RESUMO
INTRODUCTION: The oral route is widely accepted as the most physiological path for exogenous administration of insulin, as it closely mimic the endogenous insulin pathway. Thus, in this work it is proposed an innovative lipid-polymeric nanocarrier to delivery insulin orally. Areas covered: Nanoparticles were produced through a modified solvent emulsification-evaporation method, using ethyl palmitate and hydroxypropylmethylcellulose acetate succinate as matrix. Lipid-polymeric nanoparticles were around 300 nm in size, negatively charged (-20 mV) and associated insulin with efficiency higher than 80%. Differential scanning calorimetry suggested thermal stability of nanoparticles. In vitro release assays under simulated gastrointestinal conditions resulted in 9% and 14% of insulin released at pH 1.2 during 2 h and at pH 6.8 for 6 h, respectively, demonstrating the ability of those nanoparticles to protect insulin against premature degradation. Importantly, nanoparticles were observed to be safe at potential therapeutic concentrations as did not originate cytotoxicity to intestinal epithelial cells. Lastly, the permeability of nanoencapsulated insulin through Caco-2 monolayers and a triple Caco-2/HT29-MTX/Raji B cell model correlated well with slow release kinetics, and fosters the effectiveness of nanoparticles to promote intestinal absorption of peptidic drugs. Expert opinion: Lipid-polymeric nanoparticles were developed to encapsulate and carry insulin through intestine. Overall, nanoparticles provide insulin stability and intestinal permeability.
Assuntos
Sistemas de Liberação de Medicamentos , Hipoglicemiantes/administração & dosagem , Insulina/administração & dosagem , Metilcelulose/análogos & derivados , Nanopartículas/química , Ácidos Palmíticos/química , Administração Oral , Animais , Células CACO-2/efeitos dos fármacos , Varredura Diferencial de Calorimetria , Cromatografia Líquida de Alta Pressão , Humanos , Concentração de Íons de Hidrogênio , Absorção Intestinal , Metilcelulose/química , Microscopia Eletrônica de Varredura , Permeabilidade , Difração de Raios XRESUMO
Vitamin A deficiency (VAD) remains a public health problem in some regions of Brazil. Increased use of orange-fleshed sweet potato (OFSP) as a source of pro-vitamin A represents a potential strategy for prevention of VAD. We compared the pro-vitamin A content, vitamin A equivalency and bioaccessibility of ß-carotene (ßC) of two varieties of home cooked OFSP and two commercial sources of processed OFSP. Pro-vitamin A carotenoid content in home cooked, Beauregard variety of OFSP exceeded that in Amelia variety and commercial products for babies. All-trans-ßC was the most abundant carotenoid in raw, cooked and commercial OFSP. Boiling and frying OFSP generally decreased total ßC. A serving of 100 g FW Beauregard variety of cooked OFSP contained greater than 100% of the estimated average requirement (EAR) for children and women, and up to 92% EAR for lactating women. Although the efficiency of micellarization of all-trans-ßC during simulated digestion of OFSP was relatively low (4-8%) and significantly less than for cis-isomers, the quantities of trans-ßC incorporated into micelles from boiled Beauregard and fried Amelia varieties exceeded that in micelles generated by digesting commercial OFSP. The bioaccessibility of pro-vitamin A carotenoids in the micelle fraction of digested OFSP was confirmed with differentiated cultures of Caco-2 human intestinal cells. Continued development of OFSP such as the Amelia and Beauregard varieties that are rich in trans-ßC and dissemination of best practices for home cooking are encouraged to increase consumption of this food to decrease the risk of vitamin A deficiency in Brazil.
Assuntos
Células CACO-2/efeitos dos fármacos , Culinária/métodos , Ipomoea batatas/química , Vitamina A/metabolismo , beta Caroteno/farmacocinética , Disponibilidade Biológica , Células CACO-2/metabolismo , Linhagem Celular , Temperatura Alta , Humanos , Técnicas In Vitro , Raízes de Plantas/química , beta Caroteno/análiseRESUMO
Colon adenocarcinoma is a disease expanding worldwide. Cancer of colon and rectum are among the top ten most insidious types in Brazil. In vitro and in vivo studies have demonstrated the efficacy of the hormone melatonin to prevent and reduce tumor growth. However, there are only few studies addressing the action of melatonin on Caco-2 cells. Thus, the cytotoxic effect of melatonin on the ultrastructure of Caco-2 cells was investigated. The MTT colorimetric method was used to assess the cytotoxicity. A total of 2×10(6)cells/mL were seeded in microplates and incubated at 50, 25, 12.5, 6.25, 3.125, 1.56, 0.78 and 0.0 (control) µg/mL of melatonin. For ultrastructural analysis concentrations with low, medium and high cytotoxicity plus the control were used for ultrastructural analysis. The concentrations 50, 1.56 and 0.78 µg/mL of melatonin showed low, medium and high cytotoxicity, respectively. Ultrastructurally, the control tumor cells were shown to be preserved. Caco-2 cells showed morphological changes at 50 µg/mL of melatonin, with numerous vacuoles, mitochondrial degeneration and reduced glycogen. However, Caco-2 cells also showed altered morphology in treatments at 1.56 and 0.78 µg/mL of melatonin with characteristics of cells in degeneration by the presence of numerous vacuoles, absence of microvilli, mitochondrial degeneration and nuclear fragmentation. Thus, one can infer that concentrations of 1.56 and 0.78 µg/mL of melatonin promote cytotoxicity in Caco-2 cells, which can probably be related to the generation of reactive oxygen species (ROS).
Assuntos
Células CACO-2/efeitos dos fármacos , Células CACO-2/ultraestrutura , Melatonina/farmacologia , Apoptose/efeitos dos fármacos , Humanos , Microscopia Eletrônica de Transmissão , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Espécies Reativas de Oxigênio/metabolismoRESUMO
BACKGROUND: The use of antimicrobial peptides (AMPs) as therapeutic agents for periodontal infections has great advantages, such as broad spectrum of action, low toxicity, and limited bacterial resistance. However, their practical use is limited because of the large amount of peptide required to exercise the microbicidal function. METHODS: LyeTxI, LL37f, and KR12 cationic peptides were prepared with ß-cyclodextrin (ßCD) at 1:1 molar ratios. The susceptibility of Porphyromonas gingivalis, Aggregatibacter actinomycetemcomitans, and Fusobacterium nucleatum were assessed in anaerobic conditions. Cytotoxicity assays were performed using osteoblast and Caco-2 epithelial cells, and hemolytic activity was assessed on rabbit erythrocytes at an absorbance of 414 nm. Parameters of surface roughness and electrical charge were established by atomic force microscopy and zeta (ζ) potential, respectively. RESULTS: AMP/ßCDs drastically decreased the peptide concentration required for activity against the bacteria tested. Moreover, AMPs associated with ßCD were able to modify cell-surface parameters, such as roughness and ζ potential. On the other hand, AMP/ßCD did not alter the degree of hemolysis induced by the pure AMPs. The effective concentration at half-maximum values of the peptides and compounds on osteoblasts were greater than the concentrations required to achieve inhibition of bacterial growth in all the species tested. AMP/ßCDs inhibited the proliferation of Caco-2 epithelial cells in a more efficient manner than AMPs alone. CONCLUSION: AMP/ßCD compounds more effectively inhibit periodontopathogenic bacteria than AMPs alone, with the additional ability of inhibiting the proliferation of epithelial cells at concentrations that are non-cytotoxic for osteoblasts and erythrocytes.
Assuntos
Antibacterianos/farmacologia , Peptídeos Catiônicos Antimicrobianos/farmacologia , Antimitóticos/farmacologia , Sequestrantes/farmacologia , beta-Ciclodextrinas/farmacologia , Aggregatibacter actinomycetemcomitans/efeitos dos fármacos , Animais , Antibacterianos/administração & dosagem , Peptídeos Catiônicos Antimicrobianos/administração & dosagem , Antimitóticos/administração & dosagem , Células CACO-2/efeitos dos fármacos , Catelicidinas/farmacologia , Proliferação de Células/efeitos dos fármacos , Eletroquímica , Células Epiteliais/efeitos dos fármacos , Eritrócitos/efeitos dos fármacos , Fusobacterium nucleatum/efeitos dos fármacos , Hemólise/efeitos dos fármacos , Humanos , Microscopia de Força Atômica , Osteoblastos/efeitos dos fármacos , Fragmentos de Peptídeos/farmacologia , Porphyromonas gingivalis/efeitos dos fármacos , Coelhos , Sequestrantes/administração & dosagem , beta-Ciclodextrinas/administração & dosagemRESUMO
The complex of vanadyl(IV) cation with oxodiacetate, VO(oda) caused an inhibitory effect on the proliferation of the human colon adenocarcinoma cell line Caco-2 in the range of 25-100 µM (P < 0.001). This inhibition was partially reversed by scavengers of free radicals. The difference in cell proliferation in the presence and the absence of scavengers was statistically significant in the range of 50-100 µM (P < 0.05). VO(oda) altered lysosomal and mitochondria metabolisms (neutral red and MTT bioassays) in a dose-response manner from 10 µM (P < 0.001). Morphological studies showed important transformations that correlated with the disassembly of actin filaments and a decrease in the number of cells in a dose response manner. Moreover, VO(oda) caused statistically significant genotoxic effects on Caco-2 cells in the low range of concentration (5-25 µM) (Comet assay). Increment in the oxidative stress and a decrease in the GSH level are the main cytotoxic mechanisms of VO(oda). These effects were partially reversed by scavengers of free radicals in the range of 50-100 µM (P < 0.05). Besides, VO(oda) interacted with plasmidic DNA causing single and double strand cleavage, probably through the action of free radical species. Altogether, these results suggest that VO(oda) is a good candidate to be evaluated for alternative therapeutics in cancer treatment.
Assuntos
Acetatos/toxicidade , Acetatos/uso terapêutico , Células CACO-2/efeitos dos fármacos , Neoplasias/tratamento farmacológico , Vanadatos/toxicidade , Vanadatos/uso terapêutico , Acetatos/química , Actinas/metabolismo , Animais , Células CACO-2/citologia , Proliferação de Células/efeitos dos fármacos , Ensaio Cometa , Citoesqueleto/efeitos dos fármacos , Dano ao DNA , Relação Dose-Resposta a Droga , Glutationa/metabolismo , Humanos , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Vanadatos/químicaRESUMO
AIM: Statin disposition and response are greatly determined by the activities of drug metabolizing enzymes and efflux/ uptake transporters. There is little information on the regulation of these proteins in human cells after statin therapy. In this study, the effects of atorvastatin and simvastatin on mRNA expression of efflux (ABCB1, ABCG2 and ABCC2) and uptake (SLCO1B1, SLCO2B1 and SLC22A1) drug transporters in Caco-2 and HepG2 cells were investigated. METHODS: Quantitative real-time PCR was used to measure mRNA levels after exposure of HepG2 and Caco-2 cells to statins. RESULTS: Differences in mRNA basal levels of the transporters were as follows: ABCC2>ABCG2>ABCB1>SLCO1B1>>>SLC22A1>SLC O2B1 for HepG2 cells, and SLCO2B1>>ABCC2>ABCB1>ABCG2>>>SLC22A1 for Caco-2 cells. While for HepG2 cells, ABCC2, ABCG2 and SLCO2B1 mRNA levels were significantly up-regulated at 1, 10 and 20 micromol/L after 12 or 24 h treatment, in Caco-2 cells, only the efflux transporter ABCB1 was significantly down-regulated by two-fold following a 12 h treatment with atorvastatin. Interestingly, whereas treatment with simvastatin had no effect on mRNA levels of the transporters in HepG2 cells, in Caco-2 cells the statin significantly down-regulated ABCB1, ABCC2, SLC22A1, and SLCO2B1 mRNA levels after 12 or 24 h treatment. CONCLUSION: These findings reveal that statins exhibits differential effects on mRNA expression of drug transporters, and this effect depends on the cell type. Furthermore, alterations in the expression levels of drug transporters in the liver and/or intestine may contribute to the variability in oral disposition of statins.Acta Pharmacologica Sinica (2009) 30: 956-964; doi: 10.1038/aps.2009.85; published online 22 June 2009.
Assuntos
Células CACO-2/metabolismo , Linhagem Celular Tumoral/efeitos dos fármacos , Ácidos Heptanoicos/farmacologia , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Proteínas de Membrana Transportadoras/metabolismo , Pirróis/farmacologia , Animais , Atorvastatina , Células CACO-2/efeitos dos fármacos , Linhagem Celular Tumoral/metabolismo , Ácidos Heptanoicos/metabolismo , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases/metabolismo , Proteínas de Membrana Transportadoras/genética , Proteína 2 Associada à Farmacorresistência Múltipla , Pirróis/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Sinvastatina/metabolismo , Sinvastatina/farmacologiaRESUMO
The aim of this work is to study the mechanisms involved in gonyautoxins (GTXs) intestinal absorption. For this purpose, we studied the transport of GTX 2/3 epimers by intestinal epithelial cell lines (IEC-6 and Caco-2) cultured on polycarbonate filters. Specific transport was calculated by subtracting from the flux of GTX 2/3 measured at 37 degrees C that occurring at 4 degrees C, this being an indication of transcellular transport. The transcellular apical-to-basolateral (A-B) flux in Caco-2 cell monolayers, was greater than that in the opposite direction, suggesting the involvement of an active transport system favoring the absorption of the toxin. However, in IEC-6 cells the transcellular basolateral-to-apical (B-A) specific transport of the toxin was greater than that in the opposite direction. The A-B and B-A fluxes were, respectively, 127 +/- 26 and 205 +/- 23 nmol/min, suggesting the presence of a prevalent secretive process of the toxin in IEC-6 cells. The A-B transport of GTX 2/3 epimers in Caco-2 cells, but not in IEC-6 cells, was partially Na(+)-dependent and significantly inhibited by adenosine. TEA and verapamil in both Caco-2 and IEC-6 cells failed to affect the A-B and B-A transport of GTX 2/3 epimers. Cyanine in IEC-6 cells, but not in Caco-2 cells, increased the A-B flux of the toxin, suggesting the involvement of the organic cation transporter in the absorption of GTX 2/3 epimers. The mitochondrial energetic uncoupler 2,4-dinitrophenol significantly inhibited the A-B and the B-A transport in both cell lines. In conclusion, IEC-6 cells secrete actively the toxins, whereas Caco-2 cells were found to absorb the toxins in a process that was inhibited in the presence of adenosine and the absorption was dependent of Na(+).