RESUMO
From previous studies, we have shown that viable colony forming units of bacteria and bacterial biofilms are reduced after sequential treatment with a surfactant-based dressing. Here, we sought to test the impact on visible bacterial pigments and the ultrastructural impact following the sequential treatment of the same surfactant-based dressing. Mature Pseudomonas aeruginosa biofilms were grown on ex vivo porcine skin explants, and an imaging-based analysis was used to compare the skin with and without a concentrated surfactant. In explants naturally tinted by bacterial chromophores, wiping alone had no effect, while the use of a surfactant-based dressing reduced coloration. Similarly, daily wiping led to increased immunohistochemical staining for P. aeruginosa antigens, but not in the surfactant group. Confocal immunofluorescent imaging revealed limited bacterial penetration and coating of the dermis and loose pieces of sloughing material. Ultrastructural analysis confirmed that the biofilms were masking the extracellular matrix (ECM), but the surfactant could remove them, re-exposing the ECM. The masking of the ECM may provide another non-inflammatory explanation for delayed healing, as the ECM is no longer accessible for wound cell locomotion. The use of a poloxamer-based surfactant appears to be an effective way to remove bacterial chromophores and the biofilm coating the ECM fibres.
Assuntos
Surfactantes Pulmonares , Lesões dos Tecidos Moles , Infecção dos Ferimentos , Animais , Suínos , Tensoativos/farmacologia , Tensoativos/uso terapêutico , Pseudomonas aeruginosa , Infecção dos Ferimentos/tratamento farmacológico , Infecção dos Ferimentos/microbiologia , Bandagens , Pele , BiofilmesRESUMO
The purpose of this study was: (1) to characterise the association of wound area, wound exudate C-reactive protein (CRP), broad-spectrum matrix metalloprotease protein (MMPs), and symptoms of fatigue and pain in individuals with chronic venous leg ulcers (CVLUs) over time and (2) to identify factors associated with the wound healing trajectory in CVLUs. Seventy four participants with CVLU who received weekly sharp debridement were recruited from a wound care clinic during the 8-week study period. To examine associations among wound CRP, MMPs, pain, fatigue, and wound healing trajectory over time, we calculated Bayes factors (BF) based on a linear mixed model. The mean age of participants was 71.8 (SD = 9.8) and the mean wound area was 2278 mm2 (SD = 7085 mm2 ) at baseline. Higher fatigue was strongly associated with higher MMPs (BF = 9, 95% HDI: [-.05, .43]), lower CRP (BF = 11, 95% HDI: [-.02, .002]), and large areas of wound (BF = 20, 95% HDI: [-.001, .01]). Higher CRP and MMPs activity in wound exudate and higher fatigue were associated with a larger wound area. To facilitate wound healing, clinicians need to utilise the multifactorial approach, which includes wound treatment and management of symptoms such as pain and fatigue, because of the molecular and psycho-behavioural factors involved in wound healing.
Assuntos
Úlcera Varicosa , Humanos , Teorema de Bayes , Úlcera Varicosa/terapia , Cicatrização , Dor/diagnóstico , Proteína C-Reativa , Fadiga/etiologia , Fadiga/terapiaRESUMO
OBJECTIVE: To investigate the therapeutic effects of topical equine amniotic membrane (eAM) suspension following corneal wounding in a controlled experimental setting. PROCEDURES: Equine amniotic membrane was collected, gamma irradiated, homogenized for topical suspension preparation, and cryopreserved. Corneoscleral rims harvested from fresh rabbit globes were wounded via keratectomy and were maintained in an air-liquid interface ex vivo corneal culture model. Treatment groups included topical gamma irradiated eAM suspension (n = 20) and a control group (n = 20). Re-epithelialization of the wound was assessed with daily photographic evaluation of area of fluorescein uptake (mm2 ). Corneal wound haze after a 21-day period was assessed by photographic analysis of haze area (mm2 ) and pixel intensity (0-255). Histologic processing of corneal tissue was performed, and protein identification of eAM suspension using Liquid chromatography-mass spectrometry (LC-MS). RESULTS: The average day of complete corneal re-epithelialization in controls (5.5 ± 1.1) and topically treated (5.5 ± 0.6) corneas, and rates of reduction in area of fluorescein uptake over time did not significantly differ (p = .44). The corneal wound haze was significantly reduced in mean area by approximately 52% and intensity by 57% in corneas treated with topical eAM suspension (p < .05), compared to controls 21 days following wounding. Protein analysis identified numerous proteins, specifically decorin, dermatopontin, and lumican, which have previously been documented in eAM. CONCLUSIONS: Area and intensity of corneal wound haze were significantly reduced in corneas treated with gamma irradiated eAM suspension, which may be due to previously identified therapeutic proteins which promote corneal clarity.
Assuntos
Epitélio Corneano , Lagomorpha , Âmnio , Animais , Córnea , Epitélio Corneano/patologia , Cavalos , Coelhos , Reepitelização , CicatrizaçãoRESUMO
PURPOSE: The purpose of this study was to compare 3 foam dressings to (1) determine the biomechanical performance of existing negative pressure wound therapy (NPWT) foams and (2) to determine if a test foam is possibly suitable as an antimicrobial "white" foam alternative for use in NPWT. DESIGN: A comparison of mechanical performance of 3 foams used for vacuum-assisted NPWT. SUBJECTS AND SETTING: Preclinical laboratory study using an in vitro model. METHODS: The performance of a "white" foam (polyvinyl alcohol [PVA]), an antimicrobial "black" foam (polyurethane [PU]), and an antimicrobial white foam alternative (test PVA) were tested and compared using 3 mechanically relevant criteria. First, the fluid removal rate was measured for 72 hours. Next, the pressure input was compared to the pressure directly beneath the center of the foam. Finally, the spread of negative pressure beneath the foam was measured and compared. RESULTS: Significant differences were found in fluid removal rates; specifically, the PU foam removed fluids faster than the PVA and test PVA foams, and the currently available PVA foams performed similarly. Both the PU and test PVA foams were able to transmit the negative pressure through the center of the dressing, while the typical PVA foam began failing at 140 mm Hg, with 50% of the samples failing at 200 mm Hg. All PU replicate foams evenly distributed the pressure, while 47% to 60% of the test PVA foams and 7% of the typical PVA foams distributed pressures evenly. CONCLUSIONS: Study findings suggest that the test PVA foam does not mechanically interfere with NPWT and performs equivalently to currently used foams. These results suggest that the test PVA may be modified and incorporated into a vacuum-assisted NPWT device. In addition, the methods employed in these experiments provide a reproducible means to compare biomechanical compatibility of various NPWT foams, dressings, and subdrape devices.
Assuntos
Tratamento de Ferimentos com Pressão Negativa , Antibacterianos , Bandagens , Humanos , CicatrizaçãoRESUMO
In psittacine birds, round cell neoplasms that originate from lymphocytes, plasma cells, histiocytes, or mast cells are sporadic and poorly described. The lack of morphological and immunohistochemical diagnostic criteria or grading schemes make specific diagnoses and prognoses challenging. We assessed cases of psittacine birds diagnosed with round cell neoplasia from 3 North American veterinary diagnostic laboratories to describe the diagnostic features of these tumors. For all cases, demographic data, anatomic distribution, histological features, and immunoreactivity for T (CD3) and B (Pax5 and MUM-1) cell markers were assessed using tissue microarrays and whole slide mounts. Thirty-eight psittacine birds representing 14 species were included. Tumors were mainly infiltrative and multicentric, were composed of homogenous sheets of round to polygonal cells, and commonly presented with a high mitotic count (average 21 mitoses per high-power field). Based on Pax5 immunoreactivity, B-cell lymphoma was most common (19/38 [50%]), and was significantly associated with involvement of the gastrointestinal and urogenital systems. Of the 38 cases, 6 (16%) were consistent with T-cell lymphoma, 3 (8%) with plasma cell tumor, and 3 (8%) were double-reactive for both B- and T-lymphocyte markers. This is the first study to describe morphologic and immunohistochemical features of round cell neoplasia in a large number of psittacine birds, and provides benchmark data for future studies aimed at elucidating the diagnosis and prognosis of these neoplasms. These data also provide useful information about reactivity of commercially available antibodies as lymphocyte markers in tissues of multiple psittacine species.
Assuntos
Doenças das Aves , Linfoma , Papagaios , Animais , Doenças das Aves/diagnóstico , Linfoma/veterináriaRESUMO
INTRODUCTION: Matrix metalloproteinases (MMPs)-2 and -9 are present in corneal ulcers, and an imbalance between MMPs and tissue inhibitors of metalloproteinases (TIMPs) leads to further corneal degradation. Amniotic membrane homogenate (AMH) has proteolytic properties beneficial for corneal healing, but it is unknown whether AMH possesses TIMPs or effectively inhibits MMP-2 and MMP-9 activity. OBJECTIVE: To determine if bovine and equine AMH reduce in vitro MMP-2 and MMP-9 activities associated with the presence of TIMPs. PROCEDURES: Undiluted and diluted twofold series (0-fold to 16-fold dilutions) of equine amniotic membrane homogenates (EAMH, n = 8) and bovine amniotic membrane homogenates (BAMH, n = 8) were subjected to fluorescence resonance energy transfer, and the fluorescence emitted was recorded over time. Average fluorescence was calculated versus recombinant concentration. Enzyme-linked immunosorbent assays for TIMPs 1-4 were applied to quantify TIMPs in the samples. RESULTS: AMH from both species were able to inhibit MMP-2 and MMP-9 activities in vitro, and the inhibition efficacy decreased gradually with dilution. BAMH was significantly more effective than EAMH at inhibiting MMP-2 and MMP-9 in vitro. TIMPs -2 and -3 were present in EAMH and BAMH. TIMP-1 was detected only in BAMH, and TIMP-4 was not detected in any samples. CONCLUSION: Both EAMH and BAMH directly inhibited MMP-2 and MMP-9 in vitro without dilution, and BAMH showed better inhibition of MMP-2 and MMP-9 before and after dilution compared to EAMH.
Assuntos
Âmnio/metabolismo , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Animais , Bovinos , Feminino , Transferência Ressonante de Energia de Fluorescência/veterinária , Cavalos , GravidezRESUMO
Many debilitating neuropsychiatric and neurodegenerative disorders are characterized by dopamine neurotransmitter dysregulation. Monitoring subsecond dopamine release accurately and for extended, clinically relevant timescales is a critical unmet need. Especially valuable has been the development of electrochemical fast-scan cyclic voltammetry implementing microsized carbon fiber probe implants to record fast millisecond changes in dopamine concentrations. Nevertheless, these well-established methods have only been applied in primates with acutely (few hours) implanted sensors. Neurochemical monitoring for long timescales is necessary to improve diagnostic and therapeutic procedures for a wide range of neurological disorders. Strategies for the chronic use of such sensors have recently been established successfully in rodents, but new infrastructures are needed to enable these strategies in primates. Here we report an integrated neurochemical recording platform for monitoring dopamine release from sensors chronically implanted in deep brain structures of nonhuman primates for over 100 days, together with results for behavior-related and stimulation-induced dopamine release. From these chronically implanted probes, we measured dopamine release from multiple sites in the striatum as induced by behavioral performance and reward-related stimuli, by direct stimulation, and by drug administration. We further developed algorithms to automate detection of dopamine. These algorithms could be used to track the effects of drugs on endogenous dopamine neurotransmission, as well as to evaluate the long-term performance of the chronically implanted sensors. Our chronic measurements demonstrate the feasibility of measuring subsecond dopamine release from deep brain circuits of awake, behaving primates in a longitudinally reproducible manner.
Assuntos
Encéfalo/metabolismo , Dopamina/metabolismo , Eletroencefalografia/métodos , Monitorização Neurofisiológica/métodos , Animais , Encéfalo/fisiologia , Eletrodos Implantados , Eletroencefalografia/instrumentação , Feminino , Macaca mulatta , Monitorização Neurofisiológica/instrumentação , Recompensa , Fatores de TempoRESUMO
PURPOSE: The purpose of this study was to evaluate the in-gel strain and tear reduction provided by 2 skin protectant products that were applied as a liquid and allowed to dry, leaving behind a protective layer. DESIGN: Prospective, 3-group comparison cohort study using an in vitro model. METHODS: A fragile agar-based gel with an embedded bead was used in a custom device that applied variable interface pressures of 550, 1080, or 1600 Pa, respectively. The device then imparted 216 N of external shear force in 0.625-mm increments. The resulting strain in the gel was measured by digital image correlation. The strain at tearing was determined by observing the images of the gels and calculating the strain at that point. This approach was used to compare untreated gels to gels treated with one of 2 commercially available cyanoacrylate-based skin protectants. The results from the 3 groups were first analyzed by analysis of variance, followed by Tukey's Honestly Significant Difference test when indicated. RESULTS: We observed a proportional increase in interface pressure and strain that differed among the 3 groups. Specifically, the gels treated with a mixed polymer skin protectant had less pretearing strain than the control gel at both the 1080-Pa load (-15%, P = 3.64 × 10) and 1600 Pa-load (-20%, P = .03). The pure cyanoacrylate-treated gels had less strain than the control at 1080 Pa (-34%, P = 4.25 × 10) and 1600 Pa (-48%, P = 1.07 × 10); it also had less strain than the mixed polymer product at 1080 Pa (-19%, P = 5.38 × 10) and 1600 Pa (-28%, P = 3.88 × 10). In terms of protection from tearing, at an interface pressure of 1080 Pa, the control gel tore 80% of the time, the mixed polymer-treated gel tore 100% of the time, and the pure cyanoacrylate-treated gel did not tear (0/5, P = 8.84 × 10). Under a load of 1600 Pa, 100% of the control and mixed polymer-treated gels tore while none of the cyanoacrylate-treated gels did (P = 2.54 × 10). CONCLUSION: The pure cyanoacrylate-based skin protectant provided the most protection, with consistent reductions in both strain and tearing. Both skin protectants reduced the initial in-gel strain; however, only the pure cyanoacrylate-treated product protected the gel from tears under the conditions tested. These results indicate that cyanoacrylate-based skin protectants can reduce shear strain and tearing in fragile elastic materials.
Assuntos
Fenômenos Biomecânicos/efeitos dos fármacos , Embucrilato/uso terapêutico , Higiene da Pele/métodos , Estudos de Coortes , Embucrilato/farmacologia , Humanos , Estudos Prospectivos , Resistência ao Cisalhamento , Estresse MecânicoRESUMO
Biofilms are prevalent in non-healing chronic wounds and implicated in delayed healing. Tolerance to antimicrobial treatments and the host's immune system leave clinicians with limited interventions against biofilm populations. It is therefore essential that effective treatments be rigorously tested and demonstrate an impact on biofilm across multiple experimental models to guide clinical investigations and protocols. Cadexomer iodine has previously been shown to be effective against biofilm in various in vitro models, against methicillin-resistant Staphylococcus aureus biofilm in mouse wounds, and clinically in diabetic foot ulcers complicated by biofilm. Similarities between porcine and human skin make the pig a favoured model for cutaneous wound studies. Two antiseptic dressings and a gauze control were assessed against mature biofilm grown on ex vivo pig skin and in a pig wound model. Significant reductions in biofilm were observed following treatment with cadexomer iodine across both biofilm models. In contrast, silver carboxymethylcellulose dressings had minimal impact on biofilm in the models, with similar results to the control in the ex vivo model. Microscopy and histopathology indicate that the depth of organisms in wound tissue may impact treatment effectiveness. Further work on the promising biofilm efficacy of cadexomer iodine is needed to determine optimal treatment durations against biofilm.
Assuntos
Anti-Infecciosos Locais/uso terapêutico , Biofilmes/efeitos dos fármacos , Iodóforos/uso terapêutico , Pseudomonas aeruginosa/efeitos dos fármacos , Staphylococcus aureus/efeitos dos fármacos , Cicatrização/efeitos dos fármacos , Infecção dos Ferimentos/tratamento farmacológico , Animais , Doença Crônica/tratamento farmacológico , Humanos , Modelos Animais , SuínosRESUMO
Studies of neural oscillations in the beta band (13-30 Hz) have demonstrated modulations in beta-band power associated with sensory and motor events on time scales of 1 s or more, and have shown that these are exaggerated in Parkinson's disease. However, even early reports of beta activity noted extremely fleeting episodes of beta-band oscillation lasting <150 ms. Because the interpretation of possible functions for beta-band oscillations depends strongly on the time scale over which they occur, and because of these oscillations' potential importance in Parkinson's disease and related disorders, we analyzed in detail the distributions of duration and power for beta-band activity in a large dataset recorded in the striatum and motor-premotor cortex of macaque monkeys performing reaching tasks. Both regions exhibited typical beta-band suppression during movement and postmovement rebounds of up to 3 s as viewed in data averaged across trials, but single-trial analysis showed that most beta oscillations occurred in brief bursts, commonly 90-115 ms long. In the motor cortex, the burst probabilities peaked following the last movement, but in the striatum, the burst probabilities peaked at task end, after reward, and continued through the postperformance period. Thus, what appear to be extended periods of postperformance beta-band synchronization reflect primarily the modulated densities of short bursts of synchrony occurring in region-specific and task-time-specific patterns. We suggest that these short-time-scale events likely underlie the functions of most beta-band activity, so that prolongation of these beta episodes, as observed in Parkinson's disease, could produce deleterious network-level signaling.
Assuntos
Corpo Estriado/fisiologia , Macaca/fisiologia , Córtex Motor/fisiologia , Movimento , Animais , HumanosRESUMO
PURPOSE: The purpose of these experiments was to compare 2 commercially available skin protectants with different chemical compositions. DESIGN: Two materially different skin protectants were applied to ex vivo pig skin, subjected to stresses, and the resulting skin was observed and analyzed. METHODS: Using ex vivo pig skin, we sought to better understand the physical differences between a cyanoacrylate-based and a mixed cyanoacrylate/acrylic polymer-based skin protectant. A combination of imaging techniques and microscopic analyses was used to observe and quantify differences in layer thickness and the degree of steadfastness of the layers to liquid stresses. RESULTS: The experiments revealed that the solely cyanoacrylate-based protectant created a layer that was, on average, 5.1 times thicker than the mixed polymer product (p= 1.8 × 10). Observation via electron microscopy also revealed that the extent of coverage varied between the 2 products. In a final experiment, we observed that the mixed polymer product maintained a high degree of adhesiveness, which led to the removal of sheets of epithelium upon gentle blotting. CONCLUSIONS: The experiments revealed that while the 2 skin protectants share a common ingredient, both the quantity of that ingredient and the inclusion of other materials in one of them lead to substantially different properties when tested in the research setting.
Assuntos
Cianoacrilatos/farmacologia , Cianoacrilatos/normas , Pele/lesões , Animais , Cianoacrilatos/uso terapêutico , Humanos , Pele/microbiologia , Suínos/microbiologia , Adesivos Teciduais/farmacologia , Adesivos Teciduais/uso terapêuticoRESUMO
Bacteria have been found to form multicellular aggregates which have collectively been termed "biofilms." It is hypothesized that biofilm formation is a means to protect bacterial cells including protection form the immune response of humans. This protective mechanism is believed to explain persistent chronic wound infections. At times, the biofilms are abundant enough to see, and remove by simple wiping. However, recent evidence has shown that the removal of these visible portions are not sufficient, and that biofilms can continue to form even with daily wiping. In this work, we tested an approach to detect the biofilms which are present after clinically wiping or sharp wound debridement. Our method is based on a variation of impression cytology in which a nitrocellulose membrane was used to collect surface biofilm components, which were then differentially stained. In this prospective study, members of an interdisciplinary pressure ulcer team at a university hospital tested our method's ability to predict the generation of wound slough in the week that followed each blotting. A total of 70 blots collected from 23 pressure ulcers produced 27 wounds negative for staining and 43 positive. In the negative blots 55.6% were found to have decreased wound slough, while 81.4% with positive staining had either increase or unchanged wound slough generation. These results lead to an odds ratio of positive blotting cases of 9.37 (95% confidence intervals: 2.47-35.5, p = 0.001) for slough formation; suggesting that the changes in wound slough formation can be predicted clinically using a non-invasive wound blotting method.
Assuntos
Biofilmes , Úlcera por Pressão/microbiologia , Úlcera por Pressão/fisiopatologia , Colódio , Técnicas Citológicas/métodos , Desbridamento , Humanos , Membranas Artificiais , Estudos Prospectivos , Adesivos TeciduaisRESUMO
Proteinases are enzymes that can digest other proteins. In chronic wounds, a sub-class of these enzymes with the ability to degrade the extracellular matrix (matrix metalloproteinases, MMPs) have been found to both inhibit healing and to be able to aid in enzymatically debriding a wound. Enzymatic debridement using the enzymes present in a wound is generally called autolytic debridement. Clinicians seeking to employ autolytic debridement typically use occlusive materials such as medical honey, alginate dressings and other occlusive dressings. A relatively new class of gel dressings comprised of surfactants are now available for clinical use. A variety of surfactants are used in the study of MMP biochemistry. Surfactants can deactivate MMPs or can enhance their activity, depending on the surfactant. In order to begin to understand how the MMPs found in chronic wounds would respond to these new dressings, we tested a serial dilution series of two of the currently available surfactant-based dressings to determine their effects on four separate MMPs. The dose-response versus MMP activity of bacterial collagenase, host-derived MMP-8 and MMPs-2 and -9 was assessed using a simple mix-and-read fluorescent peptide activity assay. The enzyme's native activity in the absence of the gel was used to compare against the surfactant-treated samples. We found that the surfactant affected the proteinase activity differently for each enzyme. The activity of the bacterial collagenase was increased at low concentrations but slightly inhibited as the concentrations increased. The host MMP-8 collagenase responded similarly in that it was inhibited at higher concentrations. Interestingly, both MMP gelatinases presented with substantially increased activities, with MMP-2 increased to 200% of native activity, while MMP-9 presented with an increase of 300% activity over the same concentration range. MMPs appear to respond to a surfactant-based gel dressing differentially, with the MMP most commonly elevated in chronic wounds having the highest boost to activity. In wounds with elevated MMPs, our data suggest that the use of these surfactant-based dressings would be expected to enhance the activity of MMPs 2 and 9 gelatinases while simultaneously inhibiting MMP-8 collagenase. Hypothetically, this imbalanced effect would support a protection of the native dermal collagen and removal of denatured materials. However, the demonstration of these anticipated consequences is still being investigated.
Assuntos
Colagenases/metabolismo , Desbridamento/métodos , Metaloproteinases da Matriz/metabolismo , Curativos Oclusivos , Tensoativos/farmacologia , Cicatrização/fisiologia , HumanosRESUMO
Bacterial biofilms have been found in many, if not all, chronic wounds. Their excessive extracellular matrix secretion and the metabolic changes that they undergo render them highly tolerant of many antibiotic and antimicrobial treatments. Physical removal and/or disruption are a common approach to treating wounds suspected of having bacterial biofilms. While many of these techniques use mechanical energy as the primary means of removal, we have begun to investigate if surfactants could facilitate the removal of bacterial biofilms, or if they might sensitise the biofilms to antimicrobial interventions. We tested a new surfactant-based wound gel on an ex vivo porcine skin explant model infected with a functionally tolerant 3-day biofilm. The wounds were dressed with a surfactant-based gel directly on the wound or with moistened gauze. The wounds were then wiped daily with moistened gauze, and the gel or gauze was re-applied. Each day, an explant from each group was harvested and tested for total viable bacteria counts and viable biofilm-protected bacteria counts. The results show that daily wiping with moistened gauze led to an initial decrease of bacteria, but by day 3, the biofilm had been fully re-established to the same level prior to the beginning of treatment. For the surfactant-based treatment, there was no detectable functional biofilm after the first treatment. The gauze control, which was also subjected to daily wiping, still contained functional biofilms, indicating that this result was not due to wiping alone. The total bacteria in the surfactant-treated explants steadily decreased through day 3, when there were no detectable bacteria, while the wiping-only control bacteria counts remained steady. The use of a moist gauze to wipe the visually apparent slime off of a wound appears to be insufficient to reduce biofilm over a 3-day period. Daily application of the surfactant gel dressing and wiping reduced the biofilm to undetectable levels within 3 days in a skin explant model. A 3-day regimen of dressing the wound model with a surfactant gel followed by gentle removal of the gel by wiping with a moistened gauze appears to be a simple and adequate approach to removing a bacterial biofilm infection in an ex vivo model. Additional clinical evidence is needed to determine if this promising approach can perform the same in clinically infected chronic wounds.
Assuntos
Anti-Infecciosos/uso terapêutico , Bandagens , Biofilmes/efeitos dos fármacos , Infecções por Pseudomonas/tratamento farmacológico , Tensoativos/uso terapêutico , Transplantes/microbiologia , Infecção dos Ferimentos/tratamento farmacológico , Animais , Feminino , Humanos , Masculino , Pseudomonas aeruginosa/efeitos dos fármacos , Suínos , CicatrizaçãoRESUMO
Amphibious fishes often emerse (leave water) when faced with unfavourable water conditions. How amphibious fishes cope with the risks of rising water temperatures may depend, in part, on the plasticity of behavioural mechanisms such as emersion thresholds. We hypothesized that the emersion threshold is reversibly plastic and thus dependent on recent acclimation history rather than on conditions during early development. Kryptolebias marmoratus were reared for 1 year at 25 or 30°C and acclimated as adults (one week) to either 25 or 30°C before exposure to an acute increase in water temperature. The emersion threshold temperature and acute thermal tolerance were significantly increased in adult fish acclimated to 30°C, but rearing temperature had no significant effect. Using a thermal imaging camera, we also showed that emersed fish in a low humidity aerial environment (30°C) lost significantly more heat (3.3°C min(-1)) than those in a high humidity environment (1.6°C min(-1)). In the field, mean relative humidity was 84%. These results provide evidence of behavioural avoidance of high temperatures and the first quantification of evaporative cooling in an amphibious fish. Furthermore, the avoidance response was reversibly plastic, flexibility that may be important for tropical amphibious fishes under increasing pressures from climatic change.
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Aclimatação/fisiologia , Comportamento Animal/fisiologia , Regulação da Temperatura Corporal/fisiologia , Ciprinodontiformes/fisiologia , Temperatura Alta , Umidade , Animais , Ciprinodontiformes/crescimento & desenvolvimento , ÁguaRESUMO
The effects of a triple combination of siRNAs targeting key scarring genes were assessed using an ex vivo organ culture model of excimer ablated rabbit corneas. The central 6 mm diameter region of fresh rabbit globes was ablated to a depth of 155 microns with an excimer laser. Corneas were excised, cultured at the air-liquid interface in defined culture medium supplemented with transforming growth factor beta 1 (TGFB1), and treated with either 1% prednisolone acetate or with 22.5 µM cationic nanoparticles complexed with a triple combination of siRNAs (NP-siRNA) targeting TGFB1, TGFB Receptor (TGFBR2) and connective tissue growth factor (CTGF). Scar formation was measured using image analysis of digital images and levels of smooth muscle actin (SMA) were assessed in ablated region of corneas using qRT-PCR and immunostaining. Ex vivo cultured corneas developed intense haze-like scar in the wounded areas and levels of mRNAs for pro-fibrotic genes were significantly elevated 3-8 fold in wounded tissue compared to unablated corneas. Treatment with NP-siRNA or steroid significantly reduced quantitative haze levels by 55% and 68%, respectively, and reduced SMA mRNA and immunohistostaining. This ex vivo corneal culture system reproduced key molecular patterns of corneal scarring and haze formation generated in rabbits. Treatment with NP-siRNAs targeting key scarring genes or an anti-inflammatory steroid reduced corneal haze and SMA mRNA and protein.
Assuntos
Cicatriz/prevenção & controle , Córnea , Doenças da Córnea/terapia , Terapia a Laser/efeitos adversos , RNA Interferente Pequeno/uso terapêutico , Actinas/metabolismo , Análise de Variância , Animais , Anti-Inflamatórios/uso terapêutico , Cicatriz/metabolismo , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Córnea/patologia , Córnea/cirurgia , Doenças da Córnea/etiologia , Doenças da Córnea/metabolismo , Modelos Animais de Doenças , Imuno-Histoquímica , Nanopartículas/uso terapêutico , Técnicas de Cultura de Órgãos , Prednisolona/análogos & derivados , Prednisolona/uso terapêutico , RNA Interferente Pequeno/administração & dosagem , Coelhos , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Fator de Crescimento Transformador beta1/metabolismoRESUMO
Rhythmic brain activity is thought to reflect, and to help organize, spike activity in populations of neurons during on-going behavior. We report that during learning, a major transition occurs in task-related oscillatory activity in the ventromedial striatum, a striatal region related to motivation-dependent learning. Early on as rats learned a T-maze task, bursts of 70- to 90-Hz high-γ activity were prominent during T-maze runs, but these gradually receded as bursts of 15- to 28-Hz ß-band activity became pronounced. Populations of simultaneously recorded neurons synchronized their spike firing similarly during both the high-γ-band and ß-band bursts. Thus, the structure of spike firing was reorganized during learning in relation to different rhythms. Spiking was concentrated around the troughs of the ß-oscillations for fast-spiking interneurons and around the peaks for projection neurons, indicating alternating periods of firing at different frequencies as learning progressed. Spike-field synchrony was primarily local during high-γ-bursts but was widespread during ß-bursts. The learning-related shift in the probability of high-γ and ß-bursting thus could reflect a transition from a mainly focal rhythmic inhibition during early phases of learning to a more distributed mode of rhythmic inhibition as learning continues and behavior becomes habitual. These dynamics could underlie changing functions of the ventromedial striatum during habit formation. More generally, our findings suggest that coordinated changes in the spatiotemporal relationships of local field potential oscillations and spike activity could be hallmarks of the learning process.
Assuntos
Gânglios da Base/fisiologia , Ondas Encefálicas/fisiologia , Hábitos , Aprendizagem em Labirinto/fisiologia , Neurônios/fisiologia , Animais , Mapeamento Encefálico , Masculino , Ratos , Ratos Long-EvansRESUMO
Measuring the dynamic relationship between neuromodulators, such as dopamine, and neuronal action potentials is imperative to understand how these fundamental modes of neural signaling interact to mediate behavior. We developed methods to measure concurrently dopamine and extracellular action potentials (i.e., spikes) in monkeys. Standard fast-scan cyclic voltammetric (FSCV) electrochemical (EChem) and electrophysiological (EPhys) recording systems are combined and used to collect spike and dopamine signals, respectively, from an array of carbon fiber (CF) sensors implanted in the monkey striatum. FSCV requires the application of small voltages at the implanted sensors to measure redox currents generated from target molecules, such as dopamine. These applied voltages create artifacts at neighboring EPhys measurement sensors which may lead to misclassification of these signals as physiological spikes. Therefore, simple automated temporal interpolation algorithms were designed to remove these artifacts and enable accurate spike extraction. We validated these methods using simulated artifacts and demonstrated an average spike recovery rate of 84.5%. We identified and discriminated cell type-specific units in the monkey striatum that were shown to correlate to specific behavioral task parameters related to reward size and eye movement direction. Synchronously recorded spike and dopamine signals displayed contrasting relations to the task variables, suggesting a complex relationship between these two modes of neural signaling. Future application of our methods will help advance our understanding of the interactions between neuromodulator signaling and neuronal activity, to elucidate more detailed mechanisms of neural circuitry and plasticity mediating behaviors in health and in disease.
Assuntos
Potenciais de Ação , Fibra de Carbono , Dopamina , Macaca mulatta , Animais , Potenciais de Ação/fisiologia , Dopamina/metabolismo , Masculino , Neurônios/fisiologia , Neurônios/metabolismo , Eletrodos Implantados , Corpo Estriado/fisiologia , Corpo Estriado/metabolismo , Artefatos , Técnicas Eletroquímicas/métodos , Movimentos Oculares/fisiologia , RecompensaRESUMO
Measuring the dynamic relationship between neuromodulators, such as dopamine, and neuronal action potentials is imperative to understand how these fundamental modes of neural signaling interact to mediate behavior. Here, we developed methods to measure concurrently dopamine and extracellular action potentials (i.e., spikes) and applied these in a monkey performing a behavioral task. Standard fast-scan cyclic voltammetric (FSCV) electrochemical (EChem) and electrophysiological (EPhys) recording systems are combined and used to collect spike and dopamine signals, respectively, from an array of carbon fiber (CF) sensors implanted in the monkey striatum. FSCV requires the application of small voltages at the implanted sensors to measure redox currents generated from target molecules, such as dopamine. These applied voltages create artifacts at neighboring EPhys-measurement sensors, producing signals that may falsely be classified as physiological spikes. Therefore, simple automated temporal interpolation algorithms were designed to remove these artifacts and enable accurate spike extraction. We validated these methods using simulated artifacts and demonstrated an average spike recovery rate of 84.5%. This spike extraction was performed on data collected from concurrent EChem and EPhys recordings made in a task-performing monkey to discriminate cell-type specific striatal units. These identified units were shown to correlate to specific behavioral task parameters related to reward size and eye-movement direction. Synchronous measures of spike and dopamine signals displayed contrasting relations to the behavioral task parameters, as taken from our small set of representative data, suggesting a complex relationship between these two modes of neural signaling. Future application of our methods will help advance our understanding of the interactions between neuromodulator signaling and neuronal activity, to elucidate more detailed mechanisms of neural circuitry and plasticity mediating behaviors in health and in disease.