Characterization and functional analysis of fatty acid binding protein from the carcinogenic liver fluke, Opisthorchis viverrini

AbstractIn the present study, the cDNA encoding FABP (Ov-FABP) was isolated from the adult stage of Opisthorchis viverrini and characterized. The Ov-FABP protein sequence (107 amino acids) was predicted to have a molecular mass of 12.26kDa and an isoelectric point (PI) of 6.82. This sequence had a high identity and similarity to Cs-FABP of the related opisthorchid Clonorchis sinensis. Multiple sequence alignment with FABPs from other parasitic flatworms and mammals showed a number of conserved amino acids including Phe34, Gly37, Glu38, Glu39,Val50, Iso62, Gly81, Ile84, Ser87 and Arg101. In addition, the structure of Ov-FABP was predicted to have eleven β-sheets and one α-helix based on the known structures for FABPs from human (hL-FABP), rat and a schistosome. Phylogenetic analysis of amino acid sequence data revealed a close relationship of Ov-FABP with Cs-FABP and hL-FABP. Reverse transcription-PCR revealed that Ov-FABP was transcribed in the egg, metacercaria, juvenile and adult stages. The soluble form of recombinant Ov-FABP (rOv-FABP) was shown to specifically bind fatty acids, including oleic acid, palmitic acid and linoleic acid, as shown for other animals. Anti-serum against rOv-FABP (produced in mice) located the protein to parenchyma, egg, sucker musculature, testes and tegument of adult O. viverrini. Taken together, the findings suggest key functional roles for Ov-FABP in development, reproduction and/or host–parasite interactions

MIGRATORY PATTERN OF OPISTHORCHIS VIVERRINI IN HAMSTERS

Abstract. The migratory pattern of Opisthorchis viverrini was studied by feeding each of 55 hamsters orally with 50 metacercariae collected from cyprinoid fish. The route of migration was followed from the gastrointestinal (GI) to biliary tracts for 3 months after infection at 11 durations: 1, 3, 6, and 12 hours, 1, 3, 7, and 14 days, 1, 2, and 3 months. After sacrifice, the stomach, small intestine, common bile duct, gallbladder, and 3 equal portions of each lobe (right, left, median, and caudal) of the liver were examined and counted for worm burden. The newly excysted worms of O. viverrini migrated from the duodenum into the common bile duct, gallbladder and proximal bile duct of the right lobe as early as 1 hour after infection. The farthest migratory distance, the distal bile ducts of all lobes, occurred with worms aged 14 days. After reaching maturity, the number of worms remaining in the extrahepatic bile ducts was lower than the intrahepatic bile ducts. The worm recovery rates in the right and left lobes were not significantly different, while those in the median and caudal lobes were significantly lower. Less than 5% of juvenile worms could migrate far to the distal part of the bile ducts, but these decreased when the worms became adult. This finding revealed the migratory pattern of O. viverrini in hamsters from the GI to the biliary system in detail, which should be useful for better understanding the host-parasite relationships along these routes of migration. detailed information about the relationship between the route of migration and the duration of infection. The objective of this study was to investigate the migratory pattern of O. viverrini from the gastrointestinal to the biliary tracts of experimentallyinfected hamsters during 3 months of infection. MATERIALS AND METHODS Opisthorchis viverrini metacercariae were obtained from naturally-infected cyprinoid fish from an endemic area in Khon Kaen Province, northeast Thailand, by pepsin-HCl digestion and infiltration. They were identified and collected under a stereomicroscope. Each of 55 male golden Syrian hamsters, aged 6-8 weeks, were infected with 50 metacercariae by intragastric intubation. Five animals were sacrificed at each period of 1, 3, 6 and 12 hours, 1, 3, 7 and 14 days, and 1, 2 and 3 months post-infection. After sacrifice, the position of ampulla of Vater and the terminal part of the common bile duct connecting to the cystic and intrahepatic bile ducts were tied to prevent the movement of worms from the normal position. The stomach, small intestine, common bile duct, gallbladder and 4 lobes (right, left, median, and caudal) of the liver were removed from the host. The cavity structures were opened and sedimented in normal saline, and the number of worms in each part was counted. Each lobe of the liver was divided into 3 portions; proximal, median, and distal, then each was separated into small pieces by 2 forceps, sedimented in normal saline and counted for worm burden per portion

Chronic Opisthorchis viverrini Infection and Associated Hepatobiliary Disease Is Associated with Iron Loaded M2-like Macrophages

Chronic Opisthorchis viverrini-induced hepatobiliary disease is associated with significant leukocyte infiltration, including activated macrophages; however, the polarization of infiltrating macrophages remains to be fully characterized. In this study, we characterized macrophage polarization and phenotype in chronic O. viverrini-induced hepatobiliary disease in humans and hamsters using gene expression and histochemical analysis. Chronic O. viverrini infection and associated hepatobiliary diseases were associated with iron loaded M2-like macrophages in both humans and hamsters. This study provides suggestive evidence that iron loaded M2-like macrophages promote hepatobiliary disease in chronic O. viverrini infection

Ultrastructural characters of the spermatozoon of the liver fluke Opisthorchis viverrini (Poirier, 1886) (Opisthorchiidae)

The present study records the ultrastructural organization of the mature spermatozoon of Opisthorchis viverrini by means of transmission electron microscopy. The spermatozoon of O. viverrini is a filiform cell, tapered at both extremities. It exhibits the characteristics of type IV spermatozoon of digeneans, namely with two axonemes of the 9+'1' trepaxonematan pattern, external ornamentation of the plasma membrane associated with cortical microtubules that are in the posterior part of the anterior region of the sperm cell, and with two mitochondria. The maximal number of cortical microtubules is in the anterior part of the spermatozoon and arranged into two bundles. Other characteristics are spine-like bodies and a posterior extremity with only the second axoneme. Ultrastructural characters of the spermatozoon of O. viverrini are compared with those of other known digeneans belonging to the Opisthorchioidea, with particular emphasis on representatives of the family Opisthorchiidae. The main differences between O. viverrini and its congener Opisthorchis felineus are the spine-like bodies (present and absent, respectively) and the posterior spermatozoon character (axoneme and nucleus, respectively)

Suppression of NAD(P)H-quinone oxidoreductase 1 enhanced the susceptibility of cholangiocarcinoma cells to chemotherapeutic agents

La lateralidad de la jugadora con el rol de base puede ser sólo una característica individual, o bien un factor influyente en el juego de su equipo, y por tanto, una herramienta a considerar por parte del equipo rival. El trabajo de investigación se centra en la necesidad de reflejar si la lateralidad de una jugadora con rol de base en baloncesto, al iniciar la acción de juego (bote o pase) influye de manera determinante en la toma de decisiones. El contexto de trabajo utilizado es la segunda categoría de baloncesto femenino nacional, Liga Femenina 2. En la observación han participado varios equipos de dicha categoría, con total de 10 partidos analizados y un total de 170 situaciones de juego analizadas (N=170). Para establecer las relaciones entre las variables se utilizó el Coeficiente de Contingencia. Los resultados muestran una relación estadísticamente significativa de la lateralidad de la base y el lado del campo en el que se empieza a jugar (X2 (1, N=170) = 4.789, p<.05)Laterality of the point-guard can be an individual characteristic, or in the other hand, an important factor in the offensive patterns of the team. In this way, it will be a useful information for the other team. This research has focused in the laterality of the point guard, and it influences in the decision making. The League studied was the second female Spanish basketball league, LF2. 10 matches were observed and 170 game situations analyzed (N=170). Crosstabs Command and the Contingency Coefficient were used to establish the relationship between variables. The results shows a significant relationship between the laterality of the point guard and the side of the field where the attack begin (X2 (1, N=170) = 4.789, p<.05)peerReviewe

Gene discovery for the carcinogenic human liver fluke, Opisthorchis viverrini

Background: Cholangiocarcinoma (CCA) - cancer of the bile ducts - is associated with chronic infection with the liver fluke, Opisthorchis viverrini. Despite being the only eukaryote that is designated as a 'class I carcinogen' by the International Agency for Research on Cancer, little is known about its genome.\ud \ud Results: Approximately 5,000 randomly selected cDNAs from the adult stage of O. viverrini were characterized and accounted for 1,932 contigs, representing ~14% of the entire transcriptome, and, presently, the largest sequence dataset for any species of liver fluke. Twenty percent of contigs were assigned GO classifications. Abundantly represented protein families included those involved in physiological functions that are essential to parasitism, such as anaerobic respiration, reproduction, detoxification, surface maintenance and feeding. GO assignments were well conserved in relation to other parasitic flukes, however, some categories were over-represented in O. viverrini, such as structural and motor proteins. An assessment of evolutionary relationships showed that O. viverrini was more similar to other parasitic (Clonorchis sinensis and Schistosoma japonicum) than to free-living (Schmidtea mediterranea) flatworms, and 105 sequences had close homologues in both parasitic species but not in S. mediterranea. A total of 164 O. viverrini contigs contained ORFs with signal sequences, many of which were platyhelminth-specific. Examples of convergent evolution between host and parasite secreted/membrane proteins were identified as were homologues of vaccine antigens from other helminths. Finally, ORFs representing secreted proteins with known roles in tumorigenesis were identified, and these might play roles in the pathogenesis of O. viverrini-induced CCA.\ud \ud Conclusion: This gene discovery effort for O. viverrini should expedite molecular studies of cholangiocarcinogenesis and accelerate research focused on developing new interventions, drugs and vaccines, to control O. viverrini and related flukes

Helicobacter pylori in Thai patients with cholangiocarcinoma and its association with biliary inflammation and proliferation

AbstractObjectivesTo investigate whether Helicobacter spp. infection and the cagA of H. pylori are associated with hepatobiliary pathology, specifically biliary inflammation, cell proliferation and cholangiocarcinoma (CCA).MethodsHelicobacter species including H. pylori, H. bilis and H. hepaticus were detected in the specimens using the polymerase chain reaction (PCR). Biliary inflammation of the liver and gallbladders was semi-quantitatively graded on hematoxylin and eosin (H&E)-stained slides. Biliary proliferation was evaluated by immunohistochemistry using the Ki-67-labelling index.ResultsHelicobacter pylori was found in 66.7%, 41.5% and 25.0% of the patients in the CCA, cholelithiasis and control groups (P < 0.05), respectively. By comparison, H. bilis was found in 14.9% and 9.4% of the patients with CCA and cholelithiasis, respectively (P > 0.05), and was absent in the control group. The cagA gene of H. pylori was detected in 36.2% and 9.1% of the patients with CCA and cholelithiasis, respectively (P < 0.05). Among patients with CCA, cell inflammation and proliferation in the liver and gallbladder were significantly higher among those DNA H. pylori positive than negative.ConclusionsThe present findings suggest that H. pylori, especially the cagA-positive strains, may be involved in the pathogenesis of hepatobiliary diseases, especially CCA through enhanced biliary cell inflammation and proliferation

Scopoletin potentiates the anti-cancer effects of cisplatin against cholangiocarcinoma cell lines

Chemotherapy with cisplatin in cholangiocarcinoma produces adverse effects and leads to resistance development by tumors. We aimed to evaluate anti-cancer effects by co-administration of cisplatin and scopoletin in cholangiocarcinoma cells. MTT assay, median effect principle, cell cycle arrest and apoptosis assay were conducted to determine anti-cancer effects. Results revealed that treatment with cisplatin and scopoletin resulted in dose-dependent reduction of cell viability for cholangiocarcinoma cells. Combination of these agents inhibited proliferation of cells significantly more than single agent either. Combination indices reflect additive cytotoxic effect, leading to >2 times dose reduction for each agent. Both the cell cycle arrest (G0/G1) and apoptosis induction underling the enhanced cytotoxicity for the combination. Besides, single agent conferred cell cycle arresting and apoptotic effects in cholangiocarcinoma cells. By contrast, non-cancer cells were less affected with combination. Our observations suggest that cisplatin and scopoletin combination may bring positive significance in cholangiocarcinoma treatment