In the 19th century, halothane hepatitis became a sensitive and well-known subject in human anesthesiology due to the production of a noxious metabolite further discovered, trifluoroacetic acid. Subsequently, isoflurane, enflurane, and desflurane were also investigated for potentially causing hepatitis through the same metabolite. Sevoflurane, however, does not generate trifluoroacetic acid and is quickly conjugated and excreted. For more than four decades these anesthetics have been experimentally developed for intravenous injection by having added either a lipid or fluorocarbon-based carrier to produce general anesthesia with less drug and faster onset of action. The use of intravenous emulsified halogenated anesthetics as an alternative to inhalation brought contradictory findings, therefore they are still not utilized in the clinical settings of veterinary and human anesthesia. The high solubility of these anesthetic emulsions increases their tissue uptake, volume of distribution, and potency. By this means, the amount of anesthetic necessary to establish general anesthesia could be significantly reduced but would still carry the risk of causing hepatic toxicity. On the other hand, because the emulsified anesthetics have a higher tissue uptake and are liposoluble, they remain for longer periods in the cellular membrane providing cellular pre- and postconditioning effects by minimizing cellular deleterious responses to a critical environment. Emulsified isoflurane and sevoflurane are the most investigated anesthetics for this purposein the heart, brain, kidneys, liver, and central nervous system of laboratory animals and human volunteers. The focus of this study is to evaluate the cellular effects of the loaded-lipid emulsified isoflurane and sevoflurane at different concentrations on cultured primary canine hepatocytes considering their viability and apoptosis response. Specifically, the overall objective is to establish a basis for in vitro metabolism of these emulsified anesthetics on canine hepatocytes under normal oxygen tension and on canine hepatocytes exposed to extreme hypoxia (1% O2). Thus, this study is sectioned into three major chapters followed by conclusions and future studies to determine the safety and indication of these anesthetic formulations in canine hepatocytes to be further explored in the clinical setting with live animals.

Emulsões artificiais sem proteína simulando quilomicrons e remanescentes de quilomícron foram preparadas por sonicalcação de trioleína, lecitina, colesteril oleato e colesterol em solução aquosa. A seguir foram ultracentrifugadas em gradiente descontínuo de densidade. As emulsões, marcadas com 3H-trioleína e 14C-colesteril oleato foram injetadas via intra-arterial em ratos controle e em ratos tratados com Triton WR1339, sulfato de protamina e heparina, medindo-se a seguir a remoção plasmática do colesteril-ester e dos triglicérides, durante dez minutos. O Triton WR 1339 e a protamina inibiram a lipólise dos quilomícrons artificiais, diminuindo a remoção destas partículas do plasma. O Triton WR 1339 mostrou ser mais efetivo que a protamina nestes efeitos. Por outro lado, a heparina promoveu uma lipólise rápida e brusca nos quilomícrons artificiais, assim como uma aceleração na remoção destas partículas do plasma. Em contraste flagrante com esses resultados, o metabolismo dos remanescentes de quilomícron não foi consideravelmente afetado pelo tratamento com Triton e heparina. Estas experiências indicam que as emulsões artificiais reproduzem o comportamento metabólico dos quilomícrons e remanescentes de quilomícron naturais, em condições em que a atividade da lipase lipoproteica esteja alterada. / Protein-free emulsions models of chylomicrons and chylomicron remnants were prepared by sonicating triolein, lecithin., cholesteryl oleate and cholesterol in aqueous saline media, followed by ultracentrifugation in density gradient solution. The 3H-triolein and 14C-cholesteryl oleate labeled emulsions were injected into the carotid artery of control rats and rats treated with Triton WR 1339, protamine sulphate and heparin. Plasma removal of both labels was measured during ten minutes in two minutes intervals. Triton WR 1339 and protamine sulphate strongly inhibited lipolysis of chylomicron-like emulsions leading to delayed removal of the particles from blood. Triton WR 1339 de appeared to be more effective than protamine to elicit these effects. On the other hand, heparin produced instantaneous lipolysis of the chylomicron-like particles markedly enhancing its removal from plasma. Contrarily, chylomicron remnant-like emulsions were not considerably affected either by Triton WR 1339 or by heparin treatment. The above described results obtained with artificial emulsions support current concepts on the metabolic behavior of natural chylomicron and remnant submitted to changes in lipoprotein lipase action.

Made available in DSpace on 2016-12-08T16:24:08Z (GMT). No. of bitstreams: 1 PGCA10MA067.pdf: 666461 bytes, checksum: 5e9cce1bde4afb2aa807b32906354bd9 (MD5) Previous issue date: 2010-10-17 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / It is clear that different formulations of propofol have differentiated pharmacokinetic and pharmacodynamic profiles. Thus, this study aimed to evaluate the cardiovascular, respiratory, haemogasometric and metabolic effects, as well as observe possible side effects of the new formulation of propofol in nanoemulsion compared to the propofol in lipid emulsion commercially available. For this purpose, 12 healthy female cats, average weight of 2.6 ± 0.4 kg, were assigned into two groups: Nanoemulsion (NAG, n = 6) and Emulsion (EMG, n = 6), in which the animals received propofol in nanoemulsion or in lipid emulsion, respectively, as induction agent, both at a sufficient dose for intubation. Immediately after induction, the animals were intubated and supplemented with 100% oxygen through a nonrebreathing circuit. Subsequently, the propofol infusion was started with the respective formulation, at a constant rate of 0.3 mg kg-1 min-1, and maintained for 90 minutes. The parameters were evaluated at the following moments: T-10, T0, T15, T30, T45, T60, T75, T90, corresponding to the baseline evaluation and 0, 15, 30, 45, 60, 75 and 90 minutes after starting the infusion, respectively. Data were compared using repeated measures ANOVA (data over time) followed by the Student Newman Kews test when justified or t-tests as appropriate (significance taken as p&#8804;0.05). The dose required for induction was 9.5 ± 1.3 mg kg-1 and 10 ± 1 mg kg-1 for EMG and NAG, respectively. In the EMG, a reduction was observed in the heart rate, respiratory rate (f), pH, and systolic (SAP), diastolic (DAP) and mean (MAP) arterial pressure in all the moments when compared to the basal evaluation. The f decreased only in T90, and the pH in the T0, T15 and T90, when using the nanoemulsion. The PaCO2, PaO2 and SaO2 remained higher during all the infusion period in both groups in comparison to the baseline. HCO3 increased in all the moments in the EMG and in the T15, T30, T60 and T90 in the NAG. The CVP was lower than the baseline only in T60 and T90 in the NAG. Between groups, lower values occurred in all the moments evaluated for SAP, MAP, DAP and pH in the EMG in comparison to the NAG. The f was lower from T30 to T75 in the GEM, while the PaCO2 was higher between T15 and T90 in this group. No significant alterations occurred to the BIS values. The time necessary for extubation, sternal recumbency, ambulation and total recovery were 40.6±30.7, 91±37.5, 134.5±54.5, and 169.1±55.4 minutes in the NAG and 68.8±37.3, 133.3±85.3, 171.3±77.1, and 233.1±60.6 minutes in EMG. The presence of undesired effects directly related to the use of propofol was not observed for any of the formulations. There were no clinically important changes regarding the hematological parameters and the renal function, however, values higher than the physiological ones were noted for the enzyme alanine aminotransferase (ALT) from 12 to 72 hours in the EMG and from 48 to 72 hours in the NAG. We conclude that the propofol in nanoemulsion presents clinical and biochemical characteristics similar to the lipid emulsion commercially available. However, the nanoemulsion formulation provides greater cardiovascular and respiratory stability for both induction and continuous rate infusion in healthy cats / Sabe-se que diferentes formulações de propofol apresentam padrão farmacocinético e farmacodinâmico diferenciados. Desta forma, objetivou-se avaliar os efeitos cardiovascular, respiratório, hemogasométrico e metabólico, bem como os possíveis efeitos colaterais da nova formulação de propofol em nanoemulsão, em comparação ao propofol em emulsão lipídica comercialmente disponível. Para tal, foram utilizadas 12 gatas, hígidas com peso médio de 2,6±0,4kg, as quais foram alocadas em dois grupos: Grupo Nanoemulsão (GNA, n=6), os quais receberam como agente indutor propofol em nanoemulsão e Grupo Emulsão (GEM, n=6) que receberam propofol em emulsão lipídica, ambos em dose suficiente para intubação. Imediatamente após, os animais foram intubados e suplementados com oxigênio 100%, por meio de sistema sem reinalação de gases. Em ato contínuo, iniciou-se a infusão de propofol com a respectiva formulação na taxa de 0,3mg/kg/min., durante 90 minutos. Os parâmetros foram avaliados em: T-10, T0, T15, T30, T45, T60, T75, T90, correspondentes a: basal, 0, 15, 30, 45, 60, 75 e 90 minutos após início da infusão, respectivamente. Para análise estatística foi utilizado a Análise de Variância (ANOVA) com repetições múltiplas entre os tempos do mesmo grupo e para comparação entre grupos, o teste t de Student (P<0,05). A dose necessária para indução foi de 9,5±1,3mg/kg e 10±1mg/kg para GNA e GEM, respectivamente. No GEM, houve redução, em todos os momentos, quando comparados ao basal, da freqüência cardíaca, freqüência respiratória (f), pH e das pressões arteriais sistólica (PAS), diastólica (PAD) e média (PAM). A f do GNA reduziu apenas em T90 e o pH em T0, T15 e T90. A PaCO2, PaO2, e SaO2 aumentaram em ambos os grupos, em todos os momentos quando comparados ao basal. O HCO3 aumentou em todos os momentos do GEM e em T15, T30, T60 e T90 no GNA. A PVC foi menor apenas em T60 e T90 em relação ao basal, no GNA. Entre grupos, constatou-se redução, em todos os momentos, para os valores de PAS, PAM, PAD e pH no GEM em relação ao GNA, a f do GEM foi menor de T30 até T75 em relação ao GNA. A PaCO2 do GEM foi maior de T15 até T90. Não foi observada diferença significativa entre os valores do índice biespectral (BIS). Os tempos de extubação, decúbito esternal, deambulação e de recuperação total foram de 40,6±30,7; 91±37,5; 134,5±54,5; e 169,1±55,4 minutos no GNA e de 68,8±37,3; 133,3±85,3; 171,3±77,1; e 233,1±60,6 minutos no GEM, respectivamente. Não foi constatada a presença de efeitos colaterais diretamente relacionados ao uso do propofol em ambas as formulações. Não foram observadas alterações de importância clínica referentes aos parâmetros hematológicos e função renal. Observou-se aumento, acima do limite fisiológico, da enzima Alanina Aminotransferase (ALT) observados de 12 às 72h no GEM e de 48 às 72h no GNA. Conclui-se que o propofol em nanoemulsão apresenta características clínicas e bioquímicas semelhantes à formulação em emulsão lipídica comercialmente disponível. A formulação em nanoemulsão proporciona maior estabilidade cardiovascular e respiratória para indução e infusão contínua em gatas hígidas

Emulsões artificiais sem proteína simulando quilomicrons e remanescentes de quilomícron foram preparadas por sonicalcação de trioleína, lecitina, colesteril oleato e colesterol em solução aquosa. A seguir foram ultracentrifugadas em gradiente descontínuo de densidade. As emulsões, marcadas com 3H-trioleína e 14C-colesteril oleato foram injetadas via intra-arterial em ratos controle e em ratos tratados com Triton WR1339, sulfato de protamina e heparina, medindo-se a seguir a remoção plasmática do colesteril-ester e dos triglicérides, durante dez minutos. O Triton WR 1339 e a protamina inibiram a lipólise dos quilomícrons artificiais, diminuindo a remoção destas partículas do plasma. O Triton WR 1339 mostrou ser mais efetivo que a protamina nestes efeitos. Por outro lado, a heparina promoveu uma lipólise rápida e brusca nos quilomícrons artificiais, assim como uma aceleração na remoção destas partículas do plasma. Em contraste flagrante com esses resultados, o metabolismo dos remanescentes de quilomícron não foi consideravelmente afetado pelo tratamento com Triton e heparina. Estas experiências indicam que as emulsões artificiais reproduzem o comportamento metabólico dos quilomícrons e remanescentes de quilomícron naturais, em condições em que a atividade da lipase lipoproteica esteja alterada. / Protein-free emulsions models of chylomicrons and chylomicron remnants were prepared by sonicating triolein, lecithin., cholesteryl oleate and cholesterol in aqueous saline media, followed by ultracentrifugation in density gradient solution. The 3H-triolein and 14C-cholesteryl oleate labeled emulsions were injected into the carotid artery of control rats and rats treated with Triton WR 1339, protamine sulphate and heparin. Plasma removal of both labels was measured during ten minutes in two minutes intervals. Triton WR 1339 and protamine sulphate strongly inhibited lipolysis of chylomicron-like emulsions leading to delayed removal of the particles from blood. Triton WR 1339 de appeared to be more effective than protamine to elicit these effects. On the other hand, heparin produced instantaneous lipolysis of the chylomicron-like particles markedly enhancing its removal from plasma. Contrarily, chylomicron remnant-like emulsions were not considerably affected either by Triton WR 1339 or by heparin treatment. The above described results obtained with artificial emulsions support current concepts on the metabolic behavior of natural chylomicron and remnant submitted to changes in lipoprotein lipase action.

<p>Of the three components of parenteral macronutrient classes (protein, carbohydrate and lipid), the lipid class is the least understood and the fatty acid distribution in lipid emulsion products has the potential to play a critical role in the development of infant morbidities. Breast milk has long been considered the gold standard for infant nutrition, but when infants cannot tolerate enteral feeds, the use of lipid emulsions cannot be avoided despite their adverse side effects.</p> <p>In this study, fatty acid profiles from five commercially available lipid emulsion products were compared to the profile of breast milk. As well, resulting serum on each of these nutrition sources were compared to the profile of either lipid emulsion (Intralipid) or breast milk. Fatty acid profiles for matched pairs of breast milk from mother and resulting serum from infants were compared as well as the profile of normotriglyceridemic serum samples to hypertriglyceridemic ones. Results indicate that not one lipid emulsion product is like another, nor do their profiles closely resemble breast milk even though they are intended to replace the fat portion of the infant’s natural source of nutrition. Serum was not found to directly reflect the fatty acid profile of the nutrition source administered, as was expected based on literature, highlighting that there is a complex web of pathways between nutrition administration and appearance of fatty acids in the serum. Further research is necessary to define the effect of fatty acid chain length and degree of saturation on these metabolic pathways, as the very essence of interrelations such as these can complicate interpretations of results.</p> / Master of Science in Medical Sciences (MSMS)

Orientador: Carlos Augusto Araujo Valadão / Banca: Newton Nunes / Banca: Paulo Sérgio Patto dos Santos / Banca: Aury Nunes de Moraes / Banca: Nilson Oleskovicz / Resumo: A administração intravenosa de anestésicos voláteis halogenados pode ser considerada uma alternativa à aplicação pela via inalatória, e neste contexto, as formulações emulsificadas têm mostrado segurança, eficiência e bons planos anestésicos. Neste estudo, objetivou-se comparar os efeitos hemodinâmicos e respiratórios das administrações intravenosa e inalatória de isofluorano em cães, após a determinação da taxa de infusão intravenosa de 6,99 mL/kg/h deste halogenado em emulsão lipídica. Para tanto, foram utilizados oito cães, os quais formaram três grupos distintos: grupo IV - anestesia intravenosa por infusão de emulsão lipídica com isofluorano; grupo E - anestesia inalatória com isofluorano, associada à infusão intravenosa de emulsão lipídica sem o halogenado; e grupo IN - anestesia inalatória com isofluorano, associada à infusão intravenosa de solução de NaCl 0,9%. Foram evidenciadas diminuições da contratilidade cardíaca, índice cardíaco e pressão arterial sistêmica a partir de 10 e 40 minutos de anestesia, respectivamente, nos grupos IV e E. A hipotensão no grupo IV resultou em acidose metabólica, porém, hipoxemia não foi observada. O grupo IN apresentou manutenção do índice cardíaco, apesar de reduções do índice de resistência vascular sistêmica e pressão arterial. A taxa de infusão intravenosa determinada foi efetiva na manutenção de plano de anestesia cirúrgica em cães. As infusões intravenosas de isofluorano e emulsão lipídica causaram depressão cardíaca e hipotensão. Em relação à função respiratória, nenhum dos grupos mostrou efeitos deletérios. / Abstract: The intravenous administration of halogenated anesthetics can be considered an alternative to the application by the inhalation route, moreover, emulsified formulations have been showing safety, efficiency and appropriated anesthetic depth. The aim of this study was to compare the effects of isoflurane anesthesia by intravenous or inhalational route on hemodynamic and respiratory variables, after the determination of the intravenous infusion rate of 6.99 mL/kg/h of this anesthetic in lipid emulsion, in dogs. Therefore, eight dogs were distributed in three different protocols: IV group - intravenous anesthesia with emulsified isoflurane; E group - inhalational anesthesia with isoflurane associated to intravenous infusion of lipid emulsion; and IN group - inhalational anesthesia with isoflurane associated to intravenous infusion of 0.9% NaCl solution. The results regarding IV and EM groups revealed decreases of heart contractility, cardiac index and systemic blood pressure starting from 10 and 40 minutes of anesthesia, respectively. The hypotension in the IV group resulted in metabolic acidosis, however, hypoxemia was not observed. The IN group demonstrated maintenance of cardiac index, despite of reduction of the blood pressure and systemic vascular resistance index. The intravenous infusion rate determined was effective in the maintenance of an anesthetic depth of general anesthesia in dogs. The intravenous infusions of isoflurane and lipid emulsion caused cardiac depression and hypotension. In relation to respiratory function, the inhalational route and the intravenous infusions of isoflurane and lipid emulsion did not result in harmful effects. / Doutor

L'ischémie myocardique est caractérisée par la survenue de désordres ioniques et métaboliques responsables de la perte de l'intégrité structurale et fonctionnelle cellulaire, notamment au niveau des mitochondries et, en conséquence, de l'altération de l'activité électromécanique cardiaque. Dans le cadre de cette thèse, nous nous sommes intéressés : 1) à l'étude des lésions d'ischémie-reperfusion (I/R) et des moyens pharmacologiques de cardioprotection avec amlodipine (antagoniste calcique), perindorpilate (Inhibiteur de l'enzyme de conversion). Après avoir mis en évidence les différentes altérations électrophyiologiques, hémodynamiques, structurales et fonctionnelles mitochondriales induite par un épisode d'I/R, les molécules précédemment citées sont administrées seules ou en association, en bolus IV, avant l'induction de l'ischémie myocardique par ligature de l'artère interventriculaire antérieure dans sa partie distale et à une minute seulement après la reperfusion (Etude I, n=36 porcelets domestiques) ; 2) à la cardiotoxicité aigue d'un anesthésique local en l'occurrence la lévobubivacaine à la suite d'une injection intravasculaire (IV). Cette toxicité a été évaluée dans deux situations distinctes : circulation coronaire préservée ou ischémie myocardique. Ces deux situations permettent de « mimer » respectivement une intoxication à la levobupivacaïne chez un sujet sain ou coronarien ; 3) à l'effet bénéfique éventuel des émulsions lipidiques (Intralipid®) dans les mêmes conditions expérimentales. Les émulsions lipidiques ont été administrées quelques minutes après l'injection (IV) de la levobupivacaïne (Etude II, n=48 porcelets domestiques). Ces études ont été menées, in vivo, sur des porcs anesthésiés et ventilés. Les paramètres électrophysiologiques et hémodynamiques ont été déterminés tout au long de l'expérimentation à des intervalles réguliers. A la fin des expériences, les animaux ont été euthanasiés et des fragments de ventricule gauche ont été prélevés pour l'évaluation structurale cellulaire et fonctionnelle mitochondriale / Myocardial ischemia is characterized by the development of ionic and metabolic disorders that result in the loss of the structural and functional cellular integrity, especially within mitochondria and, consequently, in alterations of cardiac electromechanical activity. In the context of this thesis, the following aspects have been investigated: (1) ischemia-reperfusion (I/R) lesions and pharmacological measures of cardioprotection involving the calcium antagonist amlodipine, and the converting enzyme inhibitor perindorpilate. After describing the various electrophysiological, hemodynamic, and mitochondrial (both structural and functional) alterations induced by I/R, amlodipine and perindorpilate were administered either alone or combined, via a bolus IV injection, prior to a distal ligation of the anterior interventricular artery, and then one minute after reperfusion (Study II; n = 36 domestic piglets); (2) the acute cardiotoxicity of the local anesthetic levobupivacaine following an IV injection. Cardiotoxic effects were evaluated in two distinctive situations: preserved coronary circulation and experimental myocardial ischemia. Using both situations allows for mimicking levobupivacaine overdose in a healthy patient or a patient with coronary disease, respectively; (3) the possibly beneficial effect of lipid emulsions (Intralipid®) in both experimental conditions. Lipid emulsions were administered a few minutes following the IV injection of levobupivacaine (Study II, n=48 domestic piglets). These investigations were conducted in vivo on piglets anesthetized and ventilated. Electrophysiological and hemodynamic parameters were measured at given intervals throughout the study. At the end of the study, the animals were sacrificed and tissue samples of the left ventricles were withdrawn to measure the structure and function of mitochondria

Made available in DSpace on 2014-06-11T19:31:08Z (GMT). No. of bitstreams: 0 Previous issue date: 2008-11-28Bitstream added on 2014-06-13T20:41:23Z : No. of bitstreams: 1 almeida_rm_dr_jabo.pdf: 437070 bytes, checksum: e5cd6f576c7b9ff4974078130d3e9da5 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / A administração intravenosa de anestésicos voláteis halogenados pode ser considerada uma alternativa à aplicação pela via inalatória, e neste contexto, as formulações emulsificadas têm mostrado segurança, eficiência e bons planos anestésicos. Neste estudo, objetivou-se comparar os efeitos hemodinâmicos e respiratórios das administrações intravenosa e inalatória de isofluorano em cães, após a determinação da taxa de infusão intravenosa de 6,99 mL/kg/h deste halogenado em emulsão lipídica. Para tanto, foram utilizados oito cães, os quais formaram três grupos distintos: grupo IV – anestesia intravenosa por infusão de emulsão lipídica com isofluorano; grupo E - anestesia inalatória com isofluorano, associada à infusão intravenosa de emulsão lipídica sem o halogenado; e grupo IN - anestesia inalatória com isofluorano, associada à infusão intravenosa de solução de NaCl 0,9%. Foram evidenciadas diminuições da contratilidade cardíaca, índice cardíaco e pressão arterial sistêmica a partir de 10 e 40 minutos de anestesia, respectivamente, nos grupos IV e E. A hipotensão no grupo IV resultou em acidose metabólica, porém, hipoxemia não foi observada. O grupo IN apresentou manutenção do índice cardíaco, apesar de reduções do índice de resistência vascular sistêmica e pressão arterial. A taxa de infusão intravenosa determinada foi efetiva na manutenção de plano de anestesia cirúrgica em cães. As infusões intravenosas de isofluorano e emulsão lipídica causaram depressão cardíaca e hipotensão. Em relação à função respiratória, nenhum dos grupos mostrou efeitos deletérios. / The intravenous administration of halogenated anesthetics can be considered an alternative to the application by the inhalation route, moreover, emulsified formulations have been showing safety, efficiency and appropriated anesthetic depth. The aim of this study was to compare the effects of isoflurane anesthesia by intravenous or inhalational route on hemodynamic and respiratory variables, after the determination of the intravenous infusion rate of 6.99 mL/kg/h of this anesthetic in lipid emulsion, in dogs. Therefore, eight dogs were distributed in three different protocols: IV group – intravenous anesthesia with emulsified isoflurane; E group – inhalational anesthesia with isoflurane associated to intravenous infusion of lipid emulsion; and IN group - inhalational anesthesia with isoflurane associated to intravenous infusion of 0.9% NaCl solution. The results regarding IV and EM groups revealed decreases of heart contractility, cardiac index and systemic blood pressure starting from 10 and 40 minutes of anesthesia, respectively. The hypotension in the IV group resulted in metabolic acidosis, however, hypoxemia was not observed. The IN group demonstrated maintenance of cardiac index, despite of reduction of the blood pressure and systemic vascular resistance index. The intravenous infusion rate determined was effective in the maintenance of an anesthetic depth of general anesthesia in dogs. The intravenous infusions of isoflurane and lipid emulsion caused cardiac depression and hypotension. In relation to respiratory function, the inhalational route and the intravenous infusions of isoflurane and lipid emulsion did not result in harmful effects.

As infecções fúngicas invasivas (IFIs) são um problema de saúde pública e representam uma importante causa de morbidade e mortalidade. A Anfotericina B (AnfB) é a droga de escolha no tratamento das IFIs. Apesar da sua eficácia, a AnfB está associada com efeitos adversos como a nefrotoxicidade. Com o número elevado de pacientes suscetíveis às IFIs, estudos foram desenvolvidos e demonstraram que a nefrotoxicidade pode ser prevenida através do uso de AnfBs modificadas. Dessas modificações, a AnfB incorporada à uma emulsão lipídica (AnfB/EL) apresenta baixo custo e similar eficácia. Existe uma alta prevalência de deficiência de vitamina D (dVD) na população mundial. Sendo assim, a hipovitaminose D pode acelerar a progressão da doença renal e refletir em mau prognóstico em casos de nefrotoxicidade. Esse trabalho visa analisar se a deficiência da vitamina D pode induzir a nefrotoxicidade da AnfB/EL. Ratos Wistar foram divididos em quatro grupos: controle, animais que receberam dieta padrão por 34 dias; AnfB/EL, animais que receberam dieta padrão por 34 dias e injeção intraperitoneal de AnfB/EL (5mg/kg/dia) nos últimos 4 dias; dVD, animais que receberam dieta depletada em vitamina D por 34 dias; e dVD+AnfB/EL, animais que receberam dieta depletada em vitamina D por 34 dias e a injeção intraperitoneal de AnfB/EL (5mg/kg/dia) nos últimos 4 dias. Amostras de sangue, urina e tecido renal foram coletadas para a análise dos efeitos da droga sobre a função, hemodinâmica e morfologia renal. A associação de AnfB/EL com a hipovitaminose D levou a injúria renal, lesão tubular discreta, hiperfosfatúria, hipermagnesiúria, hipertensão e comprometimento do mecanismo de concentração urinária. Dessa forma, é essencial monitorar os níveis de vitamina D em pacientes tratados com AnfB/EL, uma vez que a deficiência dessa vitamina é um fator indutor de nefrotoxicidade associada ao uso da AnfB/EL / Invasive fungal infections (IFI) have become a worldwide serious health problem and represent a significant cause of morbidity and mortality. Amphotericin B (AmB) is the drug of choice for the treatment of IFI. Despite its efficacy, use of AmB has been associated with adverse reactions such as nephrotoxicity The increasing number of high-risk patients, who are more susceptible to IFI and are more likely to use AmB, has enabled the development of modified AmBs in order to reduce nephrotoxicity. Among these formulations, an extemporaneous lipid emulsion preparation of AmB (AmB/LE) is a lower cost alternative with similar benefits. Moreover, studies have shown a high prevalence of vitamin D deficiency (VDD) in immunocompromised individuals and patients hospitalized in intensive care units. Thus, vitamin D deficiency or insufficiency may hasten the progression of kidney disease and reflect on a worse prognosis in cases of acute kidney injury and drug-induced nephrotoxicity. In view of the high worldwide incidence of hypovitaminosis D, the aim of this study was to investigate whether vitamin D deficiency may induce AmB/LE-related nephrotoxicity. Wistar rats were divided into four groups: control, received a standard diet for 34 days; AmB/LE, received a standard diet for 34 days and AmB/LE (5mg/kg/day) intraperitoneally in the last 4 days; VDD, received a vitamin D-free diet for 34 days; and VDD+AmB/LE, received a vitamin D-free diet for 34 days and AmB/LE (5mg/kg/day) intraperitoneally in the last 4 days. Blood, urine and renal tissue samples were collected in order to evaluate the effects of the drug on renal function, hemodynamic and morphology. Association of AmB/LE and vitamin D deficiency led to impaired renal function, mild tubular injury, hypertension and urinary concentration defect. Hence, it is important to monitor vitamin D levels in AmB/LE treated patients, since vitamin D deficiency induces AmB/LE nephrotoxicity

La préparation de formulations à libération contrôlée est le domaine des sciences pharmaceutiques qui vise à modifier l’environnement immédiat des principes actifs pour en améliorer l’efficacité et l’innocuité. Cet objectif peut être atteint en modifiant la cinétique de circulation dans le sang ou la distribution dans l’organisme. Le but de ce projet de recherche était d’étudier le profil pharmacocinétique (PK) de différentes formulations liposomales. L’analyse PK, généralement employée pour représenter et prédire les concentrations plasmatiques des médicaments et de leurs métabolites, a été utilisée ici pour caractériser in vivo des formulations sensibles au pH servant à modifier la distribution intracellulaire de principes actifs ainsi que des liposomes destinés au traitement des intoxications médicamenteuses. Dans un premier temps, la PK d’un copolymère sensible au pH, à base de N-isopropylacrylamide (NIPAM) et d’acide méthacrylique (MAA) a été étudiée. Ce dernier, le p(NIPAM-co-MAA) est utilisé dans notre laboratoire pour la fabrication de liposomes sensibles au pH. L’étude de PK conduite sur les profils de concentrations sanguines de différents polymères a défini les caractéristiques influençant la circulation des macromolécules dans l’organisme. La taille des molécules, leur point de trouble ainsi que la présence d’un segment hydrophobe à l’extrémité des chaînes se sont avérés déterminants. Le seuil de filtration glomérulaire du polymère a été évalué à 32 000 g/mol. Finalement, l’analyse PK a permis de s’assurer que les complexes formés par la fixation du polymère à la surface des liposomes restaient stables dans le sang, après injection par voie intraveineuse. Ces données ont établi qu’il était possible de synthétiser un polymère pouvant être adéquatement éliminé par filtration rénale et que les liposomes sensibles au pH préparés avec celui-ci demeuraient intacts dans l’organisme. En second lieu, l’analyse PK a été utilisée dans le développement de liposomes possédant un gradient de pH transmembranaire pour le traitement des intoxications médicamenteuses. Une formulation a été développée et optimisée in vitro pour capturer un médicament modèle, le diltiazem (DTZ). La formulation liposomale s’est avérée 40 fois plus performante que les émulsions lipidiques utilisées en clinique. L’analyse PK des liposomes a permis de confirmer la stabilité de la formulation in vivo et d’analyser l’influence des liposomes sur la circulation plasmatique du DTZ et de son principal métabolite, le desacétyldiltiazem (DAD). Il a été démontré que les liposomes étaient capables de capturer et de séquestrer le principe actif dans la circulation sanguine lorsque celui-ci était administré, par la voie intraveineuse. L’injection des liposomes 2 minutes avant l’administration du DTZ augmentait significativement l’aire sous la courbe du DTZ et du DAD tout en diminuant leur clairance plasmatique et leur volume de distribution. L’effet de ces modifications PK sur l’activité pharmacologique du médicament a ensuite été évalué. Les liposomes ont diminué l’effet hypotenseur du principe actif administré en bolus ou en perfusion sur une période d’une heure. Au cours de ces travaux, l’analyse PK a servi à établir la preuve de concept que des liposomes possédant un gradient de pH transmembranaire pouvaient modifier la PK d’un médicament cardiovasculaire et en diminuer l’activité pharmacologique. Ces résultats serviront de base pour le développement de la formulation destinée au traitement des intoxications médicamenteuses. Ce travail souligne la pertinence d’utiliser l’analyse PK dans la mise au point de vecteurs pharmaceutiques destinés à des applications variées. À ce stade de développement, l’aspect prédictif de l’analyse n’a pas été exploité, mais le côté descriptif a permis de comparer adéquatement diverses formulations et de tirer des conclusions pertinentes quant à leur devenir dans l’organisme. / Drug delivery is the field of pharmaceutical sciences which focuses on altering the immediate environment of drug molecules to improve their efficacy and safety. Drug delivery systems can potentiate the effect of active principles or alleviate their side effects by modifying their circulation profiles and/or biodistribution. The objective of this research project was to investigate the role of pharmacokinetic (PK) analysis in the development of novel drug delivery systems. PK analysis is generally applied to describe and predict the blood concentration profiles of low molecular weight drugs and their metabolites. Nevertheless, it is herein used to characterize the circulation of 2 liposomal formulations: pH-sensitive liposomes designed to alter the intracellular distribution of drugs and liposomes with transmembrane pH gradient for drug detoxification. The first series of experiments were designed to study the circulation kinetics of a pH-sensitive polymer prepared with N-isopropylacrylamide (NIPAM) and methacrylic acid (MAA). The copolymer p(NIPAM-co-MAA) is used in our laboratory to prepare serum-stable, PEGylated, pH-sensitive liposomes. The circulation profiles of polymers with different characteristics were characterized. The parameters which impacted the fate of the macromolecules were the length of the polymer chain, its cloud point and the presence of a hydrophobic anchor at one extremity of the molecule. The glomerular filtration cut-off of the polymer was determined to be around 32,000 g/mol. PK analysis allowed to conclude that the complexes prepared by anchoring the polymer on the surface of the liposomes remained stable in the bloodstream. This data established that pH-sensitive vesicular formulations could be produced using a polymer which could be excreted through renal filtration. It also confirmed that the formulation remained intact in the bloodstream. The second part of this work involved the development of liposomes with a transmembrane pH gradient designed to treat cardiovascular drug intoxications. Liposomes were designed and optimized in vitro to capture a model cardiovascular drug, diltiazem (DTZ). In vitro, the liposome uptake capacity was 40-fold higher than the lipid emulsion used in the clinic. PK analysis was used to verify the stability of the formulation in vivo, and to assess the impact of the liposomes on the plasma concentration of DTZ and its principal active metabolite, deacetyl-diltiazem (DAD). It was shown that the vesicles were able to capture and sequester DTZ and DAD. Injection of liposomes 2 min prior to administration of DTZ significantly increased the area under the plasma-concentration vs. time curve of both DTZ and DAD, while lowering their clearance and volume of distribution. The impact of the changes in PK on the pharmacological effect of the drug was also investigated. Liposomes tempered the hypotensive effect of the drug when the latter was administered via an intravenous bolus or a 1-h perfusion. Throughout this work, PK analysis proved to be an efficient tool to study the ability of transmembrane pH gradient liposomes to alter the blood circulation profiles of a cardiovascular drug, and to reduce its pharmacological effect. This proof of concept establishes firm ground for the further development of this colloidal formulation to treat drug intoxications. This work pointed out the relevance of PK analysis for the development of multi-purpose, colloidal drug delivery systems. At this stage, the predictive nature of the analysis was not exploited, but its descriptive properties allowed objective comparison of the circulation profiles of distinct systems and pertinent conclusions concerning their fate in vivo.