Effect of vaporized perfluorocarbon on oxidative stress during the cold ischemia phase of lung graft preservation / Efeito do perfluorocarbono vaporizado sobre o estresse oxidativo no período de isquemia fria durante a preservação pulmonar

ABSTRACT Liquid perfluorocarbon (PFC) instillation has been studied experimentally as an adjuvant therapy in the preservation of lung grafts during cold ischemia. The objective of this study was to evaluate whether vaporized PFC is also protective of lung grafts at different cold ischemia times. We performed histological analysis of and measured oxidative stress in the lungs of animals that received only preservation solution with low-potassium dextran (LPD) or vaporized PFC together with LPD. We conclude that vaporized PFC reduces the production of free radicals and the number of pulmonary structural changes resulting from cold ischemia.

RESUMO O perfluorocarbono (PFC) líquido tem sido estudado experimentalmente como uma substância adjuvante na preservação de enxertos pulmonares durante o período de isquemia fria. O objetivo deste estudo foi avaliar se o PFC vaporizado (e não instilado) também atuaria como protetor de enxertos pulmonares em diferentes tempos de isquemia fria. Realizamos análise histológica e dosamos o estresse oxidativo em pulmões de animais que receberam somente uma solução de preservação com low-potassium dextran (LPD, dextrana com baixa concentração de potássio) ou PFC vaporizado associado a LPD. Concluímos que o PFC vaporizado reduziu a produção de radicais livres e provocou menor número de alterações estruturais pulmonares decorrentes do período de isquemia fria que o uso de LPD isoladamente.

Use of nanoparticles to monitor human mesenchymal stem cells transplanted into penile cavernosum of rats with erectile dysfunction

PURPOSE: This study was performed to examine the treatment of erectile dysfunction by use of superparamagnetic iron oxide nanoparticles-labeled human mesenchymal stem cells (SPION-MSCs) transplanted into the cavernous nerve injured cavernosa of rats as monitored by molecular magnetic resonance imaging (MRI). MATERIALS AND METHODS: Eight-week-old male Sprague-Dawley rats were divided into three groups of 10 rats each: group 1, sham operation; group 2, cavernous nerve injury; group 3, SPION-MSC treatment after cavernous nerve injury. Immediately after the cavernous nerve injury in group 3, SPION-MSCs were injected into the cavernous nerve injured cavernosa. Serial T2-weighted MRI was done immediately after injection and at 2 and 4 weeks. Erectile response was assessed by cavernous nerve stimulation at 2 and 4 weeks. RESULTS: Prussian blue staining of SPION-MSCs revealed abundant uptake of SPION in the cytoplasm. After injection of 1x10(6) SPION-MSCs into the cavernosa of rats, T2-weighted MRI showed a clear hypointense signal induced by the injection. The presence of SPION in the corpora cavernosa was confirmed with Prussian blue staining. At 2 and 4 weeks, rats with cavernous nerve injury had significantly lower erectile function than did rats without cavernous nerve injury (p<0.05). The group transplanted with SPION-MSCs showed higher erectile function than did the group without SPION-MSCs (p<0.05). The presence of SPION-MSCs for up to 4 weeks was confirmed by MRI imaging and Prussian blue staining in the corpus cavernosa. CONCLUSIONS: Transplanted SPION-MSCs existed for up to 4 weeks in the cavernous nerve injured cavernosa of rats. Erectile dysfunction recovered and could be monitored by MRI.

Physicochemical Properties and Interfacial Adaptation of Root Canal Sealers

This study compared the physicochemical properties and interfacial adaptation to canal walls of Endo-CPM-Sealer, Sealapex and Activ GP with the well-established AH Plus sealer. The following analyses were performed: radiopacity, pH variation and solubility using samples of each material and scanning electron microscopy of root-filled bovine incisors to evaluate the interfacial adaptation. Data were analyzed by the parametric and no-parametric tests (α=0.05). All materials were in accordance with the ANSI/ADA requirements for radiopacity. Endo-CPM-Sealer presented the lowest radiopacity values and AH Plus was the most radiopaque sealer (p=0.0001). Except for ActiV GP, which was acidic, all other sealers had basic chemical nature and released hydroxyl ions. Regarding solubility, all materials met the ANSI/ADA recommendations, with no statistically significant difference between the sealers (p=0.0834). AH Plus presented the best adaptation to canal walls in the middle (p=0.0023) and apical (p=0.0012) thirds, while the sealers Activ GP and Endo-CPM-Sealer had poor adaptation to the canal walls. All sealers, except for ActiV GP, were alkaline and all of them fulfilled the ANSI/ADA requirements for radiopacity and solubility. Regarding the interfacial adaptation, AH Plus was superior to the others considering the adaptation to the bovine root canal walls.

Este estudo comparou as propriedades físico-químicas e a adaptação interfacial às paredes do canal dos cimentos Endo-CPM-Sealer, Sealapex e Activ GP com o bem estabelecido cimento AH Plus. As seguintes análises foram realizadas: radiopacidade, variação de pH e de solubilidade utilizando amostras de cada material, e microscopia eletrônica de varredura utilizando incisivos bovinos obturados para avaliar a adaptação interfacial. Os dados foram analisados utilizando testes paramétricos e não-paramétricos (α=0,05). Todos os materiais estavam de acordo com os requerimentos da ANSI/ADA para radiopacidade, sendo que o Endo-CPM-Sealer apresentou os menores valores de radiopacidade e o AH Plus foi o cimento mais radiopaco (p=0,0001). Exceto o Activ GP, que foi ácido, todos os outros cimentos apresentaram natureza química básica e liberaram íons hidroxila. Com relação à solubilidade, todos os materiais estavam de acordo com as recomendações da ANSI /ADA, sem diferença significante entre os cimentos (p=0,0834). O AH Plus apresentou a melhor adaptação às paredes do canal nos terços médio (p=0,0023) e apical (p=0,0012), enquanto que os cimentos Activ GP e Endo-CPM-Sealer apresentaram uma pobre adaptação às paredes do canal. Em conclusão, todos os cimentos, exceto o Activ GP, foram alcalinos e todos preencheram os requerimentos da ANSI/ADA para radiopacidade e solubilidade. Com relação à adaptação interfacial, o AH Plus foi superior aos demais para adaptação às paredes do canal radicular de incisivos bovinos.

Evaluation of Porcine Pancreatic Islets Transplanted in the Kidney Capsules of Diabetic Mice Using a Clinically Approved Superparamagnetic Iron Oxide (SPIO) and a 1.5T MR Scanner

OBJECTIVE: To evaluate transplanted porcine pancreatic islets in the kidney capsules of diabetic mice using a clinically approved superparamagnetic iron oxide (SPIO) and a 1.5T MR scanner. MATERIALS AND METHODS: Various numbers of porcine pancreatic islets labeled with Resovist, a carboxydextran-coated SPIO, were transplanted into the kidney capsules of normal mice and imaged with a 3D FIESTA sequence using a 1.5T clinical MR scanner. Labeled (n = 3) and unlabeled (n = 2) islets were transplanted into the kidney capsules of streptozotocin-induced diabetic mice. Blood glucose levels and MR signal intensities were monitored for 30 days post-transplantation. RESULTS: There were no significant differences in viability or insulin secretion between labeled and unlabeled islets. A strong correlation (r2 > 0.94) was evident between the number of transplanted islets and T2 relaxation times quantified by MRI. Transplantation with labeled or unlabeled islets helped restore normal sustained glucose levels in diabetic mice, and nephrectomies induced the recurrence of diabetes. The MR signal intensity of labeled pancreatic islets decreased by 80% over 30 days. CONCLUSION: The transplantation of SPIO-labeled porcine islets into the kidney capsule of diabetic mice allows to restore normal glucose levels, and these islets can be visualized and quantified using a 1.5T clinical MR scanner.

Successful scale-up of human embryonic stem cell production in a stirred microcarrier culture system

Future clinical applications of human embryonic stem (hES) cells will require high-yield culture protocols. Currently, hES cells are mainly cultured in static tissue plates, which offer a limited surface and require repeated sub-culturing. Here we describe a stirred system with commercial dextran-based microcarriers coated with denatured collagen to scale-up hES cell production. Maintenance of pluripotency in the microcarrier-based stirred system was shown by immunocytochemical and flow cytometry analyses for pluripotency-associated markers. The formation of cavitated embryoid bodies expressing markers of endoderm, ectoderm and mesoderm was further evidence of maintenance of differentiation capability. Cell yield per volume of medium spent was more than 2-fold higher than in static plates, resulting in a significant decrease in cultivation costs. A total of 10(8) karyotypically stable hES cells were obtained from a unitary small vessel that needed virtually no manipulation during cell proliferation, decreasing risks of contamination. Spinner flasks are available up to working volumes in the range of several liters. If desired, samples from the homogenous suspension can be withdrawn to allow process validation needed in the last expansion steps prior to transplantation. Especially when thinking about clinical trials involving from dozens to hundreds of patients, the use of a small number of larger spinners instead of hundreds of plates or flasks will be beneficial. To our knowledge, this is the first description of successful scale-up of feeder- and Matrigel™-free production of undifferentiated hES cells under continuous agitation, which makes this system a promising alternative for both therapy and research needs.

Cambio del isotipo en la respuesta a antígenos T independientes por co-inoculación con antígenos T dependientes / Isotype switch in the response against T independent antigens by co-inoculation with T dependent antigens

Un paradigma clásico de la inmulogía plantea que para que ocurra cambio de isotipo en los anticuerpos es condición sine qua non la presentación del antígeno a un linfócito T colaborador por parte de una célula presentadora de antígenos. En el presente trabajo se diseñó un modelo animal, ratones BALB/c, de respuesta inmune frente a dos antígenos típicos. Se utilizo dextrán como antígeno T independiente (AgTI) y seroalbúmina bovina (SAB) como antígeno T dependiente (AgTD), y se estúdio la respuesta, analizando los isotipos de los anticuerpos específicos producidos. Los resultados obtenidos muestran que la respuesta a dextrán en presencia de SAB ocurre con cambio de isotipo (swith), essencialmente de IgM a IgG. Estos experimentos sugieren que la SAB genera un entorno bioquímico inductor de cambio de isotipo tanto en supropia via de procesamiento como en del dextrán. Los resultados señalan que la asociación exclusiva de los AgTDs con las respuestas em las que ocurre cambio de isotipo es incorrecta. Considerando el modelo propuesto resulta poco probable encontrar in vivo y en forma espontânea casos en los que los AgTIs ingreses al organismo aislados; en cambio, es mucho más probable que el ingreso ocurra conjuntamente con AgTDs, y en consecuencia ocurra cambio de isotipo.

Determination of macromolecular exchange and PO2 in the microcirculation: a simple system for in vivo fluorescence and phosphorescence videomicroscopy

We have developed a system with two epi-illumination sources, a DC-regulated lamp for transillumination and mechanical switches for rapid shift of illumination and detection of defined areas (250-750 æm²) by fluorescence and phosphorescence videomicroscopy. The system permits investigation of standard microvascular parameters, vascular permeability as well as intra- and extravascular PO2 by phosphorescence quenching of Pd-meso-tetra (4-carboxyphenyl) porphine (PORPH). A Pechan prism was used to position a defined region over the photomultiplier and TV camera. In order to validate the system for in vivo use, in vitro tests were performed with probes at concentrations that can be found in microvascular studies. Extensive in vitro evaluations were performed by filling glass capillaries with solutions of various concentrations of FITC-dextran (diluted in blood and in saline) mixed with different amounts of PORPH. Fluorescence intensity and phosphorescence decay were determined for each mixture. FITC-dextran solutions without PORPH and PORPH solutions without FITC-dextran were used as references. Phosphorescence decay curves were relatively unaffected by the presence of FITC-dextran at all concentrations tested (0.1 æg/ml to 5 mg/ml). Likewise, fluorescence determinations were performed in the presence of PORPH (0.05 to 0.5 mg/ml). The system was successfully used to study macromolecular extravasation and PO2 in the rat mesentery circulation under controlled conditions and during ischemia-reperfusion

Estudo da açäo antiinflamatória e potência ulcerogênica de compostos mesoiônicos, 1,4-difeniltiossemicarbazida e derivados / Study of the antiinflammatory action and ulcerogenic potency of mesoionics, 1,4-diphenylthiosemicarbazide compounds and derivatives

Os compostos 1,4-difeniltiossemicarbazida (DTSC), derivados (`IR IND. 1ï; `IR IND. 2ï e `IR IND. 3ï) e mesoiônicos (`MIR IND. 1ï, `RMIR IND. 1ï e `RMIR IND. 3ï) foram avaliados quanto à atividade antiinflamatória. os compostos DTSC, `IR IND. 1ï, `IR IND. 2ï, e `RMIR IND. 1ï inibiram significativamente o pico máximo do edema induzido por carragenina em 82 por cento, 72 por cento, 34 por cento e 78 por cento, respectivamente. Porém, somente os derivados mesoiônicos inibiram o edema induzido por dextrano. Já no edema desencadeado por histamina, somente o composto IR, não foi capaz de antagonizar de maneira significativa a ação da histamina...

Hypertonic-hyperosmotic solution modifies myoglobin levels in early reperfusion after ischemic cardiac arrest. Experimental model

To evaluate the effect of reperfusison with hypertonic-hyperosmotic solution, cardiectomy was performed in 25 New Zealand white rabbits. Seven isolated hearts were submitted to 30 min of global ischemia and reperfused with oxygenated uffer for 60 min. Myoglobin and isoensyme MB of creatine kinase concnetrations were each measured in the effluent 15 min, and values were correlated (r=0.5011, p=0.015). After this procedure, 18 isolated hearts were radomized in two groups. Hearts of group I were reperfused with hypertonic-hyperosmotic solution (NaCl 7.5 percent dextran 60,000 MW) diluted in oxygenated buffer, and group II with oxygenated buffer. Myoglobin and coronary flow were measured in both groups, group I showed lower levels of myoglobin (p=0.0069) and higher coronary flow (p = 0.028) than group II. In conclusion, changes in myoglobin concentration in the heart effluent are more sensitive than changes in isoenxyme MB of creatine kinase; thus, evaluation of this parameter may be useful in the detection of ishcemia reperfusion injury. Additionally, hypertonic-hyperosmotic solution improves the coronary flow and has a protective effect against ichemia-reperfusion injury

Pertussis toxin from Bordetella pertussis blocks neutrophil migration and neutrophil-dependent edema in response to inflammation

Pertussis toxin (Ptx) is a hexameric protein with classical AB architecture produced by Bordetella pertussis. The aim of this study was to investigate the effect og Ptx on migration of polymorphonuclear leukocytes to site of inflamation and on cell- dependent edema. Ptx was purified from the supernatant of the culture medium of B. pertussis using hydroxylapatite chromatography and fetuin affinity chromatography. Ptx induced a maximal clusterin of Chinese hamster ovary cells at concentration as low as 0.1 ng/ ml. Intravenous inection of Ptx (400 ng) significantly blocked the neutrophil migration induced by 200 ng of lipopolysaccharide (LPS from E. coli O111:B4; 2.27 ñ 0.13 vs 0.61 ñ 0.16 per 10**6 neutrophils/ml; P < 0.001; N = 5) and by 200 ng of formylmethionyl-leucyl-phenylalanine(fMLP; 2.53 ñ 0.45 vs 0.75 ñ 0.14 per 10**6 neutrophils/ml; P < 0.01; N=6) into the peritoneal cavities of male Wistar rats (eighing 150-180). In addition, Ptx (400ng) pretreatment also blocked the edema induced by intraplantar injection of 100 µg carrageenin ( increase in volume: 0.667 ñ 0.087 vs 0.313 ñ 0.058 ml; P < 0.01; N = 5) but not the edema induced by 100 µg dextran ( increase in volume: 0.537 ñ 0.06 vs 0.385 ñ 0.076 ml; P > 0.05; N = 5). These data demonstrate that Ptx blocked neutrophil migration induced by a direct f MLP stimulus of a site of inflammation. In addition, this toxin blocks the indirect stimulus of LPS on neutrophil migration. Furthermore, Ptx also inhibits the neutrophil-dependent edema induced by carrageenin, but not the edema induced by dextran that is in part dependent on basophil cells. These results warrant further studies on the mechanisms of Ptx inhibition of neutrophil-dependent edema and cell migration

Effect of acute haemodilution on right atrial type-A receptor activity and right atrial pressure in anaesthetized cats.

To determine the effects of acute haemodilution on right atrial pressure and right atrial type-A receptor activity, experiments were conducted in nine anaesthetized cats. Haemodilution was induced by replacement of blood by same volume of dextran (molecular weight-150000). Arterial blood pressure, right atrial pressure and right atrial type-A receptor activity were recorded. Hematocrit, arterial blood pO2, pCO2, and pH were measured. Mean arterial blood pressure, pO2, pCO2, and pH were not altered by haemodilution which extended to a hematocrit level of about 13%. Mean right atrial pressure, 'a' wave pressure showed significant changes after 2nd and 3rd exchange. Heart rate and 'a' wave amplitude changed significantly (P < 0.05) after 3rd exchange. The amplitude of the 'a' wave was increased from 3.57 +/- 0.93 to 5.79 +/- 1.46 cm H2O, heart rate was increased from 163.89 +/- 5.36 to 169.22 +/- 4.84 beats/min, mean right atrial pressure was increased from 2.85 +/- 0.31 to 4.61 +/- 0.27 cm H2O, initial 'a' wave pressure was increased from 1.78 +/- 0.09 to 5.06 +/- 0.44 cm H2O. The data indicate that under present experimental conditions, the activity of the type-A receptor did not show any significant change (P > 0.05) on acute haemodilution. Significant increase (P < 0.05) in the amplitude of 'a' wave, mean right atrial pressure and initial 'a' wave pressure could be attributed to increased myocardial contractility.

Efeitos da ofloxacina na inflamaçäo induzida em ratos / Effects of ofloxacin over the induced inflammation in rats

O objetivo desta pesquisa foi investigar a eventual açao antiinflamatória do antibiótico ofloxacina, em ratos. O efeito da ofloxacina foi pesquisado no edema experimental de pata por carragenina e por dextrana, na pleurisia por carragenina, na inflamaçao crônica por pelotas de algodao implantadas na parede abdominal e no granuloma de corpo estranho por fio de polipropileno implantado no dorso. Costatou-se que a ofloxacina nao influenciou a evoluçao da inflamaçao, exceto por inibir (em altas doses) a proliferaçao de células inflamatórias mononucleares do granuloma de corpo estranho formado pela presença de fio de polipropileno. Concluiu-se que a ofloxacina tem açao antiproliferativa sobre células inflamatórias de ratos.

Use of NaCl 7,5 per cent versus NaCl 7,5 per cent dextran in the resuscitation of septoc rats

Foi utilizado NaCl 7,5 por cento (HS) e NaCl 7,5 por cento dextran (HS/DX) no tratamento de choque séptico experimental em ratos. A infusäo de HS resultou numa melhora imediata e sustentada da pressäo arterial média (MAP) com um aumento da taxa de sobrevivência, comparada com animais controles ou tratados com HS/DX. A adiçäo de dextran à soluçäo de salina hipertônica näo foi capaz de aumentar o efeito protetor causado pela HS em ratos sépticos, pois estes mostraram-se alérgicos a estas macromoléculas

Specific opiate effects on dextran-induced inflammation in rats

Los efectos de los opiáceos fueron estudiados en dos modelos de inflamación aguda en la rata. La morfina (10mg/Kg, i.P.) inhibió en un 50% el edema de la pata inducido por la inyección interdigital de una solución al 1% de dextran. Dosis bajas de naltrexona, aunque no las más altas, produjeron una inhibición similar. La naltrexona (10mg/Kg, i.p.) prevno similar. La naltrexona (10mg/Kg, i,p.) previno completamete el efecto antiedematoso de la morfina. La anestesia local del miembro inferior con lidocaina no modificó el edema producido por dextran ni la acción de la morfina. La morfina (10mg/Kg, i.p.) aumentó en un 50% la permeabilidad vascular d ela piel provocada por la inyección intradérmica de una solución de dextran al 1%. Los resultados presentados sugieren un papel modulatorio específico de los opiáceos sobre las repuestas inflamatorias agudas de la rata

Do lymphocytes play a pro-inflammatory role in any pharmacological type of rat paw edema?

1. The hypothesis of a pro-inflammatory activity in lymphocytes related to the development of nonimmune acute inflammation was tested in leucopenic rats injected subcutaneously with various stimuli representing different pharmacological classes of paw edema-inducing agents into the subplantar area of one of the hind paws. 2. Amethopterin-induced leucopenia rendered animals hyporeactive to carrageenin and trypsin, but not to kaolin or dextran. 3. Administration of splenic lymphocytes to leucopenic rats to correct lymphopenia partially restored the inhibited responses to carrageenin and the early phase of responses to trypsin. 4. These results suggest that lymphocytes may play a pro-inflammatory role in nonimmune acute inflammation, with is more demonstrable in some pharmacological classes of paw edema than in others

Técnica de preparaçäo do meio de conservaçäo de córneas de McCarey-Kaufman modificado / Technique of the media preparation for corneal preservation of modified McCarey-Kaufman

Este trabalho relata a técnica de preparaçäo do meio de conservaçäo de córneas de McCarey-Kaufman modificado na Divisäo de Farmácia do Hospital das Clínicas da Faculdade de Medicina da Universidade de Säo Paulo (meio "MMK-HB"), tendo-se obtido um meio que está de acordo com os padröes estabelecidos na literatura