Nanotechnology for the treatment of deep endometriosis: uptake of lipid core nanoparticles by LDL receptors in endometriotic foci

OBJECTIVE: Rapidly dividing cells in multiple types of cancer and inflammatory diseases undergo high low density lipoprotein (LDL) uptake for membrane synthesis, and coupling an LDL-like nanoemulsion, containing lipid nanoparticles (LDE) to a chemotherapeutic agent efficiently targets these cells without significant systemic effects. This was a prospective exploratory study that evaluated the uptake of a radioactively labeled LDE emulsion by receptors of endometriotic foci and the capacity of the LDE for cellular internalization. METHODS: The lipid profile of each patient was determined before surgery, and labeled LDE were injected into fourteen patients with intestinal or nonintestinal endometriosis. The radioactivity of each tissue sample (intestinal endometriosis, nonintestinal endometriosis, healthy peritoneum, or topical endometrium) was measured. RESULTS: The group with intestinal endometriosis presented higher levels of plasma LDL but lower LDE uptake by foci than the nonintestinal group, suggesting less cell division and more fibrosis. The uptake of LDE was highest in the topical endometrium, followed by the healthy peritoneum, and lowest in the endometriotic lesion. Since the endometriotic foci showed significant LDE uptake, there was likely increased consumption of LDL by these cells, similar to cells in cancers and inflammatory diseases. Plasma cholesterol levels had no influence on LDE uptake, which showed that the direct delivery of the nanoemulsion to target tissues was independent of serum lipoproteins. There were no significant differences in the parameters (p>0.01) because of the small sample size, but the findings were similar to those of previous studies. CONCLUSION: Nanotechnology is a promising therapeutic option for surgery and hormonal blockage for deep endometriosis, with a lower complication rate and no systemic side effects.

Treatment of patients with aortic atherosclerotic disease with paclitaxel-associated lipid nanoparticles

OBJECTIVE: The toxicity of anti-cancer chemotherapeutic agents can be reduced by associating these compounds, such as the anti-proliferative agent paclitaxel, with a cholesterol-rich nanoemulsion (LDE) that mimics the lipid composition of low-density lipoprotein (LDL). When injected into circulation, the LDE concentrates the carried drugs in neoplastic tissues and atherosclerotic lesions. In rabbits, atherosclerotic lesion size was reduced by 65% following LDE-paclitaxel treatment. The current study aimed to test the effectiveness of LDE-paclitaxel on inpatients with aortic atherosclerosis. METHODS: This study tested a 175 mg/m2 body surface area dose of LDE-paclitaxel (intravenous administration, 3/3 weeks for 6 cycles) in patients with aortic atherosclerosis who were aged between 69 and 86 yrs. A control group of 9 untreated patients with aortic atherosclerosis (72-83 yrs) was also observed. RESULTS: The LDE-paclitaxel treatment elicited no important clinical or laboratory toxicities. Images were acquired via multiple detector computer tomography angiography (64-slice scanner) before treatment and at 1-2 months after treatment. The images showed that the mean plaque volume in the aortic artery wall was reduced in 4 of the 8 patients, while in 3 patients it remained unchanged and in one patient it increased. In the control group, images were acquired twice with an interval of 6-8 months. None of the patients in this group exhibited a reduction in plaque volume; in contrast, the plaque volume increased in three patients and remained stable in four patients. During the study period, one death unrelated to the treatment occurred in the LDE-paclitaxel group and one death occurred in the control group. CONCLUSION: Treatment with LDE-paclitaxel was tolerated by patients with cardiovascular disease and showed the potential to reduce atherosclerotic lesion size.

Plasma kinetics of an LDL-like nanoemulsion and lipid transfer to HDL in subjects with glucose intolerance

OBJECTIVE: Glucose intolerance is frequently associated with an altered plasma lipid profile and increased cardiovascular disease risk. Nonetheless, lipid metabolism is scarcely studied in normolipidemic glucose-intolerant patients. The aim of this study was to investigate whether important lipid metabolic parameters, such as the kinetics of LDL free and esterified cholesterol and the transfer of lipids to HDL, are altered in glucose-intolerant patients with normal plasma lipids. METHODS: Fourteen glucose-intolerant patients and 15 control patients were studied; none of the patients had cardiovascular disease manifestations, and they were paired for age, sex, race and co-morbidities. A nanoemulsion resembling a LDL lipid composition (LDE) labeled with 14C-cholesteryl ester and ³H-free cholesterol was intravenously injected, and blood samples were collected over a 24-h period to determine the fractional clearance rate of the labels by compartmental analysis. The transfer of free and esterified cholesterol, triglycerides and phospholipids from the LDE to HDL was measured by the incubation of the LDE with plasma and radioactivity counting of the supernatant after chemical precipitation of non-HDL fractions. RESULTS: The levels of LDL, non-HDL and HDL cholesterol, triglycerides, apo A1 and apo B were equal in both groups. The 14C-esterified cholesterol fractional clearance rate was not different between glucose-intolerant and control patients, but the ³H-free-cholesterol fractional clearance rate was greater in glucose-intolerant patients than in control patients. The lipid transfer to HDL was equal in both groups. CONCLUSION: In these glucose-intolerant patients with normal plasma lipids, a faster removal of LDE free cholesterol was the only lipid metabolic alteration detected in our study. This finding suggests that the dissociation of free cholesterol from lipoprotein particles occurs in normolipidemic glucose intolerance and may participate in atherogenic signaling.

Simultaneous transfer of cholesterol, triglycerides, and phospholipids to high-density lipoprotein in aging subjects with or without coronary artery disease

OBJECTIVE: To verify whether the capacity of high-density lipoprotein (HDL) to simultaneously receive nonesterified cholesterol, triglycerides, cholesteryl esters, and phospholipids changes with aging and the presence of coronary artery disease. DESIGN: Cross-sectional study with biochemical analyses. SUBJECTS: Eleven elderly patients with coronary artery disease (74±5 years) were compared with the following groups of non-coronary artery disease subjects (referred to as "healthy"): 25 young (25±5 years), 25 middle-aged (42± years), and 25 elderly subjects (75±8 years). METHODS: Plasma samples were incubated with a nanoemulsion labeled with radioactive lipids; the transfer of the lipids from the nanoemulsion to the HDL was measured in chemically precipitated HDL. HDL size and paraoxonase-1 activity were also determined. RESULTS: The transfer of cholesteryl esters and phospholipids to high-density lipoprotein was significantly greater (p<0.001) in healthy elderly subjects than in the middle-aged and younger subjects. Non-esterified cholesterol and triglyceride transfer was not different among these three groups. The HDL size was significantly greater (p<0.001) in healthy elderly subjects than in the middle-aged and younger subjects. The paraoxonase-1 activity was similar among the groups. Compared with healthy elderly subjects, coronary artery disease elderly subjects had significantly less (p<0.05) transfer of non-esterified cholesterol, triglycerides, and cholesteryl esters to the HDL and a significantly smaller (p<0.05) HDL size. CONCLUSION: Because lipid transfer is enhanced in healthy elderly subjects but not in those with coronary artery disease, increasing lipid transfer to HDL may be a protective mechanism against the disease.

Synthetic nanoemulsion resembling a protein-free model of 7-ketocholesterol containing low density lipoprotein: In vitro and in vivo studies

7-ketocholesterol (7-KC) differs from cholesterol by a functional ketone group at C7. It is an oxygenated cholesterol derivative (oxysterol), commonly present in oxidized low-density lipoprotein (LDL). Oxysterols are generated and participate in several physiologic and pathophysiologic processes. For instance, the cytotoxic effects of oxidized LDL have been widely attributed to bioactive compounds like oxysterols. The toxicity is in part due to 7-KC. Here we aimed to demonstrate the possibility of incorporating 7-KC into the synthetic nanoemulsion LDE, which resembles LDL in composition and behavior. This would provide a suitable artificial particle resembling LDL to study 7-KC metabolism. We were able to incorpórate 7-KC in several amounts into LDE. The incorporation was evaluated and confirmed by several methods, including gel filtration chromatography, using radiolabeled lipids. The incorporation did not change the main lipid composition characteristics of the new nanoparticle. Particle sizes were also evaluated and did not differ from LDE. In vivo studies were performed by injecting the nanoemulsion into mice. The plasma kinetics and the targeted organs were the same as described for LDE. Therefore, 7-KC-LDE maintains composition, size and some functional characteristics of LDE and could be used in experiments dealing with 7-ketocholesterol metabolism in lipoproteins.

Polimorfismo S447X da lipase lipoprotéica: influência sobre a incidência de doença arterial coronariana prematura e sobre os lípides plasmáticos / The S447X polymorphism of lipoprotein lipase: effect on the incidence of premature coronary disease and on plasma lipids

OBJETIVO: O objetivo deste estudo foi avaliar o efeito do polimorfismo S447X sobre os lípides plasmáticos em pacientes com doença arterial coronariana (DAC) prematura. MÉTODOS: Os lípides plasmáticos e a genotipagem foram determinados em 2 grupos: 313 pacientes com DAC prematura (<55 anos) e 150 controles sem DAC. RESULTADOS: A freqüência do polimorfismo S447X foi de 18 por cento nos pacientes com DAC e de 23 por cento no grupo controle. O polimorfismo S447X da lipase lipoprotéica está relacionado com diminuição das concentrações plasmática de triglicérides nos pacientes do sexo masculino com DAC, não havendo essa relação no sexo feminino. CONCLUSÃO: A presença do polimorfismo S447X da lípase lipoprotéica não foi associada à incidência de DAC.

OBJECTIVE: The objective of this study was to evaluate the effect of polymorphism S447X on plasma lipids of patients with premature coronary artery disease (CAD). METHODS: Plasma lipids and genotypes were determined in 2 groups: 313 patients with premature CAD (<55 years of age) and 150 controls without CAD. RESULTS: Frequency of the S447X polymorphism was 18 percent in patients with CAD and 23 percent in the control group. The S447X polymorphism of lipoprotein lipase is related to a decrease in plasma triglyceride concentrations in male patients with CAD, but this correlation is not observed in female patients. CONCLUSION: The presence of the S447X lipoprotein lipase polymorphism was not associated with the incidence of CAD.

Concentração sérica de lípides e apolipoproteína B em recém-nascidos / Concentration of serum lipids and apolipoprotein B in newborns

OBJETIVO: Determinar numa população de recém-nascidos de uma cidade do sul do Brasil as concentrações de lípides séricos e apolipoproteína B (apo-B) em neonatos normais a termo e pré-termo, analisando-se a influência da idade gestacional e do peso ao nascimento. MÉTODOS: Foram estudados 212 neonatos de ambos os sexos, deles, 142 a termo (>37 semanas de gestação) e 70 neonatos pré-termo (<37 semanas). Os neonatos a termo e pré-termo foram classificados pelo peso de nascimento em pequeno para a idade gestacional ou adequado para a idade gestacional. Utilizou-se o sangue do cordão umbilical para as análises bioquímicas. RESULTADOS: Os valores de colesterol total, LDL-C, HDL-C e de apo-B foram maiores nos neonatos pré-termo (79±34, 26±6, 45±15 e 36±14 mg/dL, respectivamente) do que nos neonatos a termo (58±19, 20±10, 31±14 e 28±7 mg/dL, respectivamente; p < 0,0001). Inversamente, os valores de triglicérides foram menores nos neonatos pré-termo (36±14 mg/dL) do que nos neonatos a termo (43±25 mg/dL; p < 0,0018). O sexo e o fato de o neonato ser pequeno ou adequado para a idade gestacional não influenciaram os valores de colesterol total e frações, de triglicérides e apo-B. CONCLUSÃO: As concentrações de lípides plasmáticos e de apo-B dos neonatos da população estudada são semelhantes às dos neonatos de outros países de continentes diversos reportados na literatura e, como esperado, é acentuadamente menor do que os valores referidos na literatura para as de crianças acima de dois anos. A maturidade fetal influencia a concentração de lípides dos neonatos, mas o peso de nascimento não tem efeito nesses parâmetros.

Influence of a cholesterol-rich emulsion and BCNU on the apoptotic indices of myeloma cells induced in BALB/C mice

Proliferating malignant cells express low-density lipoprotein (LDL) receptors, and a cholesterol-rich microemulsion (LDE) resembling the lipid portion of LDL can be bound to lipophilic drugs such as 1,3-bis (2-chloroethyl)-1-nitrosourea (BCNU) to increase the drug´s capture and decrease the drug-associated toxicity. In this work, we studied the effect of BCNU and BCNU+LDE on apoptosis in plasmacytoma-related cells obtained from BALB/c mice with myeloma. Four groups of mice (n=4 each) were treated with LDE, BCNU, BCNU in LDE or vehicle solution (control group). Morphological methods (histology and transmission electron microscopy) and immunohistochemistry (TUNEL procedure) were used to evaluate the occurrence of apoptosis. The drugs were injected intraperitoneally in the 14th month after induction of the myeloma and the mice in all groups were sacrificed six hours after this injection. The apoptotic indices of the plasmacytoma mesenteric cells evaluated in the four experimental groups revealed that the LDE emulsion significantly (p<0.05) increased the percentage of tumoral cells dying by apoptosis. All the groups with LDE, alone or in combination with BCNU showed significantly higher apoptotic indices than the controls. This enhanced cytotoxicity suggests a potential use for LDE in improving the efficacy of chemotherapeutic agents.

Ausência de efeito do captopril no metabolismo de uma emulsão lipídica artificial semelhante aos quilomícrons em pacientes hipertensos e hipercolesterolêmicos / Lack of effect of captopril on the metabolism of an artificial lipid emulsion similar to chylomicrons in hypertensive hypercholesterolemic patients

OBJETIVO: Avaliar o efeito do captopril, sobre o metabolismo dos quilomícrons e de seus remanescentes e as possíveis alterações nas concentrações dos lípides plasmáticos em hipertensos e hipercolesterolêmicos. MÉTODOS: O metabolismo dos quilomícrons foi testado pelo método da emulsão lipídica artificial de quilomícrons marcada com 3H-oleato de colesterol, foi injetada intravenosamente em 10 pacientes com hipertensão arterial leve-moderada antes e após 45 dias de tratamento com captopril (50 mg/dia). Após injeção, foram coletadas amostras de sangue durante 60min em intervalos de tempo pré-estabelecidos para determinar a curva de decaimento e a taxa fracional de remoção (TFR em min-1), bem como o tempo de residência no plasma, da emulsão lipídica artificial, por análise compartimental. As concentrações dos lípides do plasma também foram avaliadas antes e após o tratamento. RESULTADOS: A taxa fracional de remoção (em min-1) da emulsão lipídica antes e após o tratamento com captopril (0,012±0,003 e 0,011±0,003, respectivamente; p=0,85, n.s.) ou o tempo de permanência da emulsão no plasma (83,3±20,8 e 90,9± 22,5 min, n.s.) não se alteraram, mas os níveis de colesterol total e de LDL-c reduziram-se em 7 por cento e 10 por cento respectivamente (p=0,02). As concentrações de HDL-c, triglicérides, Lp(a) e apolipoproteínas AI e B não se modificaram. CONCLUSÃO: O tratamento com captopril, avaliado pelo método da emulsão lipídica artificial, não provoca alterações deletérias no metabolismo dos quilomícrons e seus remanescentes.

Hipertrigliceridemia e dislipidemia pós-prandial / Hyperlipidemia and post-prandial dyslipidemia

As lipoproteínas ricas em triglicerídeos, quilomícrons, VLDL e seus respectivos remanescentes säo as responsáveis pelo transporte das gorduras originadas na dieta e no fígado. Após sua síntese, as lipoproteínas ricas em triglicerídeos säo secretadas no plasma, adquirem apolipoproteínas (apo) como as apo C-II, C-III e apo E e sofrem acäo da lipase lipoprotéica. Após a hidrólise dos triglicerídeos, säo formados os remanescentes, que podem se transformar em outras lipoproteínas, como no caso das VLDL, que originam as IDL e LDL, ou se ligar aos receptores que reconhecem o apo E como o receptor para a LDL e o receptor LRP e serem removidas principalmente pelo fígado. As hipertrigliceridemias diagnosticadas no jejum podem ser de origem endógena (acúmulo de VLDL ou IDL), exógena (acúmulo de quilomícrons) ou podem apresentar acúmulo das VLDL e quilomícrons. Mesmo quando näo há acúmulo dos quilomícrons no período de jejum, a hipertrigliceridemia causada pelo excesso de VLDL prejudica, por competiçäo pelas vias metabólicas comuns, a remoçäo plasmática dos quilomícrons. Essa é a chamada dislipidemia pós-prandial. O acúmulo de quilomícrons e remanescentes parece ser um fator de risco para aterosclerose, independentemente dos níveis de LDL-colesterol. O uso de fibratos, como o genfibrozil, reduz significativamente a lipemia pós-prandial.

Comparação entre homens e mulheres hipercolesterolêmicos de alto risco de desenvolvimento de aterosclerose. Estudo dos fatores de risco e da resposta ao tratamento com pravastatina / Comparison of hypercholesterolemic men and women at high risk for atherosclerosis. Study of risk factors and response to pravastatin treatment

PURPOSE: To compare the prevalence of risk factors, and the response to pravastatin treatment between men and women. METHODS: We evaluated 486 men and 386 women, of these 230 men and 187 women received 10 mg of pravastatin for three months. RESULTS: There were differences between women and men in respectively: arterial hypertension (45.5vs 40.8; p = 0.0012), left ventricular hypertrophy (33.0vs 22.0; p = 0.0041), sedentarism (94.8vs 87.8; p = 0.0005), smoking (43.0vs 61.8; p < 0.0001), Framingham scores (20.0 +/- 7.1 vs 16.0 +/- 7.6 p < 0.0001), HDL-C (43.0 +/- 11.0 vs 38.0 +/- 9.0 mg/dL; p 0.001), triglycerides-TG (216.0 +/- 115.0 vs 271.0 +/- 172.0 mg/dL; p < 0.001) and Castelli's indexes (CI) I and II (7.7 +/- 2.6 vs 8.6 +/- 3.2; p = 0.002 and 5.0 +/- 1.5 vs 5.5 +/- 2.0; p = 0.015). In men under pravastatin treatment there was a greater reduction in TG (32.0 vs 21.0p < 0.05) and CI I (-41.0vs -37.0; p < 0.05) and II (-40.0vs -38.0; p < 0.05). CONCLUSION: Men and women differed in risk factors prevalence and response to treatment with pravastatin.

Lipoprotein(a) and apolipoprotein profile of patients submitted to heart transplantation

Forty-three Caucasian heart transplantation (HT) patients were evaluated in a late evolutional phase (27 mais ou menos 20 months, median 24 months). They were on a stable immunosuppressive triple drug regime and were compared with 45 controls (NL). Lipoprotein(a) [Lp(a)], apolipoprotein (apo) AI, B (n=27) and total cholesterol (TC) and tryglicerides (TG) (n=43) were determined after a 14 hour fast. None of the patients had uremia or nephrotic syndrome, uncontrolled diabetes mellitus or other situation known to increased Lp(a) levels. They were not under hypolipidemic drugs. According to the results of serial coronary angiography, HT patients were further divided in two groups: GCD (n=9) and NGCD (n=19) respectively with or without graft coronary disease. Apo AI and Lp(a) were higher in HT patients than in NL, 1.7 mais ou menos 0,3 vs. 1.1 mais ou menos 0,4 g/l respectively, p<0,0001, and medians (25-75 percentiles) 26 (13.4-60.4) vs. 15 (2.4-33.4) mg/dl, p=0.026.TC, TG and apo B were similar in both groups. There were no differences on the lipid profile of both GCD and NGCD patients. These results suggest that Lp(a) is increased in HT, contrarily to early reports that this lipoprotein may be reduced in HT

Estudo comparativo do gemfibrozil versus pravastatina no tratamento de portadores de doença arterial coronária e níveis reduzidos de HDL-colesterol / Comparative study of gemfibrozil versus pravastatin in the treatment of patients with coronary artery disease and low HDL cholesterol levels

PURPOSE--To evaluate the effects of gemfibrozil and pravastatin in coronary artery disease patients with HDL-cholesterol (HDL-C) < 35 mg/dl). METHODS--Twenty-nine patients (20 males, 60 +/- 9) were divided in a gemfibrozil group (G) (1200 mg/day n = 15) and a pravastatin group (P) (10 or 20 mg n = 10 and 4, respectively). The plasma lipid profile (LP) e.g. total cholesterol (TC), fractions and triglycerides (TG) was determined at 4 and 12 weeks of treatment. RESULTS--HDL-C was not affected in P, TC and LDL-cholesterol (LDL-C) reductions were superior to those in G (31.3 vs 13.4 and 38.7 and 11.5, p < 0.05 and < 0.01 respectively). In G HDL-C raised by 50 (12th week p < 0.01). Gemfibrozil reduced TG levels in 44.7 while in P it varied -32.2 (12th week p < 0.01 and < 0.05 respectively). CONCLUSION--Gemfibrozil is more effective in reducing TG and raising HDL-C than pravastatin. On the other hand, pravastatin was more potent in reducing LDL-C levels.

Objetivo - Comparar os efeitos do gemfibrozil e pravastatina no perfil lipídico (PL) de pacientes coronarianos com HDL-colesterol (HDL-C) <35 mg/dl Métodos - Vinte e nove pacientes (20 homens, 60 9 anos) divididos em grupo gemfibrozil (G) (1200mg/dia n=15) e grupo pravastatina (P) (10mg e 20mg, 10 e 4 pacientes, respectivamente). O perfil lipídico plasmático [colesterol total (CT), frações e triglicérides (TG)] foi avaliado a 4 e 12 semanas de tratamento. Resultados - Em P, o HDL-C não se alterou, as reduções de CT e LDL-colesterol (LDL-C) foram superiores às de G (31,3% vs 13,4% e 38,7 e 11,5%, respectivamente p<0,05 e <0,01). Em G. o HDL-C elevou-se em até 50% (12º semana, p<0,01). O gemfibrozil reduziu os TG em 44,7% (p<0,01) enquanto que em P, a variação foi de -32,2% na 12º semana (p<0,05). Conclusão - O gemfibrozil é mais eficaz que a pravastatina para reduzir os TG e elevar o HDL-C, porém a pravastatina é um redutor mais potente do LDLC

Níveis plasmáticos de lipoproteína (a) em indivíduos normais e portadores de doença coronariana confirmada por cinecoronariografia / Lipoprotein (a) plasma levels in normal subjects and patients with coronary disease carriers subjects confirmed by coronary cineangiography

Avaliar a concentraçäo plasmática da lipoproteína (a) - Lp(a) - em indivíduos com cinecoronariografia normal ou com sinais de aterosclerose. Trinta e um indivíduos com cinecoronariografia normal e 131 com alteraçöes compatíveis com aterosclerose, de ambos os sexos. Foram medidos os níveis plasmáticos de Lp(a) por radioimunoensaio e também os de colesterol, triglicérides, apolipoproteínas A, A1 e B e avaliados fatores de risco como hipertensäo arterial sistêmica, tabagismo, diabetes, além de atividade física. Os indivíuduos com doença coronariana apresentaram Lp(a) plasmática média de 41,9 mg/dl, em comparaçäo com 23,9 mg/dl no grupo normal. O risco de desenvolvimento de doença coronariana entre os com Lp(a) igual ou acima de 25 mg/dl foi de 2,3 vezes, em comparaçäo com os indivíduos com valores abaixo. Houve correlaçäo entre tabagismo e doença coronariana, o que näo foi confirmado estatísticamente no tocante aos outros fatores de risco avaliados. Confirma-se a importância da Lp(a) como fator de risco da doença coronariana