RESUMEN
In vitro and in vivo assays were conducted to investigate the effects of trans-resveratrol (RVT) on liquid-extended boar semen during 72 h of storage at 17 °C. Thirty-six ejaculates were collected from six boars, evaluated, and extended. RVT was then added at the indicated treatment concentration (0, 0.01, 0.1 or 1 mM), and the ejaculates were cooled to 17 °C and evaluated at 0, 24, 48, and 72 h. Samples were evaluated for sperm motility, kinetics, plasma and acrosome integrity, mitochondrial membrane potential, anion superoxide levels, lipoperoxidation, and antioxidant enzyme activity. In the follow-up experiment, twenty-eight gilts were fixed-time inseminated with 0 or 0.01 mM RVT liquid-extended boar semen. After five days, they were slaughtered, and their reproductive tracts were recovered. The embryos were collected, and the pregnancy, fertility, and viable embryo rates were calculated. In the in vitro assays, total motility, plasma and acrosome membrane integrity, mitochondrial membrane potential, anion superoxide levels, and lipoperoxidation did not change at any of the evaluation times with the use of RVT up to 0.01 mM. RVT decreased SOD activity without changes in GPx. RVT used at 1 mM showed harmful effects for almost all evaluated parameters. For the in vivo assay, the same pregnancy and fertility rates were observed for both groups, while the viable embryo rate was three-fold lower in the 0.01 mM group than in the 0 mM group. The results showed a dichotomous effect of RVT; a low concentration was not harmful in vitro but was catastrophic for embryo viability.
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Fertilidad/efectos de los fármacos , Resveratrol/farmacología , Preservación de Semen/veterinaria , Semen/efectos de los fármacos , Espermatozoides/efectos de los fármacos , Porcinos , Acrosoma/efectos de los fármacos , Animales , Antioxidantes/farmacología , Femenino , Inseminación Artificial/veterinaria , Masculino , Soluciones Preservantes de Órganos/farmacología , Embarazo , Análisis de Semen/veterinaria , Preservación de Semen/métodos , Motilidad Espermática/efectos de los fármacos , SuperóxidosRESUMEN
PURPOSE: To evaluate the effect of amniotic fluid in liver preservation in organ transplantation, and compare it with standard preservation solutions. METHODS: The groups consisted of Group 1: Ringer Lactate (RL) group, Group 2: HTK group, Group 3: UW group, Group 4: AF group. The livers of rats from Group 1, 2, 3, and 4 were perfused and placed into falcon tubes containing RL, HTK, UW, and AF solutions at +4 °C, respectively. The tubes were stored for 12 hours in the refrigerator at +4°C. Tissue samples were taken at the 6th and 12th hours for histopathological examinations of the perfused livers, and storage solutions for biochemical analyzes at 6th and 12th hours. RESULTS: AF was shown to maintain organ viability by reducing the number of cells undergoing apoptosis. Histopathological changes such as sinusoidal dilatation, hydropic degeneration, and focal necrosis were found to be similar to the groups in which the standard organ preservation solutions were used. Additionally, the results of INOS, IL-10, and TNF-α,which were evaluated immunohistochemically, have been shown to be similar to the UW and HTK groups. CONCLUSIONS: AF provided conservation similar to UW and HTK in the 12-hour liver SCS process. The fact that apoptosis values are comparable to standard preservation solutions supports the success of AF in the cold storage of the liver.
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Líquido Amniótico , Criopreservación/métodos , Hígado , Soluciones Preservantes de Órganos/farmacología , Animales , Glucosa/farmacología , Inmunohistoquímica , Etiquetado Corte-Fin in Situ , Interleucina-10/análisis , Hígado/irrigación sanguínea , Hígado/patología , Masculino , Manitol/farmacología , Óxido Nítrico Sintasa de Tipo II/análisis , Preservación de Órganos/métodos , Cloruro de Potasio/farmacología , Procaína/farmacología , Distribución Aleatoria , Ratas Wistar , Valores de Referencia , Daño por Reperfusión/prevención & control , Reproducibilidad de los Resultados , Solución de Ringer/farmacología , Factores de Tiempo , Supervivencia Tisular , Factor de Necrosis Tumoral alfa/análisisRESUMEN
PURPOSE: Fungal infections in lamellar keratoplasty are a growing concern. Optisol-GS does not contain an antifungal agent and supplementation with 0.255 µg/mL Amphotericin B (AmpB) has been considered. This study tested the ability of 0.255 µg/mL AmpB in Optisol-GS to eliminate yeast contamination of corneal tissue. METHODS: Three isolates of Candida albicans, 1 of Candida parapsilosis, and 1 of Candida glabrata were tested in Optisol with and without AmpB. Corneoscleral rims stored at -80°C were thawed and placed in 10 multiwell plates (4 per plate). The rims were inoculated with 4 respective loads of yeast: 0, 10, 10, and 10 colony-forming units in 2 sets of 5 for 5 yeasts. One set was filled with Optisol plus AmpB and the other with Optisol only. All 10 plates were incubated at cold storage (2°C-8°C) for 48 hours. After 48 hours, all corneal rims were placed into 10 mL of yeast extract peptone dextrose medium; a swab culture of each well was plated onto Sabouraud plates; and all plates with the remaining Optisol were incubated at 30°C. Yeast growth was monitored for 10 days. Minimum inhibitory concentration and minimum fungicidal concentration were determined. RESULTS: All corneoscleral specimens were positive regardless of fungal load or presence of AmpB. All controls remained negative. Minimum inhibitory concentrations and minimum fungicidal concentrations were equivalent and ranged between 0.5 and 2.0 µg/mL. CONCLUSIONS: AmpB at a concentration of 0.255 µg/mL in Optisol-GS at cold storage (2°C-8°C) over 48 hours did not eliminate yeast from corneal tissue.
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Anfotericina B/farmacología , Antifúngicos/farmacología , Candida/efectos de los fármacos , Sulfatos de Condroitina/farmacología , Córnea/microbiología , Dextranos/farmacología , Gentamicinas/farmacología , Soluciones Preservantes de Órganos/farmacología , Preservación de Órganos/métodos , Candidiasis/prevención & control , Mezclas Complejas/farmacología , Bancos de Ojos , Infecciones Fúngicas del Ojo/prevención & control , HumanosRESUMEN
Abstract Purpose: To evaluate the effect of amniotic fluid in liver preservation in organ transplantation, and compare it with standard preservation solutions. Methods: The groups consisted of Group 1: Ringer Lactate (RL) group, Group 2: HTK group, Group 3: UW group, Group 4: AF group. The livers of rats from Group 1, 2, 3, and 4 were perfused and placed into falcon tubes containing RL, HTK, UW, and AF solutions at +4°C, respectively. The tubes were stored for 12 hours in the refrigerator at +4°C. Tissue samples were taken at the 6th and 12th hours for histopathological examinations of the perfused livers, and storage solutions for biochemical analyzes at 6th and 12th hours. Results: AF was shown to maintain organ viability by reducing the number of cells undergoing apoptosis. Histopathological changes such as sinusoidal dilatation, hydropic degeneration, and focal necrosis were found to be similar to the groups in which the standard organ preservation solutions were used. Additionally, the results of INOS, IL-10, and TNF-α,which were evaluated immunohistochemically, have been shown to be similar to the UW and HTK groups. Conclusions: AF provided conservation similar to UW and HTK in the 12-hour liver SCS process. The fact that apoptosis values are comparable to standard preservation solutions supports the success of AF in the cold storage of the liver.
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Animales , Masculino , Criopreservación/métodos , Soluciones Preservantes de Órganos/farmacología , Líquido Amniótico , Hígado/irrigación sanguínea , Hígado/patología , Preservación de Órganos/métodos , Cloruro de Potasio/farmacología , Procaína/farmacología , Valores de Referencia , Factores de Tiempo , Supervivencia Tisular , Inmunohistoquímica , Daño por Reperfusión/prevención & control , Distribución Aleatoria , Reproducibilidad de los Resultados , Factor de Necrosis Tumoral alfa/análisis , Interleucina-10/análisis , Ratas Wistar , Etiquetado Corte-Fin in Situ , Óxido Nítrico Sintasa de Tipo II/análisis , Solución de Ringer/farmacología , Glucosa/farmacología , Manitol/farmacologíaRESUMEN
BACKGROUND: Prolonged cold ischemia is a risk factor for delayed graft function of kidney transplants, and is associated with caspase-3-mediated apoptotic tubular cell death. We hypothesized that treatment of tubular cells and donor kidneys during cold storage with a caspase inhibitor before transplant would reduce tubular cell apoptosis and improve kidney function after transplant. METHODS: Mouse tubular cells were incubated with either dimethyl sulfoxide (DMSO) or Q-VD-OPh during cold storage in saline followed by rewarming in normal media. For in vivo studies, donor kidneys from C57BL/6 mice were perfused with cold saline, DMSO (vehicle), or QVD-OPh. Donor kidneys were then recovered, stored at 4°C for 60 minutes, and transplanted into syngeneic C57BL/6 recipients. RESULTS: Tubular cells treated with a caspase inhibitor had significantly reduced capsase-3 protein expression, caspase-3 activity, and apoptotic cell death compared with saline or DMSO (vehicle) in a dose-dependent manner. Treatment of donor kidneys with a caspase inhibitor significantly reduced serum creatinine and resulted in significantly less tubular cell apoptosis, BBI, tubular injury, cast formation, and tubule lumen dilation compared with DMSO and saline-treated kidneys. CONCLUSIONS: Caspase inhibition resulted in decreased tubular cell apoptosis and improved renal function after transplantation. Caspase inhibition may be a useful strategy to prevent cold ischemic injury of donor renal grafts.
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Clorometilcetonas de Aminoácidos/farmacología , Apoptosis/efectos de los fármacos , Inhibidores de Caspasas/farmacología , Isquemia Fría , Funcionamiento Retardado del Injerto/prevención & control , Trasplante de Riñón/métodos , Riñón/efectos de los fármacos , Riñón/cirugía , Soluciones Preservantes de Órganos/farmacología , Preservación de Órganos/métodos , Quinolinas/farmacología , Animales , Biomarcadores/sangre , Caspasa 3/metabolismo , Línea Celular , Isquemia Fría/efectos adversos , Creatinina/sangre , Funcionamiento Retardado del Injerto/enzimología , Funcionamiento Retardado del Injerto/patología , Funcionamiento Retardado del Injerto/fisiopatología , Riñón/enzimología , Riñón/patología , Trasplante de Riñón/efectos adversos , Masculino , Ratones Endogámicos C57BL , Modelos Animales , NefrectomíaRESUMEN
BACKGROUND/AIMS: Tooth avulsion consists of the complete displacement of a tooth from the alveolar socket. When immediate replantation is not possible, the avulsed tooth should be kept in a storage medium capable of maintaining the viability of periodontal ligament (PDL) cells on the root surface. However, there is no consensus on the best storage medium able to prevent sequels such as ankylosis and tooth resorption. The aim of this study was to perform a systematic review to evaluate the in vivo effectiveness of different storage media for avulsed teeth. METHODS: Two reviewers performed a database search for studies published between January 1950 and December 2015 which were indexed in the PubMed, Scopus, Web of Science, and Bireme databases. An additional manual search was performed. Studies with animal models that evaluated tooth avulsion, storage media, and replantation were included. After full-text analysis of the potentially relevant studies, the selected studies were included in the systematic review. RESULTS: The database search found 157 distinct studies evaluating avulsed teeth storage media. However, only six studies met the selection criteria and were included in the review. There was a high variability in the study estimates for the parameters analyzed. When assessing the quality and level of evidence of each study, one study was rated as having a very low level of evidence, four studies had low levels of evidence, and one had a moderate level of evidence. CONCLUSION: As a result of data heterogeneity and limitations of the studies, there was insufficient evidence to determine the most effective storage medium for avulsed teeth.
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Modelos Animales , Soluciones Preservantes de Órganos/farmacología , Manejo de Especímenes/métodos , Avulsión de Diente , AnimalesRESUMEN
BACKGROUND/AIM: In cases of tooth avulsion, a minimal extra-alveolar dry storage period or the use of a suitable storage medium is crucial to maintaining the vitality of the periodontal ligament. Whey has similar properties to milk and has therefore been investigated as a storage medium for avulsed teeth. The aim of this study was to evaluate the repair process after replantation of rat teeth kept in whey and whole milk. MATERIALS AND METHODS: Thirty-six male rats were divided into four groups of nine animals. The upper right incisor was extracted under general anesthesia. In Group I, the teeth were immediately replanted without treatment (positive control). In Group II, the teeth were stored in 50 mL of sweet whey. In Group III, the teeth were kept in 50 mL of long-shelf-life whole milk (UHT, Parmalat® ). In Group IV, the teeth were kept dry (negative control). After 60 minutes, the teeth in Groups II, III, and IV were replanted into their sockets. The animals were subjected to euthanasia 60 days after replantation. The specimens were stained with hematoxylin and eosin for histomorphometric analysis. RESULTS: The organization of the periodontal ligament in Group II (whey) was similar to that in Groups I (immediate replantation) and III (whole milk) (P > .05). However, some specimens in this group exhibited periodontal fibers inserted into the bone and cementum throughout the entire length of the periodontal ligament. This occurred in the group submitted to immediate replantation, whereas this histological aspect was not seen in whole milk group. Group IV (late replantation) had a higher rate of root resorption. Regarding the root repair process, it was expected that Group I (immediate) would demonstrate more favorable repair than the other groups. However, Group III (whole milk) had better results when compared to Groups II (whey) and IV (late) (P < .05). CONCLUSION: Whey and whole milk achieved similar results and were adequate storage media for avulsed teeth.
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Leche , Soluciones Preservantes de Órganos/farmacología , Ligamento Periodontal/efectos de los fármacos , Reimplante Dental/métodos , Suero Lácteo , Animales , Bovinos , Incisivo , Masculino , Distribución Aleatoria , Ratas , Ratas WistarRESUMEN
AIM: To perform multiparametric analysis of the effects of soya milk (SM), whole milk (WM) and Hank's balanced salt solution (HBSS) on the viability of fibroblasts (HGF). The study also aimed to evaluate the influence of these solutions on bovine root dentine according to OH- and PO43- on the surface. METHODOLOGY: The HGF cytotoxicity was determined according to XTT, NR and SRB assays at 1, 3 and 6 h. Root dentine fragments were assessed by Fourier infrared (FTIR) spectrophotometer before and after immersion in the solutions for the same periods. The positive control group included cells and tooth fragments maintained in Dulbecco's modified Eagle's medium (DMEM), and the negative control included tooth fragments that were kept dry. Data were analysed using anova and Tukey's test. RESULTS: No significant difference was found in cell viability evaluated by XTT (P > 0.05). Using the NR assay, WM and HBSS had significantly lower cell viability compared to the positive control group at 6 h (P < 0.05). SM had similar cell viability to the positive control group at all periods evaluated when assessed using all three tests (P > 0.05). A significant difference was found in values of OH- for the negative control group at 1 h (P = 0.002). CONCLUSIONS: Soya milk promoted better cell viability, whereas on dentine composition, the solutions behaved similarly. The association of different assay methods is promising for improving cell viability analysis. The 1-h time-point is a crucial factor in the prognosis of dental replantation because the teeth remain more hydrated and help maintain cell viability.
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Supervivencia Celular/efectos de los fármacos , Dentina/efectos de los fármacos , Leche , Leche de Soja/farmacología , Animales , Bovinos , Fibroblastos/efectos de los fármacos , Incisivo/efectos de los fármacos , Soluciones Preservantes de Órganos/farmacología , Raíz del Diente/efectos de los fármacosRESUMEN
BACKGROUND/AIM: Natural resources, such as coconut water, propolis, and egg whites, have been examined as possible storage media for avulsed teeth. However, there is a lack of research focused on the efficacy of these three products together compared with Hank's balanced salt solution and milk. The aim of this study was to evaluate the capacity of seven storage media to maintain the viability of human periodontal ligament fibroblasts (PDLFs). MATERIAL AND METHODS: PDLFs were kept at 5°C and 20°C, in skimmed milk (SMilk), whole milk (WMilk), recently prepared Hank's balanced salt solution (HBSS), Save-A-Tooth® system's HBSS (Save), natural coconut water (Coconut), Propolis, and egg white (Egg) for 3, 6, 24, 48, 72, 96, and 120 h, through the analysis of tetrazolium salt-based colorimetric (MTT) assay. RESULTS: At 5°C, SMilk and WMilk were better than HBSS in maintaining cell viability, from 24 h onward. At 20°C, HBSS was the best storage medium at 96 and 120 h. At both temperatures, from 6 h onward, Coconut, Propolis and Egg were less effective than SMilk, WMilk, and HBSS. In general, the performance of Coconut, Propolis and Egg were not influenced by storage temperature. However, the lowest temperature undermined the effectiveness of HBSS from 24 h and favored SMilk and WMilk, from 96 and 48 h onward, respectively. Save and water were the worst storage media. CONCLUSION: SMilk was the best storage medium, followed by WMilk and HBSS. Coconut, Propolis, and Egg can be indicated for the conservation of PDLF up to 3 h. The lower temperature (5°C) undermined the effectiveness of HBSS and favored SMilk and WMilk.
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Supervivencia Celular/efectos de los fármacos , Fibroblastos/fisiología , Soluciones Preservantes de Órganos/farmacología , Ligamento Periodontal/citología , Animales , Cocos , Clara de Huevo , Humanos , Soluciones Isotónicas/farmacología , Leche , Própolis/farmacología , TemperaturaRESUMEN
AIM: To investigate the ability of newly developed powdered coconut water formulas (ACP) with different osmolarities to maintain the viability of periodontal ligament (PDL) cells over time compared with other solutions. METHODOLOGY: Dogs teeth were extracted and stored for two periods, 3 h or 24 h, in the following media: long-shelf life CW (CW), pH-adjusted long-shelf life CW (pH-CW) and powdered CW that was pH and osmolality adjusted (ACP-404-I, 250 mOsm kg-1 H2 O; pH 7.0; ACP-404-II, 372 mOsm kg-1 H2 O; pH 7.0; ACP-404-III, 300 mOsm kg-1 H2 O; pH 7.4). The positive control group (Pc) corresponded to immediate measurement after tooth extraction, and two negative controls (Nc) corresponded to 3 h and 24 h of dry time. PDL cells were extracted, and cell viability analysed by Trypan blue exclusion. Data were analysed statistically using two-way anova followed by the Tukey test and one-way anova followed by the Dunnett test (P < 0.05). RESULTS: At 3 h and 24 h, ACP-404-I had a performance similar to those of ACP-404-II and pH-CW, with significantly higher (P = 0.004) percentages of viable cells than ACP-404-III and CW. The positive control group had a significantly higher (P = 0.002) percentage of viable cells than the negative control groups, CW and ACP-404-III, irrespective of the period evaluated. CONCLUSION: Powdered coconut water formulas, ACP-404-I and ACP-404-II, preserved viability for up to 24 h.
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Supervivencia Celular/efectos de los fármacos , Cocos , Soluciones Preservantes de Órganos/farmacología , Ligamento Periodontal/citología , Animales , Perros , Concentración Osmolar , PolvosRESUMEN
Liver transplantation is currently the preferred treatment option for end-stage liver disease. Donation after cardiac death was a common practice in the early years of organ donation before brain death criteria were established. Those organs were subjected to variable periods of warm ischemia that might intensify cold ischemia/reperfusion injuries. In the present, shortage of brain dead donors has led to the reassessment of organ donation after cardiac death. Since many cytoprotective roles have been describe for H2S during ischemia/reperfusion on a variety of tissues, we hypothesized that graft exposure to this bioactive gas might improve preservation of non-heart beating donated organs. Therefore, to establish a rat model of donation post-cardiac arrest and using this approach to judge H2S delivery effects on graft hypothermic preservation, were the main objectives of this investigation. Cardiopulmonary arrest was induced in sedated rats by overload of potassium (K(+)). Livers were surgically removed and subsequently stored in HTK Solution (Histidine-tryptophan-ketoglutarate) at 0-4°C. After 24 h of hypothermic preservation, livers were rewarmed in an ex vivo model. Three experimental groups were established as follows: I--Livers procured before cardiac death and cold stored 24 h in HTK (BCD); II--Livers procured after cardiac death (45 min) and cold stored 24 h in HTK (ACD); III--Livers procured after cardiac death (45 min) and cold stored 24 h in HTK+10 µM Sodium Sulfide (Na2S) (ACD-SS). Data suggest that after 45 min of warm ischemia, viability parameters assessed during reperfusion in the ex vivo model were significantly impaired. Real time PCR revealed that after ex vivo reperfusion there is an increased expression of HO-1 and TNF-α and a modest drop in Bcl-2 mRNA, which could be interpreted as the cellular response to the hypoxic insult sustained during warm ischemia. On the other hand, warm ischemic livers exposed to H2S during cold storage, improved microcirculation, morphology and viability parameters during ex vivo reperfusion and showed significant modulation of HO-1 mRNA expression. In conclusion, HTK supplementation with Na2S arose as a potential treatment to recover non-heart beating harvested organs. Furthermore, an appropriate model of cardiac dead liver donors was successfully developed.
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Trasplante de Hígado , Hígado/metabolismo , Preservación de Órganos/métodos , Sulfuros/farmacología , Isquemia Tibia , Animales , Criopreservación/métodos , Citoprotección/efectos de los fármacos , Glucosa/farmacología , Hemo-Oxigenasa 1/biosíntesis , Hemo-Oxigenasa 1/genética , Sulfuro de Hidrógeno/química , Isquemia/patología , Masculino , Malondialdehído/análisis , Manitol/farmacología , Soluciones Preservantes de Órganos/farmacología , Cloruro de Potasio/farmacología , Procaína/farmacología , Proteínas Proto-Oncogénicas c-bcl-2/genética , ARN Mensajero/biosíntesis , Ratas , Ratas Wistar , Daño por Reperfusión/prevención & control , Factor de Necrosis Tumoral alfa/biosíntesisRESUMEN
INTRODUCTION: This work focuses on ammonia metabolism of Liver Microorgans (LMOs) after cold preservation in a normothermic reoxygenation system (NRS). We have previously reported the development of a novel preservation solution, Bes-Gluconate-PEG 35 kDa (BG35) that showed the same efficacy as ViaSpan to protect LMOs against cold preservation injury. The objective of this work was to study mRNA levels and activities of two key Urea Cycle enzymes, Carbamyl Phosphate Synthetase I (CPSI) and Ornithine Transcarbamylase (OTC), after preservation of LMOs in BG35 and ViaSpan and the ability of these tissue slices to detoxify an ammonia overload in a NRS model. MATERIAL AND METHODS: After 48 h of cold storage (0°C in BG35 or ViaSpan) LMOs were rewarmed in KHR containing an ammonium chloride overload (1 mM). We determined ammonium detoxification capacity (ADC), urea synthesis and enzyme activities and relative mRNA levels for CPSI and OTC. RESULTS: At the end of reoxygenation LMOs cold preserved in BG35 have ADC and urea synthesis similar to controls. ViaSpan group demonstrated a lower capacity to detoxify ammonia and to synthesize urea than fresh LMOs during the whole reoxygenation period which correlated with the lower mRNA levels and activities for CPSI and OTC observed for this group. CONCLUSION: We demonstrate that our preservation conditions (48 hours, BG35 solution, anoxia, 0ºC) did not affect ammonia metabolism of cold preserved LMOs maintaining the physiological and biochemical liver functions tested, which allows their future use as biological component of a BAL system.
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Amoníaco/metabolismo , Frío , Hígado/efectos de los fármacos , Hígado/metabolismo , Soluciones Preservantes de Órganos/farmacología , Preservación de Órganos/métodos , Adenosina/farmacología , Alopurinol/farmacología , Animales , Carbamoil-Fosfato Sintasa (Amoniaco)/metabolismo , Glutatión/farmacología , Insulina/farmacología , Hígado/patología , Pruebas de Función Hepática , Masculino , Modelos Animales , Ornitina Carbamoiltransferasa/metabolismo , Oxígeno/administración & dosificación , Oxígeno/metabolismo , ARN Mensajero/metabolismo , Rafinosa/farmacología , Ratas , Ratas Wistar , Reperfusión , Factores de TiempoRESUMEN
PURPOSE: To evaluate the existence of in vitro long-term antimicrobial activity of Optisol-GS against microorganisms related to corneal infection using a closed-chamber study model. METHODS: Optisol-GS was contaminated with microorganisms related to corneal infections, and different times after contamination was analyzed using a closed-chamber study model. Microbial growths were analyzed by macroscopic observation. RESULTS: For Staphylococcus aureus and Pseudomonas aeruginosa, bacterial growth was observed in samples taken 1 hour through 7 days and 14 days after contamination occurred. For Staphylococcus epidermidis, Streptococcus agalactiae, and Candida albicans, microbial growth was observed in all samples studied. For Streptococcus pneumoniae, bacterial growth was observed in samples taken 1 hour through 72 hours after contamination. For Streptococcus pyogenes, bacterial growth was observed in samples taken 1 hour through 7 days after contamination. For Escherichia coli, bacterial growth was observed in samples taken 1 hour through 48 hours after contamination occurred. CONCLUSIONS: We conclude that no in vitro antimicrobial effect for any microorganism analyzed was observed in contaminated Optisol-GS after 72 hours; however, effective antimicrobial activity was observed for S. aureus, Str. pneumoniae, Str. pyogenes, P. aeruginosa, and E. coli after 7 to 10 days.
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Antiinfecciosos/farmacología , Bacterias/efectos de los fármacos , Sulfatos de Condroitina/farmacología , Córnea , Dextranos/farmacología , Hongos/efectos de los fármacos , Gentamicinas/farmacología , Soluciones Preservantes de Órganos/farmacología , Preservación de Órganos , Bacterias/crecimiento & desarrollo , Mezclas Complejas/farmacología , Cámaras de Difusión de Cultivos , Hongos/crecimiento & desarrollo , Estreptomicina/farmacologíaRESUMEN
Hypothermia is employed as a method to diminish metabolism rates and preserve tissues and cells. However, low temperatures constitute a stress that produces biochemical changes whose extension depends on the duration and degree of cold exposure and is manifested when physiological temperature is restored. For many cellular types, cold induces an oxidative stress that is dependent on the elevation of intracellular iron, damages macromolecules, and is prevented by the addition of iron chelators. Pisum sativum Ferredoxin-NADP(H) Reductase (FNR) has been implicated in protection from injury mediated by intracellular iron increase and successfully used to reduce oxidative damage on bacterial, plant and mammalian systems. In this work, FNR was expressed in Cos-7 cells; then, they were submitted to cold incubation and iron overload to ascertain whether this enzyme was capable of diminishing the harm produced by these challenges. Contrary to expected, FNR was not protective and even exacerbated the damage under certain circumstances. It was also found that the injury induced by hypothermia in Cos-7 cells presented both iron-dependent and iron-independent components of damage when cells were actively dividing but only iron-independent component when cells were in an arrested state. This is in agreement with previous findings which showed that iron-dependent damage is also an energy-dependent process.
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Frío/efectos adversos , Ferredoxina-NADP Reductasa/genética , Pisum sativum/enzimología , Adenosina/farmacología , Alopurinol/farmacología , Animales , Células COS , Supervivencia Celular/efectos de los fármacos , Chlorocebus aethiops , Disacáridos/farmacología , Electrólitos/farmacología , Glutamatos/farmacología , Glutatión/farmacología , Histidina/farmacología , Insulina/farmacología , Hierro/farmacología , L-Lactato Deshidrogenasa/metabolismo , Peroxidación de Lípido , Malondialdehído/metabolismo , Manitol/farmacología , Soluciones Preservantes de Órganos/farmacología , Oxiquinolina/farmacología , Rafinosa/farmacologíaRESUMEN
OBJECTIVE: Advances in graft reepithelialization and revascularization have renewed interest in airway transplantation. This study aims to determine whether topically applied preservation solutions can ameliorate ischemic injury to tracheal grafts. We analyzed 1) the effects of cold ischemia on the mucociliary clearance of tracheal grafts and 2) the impact of topically applied preservation solutions on the effects of cold ischemia on mucociliary clearance. METHOD: Tracheal segments (n=217) from 109 male Wistar rats were harvested, submerged in low-potassium-dextran-glucose, histidine-tryptophan-ketoglutarate, or saline solution (saline group), and stored at 4°C for 6, 10, 16, or 24 hours. A control group (not submerged) was analyzed immediately after harvesting. In situ mucociliary transport and ciliary beating frequency were measured using a stroboscope. Epithelial integrity, cellular infiltration, and mucus storage were quantified by light microscopy and image analysis software, along with transmission electron microscopy. RESULTS: 1) The effects of cold ischemia: in situ mucociliary transport and ciliary beating frequency were greater in the control group than after cold ischemia. Microscopic analysis results were similar between groups. 2) The effects of preservation solutions: there was no difference between the low-potassium-dextran-glucose, histidine-tryptophan-ketoglutarate, and saline groups in functional or light microscopy analysis. The saline group presented stronger signs of ischemic injury with transmission electron microscopy. CONCLUSIONS: Cold ischemia diminished the mucociliary clearance of the tracheal respiratory epithelium. Topically applied preservation solutions did not ameliorate the injury caused by cold ischemia to the tracheal respiratory epithelium.
Asunto(s)
Isquemia Fría/métodos , Soluciones Preservantes de Órganos/farmacología , Mucosa Respiratoria/efectos de los fármacos , Tráquea/efectos de los fármacos , Animales , Masculino , Microscopía Electrónica de Transmisión , Depuración Mucociliar/efectos de los fármacos , Ratas , Ratas Wistar , Mucosa Respiratoria/ultraestructura , Tráquea/trasplante , Tráquea/ultraestructuraRESUMEN
OBJECTIVE: Advances in graft reepithelialization and revascularization have renewed interest in airway transplantation. This study aims to determine whether topically applied preservation solutions can ameliorate ischemic injury to tracheal grafts. We analyzed 1) the effects of cold ischemia on the mucociliary clearance of tracheal grafts and 2) the impact of topically applied preservation solutions on the effects of cold ischemia on mucociliary clearance. METHOD: Tracheal segments (n=217) from 109 male Wistar rats were harvested, submerged in low-potassium-dextran-glucose, histidine-tryptophan-ketoglutarate, or saline solution (saline group), and stored at 4°C for 6, 10, 16, or 24 hours. A control group (not submerged) was analyzed immediately after harvesting. In situ mucociliary transport and ciliary beating frequency were measured using a stroboscope. Epithelial integrity, cellular infiltration, and mucus storage were quantified by light microscopy and image analysis software, along with transmission electron microscopy. RESULTS: 1) The effects of cold ischemia: in situ mucociliary transport and ciliary beating frequency were greater in the control group than after cold ischemia. Microscopic analysis results were similar between groups. 2) The effects of preservation solutions: there was no difference between the low-potassium-dextran-glucose, histidine-tryptophan-ketoglutarate, and saline groups in functional or light microscopy analysis. The saline group presented stronger signs of ischemic injury with transmission electron microscopy. CONCLUSIONS: Cold ischemia diminished the mucociliary clearance of the tracheal respiratory epithelium. Topically applied preservation solutions did not ameliorate the injury caused by cold ischemia to the tracheal respiratory epithelium. .
Asunto(s)
Animales , Masculino , Ratas , Isquemia Fría/métodos , Soluciones Preservantes de Órganos/farmacología , Mucosa Respiratoria/efectos de los fármacos , Tráquea/efectos de los fármacos , Microscopía Electrónica de Transmisión , Depuración Mucociliar/efectos de los fármacos , Ratas Wistar , Mucosa Respiratoria/ultraestructura , Tráquea/trasplante , Tráquea/ultraestructuraRESUMEN
OBJECTIVE: To compare histopathological findings and the degree of apoptosis among rat lungs preserved with low-potassium dextran (LPD) solution, histidine-tryptophan-ketoglutarate (HTK) solution, or normal saline (NS) at two ischemia periods (6 h and 12 h) using an experimental rat model of ex vivo lung perfusion. METHODS: Sixty Wistar rats were anesthetized, randomized, and submitted to antegrade perfusion via pulmonary artery with one of the preservation solutions. Following en bloc extraction, the heart-lung blocks were preserved for 6 h or 12 h at 4 ºC and then reperfused with homologous blood for 60 min in an ex vivo lung perfusion system. At the end of the reperfusion, fragments of the middle lobe were extracted and processed for histopathological examination. The parameters evaluated were congestion, alveolar edema, alveolar hemorrhage, inflammatory infiltrate, and interstitial infiltrate. The degree of apoptosis was assessed using the TdT-mediated dUTP nick end labeling method. RESULTS: The histopathological examination showed that all of the lungs preserved with NS presented alveolar edema after 12 h of ischemia. There were no statistically significant differences among the groups in terms of the degree of apoptosis. CONCLUSIONS: In this study, the histopathological and apoptosis findings were similar with the use of either LPD or HTK solutions, whereas the occurrence of edema was significantly more common with the use of NS.
Asunto(s)
Apoptosis , Hígado/patología , Pulmón , Soluciones Preservantes de Órganos/farmacología , Preservación de Órganos/métodos , Cloruro de Potasio/farmacología , Animales , Glucosa/farmacología , Hígado/efectos de los fármacos , Masculino , Manitol/farmacología , Perfusión/métodos , Procaína/farmacología , Edema Pulmonar , Distribución Aleatoria , Ratas , Ratas WistarRESUMEN
OBJETIVO: Comparar os achados histopatológicos e de apoptose em pulmões de ratos preservados em soluções low-potassium dextran (LPD, baixo potássio dextrana), histidine-tryptophan-ketoglutarate (HTK, histidina-triptofano-cetoglutarato) ou salina normal (SN) em 6 h e 12 h de isquemia pela utilização de um modelo experimental de perfusão pulmonar ex vivo. MÉTODOS: Sessenta ratos Wistar foram anestesiados, randomizados e submetidos à perfusão anterógrada pela artéria pulmonar com uma das soluções preservadoras. Após a extração, os blocos cardiopulmonares foram preservados por 6 ou 12 h a 4ºC, sendo então reperfundidos com sangue homólogo em um sistema de perfusão ex vivo durante 60 min. Ao final da reperfusão, fragmentos do lobo médio foram extraídos e processados para histopatologia, sendo avaliados os seguintes parâmetros: congestão, edema alveolar, hemorragia alveolar, hemorragia, infiltrado inflamatório e infiltrado intersticial. O grau de apoptose foi avaliado pelo método TdT-mediated dUTP nick end labeling. RESULTADOS: A histopatologia demonstrou que todos os pulmões preservados com SN apresentaram edema alveolar após 12 h de isquemia. Não houve diferenças em relação ao grau de apoptose nos grupos estudados. CONCLUSÕES: No presente estudo, os achados histopatológicos e de apoptose foram semelhantes com o uso das soluções LPD e HTK, enquanto a presença de edema foi significativamente maior com o uso de SN.
OBJECTIVE: To compare histopathological findings and the degree of apoptosis among rat lungs preserved with low-potassium dextran (LPD) solution, histidine-tryptophan-ketoglutarate (HTK) solution, or normal saline (NS) at two ischemia periods (6 h and 12 h) using an experimental rat model of ex vivo lung perfusion. METHODS: Sixty Wistar rats were anesthetized, randomized, and submitted to antegrade perfusion via pulmonary artery with one of the preservation solutions. Following en bloc extraction, the heart-lung blocks were preserved for 6 h or 12 h at 4ºC and then reperfused with homologous blood for 60 min in an ex vivo lung perfusion system. At the end of the reperfusion, fragments of the middle lobe were extracted and processed for histopathological examination. The parameters evaluated were congestion, alveolar edema, alveolar hemorrhage, inflammatory infiltrate, and interstitial infiltrate. The degree of apoptosis was assessed using the TdT-mediated dUTP nick end labeling method. RESULTS: The histopathological examination showed that all of the lungs preserved with NS presented alveolar edema after 12 h of ischemia. There were no statistically significant differences among the groups in terms of the degree of apoptosis. CONCLUSIONS: In this study, the histopathological and apoptosis findings were similar with the use of either LPD or HTK solutions, whereas the occurrence of edema was significantly more common with the use of NS.
Asunto(s)
Animales , Masculino , Ratas , Apoptosis , Pulmón , Hígado/patología , Soluciones Preservantes de Órganos/farmacología , Preservación de Órganos/métodos , Cloruro de Potasio/farmacología , Glucosa/farmacología , Hígado/efectos de los fármacos , Manitol/farmacología , Edema Pulmonar , Perfusión/métodos , Procaína/farmacología , Distribución Aleatoria , Ratas WistarRESUMEN
The objective was to evaluate the efficiency of phosphate-buffered saline (PBS) and Minimum Essential Medium (MEM) during the transport of equine preantral and antral follicles at various temperatures and incubation interval. Equine ovaries (n = 10) from an abattoir were cut into 19 fragments; one was immediately fixed in Bouin's solution (control) and the other fragments were placed in PBS or MEM solution at 4, 20, or 39 °C for 4, 12, or 24 h. After the respective incubation periods, all fragments were fixed in Bouin's solution for 24 h and then submitted to standard histologic analysis. In total, 2567 ovarian follicles were analyzed, including 1752 primordial, 764 primary, 34 secondary and seven antral follicles. Relative to the control group, the transport of equine ovarian fragments in both solutions significantly reduced the percentage of morphologically normal follicles with increasing time and temperature. At 4 °C for 4 h, considering primordial and developing follicles, PBS had a higher (P < 0.05) rate (98.9%) of morphologically normal follicles than MEM, 48.7%. At 39 °C for 12 h, all follicles in both solutions were degenerated. Regarding the stage of follicular development, primordial follicles were less (P < 0.05) affected by preservation than primary and secondary follicles in all media, times and temperatures tested, except at 4 °C for 12 h in PBS, in which the primary and secondary follicles were less (P < 0.05) affected. Overall, 43% of antral follicles were morphologically normal when maintained in MEM at 4 °C for 4 h. In conclusion, equine follicles were successfully preserved in ovarian fragments at 4 °C in phosphate-buffered saline for up to 4 h.
Asunto(s)
Permeabilidad de la Membrana Celular/efectos de los fármacos , Caballos , Soluciones Preservantes de Órganos/farmacología , Folículo Ovárico , Temperatura , Conservación de Tejido/métodos , Algoritmos , Animales , Cruzamiento/métodos , Permeabilidad de la Membrana Celular/fisiología , Medios de Cultivo/farmacología , Femenino , Caballos/fisiología , Folículo Ovárico/efectos de los fármacos , Folículo Ovárico/ultraestructura , Estaciones del Año , Factores de Tiempo , Conservación de Tejido/veterinariaRESUMEN
PURPOSE: To evaluate the effects of ischemic preconditioning (IPC) associate with different preservation solutions, in the protecting of gut. METHODS: Four groups of 14 rats underwent laparotomy and collecting 20 cm of ileum, for preservation, at 4ºC, in Belzer (Belz), Ringer (RL), Celsior (Cs) and Custodiol (Cust) solutions, for 24 hours. Prior to collection, half of the animals in each group were subjected to IPC. During preservation, in the periods of zero, 12, 18 and 24 hours, were conducted evaluating the degree of mucosal injury and dosage of malondialdehyde acid (MDA). RESULTS: In all periods the RL group, with and without IPC, presented MDA values higher than the Belz and Cs. The degree of mucosal injury in the non-ipc RLgroup with 12h preservation was higher than the others; with 18 and 24h, the RL and Cust had higher degrees of damage than Cs and Belz. With IPC, in all periods, the group Cs and Belz had lower degrees of injury. CONCLUSION: The Celsior and Belzer solutions had better protective effects on the gut and these effects were enhanced by IPC.
OBJETIVO: Avaliar os efeitos do precondicionamento isquêmico (PCI) associado a diferentes soluções de preservação, na proteção do intestino delgado. MÉTODOS: Quatro grupos de 14 ratos Wistar, foram submetidos à laparotomia e coleta de 20 cm de íleo, para preservação, a 4ºC, nas soluções de Belzer (Belz), Ringer (RL), Celsior (Cs) e Custodiol (Cust) por 24 horas. Previamente à coleta, em metade dos animais de cada grupo, o intestino foi submetido ao PCI. Durante a preservação, nos períodos de Zero, 12, 18 e 24 horas, foram realizados avaliação do grau de lesão da mucosa e dosagem do ácido malondialdeído (MDA). RESULTADOS: Em todos os períodos o grupo RL, sem e com pci, apresentou valores maiores de MDA do que o Belz e Cs. O grau de lesão da mucosa nos grupos sem pci com preservação de 12h, no grupo RL, foi maior que nos demais; com 18h e 24h o grupo RL e Cust apresentaram maiores graus de lesão do que Cs e Belz. Com o pci, em todos os períodos, os grupos Belz e Cs apresentaram menores graus de lesão CONCLUSÃO: As Soluções Celsior e Belzer tiveram melhores efeitos na proteção do intestino e estes efeitos foram incrementados pelo precondicionamento isquêmico.