Protoscolicidal Effects Research Articles

Evaluation of the In-Vitro Effects of Albendazole, Mebendazole, and Praziquantel Nanocapsules against Protoscolices of Hydatid Cyst.

Cystic echinococcosis still remains a serious health and economic problem worldwide. The etiologic agent is Echinococcus granulosus sensu lato, giving origin to a fluid-filled cystic lesion. Therapy faces several challenges. Nanodrugs have shown promise as chemotherapeutics against hydatid cysts. The present study evaluated a highly safe lipid nano-polymeric capsule for its superior efficacy and ability to overcome drug resistance. Nanocapsule drugs were formulated into six groups: Albendazole, mebendazole, praziquantel, albendazole + mebendazole, albendazole + praziquantel, and praziquantel + mebendazole. The protoscolicidal effects of these six groups were assessed at 10, 60, and 120 min in three concentrations (1, 0.5, and 0.25 mg/mL). Drug formulations were evaluated via zeta potential, droplet size, solubility, particle size analyzer (PSA), and scanning electron microscopy. According to the PSA results, the mean size of the albendazole nanocapsules was 193.01 nm, mebendazole was 170.40 nm, and praziquantel was 180.44 nm. Albendazole + mebendazole showed the greatest protoscolicidal activity at a concentration of 1 mg/mL after 120 min. In contrast, each drug's 0.25 mg/mL single-dose times showed the least protoscolicidal activity after 120 min. With the right application of nanotechnology, it is possible to produce safe and effective drugs, such as the polymeric combination of albendazole and mebendazole, which has promising implications.

In vitro and ex vivo protoscolicidal effect of poly(amidoamine) nanoemulsion against Echinococcus granulosus

Hydatidosis causes a serious health hazard to humans and animals leading to significant economic and veterinary and public health concern worldwide. The present study aimed to evaluate the in vitro and ex vivo protoscolicidal effects of synthesized poly(amidoamine), PAMAM, nanoemulsion. In this study, PAMAM was characterized through dynamic light scattering technique to investigate the particle size and zeta potential of nanoemulsified polymer. For the in vitro and ex vivo assays, we used eosin dye exclusion test and scanning electron microscope (SEM) to evaluate the effects of the prepared and characterized PAMAM nanoemulsion against protoscoleces from Echinococcus granulosus sensu lato G6 (GenBank: OQ443068.1) isolated from livers of naturally infected camels. Various concentrations (0.5, 1, 1.5 and 2 mg/mL) of PAMAM nanoemulsion at different exposure times (5, 10, 20 and 30 min) were tested against protoscolices. Our findings showed that PAMAM nanoemulsion had considerable concentration- and time-dependent protoscolicidal effect at both in vitro and ex vivo experiments. Regarding in vitro assay, PAMAM nanoemulsion had a potent protoscolicidal effect when compared with the control group with a highest protoscolicidal activity observed at the concentration of 2 mg/mL at all exposure times, such that 100% of protoscolices were killed after 20 min of exposure. Also, the mortality of protoscolices was 100% after 30 min of exposure to 1 and 1.5 mg/mL of PAMAM nanoemulsion, in vitro. Concerning ex vivo assay PAMAM nanoemulsion recorded the highest mortality rates at the concentration of 2 mg/mL (55, 99.4 and 100% at 10, 20, 30 min, respectively). Ultrastructure examination of examined protoscolices after 20 min of exposure to PAMAM nanoemulsion showed a complete loss of rostellar hooks, disruption of suckers with disorganization of hooks with partial or complete loss of them, and damage of protoscolices tegument with loss of their integrity in the form of holes and contraction of the soma region were observed in 1.5 and 2 mg/mL of PAMAM, in vitro and ex vivo, showing more damage in the in vitro conditions. It can be concluded that PAMAM nanoemulsion is a promising protoscolicidal agent offering a high protoscolicidal effect at a short exposure time. Further in vivo studies and preclinical animal trials are required to evaluate its efficacy and clinical applications against hydatid cysts.

High potency of linalool-zinc oxide nanocomposite as a new agent for cystic echinococcosis treatment.

This current in vitro, ex vivo, and in vivo research aims to evaluate and analyze the linalool-zinc oxide nanocomposite (Lin-ZNP) for treating cystic echinococcosis. Lin-ZNP was synthesized using an ethanolic solution of polyvinyl alcohol. The protoscolicidal effects of Lin-ZNP were tested on hydatid cyst protoscoleces (PTS) in both in vitro and ex vivo by eosin exclusion test. The study also examined the impact on caspase-3 gene expression and the external structure of PTS. The in vivo effect was measured by examining hydatid cysts' quantity, dimensions, and weight in mice intraperitoneally infected with 0.5 mL of PTS solution containing 1,000 PTS. The antioxidant and inflammatory cytokine gene expression levels were examined using real-time PCR. Lin-ZNP significantly (P < 0.001) killed the PTS in both in vitro and ex vivo in a dose- and time-dependent manner. The treated PTS exhibited creases and protrusions as a result of bleb formation and upregulation in the gene expression of caspase-3. Upon treatment with Lin-ZNP, there was a significant (P < 0.001) reduction in the number, diameter, and weight of the hydatid cysts. Treatment with Lin-ZNP nanocomposite led to a significant increase in the expression of antioxidant genes and a notable decrease in oxidative stress markers, and the expression levels of IL-4 and IL-10. Lin-ZNP has the potential to act as a scolicidal agent and demonstrates promise in controlling hydatid cysts in a mouse model, attributed to its antioxidant and anti-inflammatory properties. However, additional studies in clinical trials are needed to confirm the use of Lin-ZNP for treating hydatidosis.

Green Synthesis, Characterization, and Antiparasitic Effects of Gold Nanoparticles against Echinococcus granulosus Protoscoleces.

Echinococcosis, or hydatidosis, is one of the most important zoonotic diseases, which is initiated by the larval stage in the clasts of Echinococcus granulosus. For the treatment of hydatidosis, surgery is still the preferred method and the first line of treatment for symptomatic patients. Unfortunately, most of the scolicidal agents that are injected inside cysts during hydatid cyst surgery have side effects, including leaking out of the cyst and adverse effects on the living tissue of the host, such as necrosis of liver cells, which limits their use. This work was carried out to study the lethal effect of green synthesized gold nanoparticles (Au-NCs) against hydatid cyst protoscoleces. Au-NCs were green synthesized using the Saturja khuzestanica extract. Au-NCs were characterized by UV-visible absorbance assay, electron microscopy analysis, X-ray diffraction (XRD), and Fourier transform infrared (FTIR) spectroscopy. Scolicidal properties of Au-NCs (1-5 mg/mL) were studied against protoscoleces for 10-60 min. The effect of Au-NCs on the expression level of the caspase-3 gene as well as the ultrastructural examination was studied by real-time PCR and scanning electron microscopy (SEM). The cytotoxicity of Au-NCs on hepatocellular carcinoma (HepG2) and normal embryonic kidney (HEK293) cell lines was also studied by the cell viability assay. The obtained Au-NCs are cubes and have an average size of 20-30 nm. The highest scolicidal efficacy was observed at 5 mg/mL with 100% mortality after 20 min of treatment for hydatid cyst protoscoleces. In ex vivo, Au-NCs required more incubation time, indicating more protoscolicidal effects. Au-NCs markedly upregulated the gene level of caspase-3 in protoscoleces; whereas they changed the ultra-structure of protoscoleces by weakening and disintegrating the cell wall, wrinkles, and protrusions due to the formation of blebs. We showed the effective in vitro and ex vivo scolicidal effects of Au-NCs against hydatid cyst protoscoleces by provoking the apoptosis process of caspase-3 activation and changing the ultrastructure of protoscoleces with no significant cytotoxicity against human normal cells. However, additional studies should be conducted to determine the possible harmful side effects and accurate efficacy.

In vitro and ex vivo protoscolicidal effects of hydroalcoholic extracts of Eucalyptus microtheca on protoscoleces of Echinococcus granulosus sensu stricto: A light and scanning electron microscopy (SEM) study

BackgroundCystic echinococcosis (CE) is one of the most widespread and important global helminth zoonoses. Treatment relies mainly on surgery and, or percutaneous interventions. However, spillage of live protoscoleces (PSCs) leading to recurrence is a problem during surgery. So, the application of protoscolicidal agents before surgery is required. This study aimed to investigate the activity and safety of hydroalcoholic extracts of E. microtheca against PSCs of Echinococcus granulosus sensu stricto (s.s.) both in vitro and also ex vivo, which is a simulation to Puncture, Aspiration, Injection, and Re-aspiration (PAIR) method. MethodsConsidering the effects of heat on the protoscolicidal effecacy of Eucalyptus leaves, hydroalcoholic extraction was performed by both soxhlet extraction at 80 °C and percolation at room temperature. The protoscolicidal action of hydroalcoholic extracts was assessed by in vitro and ex vivo assessments. Infected sheep livers were collected from the slaughterhouse. Then, the genotype of hydatid cysts (HCs) was confirmed by sequencing and, isolates were limited to E. granulosus s.s. In the next step, ultrastructural changes of Eucalyptus–exposed PSCs were studied by scanning electron microscopy (SEM). Finally, a cytotoxicity test was conducted by (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay to evaluate the safety of E. microtheca. ResultsThe prepared extracts by soxhlet extraction and percolation were, successfully exerted strong protoscolicidal effects in both in vitro and ex vivo tests. The results of in vitro assessment indicated that hydroalcoholic extract of E. microtheca prepared by percolation at room temperature (EMP) and hydroalcoholic extract of E. microtheca prepared by soxhlet extraction at 80 °C (EMS) killed all PSCs (100%) at concentrations of 10 and 12.5 mg/mL, respectively. Also, EMP showed 99% protoscolicidal action after 20 min in an ex vivo setting compared to EMS. SEM micrographs confirmed potent protoscolicidal and destructive effects of E. microtheca against PSCs. The cytotoxicity of EMP was tested on the HeLa cell line using MTT assay. The value of 50% cytotoxic concentration (CC50) was calculated at 46.5 μg/mL after 24h. ConclusionBoth hydroalcoholic extracts showed potent protoscolicidal activity and, especially EMP produced remarkable protoscolicidal effects compared to the control group.

Protoscolicidal effects of curcumin nanoemulsion against protoscoleces of Echinococcus granulosus

BackgroundThe aim of the present study was to assess in vitro protoscolicidal effects of curcumin nanoemulsion (CUR-NE) against protoscoleces of cystic echinococcosis (CE)/hydatid cysts.MethodsThe CUR-NE was prepared via spontaneous emulsification of soybean as the oil phase, a mixture of Tween 80 and Tween 85 as the surfactant, ethanol as the co-surfactant and distilled water. Various concentrations of CUR-NE (156, 312, 625 and 1250 µg/ml) were exposed to collected protoscoleces of infected sheep liver hydatid cysts for 10, 20, 30, 60 and 120 min. Viability of the protoscoleces were assessed using eosin exclusion test. Morphological changes of the protoscoleces were observed using differential interference contrast (DIC) microscopy.ResultsThe mean particle size and zeta potential of CUR-NE included 60.4 ± 14.8 nm and − 16.1 ± 1.1 mV, respectively. Results showed that the viability of the protoscoleces decreased significantly with increases in CUR-NE concentrations (p < 0.001). The mortality rates of protoscoleces with exposure to concentrations of 1250 and 625 µg/ml of CUR-NE for 60 min were 94 and 73.33%, respectively. Mortality of the protoscoleces was 100% after 120 min of exposure to 1250 and 625 µg/ml concentrations of CUR-NE. Using NIC microscopy, extensively altered tegumental surface protoscoleces was observed after protoscoleces exposure to CUR-NE.ConclusionThe findings of the present study revealed the in vitro protoscolicidal potential of CUR-NE. Therefore, CUR-NEs are addressed as novel protoscolicidal agents, which can be used as an alternative natural medicine to kill the protoscoleces, owing to their low toxicity and significant inhibition potency. However, further studies are necessary to investigate pharmacologic and pharmacokinetics of CUR-NEs.

High potency of magnetic iron oxide nanoparticles covered by piroctone olamine against cystic echinococcosis

This study examined the synthesis of magnetic iron oxide nanoparticles coated with PO (FOMNPsP) and assessed their in vitro, ex vivo, and in vivo effects against cystic echinococcosis. The FOMNPsP was synthesized through the alkalization of iron ions in a deoxygenated form. In vitro and ex vivo protoscolicidal effects of FOMNPsP (100–400 μg/mL) were evaluated on hydatid cyst protoscoleces by the eosin exclusion test for 10–60 min. The effect of FOMNPsP on caspase-3 gene expression and exterior ultra-structural of protoscoleces was assessed by real-time PCR and scanning electron microscopy (SEM), respectively. In vivo effects were assessed by evaluating the number, size, and weight of hydatid cysts among infected mice. The FOMNPsP size was < 55 nm, and the most frequent particles were in the 15–20 nm range. In vitro and ex vivo assays revealed that the highest protoscolicidal effect was observed at 400 µg/mL with 100% lethality. After exposure of protoscoleces with FOMNPsP, the level of gene expression of caspase-3 was dose-dependently increased (p < 0.05). By SEM, the FOMNPsP-treated protoscoleces showed wrinkles and bulges resulting from the formation of blebs. FOMNPsP significantly decreased (p < 0.01) the mean number, size, and weight of the hydatid cyst. FOMNPsP revealed the potent protoscolicidal traits through disrupting the cell wall and apoptosis induction. The results also indicated the promising effect of FOMNPsP in controlling hydatid cysts in the animal model. Although FOMNPsP is safe for human normal cells, more investigations are required to clarify its toxicity and precise mechanisms of action.

In vitro protoscolicidal effects of lithocholic acid on protoscoleces of Echinococcus granulosus and its mechanism

Surgery has been found to be the best choice of treatment for hydatidosis. However, leakage of cyst contents during surgery is the foremost reason for recurrence of hydatidosis. In this study, we investigated the in vitro efficacy of lithocholic acid (LCA) against Echinococcus granulosus protoscoleces. The protoscoleces were divided into a control group, an albendazole (ABZ) positive control group and LCA intervention groups at concentrations of 0.5, 1, 2, and 3 mmol/L and stained with 0.1% eosin for observation using an inverted microscope; the protoscolecal ultrastructure was examined with SEM and TEM; the activities of ROS, SOD, and caspase-3 were investigated using an ROS kit, SOD kit, and caspase-3 kit, respectively; the contents of HO-1 and NQO-1 were analyzed by enzyme-linked immunosorbent assay; and the expression level of cytochrome c (Ctyc) was analyzed by western blotting. Results: As the concentration of LCA increased, the survival rate of protoscoleces gradually decreased. The microstructure shows that the external shape and internal structure were gradually deformed and collapse. SOD, GSH, HO-1 and NQO-1 decreased more significantly in the 3 mmol/L LCA group. However, ROS levels gradually increased. LCA treatment for 3 days at all concentrations significantly increased caspase-3 activity and expression in a dose-dependent manner. LCA decreased the level of Ctyc protein in vitro. LCA demonstrated a parasiticidal effect on the protoscoleces of Echinococcus granulosus in vitro. LCA may induce apoptosis of E. granulosus protoscoleces by oxidative stress and mitochondrial pathways.

Effects of green synthesized zinc nanoparticles alone and along with albendazole against hydatid cyst protoscoleces

BackgroundThe current investigation aims to green synthesized the zinc nanoparticles (ZnNPs) using Lavandula angustifolia extract by microwave technique and its protoscolicidal effects alone and combined with albendazole against hydatid cyst protoscoleces. MethodsDifferent concentrations of the ZnNPs (50, 100, and 200 μg/ml) alone and combined with albendazole (ALZ, 100 μg/ml) were treated with hydatid cyst protoscoleces obtained from liver of infected sheep for 5–60 min in vitro and ex vivo. Eosin exclusion examination was used to assess the viability of protoscoleces. The induction of apoptosis in hydatid cyst protoscoleces was assessed by measurement of the Caspase-3 activity of protoscoleces treated with various concentrations of ZnNPs. ResultsThe size of green synthesized ZnNPs was ranged from 30 to 80 nm, most of these nanoparticles were between 50 and 60 nm in size. In vitro, the highest scolicidal effect of ZnNPs was observed at the concentration of 200 μg/ml, where it killed 81.6% of protoscolices. While the combination of these nanoparticles with ALZ, especially at the concentration of 200 μg/ml, completely killed the protoscolices after 10 min’ exposure. However, compared to in vitro assay, the drugs tested took longer to show their protoscolicidal effect. ConclusionBased on the obtained results, ZnNPs particularly in combination with albendazole displayed the potent protoscolicidal in vitro and ex vivo as an intraperitoneal model of administration of agents to hydatid cyst treatment; nevertheless, additional investigations are mandatory to evaluate the efficacy and safety Zn NPs as a favorable protoscolicidal agent in clinical setting.

Antiparasitic effects of Elettaria cardamomum L. essential oil and its main compounds, 1-8 Cineole alone and in combination with albendazole against Echinococcus granulosus protoscoleces

BackgroundThe present investigation aims to determine the chemical structure and protoscolicidal effects of Elettaria cardamomum L. essential oil (ECEO) and its main compounds 1–8 cineole alone and along with albendazole (ALZ) against Echinococcus granulosus protoscoleces in vitro and ex vivo. We also decided to evaluate some cellular mechanisms such as the apoptotic activity and the permeability of plasma membrane of protoscoleces treated with ECEO and 1–8 cineole. MethodsHydatid cyst protoscoleces were divided into seven groups including protoscoleces treated with ECEO 50 µl/mL (T1), protoscoleces treated with ECEO 100 µl/mL (T2), protoscoleces treated with ECEO 200 µl/mL (T3), protoscoleces treated with 1–8 cineole 100 µg/mL (T4), protoscoleces treated with 1–8 cineole 200 µg/mL (T5), protoscoleces treated with 1–8 cineole 100 µg/mL + albendazole 50 µg/mL (T6), and protoscoleces treated with 1–8 cineole 200 µg/mL + albendazole ALZ-50 µg/mL (T7). The viability of protoscoleces were recorded by eosin staining examination. Moreover, the induction of apoptosis and the plasma membrane permeability of the protoscoleces treated with ECEO and 1–8 cineole were evaluated. ResultsThe highest protoscolicidal effect of ECEO was observed at the dose of 200 µl/ml (T3). 1,8-Cineole alone and combined with ALZ, particularly at the dose of 200 µg/ml (T5 and T7), destroyed the 100% protoscolices after 10 min incubation. The ECEO (T1-T3) and 1–8 cineole alone (T4 and T5) and in combination with ALZ (T6 and T7) took longer to display their protoscolicidal effect ex vivo. The obtained results of relative fuorescent items exhibited that the protoscoleces incubated with ECEO and 1,8-Cineole, alter the permeability of plasma membrane by Sytox Green with increasing the concentration. The findings revealed exhibited that ECEO and 1,8-Cineole increasingly and dose-dependently induced activation of caspase-3 enzyme ranging from 6.8 to 23.3%. ConclusionOur obtained results revealed that ECEO and its main compound, 1,8-Cineole exhibited the potent protoscolicidal in vitro and ex vivo; and if more research is done on their efficacy and toxicity in animal models and even clinical setting, it can be suggested as a protoscolicidal agent to use during hydatid cyst surgery.

In Vitro Study on the Protoscolicidal Effect of Synthesized Novel Azole Derivatives on Protoscoleces of Cystic Echinococcosis

Objective: Echinococcosis is a zoonotic parasitic infection caused by Echinococcus granulosus sesu lato (E. granulosus s.l.) with world-wide distribution. In human, E.granulosus is found in the larval form, while the adult form is usually found in dogs. Surgery is still the treatment of choice for treatment of CE. However, chemotherapy using drugs like Albendazole, Mebendozole and Flubendazole has been used for a long time. There is still no agreement on their usage because of their high adverse effects, long-term therapy and low response. Using the new and novel drugs on protoscoleces is always interesting. Materials and Methods: We applied in vitro protoscolicidal effect of a group of azole derivatives on protoscoleces of CE cyst. Sheep hepatic cysts were collected from Shiraz abattoir and transferred to the parasitology department at Shiraz medical school, where protoscoleces were separated aseptically for protoscolicidal studies. The protoscoleces were incubated against 7 new synthesized azole derivatives using LC50 of the compounds. Results: Our results showed that ligand no.5 with a concentration of 50 µgr/ml and 10 µgr/ml, and the ligand no.3 in concentration of 50µgr/ml had the most protoscolicidal effects. The lowest protoscolicidal effect was observed for ligand no.7 with 44.30±22.1 and 53.46±28.63 percent destroying effect, respectively. Conclusions: Regarding the high protoscolecidal effect of the ligands no. 3 and 5, these compounds have in vitro protoscolicidal effects. Further ex vivo and, in vivo investigations on these compounds is suggested.

Cystic Echinococcosis and Host Interleukin-2, Interleukin-9 and Interleukin-10 as Biomarkers for Evaluation of Albendazole and Praziquantel Treatment in Experimentally Infected Mice

Echinococcus granulosus sensu lato larvae cause cystic echinococcosis, a highly contagious zoonotic illness that poses a worldwide threat to human health and development. The purpose of this study was to determine the therapeutic effectiveness of albendazole and praziquantel, each alone and in combination, in vivo and to estimate the serum levels of interleukin-2, interleukin-9 and interleukin-10 as biomarkers in evaluating the responsiveness to albendazole and praziquantel treatment in experimentally infected mice. To assess treatment efficacy in vivo, mice were intraperitoneally inoculated with viable protoscolices and then gavaged daily for 1 mo with albendazole (15 mg/kg/d) and praziquantel (25 mg/kg/d), separately and in combination (albendazole 15 mg/kg/d and praziquantel 25 mg/kg/d. The mice were euthanized on d 120 of the experiment and the number, size and location of the generated secondary hydatid cysts were investigated and serum levels of interleukin-2, interleukin-9 and interleukin-10 were estimated. The size and number of hydatid cysts were decreased upon treatment with each drug, but the decrease was significant (p<0.05) in isolates 2, 3, 4, 6, 8, 9 and 10, and 2 and 3, respectively. The most often infected organ was the liver (44.91 %), followed by a peritoneal cavity (30.3 %), subcutaneous (21.1 %), spleen (3.29 %) and lungs (0.38 %). Interleukin-2 level was significantly elevated in mice inoculating with most parasite isolates, whereas interleukin-9 and interleukin-10 levels were not altered considerably experimentally in infected and treated mice compared with untreated control. Albendazole and praziquantel markedly reduced the number and size of secondary hydatid cysts compared to the untreated control group in Echinococcus granulosus-infected mice and stimulated the production of interleukin-2, which is helpful for parasite elimination. These results indicate that albendazole and praziquantel have protoscolicidal effects and the combination of albendazole and praziquantel has significant additive effects.

Anthelmintic activity of Stevia multiaristata extract against Echinococcus granulosus sensu stricto.

Cystic echinococcosis is a zoonotic disease caused by the larval stage of the parasite Echinococcus granulosus sensu lato. The available anti-parasitic treatment is mostly limited to a continuous administration of albendazole. However, due to its numerous side-effects and efficacy of around 50%, there is a need to find new drugs to improve the treatment for this disease. In the current study, the in vitro and in vivo efficacy of a Stevia multiaristata extract against E. granulosus sensu stricto (s.s.) was demonstrated. Stevia multiaristata extract (100 and 50 μg mL−1) caused a quick viability decrease on protoscoleces which was consistent with the observed tegumental alterations. Loss of turgidity was detected in 95 ± 3.4% of cysts incubated with S. multiaristata extract during 2 days (100 μg mL−1) and the collapse of the germinal layer was observed in 60 ± 9.3% of cysts treated with 100 μg mL−1 of the S. multiaristata extract during 4 days. The half maximal effective concentration value was 69.6 μg mL−1 and the selectivity index for E. granulosus s.s. cysts was 1.9. In this clinical efficacy study, the treatment of infected mice with the S. multiaristata extract (50 mg kg−1) caused a significant decrease in the weight of the cysts compared with the control group. These results coincided with the tissue damage observed in the cysts at the ultrastructural level. In conclusion, we observed high protoscolicidal and cysticidal effects, and significant reduction in the weight of the cysts in experimentally infected mice following treatment with the S. multiaristata extract.

Evaluation of the scolicidal activities of eugenol essential oil and its nanoemulsion against protoscoleces of hydatid cysts.

BackgroundCystic echinococcosis caused by the cestode Echinococcus granulosus remains a serious helminthic zoonosis affecting humans and animals in many endemic developing countries. Surgical intervention is the best management choice, although it is associated with high recurrence rates and serious complications. Also, the commonly used chemotherapeutics exhibited serious side effects. This study aimed to evaluate the protoscolicidal effects of eugenol (Eug) essential oil and its nanoemulsion (Eug-NE) against protoscoleces (PCs) of hydatid cysts in vitro.MethodsEug-NE was prepared and characterized. Their cytotoxicity on macrophages was assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. E. granulosus PCs were treated with various concentrations of Eug and Eug-NE at different exposure times. The viability of protoscoleces was evaluated by the eosin exclusion test, and the changes in the morphology of protoscoleces were assessed. Albendazole (ABZ) was used as a positive control.ResultsThe cellular cytotoxicity of Eug and Eug-NE on macrophage cells, in minimum and maximum concentrations (0.2 and 1 μl/mL), were nearly negligible ranging from 4.7% to 8.3% and 3.7% to 7.2%, respectively. The results showed highly significant activity of Eug-NE and Eug against hydatid PCs compared to ABZ (P < 0.05). Eug and Eug-NE have similar protoscolicidal effects at all used concentrations. Their highest scolicidal activity (100% mortality rate) was recorded at 1 μl/ml after 30 min incubation (LC50 = 0.298—LC90 = 0.521 and LC50 = 0.309—LC90 = 0.646, respectively). Both formulations showed time- and dose-dependent effects.ConclusionsThis study suggested the potent scolicidal activities of Eug and Eug-NE as promising alternative scolicidal agents. Future studies are recommended to explore the mechanism of action and treatment response in vivo and clinical settings.

The effect of piperine and curcumin on the calmodulin gene of Echinococcus granulosus protoscolices in vitro

Cystic echinococcosis (CE) has been defined as an illness that results from the larval Echinococcus granulosus stage. Treating cystic echinococcosis is fundamentally dependent upon using Albendazole that may typically result in negative side effects, which is why, more sufficient options of treatment being necessary. The re-purposing of the drugs has been considered as a beneficial method to advance the drug development. In vitro proto-scolicidal effect of the piperine and curcumin have been assessed in E. granulosus and expression of calmodulin (CaM) genes, both of them were linked to the cellular signaling activities. Based on the results, curcumin and piperine’s have dose-dependent proto-scolicidal effects to achieve the optimal efficacy that the isolated piperine 20mg/k and 2g/k curcumin showed significant anthelminthic activity compared to the standard anthelminthic drug enhancing the bioavailability of curcumin. Calmodulin, as a dynamic Ca 2+ sensor, mediates various activities of the cellular signaling from E. granulosus will be possibly involved in a significant biological function.

In vitro and Ex vivo Antiparasitic Effect of Rheum ribes L. Extract Against the Hydatid Cyst Protoscoleces.

Cystic echinococcosis is a zoonotic infection in humans and herbivorous animals occurring worldwide, which is caused by the larva stage of Echinococcus granulosus. Rhubarb (Rheum ribes L.) as an herbal medicine has various therapeutic properties such as antioxidant, anticancer, and antimicrobial. With respect to the potential of the biological activities of this plant in traditional and modern medicine, we aim to examine its protoscolicidal effects against E. granulosus protoscoleces in vitro and ex vivo. Collected protoscoleces from liver hydatid cysts of infected sheep were exposed to the different concentrations of the extract (225, 450, 900 mg/mL) for 5-60 min in vitro and ex vivo. Then, by using the eosin exclusion assay, the viability of protoscoleces was studied. R. ribes extract had a potent protoscolicidal activity in vitro ; at 450 and 900 mg/ml, it killed 56.3 and 100% of protoscoleces, respectively, after 10 min of exposure. In ex vivo assay, the extract needed more time to kill the protoscoleces than the in vitro; so that at the concentration of 900 mg/mL, all protoscoleces were killed after 15 minutes. The obtained results exhibited the potent protoscolicidal effects of R. ribes extract, particularly at the concentration of 900 mg/ml, which completely killed the parasite after <15 min of exposure. However, more supplementary studies are required to verify these findings by assessing animal models and clinical subjects.

Iranian Herbal Medicines Against Hydatid Cyst Protoscoleces: A Systematic Review.

In recent years, more attention has been focused in the practice of both crude extract of medicinal plants, and the screening plant-derived compounds as substitute scolicidal agents during hydatid cyst surgery. The present study was designed to review the protoscolicidal effects of some Iranian herbal medicines against hydatid cyst protoscoleces. English databases, including PubMed, Google Scholar, Web of Science, EBSCO, Science Direct, and Scopus were searched for publications worldwide related to protoscolicidal effects of Iranian herbal medicines without date limitation so that identify all published articles (in vitro, in vivo, clinical and case-control) have studied. Keywords included "Protoscolicidal", "Scolicidal", "Herbal medicines", "Extract", "Essential oil", "Plant", "In vitro", and "Iran". Moreover, the language of data collection was limited to English. In total, 40 papers up to 2020 were included in the present systematic review. The most studies were conducted on protoscolicidal activity of methanolic extracts (17 studies) followed by essential oils (15 studies), and aqueous extract (3 studies). The most commonly used part of herbs were leaves (21 herbs), seeds (8 herbs), and fruit (6 herbs), respectively. Moreover, the most prevalent herbal family was Lamiaceae (6 studies), followed by Apiaceae (5 studies) and Liliaceae (5 studies). Considering the high efficacy of Iranian herbal medicines against hydatid cyst protoscoleces, it can be concluded that Iranian herbal medicines have ability to consider as new alternative protoscolicidal agents during hydatid cyst surgery; however, more clinical studies are required to discover the precise protoscolicida activity of Iranian medicines in animal and human subjects.

In vitro scolicidal effects of Sideritis perfoliata extract against Echinococcus granulosus.

Cystic echinococcosis, caused by helminths within the genus Echinococcus, is mainly localised in the liver and lungs of affected hosts. Surgery has been the best choice for the treatment of hydatidosis and using effective scolicidal agents during hydatid surgery is required to prevent secondary infection. Several plant extracts have been shown to exert scolicidal efficacy. This study was designed to investigate the in vitro scolicidal activity of methanol extract of Sideritis perfoliata against the protoscolices of hydatid cysts. The protoscolices were collected from a liver of a sheep slaughtered in Adiyaman city slaughter, Turkey. Three concentrations of the aerial part extract of S perfoliata (0.1, 0.2 and 0.4mg/mL) were assessed at three different exposure periods. All tests were carried in duplicate. The viability of protoscolices was assessed by the eosin exclusion test (0.1% eosin staining). Scolicidal effect of S perfoliata extract at exposure periods of 10, 20 and 30minutes was 29.6%, 32.5% and 43.6% at the concentration of 0.1%, 37.8%, 50% and 58.1% at concentration of 0.2mg/mL, and 57.9%, 71.8% and 79.1% at the concentration of 0.4mg/mL, respectively; indicating a longer time is required to display protoscolicidal effects. LC-MS/MS analysis showed that some phenolic acids, such as fumaric acid (260.13mg/L), syringic acid (27.92mg/L) and caffeic acid (26.84mg/L), and a flavonoid, luteolin (11.23mg/L) were detected in high concentrations in the extract. This study has demonstrated that the methanol extract of S perfoliata has high scolicidal activity in vitro. However, research on the in vivo efficacy of S perfoliata extract and its potential side effects is required.

In vitro effects of tropisetron and granisetron against Echinococcus granulosus (s.s.) protoscoleces by involvement of calcineurin and calmodulin

BackgroundCystic echinococcosis (CE) is a disease caused by the larval stage of Echinococcus granulosus sensu lato (s.l.). The treatment of CE mainly relies on the use of benzimidazoles, which can commonly cause adverse side effects. Therefore, more efficient treatment options are needed. Drug repurposing is a useful approach for advancing drug development. We have evaluated the in vitro protoscolicidal effects of tropisetron and granisetron in E. granulosus sensu stricto (s.s.) and assessed the expression of the calcineurin (CaN) and calmodulin (CaM) genes, both of which have been linked to cellular signaling activities and thus are potentially promising targets for the development of drugs.MethodsProtoscoleces (PSC) of E. granulosus (s.s.) (genotype G1) obtained from sheep hepatic hydatid cysts were exposed to tropisetron and granisetron at concentrations of 50, 150 and 250 µM for various periods of time up to 10 days. Cyclosporine A (CsA) and albendazole sulfoxide were used for comparison. Changes in the morphology of PSC were investigated by light microscopy and scanning electron microscopy. Gene expression was assessed using real-time PCR at the mRNA level for E. granulosus calcineurin subunit A (Eg-CaN-A), calcineurin subunit B (Eg-CaN-B) and calmodulin (Eg-CaM) after a 24-h exposure at 50 and 250 µM, respectively.ResultsAt 150 and 250 µM, tropisetron had the highest protoscolicidal effect, whereas CsA was most effective at 50 µM. Granisetron, however, was less effective than tropisetron at all three concentrations. Examination of morphological alterations revealed that the rate at which PSC were killed increased with increasing rate of PSC evagination, as observed in PSC exposed to tropisetron. Gene expression analysis revealed that tropisetron at 50 μM significantly upregulated Eg-CaN-B and Eg-CaM expression while at 250 μM it significantly downregulated both Eg-CaN-B and Eg-CaM expressions; in comparison, granisetron decreased the expression of all three genes at both concentrations.ConclusionsTropisetron exhibited a higher efficacy than granisetron against E. granulosus (s.s.) PSC, which is probably due to the different mechanisms of action of the two drugs. The concentration-dependent effect of tropisetron on calcineurin gene expression might reflect its dual functions, which should stimulate future research into its mechanism of action and evaluation of its potential therapeutical effect in the treatment of CE.Graphical

Chemical Composition, Apoptotic Activity, and Antiparasitic Effects of Ferula macrecolea Essential Oil against Echinococcus granulosus Protoscoleces

Background: Today, the present protoscolicidals used to minimize the serious risks during hydatid cyst surgery are not completely safe and have various adverse side effects. The present study aimed to evaluate the chemical composition and apoptotic activity of Ferula macrecolea essential oil (FMEO) as well as its in vitro and ex vivo protoscolicidal effects against hydatid cyst protoscoleces. Methods: Gas chromatography/mass spectrometry (GC/MS) analysis was performed to determine the chemical composition of FMEO. Protoscoleces of hydatid cysts were collected from liver fertile hydatid cysts of infected sheep and were then treated with various concentrations of the essential oil (75, 150, and 300 µL/mL) for 5–60 min in vitro and ex vivo. Then, by using the eosin exclusion test, the viability of the protoscoleces was studied. The caspase-3-like activity of the FMEO-treated protoscoleces was also evaluated through the colorimetric protease assay Sigma Kit based on the manufacturer’s instructions. Results: According to GC/MS, the main constituents of the essential oil were terpinolene (77.72%), n-nonanal (4.47%), and linalool (4.35%), respectively. In vitro, the maximum protoscolicidal activity of FMEO was observed at the concentrations of 150 and 300 µL/mL, such that 100% of the protoscoleces were killed after 30 and 20 min of exposure, respectively. Based on the obtained findings, the results demonstrate that FMEO required a longer time to kill protoscoleces ex vivo; after 12 min of exposure to FMEO, only 13.4% of the protoscoleces remained alive. After 48 h of the treatment of protoscoleces, FMEO, in a dose-dependent manner and at doses of 75, 150, and 300 µL/mL, induced the activation of the caspase enzyme by 24.3, 35.3, and 48.3%, respectively. Conclusions: Our findings demonstrate the potent protoscolicidal effects of FMEO in vitro and ex vivo; however, further studies are required to assess the safety and the efficiency of FMEO as a promising scolicidal agent in a preclinical model and clinical setting.