Ibuprofen and diclofenac differentially affect cell viability, apoptosis and morphology changes of human cholangiocarcinoma cell lines.

Objectives: Cholangiocarcinoma is a malignant biliary epithelial duct neoplasm caused by chronic inflammation after liver fluke infection. It is a major public health concern in the Greater Mekong sub-region in northeast Thailand. Herein, the effects of the non-steroidal anti-inflammatory drugs (NSAIDs) ibuprofen and diclofenac on the cell proliferation activity of the human cholangiocarcinoma cell lines KKU-M139 and KKU-213B were studied. Methods: Cell viability was assessed with MTT assays. Inverted phase-contrast light microscopy, scanning electron microscopy and transmission electron microscopy were used to investigate the cells' morphological alterations. Caspase 3/7 and Annexin V/PI were detected with a multimode microplate reader. Results: Ibuprofen and diclofenac decreased viability in both cell lines, and ibuprofen-treated cells exhibited reversible cell injury. In both KKU-M139 and KKU-213B cell lines, the diclofenac-treated cells had the greatest injury. The cells exhibited features of irreversible cell injury. In addition, caspase 3/7 and Annexin V/PI detection revealed early cell apoptotic characteristics. Conclusion: These findings suggest that NSAIDs may potentially suppress cell viability. Ibuprofen and diclofenac both induced morphological changes and apoptosis.

The Effects of Ibuprofen, Naproxen and Diclofenac on cell Apoptosis, Cell Proliferation and Histology Changes in Human Cholangiocarcinoma Cell Lines.

OBJECTIVE: To examine the effects of ibuprofen, naproxen and diclofenac, non-steroidal anti-inflammatory drugs (NSAIDs) on cell proliferation activity of the human CCA cell lines. METHODS: KKU-M139 and KKU-213B cell lines were used in this study. The cell viability was assessed by the MTT assay. Lipid synthesis determined by Oil red O staining and colorimetric assay. An inverted phase-contrast light microscope was used to investigate the histological change of the cells. Caspases 3/7 activity and AnnexinV/PI were used to assess apoptosis by multiple microplate reader. RESULTS: The results showed that ibuprofen, naproxen and diclofenac suppressed the viability of the KKU-M139 and KKU-213B cells in a dose-dependent manner, as measured especially diclofenac. However, these three NSAIDs slightly decreased lipid synthesis determined by Oil red O staining and colorimetric assay. The histological change observations showed the shrinking cell and become star-shaped in high dose treated groups. Interestingly, these NSAIDs exhibited in both of KKU-M139 and KKU-213B cell lines, the diclofenac-treated cells had the most injury cells. The cells exhibited cell injury features. In addition, the detection of caspase 3/7 and AnnexinV/PI in this investigation revealed early cell apoptotic characteristics. CONCLUSION: These finding suggest that ibuprofen, naproxen and diclofenac suppress cell viability. The results reveal that ibuprofen, naproxen and diclofenac, which induce the histological change and apoptosis. This study indicates that these NSAIDs may be used as an anti-proliferation agent for the treatment of CCA in the future.

Synergistic Toxicity of Plant Essential Oils Combined with Pyrethroid Insecticides against Blow Flies and the House Fly.

Blow flies (Diptera: Calliphoridae) and the house fly (Diptera: Muscidae) are filth flies of medical importance, and control of their population is needed. As insecticide applications have resulted in fly resistance, and the exploration of plant essential oils (EOs) has increased against filth flies, this study assessed the combination of EOs with pyrethoids to enhance toxic efficacy. The EOs of five effective plants were screened initially against the house fly (Musca domestica L.). Their chemical constituent was performed using gas chromatography-mass spectrometry (GC-MS) analysis. The main components of Boesenbergia rotunda (Zingiberaceae) rhizome, Curcuma longa (Zingiberaceae) rhizome, Citrus hystrix (Rutaceae) fruit peel, Ocimum gratissimum (Lamiaceae) seed, and Zanthoxylum limonella (Rutaceae) fruit were δ-3-caren (35.25%), ß-turmerone (51.68%), ß-pinene (26.56%), p-cumic aldehyde (58.21%), and dipentene (60.22%), respectively. The screening test revealed that the three most effective plant EOs were from B. rotunda, C. longa and O. gratissimum, which were selected for the combination with two pyrethroid insecticides (permethrin and deltamethrin), in order to enhance their synergistic efficacy against the blow flies, Chrysomya megacephala Fabricius, Chrysomya rufifacies Macquart, and Lucilia cuprina Wiedemann, and the house fly. Synergistic action was presented in almost all of the flies tested with permenthrin/deltamethrin/EOs mixtures. It was interesting that the combination of deltamethrin with three EOs showed a synergistic effect on all of the tested flies. However, an antagonistic effect was observed in C. megacephala and M. domestica treated with permethrin-B. rotunda and C. megacephala treated with permethrin-O. gratissimum. The LD50 of insecticides decreased when combined with plant EOs. This alternative strategy will be helpful in developing a formula for effective fly control management.

Trichinosis: epidemiology in Thailand.

Trichinosis is one of the most common food-borne parasitic zoonoses in Thailand and many outbreaks are reported each year. This paper reviews the history, species, and epidemiology of the disease and food habits of the people with an emphasis on the north, northeast, central and south regions of Thailand. The earliest record of trichinosis in Thailand was in 1962 in the Mae Sariang District, Mae Hong Son Province. Since then, about 130 outbreaks have been reported involving 7392 patients and 97 deaths (1962-2005). The highest number of cases, 557, was recorded in 1983. The annual epidemiological surveillance reports of the Bureau of Epidemiology, Department of Disease Control, Ministry of Public Health, Thailand, show that trichinosis cases increased from 61 in 1997 to 351 in 1998. In contrast to these figures, the number of reported cases decreased to 16 in 1999 and 128 cases in 2000. There was no record of trichinosis in 2001, but then the figures for 2002, 2003 and 2004 were 289, 126 and 212 respectively. The infected patients were mostly in the 35-44 years age group and the disease occurred more frequently in men than women at a ratio of 1.7-2.0:1. There were 84 reported cases of trichinosis in Chiang Rai, Nan, Chiang Mai, Si Sa ket, Nakhon Phanom, Kalasin, Nakhon Ratchasima, Nakhon Nayok, Nakhon Pathom and Surat Thani, provinces located in different parts of Thailand in 2005. The outbreaks were more common in the northern areas, especially in rural areas where people ate raw or under-cooked pork and/or wild animals. This indicates the need for health education programs to prevent and control trichinosis as soon as possible in the high-risk areas.

Effects of Asiatic Acid on Spatial Working Memory and Cell Proliferation in the Adult Rat Hippocampus.

Asiatic acid is a pentacyclic triterpene from Centella asiatica. Previous studies have reported that asiatic acid exhibits antioxidant and neuroprotective activities in cell culture. It also prevents memory deficits in animal models. The objective of this study was to investigate the relationship between spatial working memory and changes in cell proliferation within the hippocampus after administration of asiatic acid to male Spraque-Dawley rats. Control rats received vehicle (propylene glycol) while treated rats received asiatic acid (30 mg/kg) orally for 14 or 28 days. Spatial memory was determined using the novel object location (NOL) test. In animals administered asiatic acid for both 14 and 28 days, the number of Ki-67 positive cells in the subgranular zone of the dentate gyrus was significantly higher than in control animals. This was associated with a significant increase in their ability to discriminate between novel and familiar object locations in a novel object discrimination task, a hippocampus-dependent spatial memory test. Administration of asiatic acid also significantly increased doublecortin (DCX) and Notch1 protein levels in the hippocampus. These findings demonstrate that asiatic acid treatment may be a potent cognitive enhancer which improves hippocampal-dependent spatial memory, likely by increasing hippocampal neurogenesis.

Overexpression of matrix metalloproteinase 11 in Thai prostatic adenocarcinoma is associated with poor survival.

BACKGROUND: The incidence of prostate cancer, one of the most common cancers in elderly men, is increasing annually in Thailand. Matrix metalloproteinase 11 (MMP-11) is a member of the extracellular matrix metalloproteases which has been associated with human tumor progression and clinical outcome. AIM: To quantify MMP-11 expression in prostatic adenocarcinoma tissues and to determine whether its overexpression correlates with survival outcome, and to assess its potential as a new prognostic marker. MATERIALS AND METHODS: Expression of MMP-11 was analyzed using immunohistochemistry in 103 Thai patients with prostatic adenocarcinoma. Overall survival was analyzed using Kaplan-Meier methods and Cox regression models. RESULTS: Immunoreactivity of MMP-11 was seen in the stroma of prostatic adenocarcinoma tissue samples, high expression being significantly correlated with poor differentiation in Gleason grading, pathologic tumor stage 4 (pT4), and positive-bone metastasis (p<0.05), but not age and prostatic-specific antigen (PSA) level. Patients with high levels of MMP-11 expression demonstrated significantly shorter survival (p<0.001) when compared to those with low levels. Multivariate analysis showed that MMP-11 expression and pT stage were related with survival in prostatic adenocarcinoma [hazard ratio (HR)=0.448, 95% confidence interval (95%CI)=0.212-0.946, HR=0.333, 95%CI=0.15-0.74, respectively]. CONCLUSIONS: Expression of MMP-11 is significantly associated with survival in prostatic adenocarcinoma. High levels may potentially be used for prediction of a poor prognosis.

Organization of the nervous system in Opisthorchis viverrini investigated by histochemical and immunohistochemical study.

The structure and organization of the nervous system has been documented for various helminth parasites. However, the neuroanatomy of the carcinogenic liver fluke, Opisthorchis viverrini has not been described. This study therefore investigated the organization of the nervous system of this fluke using cholinesterase activity, aminergic and peptidergic (FMRFamide-like peptides) immunostaining to tag major neural elements. The nervous system, as detected by acetylcholinesterase (AchE) reaction, was similar in newly excysted metacercariae, migrating juveniles and adult parasites. In these stages, there were three pairs (dorsal, ventral and lateral) of bilaterally symmetrical longitudinal nerve cords and two cerebral ganglia. The ventral nerve cords and the cerebral ganglia were well-developed and exhibited strong AchE reactivity, as well as aminergic and FMRFamide-like immunoreactivity. Numerous immunoreactive nerve cell bodies were observed around the inner surface of the ventral sucker. Fine FMRFamide-like peptides immunopositive nerve fiber was rarely observed. Overall, the organization of the nervous system of O. viverrini is similar to other trematodes.