesting it is a valid surrogate for the histological sequelae of focal cerebral ischemia. In addition, the finding of almost universal completed cortical infarction at 28 days differed from our previous findings at 14-day survival using the same model and rat strain, where SNL was the almost exclusive outcome, possibly representing delayed infarct maturation. Prospective studies are needed to investigate this interesting possibility.

Vehicular air pollution is a mounting health issue of the modern age, particularly in urban populations of the developing nations. Auto-rickshaws are not considered eco-friendly as to their inefficient engines producing large amount of particulate matter (PM), thus posing significant environmental threat. The present study was conducted to ascertain the cytotoxic, phytotoxic, and mutagenic potential of PM from gasoline-powered two-stroke auto-rickshaws (TSA) and compressed natural gas-powered four-stroke auto-rickshaws (FSA). Based on the increased amount of aluminum quantified during proton-induced X-ray emission analysis of PM from TSA and FSA, different concentrations of aluminum sulfate were also tested to determine its eco-toxicological potential. The MTT assay demonstrated significant (p < 0.001) dose-dependent cytotoxic effects of different concentrations of TSA, FSA, and aluminum sulfate on BHK-21 cell line. LC50 of TSA, FSA, and aluminum sulfate was quantified at 16, 11, and 23.8 μg/ml, respectively, establishing PM from FSA, a highly cytotoxic material. In case of phytotoxicity screening using Zea mays, the results demonstrated that all three tested materials were equally phytotoxic at higher concentrations producing significant reduction (p < 0.001) in seed germination. Aluminum sulfate proved to be a highly phytotoxic agent even at its lowest concentration. Mutagenicity was assessed by fluctuation Salmonella reverse mutation assay adopting TA100 and TA98 mutant strains with (+S9) and without (−S9) metabolic activation. Despite the fact that different concentrations of PM from both sources, i.e., TSA and FSA were highly mutagenic (p < 0.001) even at lower concentrations, the mutagenic index was higher in TSA. Data advocate that all tested materials are equally ecotoxic, and if the existing trend of atmospheric pollution by auto-rickshaws is continued, airborne heavy metals will seriously affect the normal growth of local inhabitants and increased contamination of agricultural products, which will amplify the dietary intake of the toxic elements and could result in genetic mutation or long-term health implications.

Vehicles are a major source of air pollution, especially particulate matter (PM) pollution, throughout the world and auto-rickshaws are considered main contributors to this air pollution. PM, in addition to causing respiratory and cardiovascular disorders, has potential to gain access to the brain and could induce neuroinflammation leading to different neurological disorders. Therefore, in the current project, MRI and immunohistochemistry techniques were adopted to ascertain the neurotoxic potential of the chronic exposure to different PM generated by two-stroke auto-rickshaws (TSA), four-stroke auto-rickshaws (FSA), and aluminum sulfate (AS) solution in rats. The results highlighted that all treated groups followed a pattern of dose-dependent increase in pure cortical neuronal loss, selective neuronal loss (SNL), nuclear pyknosis, karyolysis, and karyorrhexis. Mild to moderate areas of penumbra were also observed with increase in the population of activated microglia and astrocytes, while no alteration in the intensities of T2W MRI signals was perceived in any group. When comparing the findings, TSA possess more neurotoxic potential than FSA and AS, which could be associated with increased concentration of certain elements in TSA emissions. The study concludes that chronic exposure to PM from TSA, FSA, and AS solutions produces diverse neuropathies in the brain, which may lead to different life-threatening neurological disorders like stroke, Alzheimer's, and Parkinson's disorders. Government and environmental agencies should take serious notice of this alarming situation, and immediate steps should be implemented to improve the standards of PM emissions from auto-rickshaws.

The aim of this novel research was to determine the toxic burden of increased elements in water resources on the inhabitant wild animals (squirrels, turtles, bats), using particle induced x-ray emission (PIXE) and histopathological approaches. PIXE analysis of skin, muscle, lung, liver and kidney revealed significant increase in Al, Cl, Fe, Mg, Mn, Si and V. Moreover, data clearly reflect a significant (P < 0.001) deposition of toxic elements (Al, Cl, Fe and K) in the lung producing interstitial/proliferative pneumonitis, intra-alveolar hemorrhages, and thickening of alveolar capillary walls. The results obtained from the liver samples emphasized that majority of the animals were intoxicated with Cl, Mg, S, Si and V, which have produced profound deterioration and swelling of the hepatocytes. Likewise, histopathology of the kidney sections spotlighted severe nephritis and degenerative changes, which could be associated with the elevated amount of Al, Cl and Mg. This data undoubtedly provide relevant information on the heavy burden of toxic elements and their pathological outcomes in wild animals and highlight their potential risks for human exposure. Thus, the information provided is critical for developing effective strategies in dealing with health hazards associated with elemental exposures.