Normobaric hyperoxia markedly reduces brain damage and sensorimotor deficits following brief focal ischaemia.

'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.

Cortical selective neuronal loss, impaired behavior, and normal magnetic resonance imaging in a new rat model of true transient ischemic attacks.

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.

Monitoring impacts of air pollution: PIXE analysis and histopathological modalities in evaluating relative risks of elemental contamination.

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.

Mutagenic and cytotoxic potential of Endosulfan and Lambda-cyhalothrin - in vitro study describing individual and combined effects of pesticides.

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.

Characterizing infarction and selective neuronal loss following temporary focal cerebral ischemia in the rat: a multi-modality imaging study.

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 T(2)-weighted MRI in the chronic post-stroke stage is considered a reliable surrogate of tissue infarction, SNL is not associated with T(2)W abnormal signal. In the clinical setting, the neuron-specific PET ligand (11)C-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 T(2)W 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 T(2)W-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 T(2)W 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.

Vascular and morphogenetic abnormalities associated with exposure of cigarette smoke condensate during chicken and murine embryogenesis.

OBJECTIVE: Embryonic movements (EM) and angiogenesis pathways are evolutionarily conserved mechanisms which are essential for proper embryonic development. Deviations in these processes by exposure to cigarette smoke condensate (CSC) may cause vascular and morphogenetic disorders. METHODS: Using chicken and mouse embryos, we have demonstrated the in vivo effects of CSC on EM, vascular development, and organogenesis. RESULTS: Examination of the CSC exposed chicken embryos revealed a significant reduction in EM, stunted growth, deviated pattern of blood vessels, hemorrhages, and localized necrosis. Likewise, mouse embryos that were exposed to CSC at E8.5 and E9.5 died between E11.5 and E12.5, respectively. These mouse embryos showed defects in morphogenesis and remodeling of the embryonic vasculature, while littermate controls showed normal development. CONCLUSIONS: Cigarette smoking during pregnancy is fatal for growing embryos. CSC may induce the remodeling of embryonic vasculature, leading to various pathologies.

Granulomatous pneumonia in a captive freshwater crocodile (Crocodylus johnstoni) caused by Mycobacterium szulgai.

A 25-yr-old male freshwater crocodile (Crocodylus johnstoni) was diagnosed with pulmonary mycobacteriosis caused by Mycobacterium szulgai. Necropsy revealed fibrinous exudate in the right pleural cavity and white miliary nodules in the right lung lobe. Histopathologic examination revealed well-demarcated granulomas consisting of multinucleated giant cells and epithelioid cells surrounded by fibrous connective tissue. Atypically, lymphocytes had accumulated in the outer region of fibrous connective tissue. Mycobacterial infection was confirmed by nested polymerase chain reaction targeting the hsp65 gene and by Fite's method for detection of acid-fast bacilli within formalin-fixed, paraffin-embedded lung tissue. Sequence analysis of the DNA amplicon revealed that the species of mycobacterium shared 98% homology with the gene encoding the hsp65 gene of M. szulgai. This is the first report of M. szulgai as the causative agent of mycobacteriosis in a reptile.

Effect of aged garlic extract on wound healing: a new frontier in wound management.

Successful wound healing depends upon angiogenesis, and impaired angiogenesis is a hallmark of the chronic wounds encountered with diabetes and venous or arterial insufficiency. To intervene and improve wound closure, it is essential to investigate the effects of different natural remedies in wound healing. The chicken dorsum skin excisional wound assay was used to investigate the influence of different concentrations of aged garlic solution (AGS) on wound healing. Gross, histopathology, scanning electron microscopy (SEM) and computer-based three-dimensional (3D) image-probing techniques were utilized to determine the effects of AGS on wound closure, re-epithelialization, dermal matrix regeneration, and angiogenesis. Ninety chicks, aged 1 week and divided in 6 groups, were topically exposed to different concentrations of AGS for 6 days: control (group A), 1% (group B), 5% (group C), 10% (group D), 15% (group E), and skin lotion (group F). Different patterns, ranging from incomplete to almost complete wound closure, were observed among different groups with highly significant results (P < 0.001) in group E. Histological investigations revealed a positive augment in the re-epithelialization of all AGS exposed wounds. An increase in the number of new loosely packed collagen and maturation of collagen bundles was observed in all treated wounds at days 4 and 6 post-wounding, respectively. Similar results were achieved through SEM of treated wounds. Histological investigations revealed the profuse dose-dependent neovascularization among AGS-treated wounds. Abbott curve, angular spectrum, and different parameters of 3D surface roughness of wounds were also measured for the precise quantification of angiogenesis. A very highly significant (P < 0.001) increase in angiogenesis was observed among all treated groups. No significant change was observed among control and skin lotion-treated groups. These observations substantiate the beneficial use of AGS in the treatment of wounds. Additional studies are needed to study the specific wound-healing mechanisms of chemical, or group of chemicals, present in AGS.

Total particulate matter and wound healing: an in vivo study with histological insights.

OBJECTIVES: Wound healing in the skin is a multifarious orchestration of cellular processes and cigarette smoking may be a cause for delayed wound healing. The aim of this study was to investigate the plausible association between exposures of cigarette total particulate matter (TPM) and wound healing. METHODS: An in vivo wound healing model of mice was established for determination of assorted events of wound healing, dermal matrix regeneration, re-epithelialization, and neovascularization. A total of 72 adult mice, separated in eight groups, were exposed to TPM for 12 days. RESULTS: A highly considerable diminution in wound closure (P < 0.001) was pragmatic among all TPM-treated mice from day 6 to day 8 post-wounding. Histological investigations unveiled a noteworthy impede in the outcome of re-epithelialization, dermal matrix regeneration and maturation of collagen bundles among all TPM-exposed wounds. Delayed commencement of neovascularization was pragmatic among all TPM-treated mice, on day 12 post wounding. Abbot curve, angular spectrum, and other different parameters of 3D surface behavior of wounds revealed a very highly significant reduction (P < 0.001) in angiogenesis on days 6 and 8 post-wounding, which points that application of TPM instigates extensive delay in trigging the progression of angiogenesis, resulting in delayed onset of wound healing. CONCLUSION: Our annotations validate the damaging effects of TPM on wound healing and excessive use of TPM may lead to the production of chronic wounds and oral ulcers.

Anti-angiogenic activities associated with exposure of environmental smoke solutions from 2-stroke auto-rickshaw.

Angiogenesis, the formation of new blood vessels, is vital for embryonic development and disruption of this process can be a powerful mechanism of abortion. Over the last few decades there has been increasing global concern regarding the public health impact attributed to environmental smoke pollution. However, no study has yet examined the relation between exhaust from 2-stroke auto-rickshaws and angiogenesis. The current experiment was carried out to elucidate the possible detrimental effects of 2-stroke auto-rickshaw smoke solutions (2SARSS) on physiological angiogenesis, using a well-defined chicken chorioallantoic membrane (CAM) assay. Gross computer based 3D image probing and histopathologic modalities were utilized to quantify different detrimental effects of 2SARSS on the fundamental processes of angiogenesis. Macroscopic investigations of 2SARSS treated CAMs revealed severe disruption in the orientation and normal branching pattern of the blood vessels with profound disorganization. Application of 2SARSS caused substantial decrease in the total vascular area of CAM (p<0.001) diameters of the primary, secondary (p<0.01) and tertiary blood vessels (p<0.001) as well as capillary plexuses formation (p<0.001). Evaluation of different 3D parameters of 2SARSS treated CAMs unveiled diminished surface roughness, angular distribution, and height of the Abbott curves. Moreover, histological evaluations of 2SARSS treated CAMs also revealed disruption of the normal architecture of the blood vessel with marked thinning of ectodermal layer and mesodermal extracellular matrix. The anti-angiogenic effects of 2SARSS clearly demonstrate its toxicity to those travelling and/or living in the vicinity of these vehicles and these populations may suffer from several angiogenesis related pathologies.

Toxicological evaluation of the effects of 2-stroke auto-rickshaw smoke solutions on wound healing.

Vehicle exhaust from traffic is a widespread air pollutant. The use of 3-wheel auto-rickshaws powered by a 2-stroke engine is widespread in south Asia; exhaust from these vehicles may cause different types of toxicities resulting in different pathologies. The aim of this study was to explore the association between exposure to 2-stroke auto-rickshaw smoke solution (2SARSS) and wound healing. The in vivo model of wound healing was customized to evaluate different stages of wound healing: dermal matrix regeneration, re-epithelialization, and neovascularization. A total of 72 adult mice were divided into 8 groups and exposed to 2SARSS for 12 days. A highly significant reduction (p<0.001) in wound closure was observed among all 2SARSS-treated groups at day 8 post-wounding. Histological examination revealed a significant delay in the outcome of re-epithelialization, dermal matrix regeneration, and maturation of collagen bundles among all 2SARSS-exposed wounds. Delayed activation of neovascularization was seen in the 2SARSS-treated groups at day 12 post-wounding. The Abbot curve, angular spectrum, and several other 3D surface parameters of reverse wound topographies revealed a highly significant reduction (p<0.001) in angiogenesis. These results demonstrate that application of 2SARSS causes a substantial delay in the progression of angiogenesis, resulting in delayed onset of wound healing. These observations validate the damaging effects of 2SARSS on wound healing. Thus, people who are directly or indirectly exposed to this toxic exhaust are expected to have delayed wound healing, which could result in chronic wounds.

Assessing the Effects of Cytoprotectants on Selective Neuronal Loss, Sensorimotor Deficit and Microglial Activation after Temporary Middle Cerebral Occlusion.

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.

Importance of pericytes and mechanisms of pericyte loss during diabetes retinopathy.

Pericytes are distinctive regulators of angiogenesis and are adumbrated to provide vessel stability and control of endothelial proliferation. The present article spotlights the persona of pericytes in physiological angiogenesis, recruitment of pericytes and different mechanisms of pericyte depletion. Developing retina appears particularly dependent on pericytes, and pericyte loss is considered as hallmark of early diabetic retinopathies. Several factors are contemplated to be engaged in pericyte conscription including angiopoietin-1 and its receptor tyrosine kinase Tie-2, vascular endothelial growth factor-A and its receptor flk-1 and the platelet-derived growth factor PDGF-B/PDGF-beta system. At present, the mechanisms by which diabetes persuade apoptosis in the retinal microvasculature remain indecisive, albeit oxidative stress, formation of advanced glycation end products , upregulation of protein kinase C, increased polyol pathway flux and focal leukostasis may be important. In this context, accelerated microvascular cell death may become a constructive surrogate end-point in pharmacological studies of experimental diabetic.

Cytotoxicity of fumonisin B(1) in spheroid and monolayer cultures of rat hepatocytes.

Fumonisin B(1) (FB(1)), the most prevalent member of toxins produced by several species of Fusarium molds, which occur mainly in maize, causes several fatal hepatopathies and nephropathies of animals. The current study was scrutinized to ascertain different cytotoxic and morphological transformations in rat hepatocytes induced by the treatments of diverse concentrations (300, 500, or 1000 microM) of fumonisin B(1) in vitro, using both monolayer and spheroid cultures. In each hepatocyte culture, the cytotoxicity of FB(1) was augmented in dose- and time-response manners. Morphological transformations among FB(1)-treated groups integrated accumulation of lipid droplets, cytoplasmic vacuolation in hepatocyte monolayers, and bleb formation in the hepatocyte spheroids. Additionally, electron microscopy revealed the loss of microvilli, mitochondrial swelling, and formation of lamellar membranous whorl in the vacuoles and bile canaliculi-like structures. Appearance of electron dense bodies in the monolayers, and loss of cell-to-cell contact in spheroids were depicted in 1000 microM FB(1)-treated hepatocytes. These outcomes insinuate different vital events in explaining morphological transformations in the cell membrane and organelles, induced by fumonisins in rat hepatocytes.

Effects of hyperoxia on 18F-fluoro-misonidazole brain uptake and tissue oxygen tension following middle cerebral artery occlusion in rodents: Pilot studies.

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.

Brain hypoxia mapping in acute stroke: Back-to-back T2' MR versus 18F-fluoromisonidazole PET in rodents.

Background Mapping the hypoxic brain in acute ischemic stroke has considerable potential for both diagnosis and treatment monitoring. PET using 18F-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.

Impaired wound healing by exposure of different mainstream whole smoke solutions of commercial cigarettes.

Cigarette smoke has been shown to potentiate wound damage and delayed ulcer healing. The chicken dorsum excisional wound assay was used to elucidate the deleterious effects of different mainstream whole smoke solutions (MSWSS) on the fundamental processes of wound healing. Gross, histopathology, SEM and computer based 3D image probing techniques were utilized to quantify different toxic effects of MSWSS on wound healing. A total of 160 chicks, aged 1 week, divided in eight groups were exposed to MSWSS with different nicotine concentration; 0.2mg (group A), 0.3mg (group B), 0.5mg (group C), 0.6mg (group D), 0.7mg (group E) and 1mg (group F). A very highly significant reduction (P<0.001) in wound closure was observed among all MSWSS treated groups at day 8 post-wounding. Histological investigations revealed a significant impede outcome in the re-epithelialization of all MSWSS exposed wounds. Delayed dermal matrix regeneration and maturation of collagen bundles were observed among all MSWSS treated wounds. Similar results were achieved through SEM of treated wounds. Histological and image probing analysis unveiled the scanty neovascularization among MSWSS treated wounds. Abbot curve, angular spectrum and different other parameters of 3D surface topographies of wounds revealed a very highly significant reduction (P<0.001) in angiogenesis among all MSWSS treated groups. These annotations validate the damaging effects of MSWSS on the healing of wounds.

Impact of sidestream whole smoke solutions on the outcome of wound repair and related angiogenesis.

Wound angiogenesis is essential to support the regenerating tissue and any setback in angiogenesis may result in retarded wound repair. Cigarette smoking causes numerous adverse effects, some of which are associated with poor healing. The current experiment was carried out to elucidate the possible detrimental effects of sidestream whole smoke solutions (SSWSS) on wound healing and related angiogenesis, using a well-defined chicken dorsum excision wound assay. Gross, histopathologic, SEM and computer based 3D image-probing modalities were utilized to quantify different detrimental effects of SSWSS on the fundamental processes of wound healing. A total of 160 chicks, aged 1 week, divided in eight groups were topically exposed for 8 days to SSWSS with different nicotine concentrations. At day 6 and day 8 post-wounding, very highly significant reduction (P<0.001) in wound closure was observed among all SSWSS treated groups. Histological and SEM evaluation of SSWSS treated wounds unveiled deteriorated dermal matrix, delayed re-epithelialization and retarded neovascularization. Moreover, image-probing exploration of SSWSS treated wounds also divulge a very highly significant decrease (P<0.001) in angular spectrum, Sa, Sy and Sci, at day 6 post-wounding. Our study suggests that the cumulative effect of different components of SSWSS has a negative impact on wound healing and related angiogenesis. Furthermore, our study demonstrates the effects that can contribute to abnormal healing and may explain why people who are consistently exposed to sidestream smoke suffer from slow healing and excessive scarring of wounds, much like the smokers themselves.

A novel model of image acquisition and processing for holistic quantification of angiogenesis disrupted by application of mainstream and sidestream cigarette smoke solutions.

Angiogenesis is a vital process in the growth of new blood vessels from pre-existing vasculature. Among several approaches being used for studies related to angiogenesis, chicken chorioallantoic membrane assay (CAM) is an excellent model system. However, its utility has been limited due to difficulty in quantifying putative angiogenic and anti-angiogenic response to an experimental compound in an objective and quantifiable manner. Herein, we report a novel approach of image acquisition and processing for better evaluation of neovascularization. The effects of mainstream cigarette smoke solutions (MSCSS) and sidestream cigarette smoke solutions (SSCSS) from different commercially available cigarettes on angiogenesis were quantified, using CAM assay. Different gross and nanometer scale topographies of CAMs were quantified, which are vital for 3D image scrutiny and can precisely enumerate angiogenesis. Pattern formation of blood vessels, diameter, area and 3D surface roughness of CAMs were substantially disrupted by application of cigarette-smoke extracts. An important point revealed in our study that SSCSS appeared to be significantly more toxic than MSCSS with respect to their effects on angiogenesis. This new imaging technique combined with other modalities, will provide a robust platform to optimize trial design and more patent studies in angiogenesis.

Toxicological overview of cigarette smoking on angiogenesis.

Angiogenesis is the process of generating new capillary blood vessels. It occurs under tight regulation in the female reproductive system, during wound healing and during embryogenesis. Angiogenesis also plays an important role in the pregnancy-associated changes in the reproductive tract. Cigarette smoke inhibits processes that may hinder normal process of angiogenesis resulting in abnormal blood supply to tissues, decreased repair and remodeling. This report summarizes the evidences of the causal association between tobacco smoking and disruption of angiogenesis. Application of small amount of nicotine on day 5 old chorioallantoic membranes (CAMs) did not disrupt the process of angiogenesis, while application of mainstream smokes (MSS) solutions to CAMs caused varying levels of disruption on normal process of angiogenesis and adversely affect capillary plexus formation, diameters of secondary and tertiary vessels. We have also observed that at equivalent doses, sidestream smoke (SSS) can significantly be more potent than MSS and can alter the normal process of angiogenesis more drastically than MSS. It suggests that SSS either contains a toxicant(s) not present in MSS or that the toxicant(s) that produces these effects is present in higher concentration in SSS than in MSS. Therefore, it is undisputed that smoking can interfere the normal process of angiogenesis, which is a vital process to maintain pregnancy and development of fetus. Smoking during pregnancy is harmful to fetal development and is associated with an increased risk of miscarriage, perinatal death and sudden infant death syndrome. Smoking-cessation programs remain a crucial strategy for preventing poor birth outcomes and decreasing the social and financial costs of smoking during pregnancy.