Immunomodulatory effects of albendazole, trichlorobendazole and wortmannilactone F on Clonorchis sinensis infection

doi:10.21203/rs.3.rs-4722979/v1

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Immunomodulatory effects of albendazole, trichlorobendazole and wortmannilactone F on Clonorchis sinensis infection

https://doi.org/10.21203/rs.3.rs-4722979/v1

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Background

Albendazole, trichlorobendazole and wortmannilactone F are widely used anti-trematode drugs to treat fluke worm infections. However, their immunomodulatory effects and hepatic toxicity in Clonorchis sinensis infection treatment are unknown. This study evaluated the regulatory effects of these anti-helminthic drugs on hepatic fibrosis and immune responses in the rat model of Clonorchis sinensis infection.

Methods

Sprague-Dawley rats were infected by Clonorchis sinensis thorough gavage. Hematoxylin and Eosin (H&E) and Masson staining were performed to examine the degree of hepatic injury and fibrosis. Plasma levels of glutamic-pyruvic transaminase and albumin were analyzed by ELISA. Flow cytometry was used to detect the frequency of immune cells in the blood sample upon different drug treatments.

Results

Fecal examination showed that the administration of trichlorobendazole and wortmannilactone F could effectively eliminate the parasites, and albendazole was also able to reduce the parasite load. Albendazole had little damage to the liver tissues, while wortmannilactone and triclorobendazole could induce severe hepatic damages. Masson staining further revealed high level of hepatic fibrosis after wortmannilactone F and triclorobendazole treatment, which was accompanied by the increased CK-19 keratin expression after wortmannilactone F treatment. Albendazole administration was able to reduce the plasma level of IL-2 and decrease the proportion of CD4 + and CD8 + T lymphocytes in the blood, while the other two drugs failed to do so.

Conclusion

Wortmannilactone F and triclorobendazole showed strong anti-parasitic effect against Clonorchis sinensis infection, but induced severe liver damages. Albendazole could reduce parasite load and also showed immunomodulatory effect.

Clonorchis sinensis

albendazole

trichlorobendazole

wortmannilactone F

hepatic damage

immunomodulation

Liver fluke disease, also known as clonorchiasis, is caused by the parasitic infection of Clonorchis sinensis in the intrahepatic bile ducts [1]. The eggs of the parasite enter the intestine with bile juice, are excreted from the body by feces in the pond. If swallowed by freshwater animals, the eggs develop into cercariae and invade the muscles of freshwater fish or shrimp to form cysts [2]. When humans or other livestock animals swallow uncooked fish or shrimp with cysts, they become infected with Clonorchis sinensis. During mild to moderate infections, the pathological changes in the bile ducts and hepatic tissues are not obvious. Heavy infections can lead to the mechanical blockage of bile ducts, biliary epithelial hyperplasia and excessive intrahepatic bile duct mucus secretion [3]. During acute severe infection, there may be inflammatory changes such as immune cell infiltration and activation around the bile ducts and in the liver tissues [4]. Chronic infection can induce hepatic damages, steatosis and fibrosis, which can also contribute to the development of cholangiocarcinoma and hepatocellular carcinoma [5–8]. Clonorchiasis is also related to necrotizing hepatitis, in which immature liver flukes migrate and release toxins to damage the hepatocytes and lead to necrotic cell death [9]. Overall, the infection of Clonorchis sinensis is the primary factor in the development of a wide variety of typical hepatobiliary disorders, including persistent inflammation and perituctal fibrosis.

Albendazole is a kind of broad-spectrum anti-helminthic drug with high efficiency and low toxicity, which can be used to treat the infection of pinworm [10], tapeworm [11], whipworm [12], hookworm [13] and Strongylodes faecalis [14]. At the same time, albendazole is used as an clinical anti-helminthic drug for Clonorchis sinensis infection [15, 16]. It has been reported that albendazole therapy can promote the clearance of parasitic infections by activating the immune system and regulating peripheral blood CD4 + T cell ratio or other immune cells in helminth-infected tuberculosis and HIV patients [17–19]. Albendazole can also regulate immune responses towards Angiostrongylus cantonensis-induced meningoencephalitis and Echinococcus multilocularis infection [20, 21].

Triclobendazole, a novel imidazole anti-trematode with similar chemical structure to albendazole, has been approved by FDA for the treatment of fasciolosis [22]. Accumulating evidence has highlighted its efficacy in treating a wide range of human fascioliasis in different countries [23–25]. Perez et al. reported that triclabendazole can regulate the hepatic infiltration of CD4 + and CD8 + T cells after Fasciola hepatica infection in lambs [26]. However, its potential anti-helminthic effect on Clonorchis sinensis infection has not been reported.

Wortemannilactones are a newly discovered compound that has been shown to act as a potential inhibitor of helminth NDAH-fumarate reductase, a key enzyme functioning in the respiratory chain of the mitochondrial inner membrane [27, 28]. Currently, the effect of wortemannilactones against zoonotic worm infection has been reported in Trichinella spiralis [29, 30]. Nevertheless, the potential therapeutic ability of wortmannilactone F against Clonorchis sinensis infection and whether it can regulate the immune response triggered by Clonorchis sinensis infection remain largely unknown.

In this study, we evaluated the anti-helminthic effect of albendazole, Trichlorobendazole and wortmannilactone F against Clonorchis sinensis infection in rats. The degree of hepatic injury, lover fibrosis and plasma levels of glutamic-pyruvic transaminase and albumin were analyzed. We also compared the potential immunomodulatory effect of these drugs in the blood samples.

Animals and treatment

The liver trematode (Clonorchis sinensis) used in this study was from Guangdong Institute of Parasitic Diseases (Guangzhou, China). Sprague-Dawley rats (4 weeks old, weighing between 120 and 140g) were obtained from the Animal Center of Kunming Medical University (Kunming, China). The rats were randomly assigned into the control (without infection), Clonorchis sinensis infection, Albendazole, Trichlorobendazole and wortmannilactone F treatment groups (n = 6 in each group). Albendazole, Trichlorobendazole and wortmannilactone F were purchased from MedChemExpress (Shanghai, China). In the infection group, each rat was gavaged with 200 metacercariae, and feces test was performed daily. Dru treatment was initiated after the first microscopic examination of Clonorchis sinensis eggs. All the drugs were gavaged at 10mg/kg/day for 8 weeks. All the rats were euthanized by CO2 asphyxiation in an euthanizing chamber, which was connected to a CO2 cylinder and the flow rate was adjusted to displace 40% of the cage volume per minute. Rats were placed into the chamber for 10 min and the death was assured by subsequent cervical dislocation. The liver tissues was collected and fixed in 4% paraformaldehyde before further analysis. All the animal protocols were approved by the animal use committee of Medical Ethics Committee of Yunnan First People's Hospital.

Hematoxylin&Eosin staining

H&E staining was performed using H&E Stain Kit (ab245880, Abcam, Cambridge, UK). Deparaffinized/hydrated section was incubated in adequate Hematoxylin, Mayer’s (Lillie’s Modification) to completely cover tissue sections for 5 min. After rinsing with distilled water to remove excess stain, the section was further stained with adequate Bluing Reagent for 2 min. After washing with distilled water, the section was dehydrated in absolute alcohol, followed by staining with Eosin Y Solution for 5 min. The section was rinsed twice using absolute ethanol and then mounted to a slide, and the images were collected under an inverse microscope.

Masson staining

Masson staining was performed using the masson staining kit (Xinyu Biotechnology, Shanghai, China). Deparaffinized/hydrated section was first incubated with Mordant in Bouin’s solution at 58^o C for 15 min. The section was then washed with distilled water twice to remove the yellow color, followed by the staining with Modified Weigert’s iron hematoxylin for 5 min. After washing the section was placed in 0.5% hydrochloric acid in 70% alcohol for 15 seconds, and then wahsed in running tap water for 30 seconds and rinsed twice with distilled water. Next, the section was incubated with Biebrich scarlet-acid fuchsin solution at ambient temperature for 5 min, followed by the incubation with Phosphomolybdic-phosphotungstic acid solution for 5 min. After further staining with Aniline blue solution for 20 min, the section was incubated with Acetic acid solution for 10 seconds and rinsed twice using absolute ethanol and then mounted to a slide with synthetic resin, and the images were collected under an inverse microscope.

Analysis of glutamic-pyruvic transaminase and albumin in peripheral blood of rats

Peripheral blood was extracted from SD rats in different groups after drug treatment for 8 weeks. The level of glutamic-pyruvic transaminase and albumin in the plasma were determined using Glutamic–pyruvic transaminase 1 ELISA Kit (Xinyu Biotechnology, Shanghai, China) and Albumin ELISA kit (Abcam, Cambridge, UK) based on the supplier’s instruction.

ELISA analysis of plasma IL-2 and IL-4

The IL-2 and IL-4 levels were determined using IL-2 ELISA kit (Ruixin Biotechnology, Beijing, China) and IL-4 ELISA kit (Xinbo Biotechnology, Nanjing, China). Briefly, the serum sample was diluted 2-fold in 1X Assay Diluent D buffer. Lyophilized IL-2 or IL-4 protein standards were diluted in 1X Assay Diluent D buffer to prepare 5-fold dilution series. 100 µL standards or diluted plasma samples were added to a 96-well plate coated with corresponding capture antibody for 2 hours at room temperature. After washing, 100 µL biotin-labeled detection antibody was added for 1 hour-incubation, followed by 45-min incubation with streptavidin–HRP detection solution. After 4 washes, the chemiluminescent detection reagent (TMB solution) was added for signal development in the dark for 30 min, and the absorbance of samples and standards was measured at 450 nm using a microplate reader (Infinite 200 PRO; Tecan). The concentration of each cytokine was determined using the linear regression of the standards.

Immunohistochemistry (IHC) of CK-19

The deparaffinized/hydrated section was buffered in citric acid unmasking solution (Beyotime, Beijing, China) at 95^oC for 10 min. The section was washed three times with distill water then incubated in 3% hydrogen peroxide for 10 min. After the incubation in TBST buffer for 5 min, the section was blocked with 5% bovine serum albumin for 30 min. The section was then incubated with primary antibody against CK-19 (Ab52635, Abcam, dilution 1:500) overnight at 4^oC. Then antibody solution was removed and the section was washed three times using TBST buffer. The section further labeled with SignalStain® Boost Detection Reagent (HRP, Rabbit #8114, Cell Signaling Technologies, CA, USA) for 30 min at room temperature. Signal development was performed using the DAB substrate (ZSGB-BIO, Beijing, China) for 5 min. The section was washed in distill water two times for 5 min and then dehydrated in alcohol gradient. The Section was mounted with coverslip using the mounting medium (Beyotime, Beijing, China) before imaging.

Detection of Clonorchis sinensis eggs

The feces samples in different groups of SD rats infected with Clonorchis sinensis were collected twice per week. 100 mg sample was diluted with 1 mL PBS, and 50 µL sample was placed on the slide for observation. Eggs were examined and quantified under a light microscope at 200 X magnification.

Flow cytometry analysis of CD4 + and CD8 + T cells

The venous blood was collected in vacuum heparin sodium anticoagulant tubes from different groups of rats. 500 µL of whole blood was mixed with 2 mL of red blood cell lysis buffer (Beyotime, Beijing, China) to remove the red bloo cells for 10 min. The samples was then centrifuged at 500xg for 5 min, and the supernatant was discarded and the cell pellet was suspended in 500 µL PBS. The samples were divided into two portions, one was stained with FITC-labeled anti-CD4 antibody (ab218745, Abcam, 1:200 dilution) and the other portion was stained with CD8-FITC antibody (ab22504, Abcam, 1:200 dilution) for 25 min in the dark. After incubation, the cells were washed twice with PBS by centrifugation and the cells were re-suspended in 400 µL PBS. The percentages of CD4 + and CD8 + cells in the samples were detected by a flow cytometry (BD FACS CantoTM II Flow Cytometer).

Statistics

Statistical analyses were performed with GraphPad Prism 8.0 (Graphpad software, NY, USA). All data were expressed as the mean ± SD. The statistical difference among multiple groups were analyzed using one-way analysis of variance (ANOVA) with Tukey’s post-hoc test for pairwise comparison. P < 0.05 was was used as the threshold for statistical significance.

Effects of albendazole, trichlorobendazole and wortmannilactone F on Clonorchis sinensis infection

To assess the anti-hilminthic effect of albendazole, trichlorobendazole and wortmannilactone F on Clonorchis sinensis infection, the rats were gavaged with metacercariae. The drug administration was initiated after the detection of Clonorchis sinensis eggs in the feces. After 8 weeks, the presence of egg in the feces in different groups were determined. We found that both trichlorobendazole and wortmannilactone F eliminated Clonorchis sinensis infection since there is no egges being detected in the feces, and albendazole also reduce the egg load in the feces in comparison to the infection group (Fig. 1A and 1B).

Effects of albendazole, trichlorobendazole and wortmannilactone F on hepatic damages

To examine whether these drugs may cause toxic effect in hepatic tissues, we performed H&E staining in the hepatic tissues. Compared to the control group, Clonorchis sinensis infection led to vacuolation in the hepatic tissues. There was almost no tissue damage in infected rats after albendazole treatment (Fig. 2A). Although trichlorobendazole and wortmannilactone F treatment eliminates Clonorchis sinensis infection, the hepatic tissues exhibited higher level of damages (Fig. 2A). The examination of glutamic-pyruvic transaminase (GPT) and albumin levels in peripheral blood of rats revealed that Clonorchis sinensis infection elevated the GPT level and reduced plasma albumin level; Albendazole treatment in the infected animal reduced GPT level and increased albumin level, while trichlorobendazole and wortmannilactone F further increased GPT level and reduced albumin level in the infected animals (Fig. 2B and 2C). Since high level of GPT and reduced albumin production are sign of hepatic damages [31, 32], these data suggest that albendazole treatment could alleviate Clonorchis sinensis infection without causing toxic effect in liver, while trichlorobendazole and wortmannilactone F treatment could aggravate hepatic damages.

Effects of albendazole, trichlorobendazole and wortmannilactone F on hepatic fibrosis and CK-19 expression

We next sought to further analyze the level of hepatic fibrosis by performing Masson staining. In agreement with the hepatic damages, albendazole treatment relieved hepatic fibrosis induced by Clonorchis sinensis infection, while trichlorobendazole and wortmannilactone F treatment exacerbated hepatic fibrosis (Fig. 3A). Since CK-19 keratin is w well-recognized marker of hepatocellular carcinoma [33, 34], we wondered whether Clonorchis sinensis infection impacts on CK-19 expression. IHC analysis of CK-19 revealed that there is an elevated expression of CK-19 in hepatic tissues upon Clonorchis sinensis infection. Albendazole and trichlorobendazole treatment reduced hepatic expression of CK-19, while wortmannilactone F treatment showed an increased level of CK-19 staining in the hepatic tissues (Fig. 3B). These data imply the potential carcinogenic effect of wortmannilactone F administration in liver tissues.

Effects of albendazole, trichlorobendazole and wortmannilactone F on CD4 + cell population

Since these chemicals were reported to regulate the immune cell populations [20, 26], we then determined the ratio of CD4 + T lymphocytes in the blood samples in different groups. Clonorchis sinensis infection resulted in an expansion of CD4 + cell population, indicating the activation of immune response. Albendazole treatment dampened the activation of CD4 + cell population, while trichlorobendazole and wortmannilactone F treatment showed no effect (Fig. 4A and 4B).

Effects of albendazole, trichlorobendazole and wortmannilactone F on CD8 + cell population

Similar to CD4 + T cells, the analysis of CD8 + cells in the blood samples revealed an elevation of the ratio of CD8 + cells upon Clonorchis sinensis infection. Albendazole treatment attenuated the expansion of CD8 + cell population, while trichlorobendazole and wortmannilactone F treatment showed no significant effect (Fig. 5A and 5B).

Effects of albendazole, trichlorobendazole and wortmannilactone F on IL-2 and IL-4 level in the plasma

Lastly, we analyzed the plasma level of IL-2 (a key cytokine for T cell expansion) and IL-4 (a cytokine inducing Th2 cell differentiation) level. Consistent with the results of CD4 + and CD8 + cell ratio analysis, there was an significant increase of IL-2 level in animals infected with Clonorchis sinensis. Albendazole treatment reduced plasma IL-2 level, while trichlorobendazole and wortmannilactone F treatment showed no significant effect (Fig. 6A). On the contrary, in all experimental groups, the plasma level of IL-4 remained unchanged (Fig. 6B). The elevated ratio of IL-2/IL-4 upon Clonorchis sinensis infection and reduced ratio after albendazole treatment (Fig. 6C) were in agreement with the T cell population changes induced by Clonorchis sinensis infection and Albendazole treatment.

Clonorchis sinensis is a fish-borne parasitic worm that inhabits the bile ducts of mammals. The disease is widespread in China, South Korea and Vietnam, affecting an estimated 15 to 20 million people [35]. Clonorchis sinensis infection can trigger hepatic inflammation, which can ultimately lead to cholangitis, periductal hepatic fibrosis, liver cirrhosis, and even cholangiocarcinoma [4–7]. Although there are existing anti-helminthic drugs for treating Clonorchis sinensis infection (such as praziquantel and tribendimidine), side effects have been reported during tribendimidine treatment and the overuse of these drugs could lead to drug resistance [36]. Since there is no vaccine available for Clonorchis sinensis infection, identification of novel therapeutics is of great importance for the patients infected with drug-resistant parasites.

Albendazole, wortmannilactone F and trichlorobendazole are well-reported anti-trematode drugs to treat fluke worm infections [15, 16, 22, 29]. However, their potential anti-helminthic activity against Clonorchis sinensis infection and side effects have not been comprehensively studied. In this study, rats with Clonorchis sinensis infection were treated with albendazole, wortmannilactone F and trichlorobendazole. All three drugs showed anti-helminthic effect in Clonorchis sinensis infection, and wortmannilactone F and trichlorobendazole almost eliminate the Clonorchis sinensis infection in the fecal samples. These observation was consistent with a previous report regarding the joint effect of wortmannilactone F and G31P on killing Clonorchis sinensis worms [37]. However, wortmannilactone F and trichlorobendazole seemed to induce high level of hepatic damages, as manifested by the increased plasma level of GPT, augmented hepatic fibrosis, and reduced level of plasma albumin. There is no well-documented tissue toxicity of wortmannilactone F in the literature. Nevertheless, there is evidence of hepatotoxicity induced by trichlorobendazole [38]. There is also evidence that trichlorobendazole could trigger pyroptosis by activating caspase-3 [39]. Therefore, extra attention should be paid to the dosage being administrated in the clinical usage of wortmannilactone F and trichlorobendazole.

CK-19 keratin has been shown to be a marker for the diagnosis and identification of hepatocellular carcinoma cells [30]. Recent evidence pinpoints that Clonorchis sinensis infection could lead to the increased expression of CK-19 in hepatocyte and hepatobiliary tissues [8, 41]. In agreement with these report, we also demonstrated that the expression level of CK-19 keratin in hepatic tissue was elevated in rat with chronic Clonorchis sinensis infection. Trichlorobendazole and albendazole treatment could reduce CK-19 expression. Surprisingly, wortmannilactone F treatment led to the highest expression level of CK-19 keratin. Therefore, although wortmannilactone F has strong anti-helminthic effect against the fluke infection, there may be an increased risk of liver damage, fibrosis and hepatocellular carcinoma predisposition. Future work is required to evaluate the systemic side effect of long-term administration of wortmannilactone F in animal models.

To explore whether these the drugs affect the immune response, we detect the CD4 + and CD8 + cell ratio in the blood and analyzed the T cell expansion cytokine (IL-2) and Th2-polairzing cytokine (IL-4). We showed the increased ratio of CD8 + and CD4 + cells upon Clonorchis sinensis infection. Albendazole treatment attenuated the expansion of CD4 + and CD8 + cell population, while trichlorobendazole and wortmannilactone F treatment showed no significant effect. Clonorchis sinensis infection is capable of triggering the host immune response towards a strong Th1 immune response in the acute phase, followed by shift to a Th2 responses [42, 43]. In the rat model, the unattended infection seems to continue to boost the T cell based immunity. The effect of albendazole on attenuating T cell expansion of IL-2 production may dampen the inflammatory damage to hepatic tissues, while trichlorobendazole and wortmannilactone F treatment did not show these effect. Nevertheless, the persistent expansion of T cell population and elevated IL-2 level may also be resulted from hepatic damages directly induced by trichlorobendazole and wortmannilactone F treatment. Future work is warranted to further investigate the mechanism by which albendazole alleviates the immune responses in the treatment of Clonorchis sinensis infection.

In summary, albendazole, wortmannilactone F and trichlorobendazole all showed anti-helminthic effect against Clonorchis sinensis infection. Wortmannilactone F and trichlorobendazole could eliminate the Clonorchis sinensis infection but induced hepatic damages, which should be used with caution. A possible approach is to use a combination of these drugs in lower doses to achieve optimal anti-helminthic effect and at the same time minimize hepatotoxicity and drug resistance development.

Ethical Approval

This study was performed in line with the principles of the Declaration of Helsinki. Approval was granted by Medical Ethics Committee of Yunnan First People's Hospital (No.2017YY230)

Conflict of Interest

The authors have no relevant financial or non-financial interests to disclose

Funding

This work was supported by National Natural Science Foundation of China(No.81760110),Joint Foundation of Kunming Medical University and Yunnan Provincial Science and Technology Department (No. 2019FE001(-121)).

Author Contribution

All authors were involved in the conceptualization and design of the study. Zhengji Song provided experimental ideas and financial support. Guihua Duan, Baoyue Zhang completed the first draft of the manuscript. Xiarong Gong and Linting Xun performed the material preparation, data collection and analysis. Xueru Zhao and Yongli Li were mainly responsible for reviewing the final manuscript. All authors participated in the editorial revision of the manuscript. All authors read and approved the final manuscript.

Acknowledgment

The following funds are gratefully acknowledged for supporting this study: National Natural Science Foundation of China(No.81760110),Joint Foundation of Kunming Medical University and Yunnan Provincial Science and Technology Department (No. 2019FE001(-121)).

Availability of Data and Materials

The data generated in this study are available upon request to the corresponding author.

Saijuntha W, Sithithaworn P, Kiatsopit N, Andrews RH, Petney TN. Liver Flukes: Clonorchis and Opisthorchis. Adv Exp Med Biol. 2019;1154:139–180.
Hong ST, Fang Y. Clonorchis sinensis and clonorchiasis, an update. Parasitol Int. 2012;61(1):17–24.
Lim JU, Joo KR, Shin HP, Cha JM, Lee JI, Lim SJ. Obstructive jaundice caused by Clonorchiasis-associated duodenal papillitis: a case report. J Korean Med Sci. 2011;26(1):135–7.
Kim EM, Kwak YS, Yi MH, Kim JY, Sohn WM, Yong TS. Clonorchis sinensis antigens alter hepatic macrophage polarization in vitro and in vivo. PLoS Negl Trop Dis. 2017;11(5):e0005614.
Wang Y, Zhang X, Wang X, Zhang N, Yu Y, Gong P, Zhang X, Ma Y, Li X, Li J. Clonorchis sinensis aggravates biliary fibrosis through promoting IL-6 production via toll-like receptor 2-mediated AKT and p38 signal pathways. PLoS Negl Trop Dis. 2023;17(1):e0011062.
Kim TS, Pak JH, Kim JB, Bahk YY. Clonorchis sinensis, an oriental liver fluke, as a human biological agent of cholangiocarcinoma: a brief review. BMB Rep. 2016;49(11):590–597.
Wu YJ, He Q, Shang M, Yin YX, Li Y, Du X, Li XR. The NF-κB signalling pathway and TM7SF3 contribute to liver fibrosis caused by secreted phospholipase A2 of Clonorchis sinensis. Parasit Vectors. 2021;14(1):152.
Qi Y, Hu J, Liang J, Hu X, Ma N, Xiang B. Clonorchis sinensis infection contributes to hepatocellular carcinoma progression in rat. Parasitol Res. 2022;121(12):3403–3415.
Li W, Dong H, Huang Y, Chen T, Kong X, Sun H, Yu X, Xu J. Clonorchis sinensis Co-infection Could Affect the Disease State and Treatment Response of HBV Patients. PLoS Negl Trop Dis. 2016;10(6):e0004806.
Shimizu H, Ito S. Successful Resolution of Recurrent Vaginal Pinworm Infection With Intermittent Albendazole Administration. Pediatr Infect Dis J. 2020;39(3):254–255.
Jagota SC. Albendazole, a broad-spectrum anthelmintic, in the treatment of intestinal nematode and cestode infection: a multicenter study in 480 patients. Clin Ther. 1986;8(2):226–31.
Adams VJ, Lombard CJ, Dhansay MA, Markus MB, Fincham JE. Efficacy of albendazole against the whipworm trichuris trichiura–a randomised, controlled trial. S Afr Med J. 2004;94(12):972–6.
Anto EJ, Nugraha SE. Efficacy of Albendazole and Mebendazole With or Without Levamisole for Ascariasis and Trichuriasis. Open Access Maced J Med Sci. 2019;7(8):1299–1302.
Schulz JD, Neodo A, Coulibaly JT, Keiser J. Pharmacokinetics of Albendazole, Albendazole Sulfoxide, and Albendazole Sulfone Determined from Plasma, Blood, Dried-Blood Spots, and Mitra Samples of Hookworm-Infected Adolescents. Antimicrob Agents Chemother. 2019;63(4):e02489-18.
Xiao SH, Xue J, Xu LL, Zhang YN, Qiang HQ. Comparative effect of mebendazole, albendazole, tribendimidine, and praziquantel in treatment of rats infected with Clonorchis sinensis. Parasitol Res. 2011;108(3):723–30.
Ying-Yan Z, Ting-Jun X, Man W, Yue-Yi F, Le L. [Prevalence of Clonorchis sinensis infection and effect of albendazole treatment among residents in two communities of Zhongshan City]. Zhongguo Xue Xi Chong Bing Fang Zhi Za Zhi. 2018;30(2):219–221.
Walson JL, Otieno PA, Mbuchi M, Richardson BA, Lohman-Payne B, Macharia SW, Overbaugh J, Berkley J, Sanders EJ, Chung MH, John-Stewart GC. Albendazole treatment of HIV-1 and helminth co-infection: a randomized, double-blind, placebo-controlled trial. AIDS. 2008;22(13):1601–9.
Abate E, Elias D, Getachew A, Alemu S, Diro E, Britton S, Aseffa A, Stendahl O, Schön T. Effects of albendazole on the clinical outcome and immunological responses in helminth co-infected tuberculosis patients: a double blind randomised clinical trial. Int J Parasitol. 2015;45(2–3):133–40.
Chachage M, Podola L, Clowes P, Nsojo A, Bauer A, Mgaya O, Kowour D, Froeschl G, Maboko L, Hoelscher M, Saathoff E, Geldmacher C. Helminth-associated systemic immune activation and HIV co-receptor expression: response to albendazole/praziquantel treatment. PLoS Negl Trop Dis. 2014;8(3):e2755.
Lam HYP, Liang TR, Jiang SJ, Peng SY. Albendazole-Schisandrin B Co-Therapy on Angiostrongylus cantonensis-Induced Meningoencephalitis in Mice. Biomolecules. 2020;10(7):1001.
Ricken FJ, Nell J, Grüner B, Schmidberger J, Kaltenbach T, Kratzer W, Hillenbrand A, Henne-Bruns D, Deplazes P, Moller P, Kern P, Barth TFE. Albendazole increases the inflammatory response and the amount of Em2-positive small particles of Echinococcus multilocularis (spems) in human hepatic alveolar echinococcosis lesions. PLoS Negl Trop Dis. 2017;11(5):e0005636.
Gandhi P, Schmitt EK, Chen CW, Samantray S, Venishetty VK, Hughes D. Triclabendazole in the treatment of human fascioliasis: a review. Trans R Soc Trop Med Hyg. 2019;113(12):797–804.
Morales ML, Tanabe MB, White AC Jr, Lopez M, Bascope R, Cabada MM. Triclabendazole Treatment Failure for Fasciola hepatica Infection among Preschool and School-Age Children, Cusco, Peru. Emerg Infect Dis. 2021;27(7):1850–1857.
Ramadan HK, Hassan WA, Elossily NA, Ahmad AA, Mohamed AA, Abd-Elkader AS, Abdelsalam EMN, Khojah HMJ. Evaluation of nitazoxanide treatment following triclabendazole failure in an outbreak of human fascioliasis in Upper Egypt. PLoS Negl Trop Dis. 2019;13(9):e0007779.
Collett CF, Morphew RM, Timson D, Phillips HC, Brophy PM. Pilot Evaluation of Two Fasciola hepatica Biomarkers for Supporting Triclabendazole (TCBZ) Efficacy Diagnostics. Molecules. 2020;25(15):3477.
Pérez J, Ortega J, Bravo A, Díez-Baños P, Morrondo P, Moreno T, Martínez-Moreno A. Phenotype of hepatic infiltrates and hepatic lymph nodes of lambs primarily and challenge infected with Fasciola hepatica, with and without triclabendazole treatment. Vet Res. 2005 Jan-Feb;36(1):1–12.
Liu WC, Wang YY, Liu JH, Ke AB, Zheng ZH, Lu XH, Luan YS, Xiu ZL, Dong YS. Wortmannilactones I-L, new NADH-fumarate reductase inhibitors, induced by adding suberoylanilide hydroxamic acid to the culture medium of Talaromyces wortmannii. Bioorg Med Chem Lett. 2016;26(21):5328–5333.
Dong Y, Yang J, Zhang H, Lin J, Ren X, Liu M, Lu X, He J. Wortmannilactones A-D, 22-membered triene macrolides from Talaromyces wortmannii. J Nat Prod. 2006;69(1):128–30.
Qin Y, Ren Y, Yi C, Chileshe N, Chen Y, Zheng L, Dai X, Dong Y, Cui Y. Effect of Wortmannilactone F on Trichinella spiralis Enteral in Mice. Vector Borne Zoonotic Dis. 2020;20(3):205–211.
Ren Y, Qin Y, Zhang X, Zheng L, Dai X, Wu H, Dong Y, Cui Y. Killing the muscular larvae of Trichinella spiralis and the anti-fibrotic effect of the combination of Wortmannilatone F and recombinant G31P in a murine model of trichinellosis. Biomed Pharmacother. 2018;108:934–940.
Rasyid SA, Armayani, Yuniati, Lio TMP. Analysis of serum glutamic pyruvic transaminase and serum glutamic oxaloacetic transaminase levels in tuberculosis patients who are undergoing oat treatment in Kendari City General Hospital, Kota Kendari, Indonesia. Infect Dis Rep. 2020;12(Suppl 1):8737.
Jagdish RK, Maras JS, Sarin SK. Albumin in Advanced Liver Diseases: The Good and Bad of a Drug! Hepatology. 2021;74(5):2848–2862.
Zhuo JY, Lu D, Tan WY, Zheng SS, Shen YQ, Xu X. CK19-positive Hepatocellular Carcinoma is a Characteristic Subtype. J Cancer. 2020;11(17):5069–5077.
Uenishi T, Kubo S, Yamamoto T, Shuto T, Ogawa M, Tanaka H, Tanaka S, Kaneda K, Hirohashi K. Cytokeratin 19 expression in hepatocellular carcinoma predicts early postoperative recurrence. Cancer Sci. 2003;94(10):851–7.
Na BK, Pak JH, Hong SJ. Clonorchis sinensis and clonorchiasis. Acta Trop. 2020;203:105309.
Koda S, Zhu XQ, Zheng KY, Yan C. Molecular Mechanisms of Clonorchis sinensis-Host Interactions and Implications for Vaccine Development. Front Cell Dev Biol. 2022;9:781768.
Qin Y, Zhang Z, Zheng L, Wu H, Dai X, Dong Y, Cui Y, Ren Y. Impact of Wortmannilactone F and G31P on Clonorchis Sinensis-infected mice. Int Immunopharmacol. 2020;85:106512.
Gandhi P, Schmitt EK, Chen CW, Samantray S, Venishetty VK, Hughes D. Triclabendazole in the treatment of human fascioliasis: a review. Trans R Soc Trop Med Hyg. 2019;113(12):797–804.
Yan L, Liu Y, Ma XF, Hou D, Zhang YH, Sun Y, Shi SS, Forouzanfar T, Lin HY, Fan J, Wu G. Triclabendazole Induces Pyroptosis by Activating Caspase-3 to Cleave GSDME in Breast Cancer Cells. Front Pharmacol. 2021;12:670081.
Sun DW, Zhang YY, Sun XD, et al. Prognostic value of cytokeratin 19 in hepatocellular carcinoma: A meta-analysis. Clin Chim Acta. 2015;448:161–169.
Liu JX, Liu M, Yu GZ, Zhao QQ, Wang JL, Sun YH, Koda S, Zhang B, Yu Q, Yan C, Tang RX, Jiang ZH, Zheng KY. Clonorchis sinensis infection induces hepatobiliary injury via disturbing sphingolipid metabolism and activating sphingosine 1-phosphate receptor 2. Front Cell Infect Microbiol. 2022;12:1011378.
Yan C, Li XY, Li B, Zhang BB, Xu JT, Hua H, Yu Q, Liu ZZ, Fu LL, Tang RX, Zheng KY. Expression of Toll-like receptor (TLR) 2 and TLR4 in the livers of mice infected by Clonorchis sinensis. J Infect Dev Ctries. 2015;9(10):1147–55.
Yan C, Zhang BB, Hua H, Li B, Zhang B, Yu Q, Li XY, Liu Y, Pan W, Liu XY, Tang RX, Zheng KY. The Dynamics of Treg/Th17 and the Imbalance of Treg/Th17 in Clonorchis sinensis-Infected Mice. PLoS One. 2015;10(11):e0143217.

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Immunomodulatory effects of albendazole, trichlorobendazole and wortmannilactone F on Clonorchis sinensis infection

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Hematoxylin&Eosin staining

Masson staining

Analysis of glutamic-pyruvic transaminase and albumin in peripheral blood of rats

ELISA analysis of plasma IL-2 and IL-4

Immunohistochemistry (IHC) of CK-19

Flow cytometry analysis of CD4 + and CD8 + T cells

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