Apoptotic and Anti-metastatic Effects of Atractylodes lancea (Thunb.) DC. in a Hamster Model of Cholangiocarcinoma.

OBJECTIVES: Cholangiocarcinoma (CCA) is a highly aggressive tumor with a greater risk of distant metastasis. The promising anti-CCA activity and safety profile of Atractylodes lancea (AL) have previously been reported in a series of in vitro, in vivo and clinical studies. The present study investigated the effect of AL extract on apoptosis and metastasis signaling pathways in the Opisthorchis viverrini/dimethylnitrosamine (OV/DMN)-induced CCA hamster model. MATERIALS AND METHODS: Hamster liver tissues were obtained from the four groups (n = 5 per group), i.e., (i) 5-FU treated CCA (40 µg/mL); (ii) CCA; (iii) AL-treated CCA (5,000 mg/kg), and (iv) normal hamsters. Total RNA was isolated, and the expression levels of apoptosis-related and metastasis-related genes were determined by qRT-PCR analysis. RESULTS: The expression levels of p16, caspase-3, caspase-8, caspase-9, Apaf-1, p53 and Eef1a1 were downregulated, while that of the remaining genes were upregulated in CCA hamsters compared with normal hamsters. AL treatment increased the expression of p16, caspase-9, caspase-3, Apaf-1, p53 and E-cadherin and decreased the expression of cyclin D1, cdk4, Bax, Akt/PKB, Bcl-2, Mfge-8, Lass4, S100A6, TGF-ß, Smad-2, Smad-3, Smad-4, MMP-9, and N-cadherin. The expression of Eef1a1 was unchanged. CONCLUSION: The anti-CCA activity of AL in OV/DMN-induced CCA hamsters could be due to the induction of cell cycle arrest at the G1 phase and activation of the apoptosis pathway, resulting in cancer cell death. The activation of the apoptosis pathway mainly involved the intrinsic pathway (activation of caspase-3 and caspase-9 through p53 and Mfge-8 modulation and downregulation of anti-apoptotic genes Akt and Bcl-2). In addition, AL could also inhibit the canonical TGF-ß signaling pathway, MMP-9 and N-cadherin to suppress tumor metastasis.

Discovering proteins for chemoprevention and chemotherapy by curcumin in liver fluke infection-induced bile duct cancer.

Modulation or prevention of protein changes during the cholangiocarcinoma (CCA) process induced by Opisthorchis viverrini (Ov) infection may become a key strategy for prevention and treatment of CCA. Monitoring of such changes could lead to discovery of protein targets for CCA treatment. Curcumin exerts anti-inflammatory and anti-CCA activities partly through its protein-modulatory ability. To support the potential use of curcumin and to discover novel target molecules for CCA treatment, we used a quantitative proteomic approach to investigate the effects of curcumin on protein changes in an Ov-induced CCA-harboring hamster model. Isobaric labelling and tandem mass spectrometry were used to compare the protein expression profiles of liver tissues from CCA hamsters with or without curcumin dietary supplementation. Among the dysregulated proteins, five were upregulated in liver tissues of CCA hamsters but markedly downregulated in the CCA hamsters supplemented with curcumin: S100A6, lumican, plastin-2, 14-3-3 zeta/delta and vimentin. Western blot and immunohistochemical analyses also showed similar expression patterns of these proteins in liver tissues of hamsters in the CCA and CCA + curcumin groups. Proteins such as clusterin and S100A10, involved in the NF-κB signaling pathway, an important signaling cascade involved in CCA genesis, were also upregulated in CCA hamsters and were then suppressed by curcumin treatment. Taken together, our results demonstrate the important changes in the proteome during the genesis of O. viverrini-induced CCA and provide an insight into the possible protein targets for prevention and treatment of this cancer.

Nanoencapsulated curcumin and praziquantel treatment reduces periductal fibrosis and attenuates bile canalicular abnormalities in Opisthorchis viverrini-infected hamsters.

This study investigated the effects of nanoencapsulated curcumin (NEC) and praziquantel (PZQ) treatment on the resolution of periductal fibrosis (PDF) and bile canalicular (BC) abnormalities in Opisthorchis viverrini infected hamsters. Chronic O. viverrini infection (OV) was initially treated with either PZQ (OP) and subsequently treated with NEC (OP+NEC), curcumin (OP+Cur) or unloaded carriers (OP+carrier) daily for one month. OP+NEC treatment reduced the PDF by suppression of fibrotic markers (hydroxyproline content, α-SMA, CTGF, fibronectin, collagen I and III), cytokines (TGF-ß and TNF-α) and TIMP-1, 2, 3 expression and upregulation of MMP-7, 13 genes. Higher activity of NEC in reducing fibrosis compared to curcumin was also demonstrated in in vitro studies. Moreover, OP+NEC also prevented BC abnormalities and upregulated several genes involved in bile acid metabolism. These results demonstrate that NEC and PZQ treatment reduces PDF and attenuates BC defect in experimental opisthorchiasis. From the Clinical Editor: Infection by Opisthorchis viverrini leads to liver fibrosis and affects population in SE Asia. Currently, praziquantel (PZQ) is the drug of choice but this drug has significant side effects. In this study, the authors combined curcumin (NEC) and praziquantel in a nanocarrier to test the anti-oxidative effect of curcumin in an animal model. The encouraging results may pave a way for better treatment in the future.

A combination of praziquantel and the traditional medicinal plant Thunbergia laurifolia on Opisthorchis viverrini infection and cholangiocarcinoma in a hamster model.

Cholangiocarcinoma (CCA) associated by Opisthorchis viverrini remains a health problem in Southeast Asia including Thailand. At present, there is still no efficient treatment for CCA. Thunbergia laurifolia is a traditionally used medicinal plant; its aqueous leave extract possesses the antioxidant activity and anti-inflammatory on hamster opisthorchiasis had been reported previously. Here, we demonstrate the combined effects of the T. laurifolia extract plus antihelminthic drug, praziquantel (PZ) on hamsters with opisthorchiasis and hamsters with opisthorchiasis related-cholangiocarcinoma through light microscopic observations of histopathological changes, as well as liver function tests for alanine transaminase (ALT) and alkaline phosphatase, and kidney function tests for blood urea nitrogen and creatinine. Results showed T. laurifolia extract combined with praziquantel reduced inflammatory cell aggregation and inhibiting CCA development, which were correlated to the serum ALT level. These present studies suggest that administration of T. laurifolia after praziquantel treatment clearly improve the hepatobiliary system and could reduce the risk of subsequent CCA development in human.

Anti-inflammatory, antioxidant and hepatoprotective effects of Thunbergia laurifolia Linn. on experimental opisthorchiasis.

Thunbergia laurifolia Linn (Rang Chuet) possesses antioxidant and anti-inflammatory properties as well as anticancer activities. The aim of the present study was to evaluate the efficacy of T. laurifolia in reducing inflammation from pathological changes in Syrian hamsters infected with the human liver fluke Opisthorchis viverrini. Hamster groups were also administered N-nitrosodimethylamine (NDMA) and treated with T. laurifolia. Light microscopic observation of histopathological changes, liver function tests for alanine transaminase (ALT) and alkaline phosphatase (ALP) and kidney function tests for blood urea nitrogen (BUN) and creatinine were performed. Antioxidant effects of both fresh and dried Rang Chuet solutions were observed. Analysis of the histopathological changes showed anti-inflammatory properties, both in the case of O. viverrini infection or with NDMA administration, by reducing the aggregation of inflammatory cells surrounding the hepatic bile ducts as indicated by normal serum ALT, ALP, BUN and creatinine levels in treated Syrian hamsters. The present study found that fresh and dried Rang Chuet solutions clearly reduced the inflammatory cells in both O. viverrini-infected and NDMA-administered groups and was correlated with the total antioxidant capacity. These findings suggest that T. laurifolia possesses antioxidant and anti-inflammatory properties and that its application may be useful for prevention of the inflammatory process, one of the risk factors of O. viverrini-associated cholangiocarcinoma (CCA).

Effect of fingerroot on reducing inflammatory cells in hamster infected with Opisthorchis viverrini and N-nitrosodimethylamine administration.

The active compound in fingerroot is effective in the treatment of many inflammatory diseases. The aim of our present study was to evaluate the efficacy of fingerroot on reducing histopathological changes in hamsters that were infected with the liver fluke Opisthorchis viverrini or were administered N-nitrosodimethylamine (NDMA), and then treated with fingerroot. Light microscopic observation and liver function tests for alanine transaminase (ALT), alkaline phosphatase (ALP), and direct bilirubin were investigated. The results of histopathological changes show that fingerroot has anti-inflammatory properties--in the case of N-nitrosodimethylamine administration on day 30--by reducing the aggregation of inflammatory cells surrounding the hepatic bile ducts, which correlates with a decreased serum ALT level. The decrease of direct bilirubin levels in hamsters treated with fingerroot suggests that fingerroot may enhance biliary contraction. The present study found that fingerroot clearly reduces the inflammatory cells in hamsters that were administered NDMA, but not in the case of O. viverrini infection. This finding suggests that fingerroot has anti-inflammatory property, but not in the case of hamster opisthorchiasis.

Turmeric reduces inflammatory cells in hamster opisthorchiasis.

The curcumin compound from turmeric is effective in the treatment of many inflammatory diseases. The aim of our present study was to evaluate the efficacy of turmeric on reducing the histopathological changes of hamster opisthorchiasis. Hamsters were infected with Opisthorchis viverrini and then administered turmeric. Using light microscopic observation, liver function tests for alanine transaminase (ALT), alkaline phosphatase, and direct bilirubin were investigated. The resulting histopathological changes show that turmeric has anti-inflammatory properties--during both N-nitrosodimethylamine administration and O. viverrini infection--by reducing the aggregation of inflammatory cells surrounding the hepatic bile ducts, which correlates with a decreased serum ALT level. The decrease in direct bilirubin levels in the hamsters treated with turmeric suggests that turmeric may enhance biliary contraction. The present study found that turmeric clearly reduces the inflammatory cells in hamster opisthorchiasis at an early stage. This finding may be connected with a reduction in the risk factors of cholangiocarcinoma development.

[Morphological criteria of chronic opisthorchiasis treatment].

Morphological examinations of the liver in chronic opisthorchiasis have both diagnostic and prognostic importance. Significant lowering of the number of cells of the portal tract inflammatory infiltrate per area unit (1 mmI), decreased number of neutrophilic and eosinophilic leukocytes, elevated count of monocytes are not only morphological markers of successful dehelminthization but they also reflect objective improvement of the liver state this being an absolute criterion of effective therapy.

[The body's responsiveness and the efficiency of antihomotoxic therapy for chronic opisthorchiasis: an aggregate clinical estimate].

The authors have proposed an extended clinical evaluation system that calls for a study of the mechanisms of development and relationships of any symptoms and diseases, including those unrelated to opisthorchiasis. Forty patients who had been diagnosed as having chronic opisthorchiasis and had a long history were examined. There was a drastic reduction in the body's susceptibility to acute inflammations during progressive chronic degenerative diseases just at the age of 17-30 years, which is indicative of systemic responsiveness abnormalities. The practical application of the proposed aggregate clinical estimate system makes it possible to apply a holistic approach to analyzing the patients' health status, to substantially upgrade the quality of clinical diagnosis, and extend the capabilities of a differential approach to evaluating the severity of disease and planning therapy.

Modulation of dihydroxy-di-n-propylnitrosamine-induced liver lesion development in Opisthorchis-infected Syrian hamsters by praziquantel treatment in association with butylated hydroxyanisole or dehydroepiandrosterone administration.

The effects of praziquantel coupled with dehydroepiandrosterone (DHEA) or butylated hydroxyanisole (BHA) administration 16 weeks subsequent to dihydroxy-di-n-propylnitrosamine (DHPN) treatment and infection with Opisthorchis viverrini (OV) on lesion development in the liver of Syrian hamsters were investigated. Animals were given 80 OV metacercariae and then two i.p. injections of DHPN (500 mg/kg body weight) 4 and 5 weeks thereafter. At week 16, groups received praziquantel (250 mg/kg, i.g.) and were placed on normal diet or diet supplemented with BHA (1%) or DHEA (0.6%) until they were killed at week 24. Histopathological assessment revealed that, whereas antihelminthic treatment alone resulted in a clear reduction in hepatocellular lesion development, effects on cholangiocellular lesions were equivocal. BHA and DHEA, in contrast, were both associated with a significant reduction in frequency of cholangiofibrosis and cholangiocellular carcinoma. The former chemical, however, increased the numbers of liver nodules while the hormone brought about a decrease as well as a shift in the phenotype of the lesions. The results thus indicate that although cholangiocellular lesion development may, unlike generation of hepatocellular nodules, be to a certain extent independent of the continued presence of parasite, it can be influenced by exogenous treatments.

[Pathomorphology of the reparative processes of the liver following praziquantel treatment of experimental opisthorchiasis]. / Patomorfologiia vosstanovitel'nykh protsessov v pecheni posle lecheniia éksperimental'nogo opistorkhoza prazikvantelem.

The authors attempted to create a model of reparative processes in the liver in the residual period of experimental opisthorchiasis. The experiment was performed in 150 Opisthorchis invaded Syrian hamsters treated with praziquantel in a single dose of 400 mg/kg body weight on days 7-180 of postinfection. The studies of morphological, histochemical and morphometric alterations in the animal liver allowed one to distinguish the stages of additional sensitization, early and late rehabilitative changes and the stage of residual changes. An inverse relationship was found between the duration of last three stages with the invasion duration prior to treatment and with increased duration of the first stage of the disease in early therapy.