Patricia Pensel

Could thymol have effectiveness on scolices and germinal layer of hydatid cysts

Scolicidal solutions remain indispensable in the treatment of hydatid cyst disease. Properties of an ideal solution would be inexpensiveness and the promotion of a rapid and complete scolicidal effect with an absence of local and systemic side effects. From this point of view, no ideal solution and agents have been described yet. The aim of the present work was to determine the in vitro effect of high concentrations of thymol against protoscoleces, microcyst and cyst of Echinococcus granulosus and to evaluate its possible role as a scolicidal agent during surgery or PAIR. After short exposure times, a rapid effect was observed depending on the parasitic material. After 2 min of exposure to thymol, viability of protoscoleces was approximately 1.3% at a concentration of 250 μg/ml. The protoscolicidal effect is dose and time dependent. The results of the in vitro treatment with thymol were similar in both microcysts and secondary murine cysts. The employment of SEM and TEM allowed us to examine, at an ultrastructural level, the effects induced by thymol on E. granulosus protoscoleces, microcysts and murine cysts. In conclusion, the data obtained clearly demonstrated that thymol caused severe damages to the parasite even after short incubation times. This fact and the lack of toxicity at the evaluated concentrations, allow us to propose it as a possible scolicidal agent during hydatid cysts surgery and/or PAIR.

Cystic echinococcosis therapy: Albendazole-loaded lipid nanocapsules enhance the oral bioavailability and efficacy in experimentally infected mice

Therapeutic failures attributed to medical management of cystic echinococcosis (CE) with albendazole (ABZ) have been primarily linked to the poor drug absorption rate resulting in low drug level in plasma and hydatid cysts. Lipid nanocapsules (LNCs) represent nanocarriers designed to encapsulate lipophilic drugs, such as ABZ. The goals of the current work were: (i) to characterize the plasma and cyst drug exposure after the administration of ABZ as ABZ-LNCs or ABZ suspension (ABZ-SUSP) in mice infected with Echinococcus granulosus, and ii) to compare the clinical efficacies of both ABZ formulations. Enhanced ABZ sulphoxide (ABZ-SO) concentration profiles were obtained in plasma and cysts from ABZ-LNC treated animals. ABZSO exposure (AUC0-LOQ) was significantly higher in plasma and cyst after the ABZ-LNC treatments, both orally and subcutaneously, compared to that observed after oral administration of ABZ-SUSP. Additionally, ABZSO concentrations measured in cysts from ABZ-LNC treated mice were 1.7-fold higher than those detected in plasma. This enhanced drug availability correlated with an increased efficacy against secondary CE in mice observed for the ABZ-LNCs, while ABZ-SUSP did not reach differences with the untreated control group. This new pharmacotechnically-based strategy could be a potential alternative to improve the treatment of human CE.

Enhanced chemoprophylactic and clinical efficacy of albendazole formulated as solid dispersions in experimental cystic echinococcosis.

Cystic echinococcosis is a chronic, complex, and still neglected disease. Although albendazole has demonstrated efficacy, only about one-third of patients experience complete remission or cure and 30-50% of treated patients develop some evidence of a therapeutic response. Different strategies have been developed in order to improve the albendazole water solubility and dissolution rate. The aim of the current work was to investigate the chemoprophylactic and clinical efficacy of an albendazole:poloxamer 188 solid dispersion formulation on mice infected with Echinococcus granulosus metacestodes. Albendazole formulated as solid dispersion had greater chemoprophylactic and clinical efficacy than albendazole alone. The improved in therapeutic efficacy could be a consequence of the increase in the systemic availability of albendazole sulfoxide. The work reported here demonstrates that in vivo treatment with albendazole:poloxamer 188 impairs the development of the hydatid cysts. This new pharmacotechnically based strategy could be a suitable alternative for treating cystic echinococcosis in humans.

Efficacy of Essential Oils of Thymus vulgaris and Origanum vulgare on Echinococcus granulosus

The aim of the present work was to determine the in vitro effect of T. vulgaris and O. vulgare essential oils against E. granulosus protoscoleces and cysts. Essential oils were added to the medium resulting in thymol final concentrations of 10 μg/mL. The essential oils had a time-dependent effect provoking the complete loss of protoscolex viability after 72 days of postincubation. The results were confirmed at the ultrastructure level. Loss of infectivity in protoscoleces incubated with O. vulgare after 60 days was observed. On the other hand, the weight of cysts recorded in mice inoculated with T. vulgaris treated protoscoleces was significantly lower than that obtained in control group. Gamma-glutamyl-transpeptidase activity was readily detected in the culture supernatant of protoscoleces treated either with the essential oils or thymol. T. vulgaris and O. vulgare essential oils and thymol can induce cell apoptosis of protoscoleces after short incubation times. The efficacy of T. vulgaris and O. vulgare essential oils was also demonstrated in vitro on E. granulosus murine cysts. Our data suggest that essential oils of T. vulgaris and O. vulgare have anthelmintic effect against protoscoleces and cysts of E. granulosus.

In vitro effect of 5-fluorouracil and paclitaxel on Echinococcus granulosus larvae and cells

Human cystic echinococcosis is a zoonosis caused by the metacestode stage of the tapeworm Echinococcus granulosus. Although benzimidazole compounds such as albendazole and mebendazole have been the cornerstone of chemotherapy for the disease, there is often no complete recovery after treatment. Hence, in searching for novel treatment options, we examined the in vitro efficacies of 5-fluorouracil (5-FU) and paclitaxel (PTX) against E. granulosus germinal cells, protoscoleces and cysts. 5-FU or PTX inhibited the growth of E. granulosus cells in a time dependent manner. Although both treatments had a protoscolicidal effect, 5-FU had a considerably stronger effect than PTX. 5-FU produced a dose- and time-dependent effect, provoking the complete loss of viability after 24 days of incubation. Moreover, cysts did not develop following the inoculation of treated protoscoleces into mice. The loss of viability was slower in PTX treated protoscoleces, reaching to approximately 60% after 30 days. The results of the in vitro treatment with 5-FU and PTX were similar in secondary murine cysts. The employment of SEM and TEM allowed us to examine, at an ultrastructural level, the effects induced by 5-FU and PTX on E. granulosus germinal cells, protoscoleces and murine cysts. In conclusion, the data obtained clearly demonstrated that 5-FU and PTX at clinically achievable concentrations inhibit the survival of larval cells, protoscoleces and metacestodes. In vivo studies to test the antiparasitic activities of 5-FU and PTX are currently being undertaken on the murine model of cystic echinococcosis.

In vivo activity of albendazole in combination with thymol against Echinococcus multilocularis.

Human alveolar echinococcosis (AE) is caused by the fox tapeworm Echinococcus multilocularis and is usually lethal if left untreated. The current strategy for treating human AE is surgical resection of the parasite mass complemented by chemotherapy with benzimidazole compounds. However, reliable chemotherapeutic alternatives have not yet been developed stimulating the research of new treatment strategies such as the use of medicinal plants. The aim of the current study was to investigate the efficacy of the combination albendazole (ABZ)+thymol on mice infected with E. multilocularis metacestodes. For this purpose, mice infected with parasite material were treated daily for 20 days with ABZ (5 mg/kg), thymol (40 mg/kg) or ABZ (5 mg/kg)+thymol (40 mg/kg) or left untreated as controls. After mice were euthanized, cysts were removed from the peritoneal cavity and the treatment efficacy was evaluated by the mean cysts weight, viability of protoscoleces and ultrastructural changes of cysts and protoscoleces. The application of thymol or the combination of ABZ+thymol resulted in a significant reduction of the cysts weight compared to untreated mice. We also found that although ABZ and thymol had a scolicidal effect, the combination of the two compounds had a considerably stronger effect showing a reduction in the protoscoleces viability of 62%. These results were also corroborated by optical microscopy, SEM and TEM. Protoscoleces recovered from ABZ or thymol treated mice showed alterations as contraction of the soma region, rostellar disorganization and presence of blebs in the tegument. However both drugs when combined lead to a total loss of the typical morphology of protoscoleces. All cysts removed from control mice appeared intact and no change in ultrastructure was detected. In contrast, cysts developed in mice treated with ABZ revealed changes in the germinal layer as reduction in cell number, while the treatment with thymol or the ABZ+thymol combination predominantly showed presence of cell debris. On the other hand, no differences were found in alkaline phosphatase (AP), glutamate oxaloacetate transaminase (GOT) and glutamate pyruvate transaminase (GPT) activities between control and treated mice, indicating the lack of toxicity of the different drug treatments during the experiment. Because combined ABZ+thymol treatment exhibited higher treatment efficiency compared with the drugs applied separately against murine experimental alveolar echinococcosis, we propose it would be a useful option for the treatment of human AE.

In vitro and in vivo efficacy of carvacrol against Echinococcus granulosus

Currently, benzimidazoles are used as chemotherapeutic agents and as a complement to surgery and PAIR in the treatment of cystic echinococcosis (CE). They are generally applied at high doses causing side effects and, 50% of cases do not respond favorably to such chemotherapy. The use of essential oils obtained by distillation from aromatic plants would be an effective alternative or complementary to the synthetic compounds, because would not bring the appearance of side effects. Carvacrol and his isomer thymol are the main phenolic components from essential oils of Origanum vulgare (oregano) and Thymus vulgaris (thyme). The aim of the present work was to evaluate the in vitro and in vivo efficacy of carvacrol against Echinococcus granulosus metacestodes. For the in vitro assay, protoscoleces and cysts of E. granulosus were incubated with carvacrol at the following final concentrations: 10, 5 and 1μg/ml of carvacrol. The maximum protoscolicidal effect was found with 10μg/ml of carvacrol. Results of viability tests were consistent with the structural and ultrastructural damage observed in protoscoleces. Ultrastructural studies revealed that the germinal layer of cysts treated with carvacrol lost the multicellular structure feature. In the clinical efficacy study, a reduction in cyst weight was observed after the administration of 40mg/kg of carvacrol during 20days in mice with cysts developed during 4 months, compared to that of those collected from control mice. Given that the in vivo effect of carvacrol was comparable with the treatment of reference with ABZ and the fact that is a safe compound, we postulated that carvacrol may be an alternative option for treatment of human CE.

Chemoprophylactic and therapeutic efficacy of thymol in murine cystic echinococcosis

Cystic echinococcosis is a zoonotic disease caused by the larval stage of the cestode Echinococcus granulosus. The drugs commonly used against cystic echinococcosis are benzimidazoles. Unfortunately, 20%-40% of cases do not respond favorably to such chemotherapy. Consequently, the search of new therapeutic alternatives such as the use of traditional medicinal plants has been increased. The aim of the current experimental work was to investigate the chemoprophylactic and clinical efficacy of thymol on mice infected with E. granulosus metacestodes. Thymol (40 mg/kg) was administered under two different therapeutic schemes: dosing every 24h over 20 days and treatment every 12h for 10 days. Thymol demonstrated efficacy against experimental murine cystic echinococcosis. The chemoprophylactic and therapeutic effects of thymol were comparable to that of albendazole. Due to the lack of toxicity observed in mice at the tested doses; we consider that thymol is a potential alternative to be applied for the treatment of human hydatid disease.

Albendazole nanocrystals in experimental alveolar echinococcosis: Enhanced chemoprophylactic and clinical efficacy in infected mice

Human alveolar echinococcosis is caused by the fox tapeworm Echinococcus multilocularis and is usually fatal if left untreated. Medical treatment with albendazole (ABZ) remains an effective option. However, due to its low aqueous solubility, ABZ is poorly and erratically absorbed following oral administration resulting in low drug levels in plasma and liver distribution. Thus, there arises the need to find a simple, efficient and scalable method to produce new ABZ formulations with increased bioavailability. Bearing this in mind, ABZ nanocrystals (ABZ-NCs) appears to be a useful tool to achieve this goal. The aim of the current study was to investigate the chemoprophylactic and clinical efficacy of an ABZ-NC formulation on mice infected with E. multilocularis. In the chemoprophylactic efficacy study, mean weight of the cysts recovered from the ABZ-NC group was 50% lower than that recorded from untreated mice, whereas the treatment with ABZ suspension did not show preventive effect. The viability of protoscoleces isolated from ABZ-NC treated mice was significantly lower than control groups. In the clinical efficacy studies, both ABZ formulations resulted in a reduction in the mean weight of the cysts obtained from mice, however only the treatment with the nanosuspension revealed significant differences (P < 0.05) compared to the control groups. Treatment with ABZ-NCs reduced the weight of the cysts by 77% and the viability of their protoscoleces to 34%. All these results coincided with the tissue damage determined at the ultrastructural level. The enhanced chemoprophylactic and clinical efficacy of ABZ-NCs observed in this study could be attributed to an increase in the oral bioavailability of the drug. In a next step, we will characterize the cyst concentration profile after the administration of ABZ-NCs in mice infected with E. multilocularis.

Could beta-myrcene be an alternative to albendazole for the treatment of experimental cystic echinococcosis?

Echinococcus granulosus causes hydatidosis or cystic echinococcosis in humans and livestock. In humans, this disease can be managed with surgery, percutaneous treatment, chemotherapy and/or observation. The chemotherapeutic agents used and approved for treatment of hydatidosis are benzimidazoles. Because of the difficulties in achieving successful treatment, considerable efforts have been made to find new natural compounds against hydatid disease. Beta-myrcene is a monoterpene presented in the essential oils of different plants. It is the principal component of essential oil of Rosmarinus officinalis (rosemary). The goal of the present study was to evaluate the in vitro effects of beta-myrcene against germinal cells, protoscoleces and murine cyst of E. granulosus, as well also, investigate its chemoprophylactic activity in a murine model of cystic echinococcosis. For the in vitro assays, the parasites were incubated with beta-myrcene at 10, 5 and 1 μg/mL. The treatments were dose and time-dependent, and consistent with the observed morphological alterations. In the chemoprophylactic efficacy study, the effect of beta-myrcene was similar to albendazole, the reference drug for human echinococcosis treatment.