Kanlayanee Sawanyawisuth

Treatment of angiostrongyliasis

Angiostrongyliasis, caused by Angiostrongylus cantonensis, is endemic in northeastern Thailand and southern and eastern Taiwan and is also reported throughout the world. Humans get infected by eating raw freshwater snails or other paratenic hosts. The three main clinical forms of angiostrongyliasis are: eosinophilic meningitis (EoM), eosinophilic encephalitis (EoE) and ocular angiostrongyliasis. EoM, the most common form, causes acute severe headache, and corticosteroid is the cornerstone treatment. EoE is rare but fatal and has no effective treatment. The clinical presentations are coma and cerebrospinal fluid eosinophils without any other causes of the deterioration of consciousness, such as infection or metabolic derangements. Ocular angiostrongyliasis is very rare and causes a permanent visual impairment and a wide range of ocular inflammation, depending on the worms route. It can occur with or without EoM. An identification of a living worm, usually a single worm in any part of an eye, is an important diagnostic clue. The treatment options are surgical removal or laser therapy. Corticosteroids may be necessary in the case of coexistence of EoM or other ocular inflammations such as retinitis or optic neuritis. The visual outcome is poor and depends on the initial visual acuity.

Clinical factors predictive of encephalitis caused by Angiostrongylus cantonensis.

Angiostrongylus cantonensis is mainly caused eosinophilic meningitis in humans, whereas a minority of patients develop encephalitic angiostrongyliasis (EA). EA is an extremely fatal condition, and the clinical factors predictive of EA have never been reported. A comparison study was conducted in a hospital situated in an endemic area of Thailand. We enrolled 14 and 80 angiostrongyliasis patients who developed encephalitis and meningitis, respectively. Logistic regression analysis was used to assess the clinical variables predictive of encephalitis. Age (adjusted odds ratio [OR], 1.22; 95% confidence interval [CI], 1.05-1.42), duration of headache (adjusted OR, 1.26; 95% CI, 1.03-1.55), and fever > 38.0 degrees C (adjusted OR, 37.05; 95% CI, 1.59-862.35) were identified as statistically significant factors for EA prediction. Elderly patients with angiostrongyliasis experiencing fever and prolonged headaches were at the highest risk of developing EA.

Decreased expression of galectin-3 is associated with metastatic potential of liver fluke-associated cholangiocarcinoma.

Galectin-3, a beta-galactoside-binding lectin, is a multifunctional protein implicated in a variety of biological functions, including tumour cell adhesion, proliferation, differentiation, cancer progression and metastasis. The present study was performed to clarify the impact of galectin-3 expression on patients with liver fluke-associated cholangiocarcinoma. Galectin-3 expression was examined immunohistochemically in 53 patients with intrahepatic cholangiocarcinoma, who had undergone surgery without pre-operative therapy. All bile duct epithelium expressed galectin-3 with different intensities, according to the different histological subtypes. The poorly-differentiated type expressed galectin-3 less intensely than the papillary, well- to moderately-differentiated types (P=0.012). We observed the association of low galectin-3 expression with lymphatic invasion (P=0.002). Suppression of galectin-3 expression in two human cholangiocarcinoma cell lines using siRNA targeted to galectin-3 significantly increased cell migration and invasion without alterations in cell proliferation. Regulation of galectin-3 expression may therefore be an alternative therapeutic approach to control metastasis of cholangiocarcinoma.

Methionine aminopeptidase 2 over-expressed in cholangiocarcinoma: Potential for drug target

Methionine aminopeptidases (MetAP) are proteases which remove the N-terminal methionine from newly synthesized proteins. Associations of MetAP2 with tumor progression of different cancers have been repeatedly reported. We aim to determine if MetAP2 is expressed in cholangiocarcinomas (CCA) and investigate to see if it would be a useful therapeutic target. We evaluated MetAP2 expression by immunohistochemistry in 82 patients of intrahepatic CCA. MetAP2 was expressed in bile ducts to various degrees. It was occasionally expressed with weak staining in normal bile duct epithelium but was strikingly over-expressed in dysplastic bile duct epithelia, primary and metastatic CCA tissues (p < 0.001). The increased expression of MetAP2 in proliferating bile duct was evident. All metastatic tumors had stronger expression of MetAP2 than the corresponding primary tumors. Fumagillin, a MetAP2 specific inhibitor, significantly inhibited cell proliferation in dose dependent manner and the degree of growth inhibition was dependent on the amount of cellular enzyme. The present study highlights the involvement of MetAP2 in an early event of carcinogenesis of CCA. The findings represent the first description of increased MetAP2 expression in CCA. The inhibition of enzyme activity using MetAP2 inhibitors may be a potential strategy for long-term control of tumor development and progression in CCA patients.

High glucose levels boost the aggressiveness of highly metastatic cholangiocarcinoma cells via O-GlcNAcylation

Increased glucose utilization is a feature of cancer cells to support cell survival, proliferation, and metastasis. An association between diabetes mellitus and cancer progression was previously demonstrated in cancers including cholangiocarcinoma (CCA). This study was aimed to determine the effects of high glucose on protein O-GlcNAcylation and metastatic potentials of CCA cells. Two pairs each of the parental low metastatic and highly metastatic CCA sublines were cultured in normal (5.6 mM) or high (25 mM) glucose media. The migration and invasion abilities were determined and underlying mechanisms were explored. Results revealed that high glucose promoted migration and invasion of CCA cells that were more pronounced in the highly metastatic sublines. Concomitantly, high glucose increased global O-GlcNAcylated proteins, the expressions of vimentin, hexokinase, glucosamine-fructose-6-phosphate amidotransferase (GFAT) and O-GlcNAc transferase of CCA cells. The glucose level that promoted migration/invasion was shown to be potentiated by the induction of GFAT, O-GlcNAcylation and an increase of O-GlcNAcylated vimentin and vimentin expression. Treatment with a GFAT inhibitor reduced global O-GlcNAcylated proteins, vimentin expression, and alleviated cell migration. Altogether, these results suggested the role of high glucose enhanced CCA metastasis via modulation of O-GlcNAcylation, through the expressions of GFAT and vimentin.

Mechanistic insights of O-GlcNAcylation that promote progression of cholangiocarcinoma cells via nuclear translocation of NF-κB.

O-GlcNAcylation, an O-linked protein glycosylation with a single molecule of N-acetylglucosamine (GlcNAc), is reversibly controlled by O-GlcNAc transferase (OGT) and N-acetyl D-glucosaminidase (OGA). Aberrant O-GlcNAcylation contributes an important role in initiation and progression of many human cancers. Elevation of O-GlcNAcylation in tumor tissues and poor prognosis of cholangiocarcinoma (CCA) patients have been reported. In this study, the role of O-GlcNAcylation in promoting tumor progression was further investigated in CCA cell lines. Suppression of O-GlcNAcylation using small interfering RNAs of OGT (siOGT) significantly reduced cell migration and invasion of CCA cells whereas siOGA treated cells exhibited opposite effects. Manipulating levels of O-GlcNAcylation did affect the nuclear translocation of NF-κB and Akt-phosphorylation together with expression of matrix-metalloproteinases (MMPs). O-GlcNAcylation and nuclear translocation of NF-κB, the upstream signaling cascade of MMP activation were shown to be important for MMP activation. Immunoprecipitation revealed the elevation of O-GlcNAc-modified NF-κB with increased cellular O-GlcNAcylation. Involvement of O-GlcNAcylation in MMP-mediated migration and invasion of CCA cells was shown to be via O-GlcNAcylation and nuclear translocation of NF-κB. This information indicates the significance of O-GlcNAcylation in controlling the metastatic ability of CCA cells, hence, O-GlcNAcylation and its products may be new targets for treatment of metastatic CCA.

Suppression of thymosin β10 increases cell migration and metastasis of cholangiocarcinoma

BackgroundThymosin β10 (Tβ10) expression is associated with malignant phenotypes in many cancers. However, the role and mechanisms of Tβ10 in liver fluke-associated cholangiocarcinoma (CCA) are not fully understood. In this study, we investigated the expression of Tβ10 in CCA tumor tissues and cell lines as well as molecular mechanisms of Tβ10 in tumor metastasis of CCA cell lines.MethodsTβ10 expression was determined by real time RT-PCR or immunocytochemistry. Tβ10 silence or overexpression in CCA cells was achieved using gene delivery techniques. Cell migration was assessed using modified Boyden chamber and wound healing assay. The effect of silencing Tβ10 on CCA tumor metastasis was determined in nude mice. Phosphorylation of ERK1/2 and the expression of EGR1, Snail and matrix metalloproteinases (MMPs) were studied.ResultsTen pairs of CCA tissues (primary and metastatic tumors) and 5 CCA cell lines were studied. With real time RT-PCR and immunostaining analysis, Tβ10 was highly expressed in primary tumors of CCA; while it was relatively low in the metastatic tumors. Five CCA cell lines showed differential expression levels of Tβ10. Silence of Tβ10 significantly increased cell migration, invasion and wound healing of CCA cells in vitro; reversely, overexpression of Tβ10 reduced cell migration compared with control cells (P<0.05). In addition, silence of Tβ10 in CCA cells increased liver metastasis in a nude mouse model of CCA implantation into the spleen. Furthermore, silence of Tβ10 activated ERK1/2 and increased the expression of Snail and MMPs in CCA cell lines. Ras-GTPase inhibitor, FPT inhibitor III, effectively blocked Tβ10 silence-associated ERK1/2 activation, Snail expression and cell migration.ConclusionsLow expression of Tβ10 is associated with metastatic phenotype of CCA in vitro and in vivo, which may be mediated by the activation of Ras, ERK1/2 and upregulation of Snail and MMPs. This study suggests a new molecular pathway of CCA pathogenesis and a novel strategy to treat or prevent CCA metastasis.

Can workplaces be predictors for recent onset latent tuberculosis in health care workers

ObjectiveTo study the association of workplaces and recent onset latent tuberculosis (LTB) in health care workers (HCW).MethodsA case-control study was conducted at Srinagarind Hospital, Khon Kaen University, Thailand. We recruited HCW who had results of tuberculin test within 2 consecutive years from 2001–2008 and also had fixed workplaces (working hours more than 40 hours/week). Cases were subjects with tuberculin conversion, while controls were subjects with negative results of tuberculin test in two consecutive years. Tuberculin conversion was defined if a subject had a negative baseline tuberculin test and a positive tuberculin test in the next consecutive years. Baseline characteristics, workplaces (office, in-patient unit, out-patient unit, intensive care, operating room, and laboratory unit), tuberculosis related variables, and prevention strategies were studied. Multiple logistic regression analysis was used to identify predictors for tuberculin conversion.ResultsThere were 624 subjects who met the criteria and 163 subjects had tuberculin conversion (26.1%; case group). The median age and male/female ratio of both groups were 39 years old and about 1:4. The cases group had higher percentage of subjects who worked at in- and out-patient department (30.7 vs 20.2 and 17.2 vs 12.2, respectively), had history of tuberculosis exposure in the past year (32.1 vs 16.1), and had history of prevention by any method and by surgical mask (49.4 vs 37.0 and 54.3 vs 38.3, respectively). Workings at in- and out-patient unit and history of tuberculosis exposure in the past year were significant predictors for tuberculin conversion (adjusted odds ratio and [95% confidence interval] of 1.99 [1.25–3.17], 1.91 [1.10–3.17], and 2.26 [1.47–4.96], respectively).SummaryWorkplaces in health care facilities do increase risks of LTB in HCW, particularly in in- and out-patient unit. Policy development regarding tuberculosis infection control programs focused on workplace prevention in health care facilities in Thailand is needed.

Specificity of immunoblotting analyses in eosinophilic meningitis

Angiostrongylus cantonensis and Gnathostoma spinigerum are the two most common causative parasites of eosinophilic meningitis (EOM). Serological tests are helpful tools for confirming the identity of the pathogen. Recent reports determined the specificity of such tests by using normal healthy controls. There have been limited studies done to rule out the cross-reactivity between these two causative parasites of EOM. This study aims to assess the specificity of the serological test in EOM by using each condition as a control for the other. Thirty-three patients with a diagnosis of EOM were enrolled. Sera from 22 patients with a positive 29-kDa antigenic diagnostic band of A. cantonensis were tested for the 21 and 24-kDa antigenic bands of G. spinigerum. Similarly, sera of 11 gnathostomiasis patients were tested for the 29-kDa diagnostic band for A. cantonensis. Only one patient in the angiostrongyliasis group had a positive result for the 21 and 24-kDa antigenic bands of G. spinigerum, while no gnathostomiasis patients showed a positive result for the 29-kDa antigenic band of A. cantonensis. The specificity of the 21 and 24-kDa antigenic bands for gnathostomiasis and the 29-kDa antigenic band for A. cantonensis was 95.5% and 100%, respectively. The antigenic bands for the diagnosis of gnathostomiasis and angiostrongyliasis in EOM were highly specific.

Improve discrimination power of serum markers for diagnosis of cholangiocarcinoma using data mining-based approach

OBJECTIVE Cholangiocarcinoma (CCA) is usually fatal because of the absence of tests for early detection and lack of effective therapy. Tumor markers with adequate diagnostic values are of clinical significance. This study is aimed to improve the diagnostic power of serum markers using the computational data mining technique to develop a combined diagnostic model that yielded the best diagnostic values for CCA. DESIGN AND METHODS Eight CCA-associated markers-carcinoembryonic antigen, carbohydrate antigen 19-9, alkaline phosphatase (ALP), and gamma glutamyl transferase, biliary-ALP, mucin5AC, CCA-associated carbohydrate antigen (CCA-CA) and CA-S27-were used as the inputs for the C4.5 decision tree classification model and the selected model was confirmed by ANN analyses. Eight serum markers for CCA were determined in the training set of 85 histologically proven-CCA patients and 82 control subjects. The chosen set of combined markers that gave the best diagnostic values for CCA was then validated in the testing set of 22 CCA patients and 60 controls. RESULTS A decision tree diagram built by the C4.5 algorithm suggested the serial analysis of CCA-CA and ALP for distinguishing CCA patients from non-CCA subjects with all diagnostic parameters ≥95%. The combined tests showed a precise diagnosis in the testing set. CONCLUSIONS The C4.5 model indicates the combined markers of CCA-CA and ALP that produced the more precise diagnosis for CCA.