Jureerut Daduang

Serum antioxidant levels and nutritional status in early and advanced stage lung cancer patients

OBJECTIVE Malnutrition frequently occurs in lung cancer patients. We aimed to determine nutritional status and antioxidant and mineral levels in Thai patients with lung cancer. METHODS A prospective study with matched case-control was conducted. Nutritional status was assessed by body mass index (BMI) and subjective global assessment (SGA). Eastern Cooperative Oncology Group (ECOG) performance status was used to assess the performance. The serum antioxidant and mineral levels were determined. RESULTS Forty-nine patients with a mean age of 58.8 (range, 35-82) who were first diagnosed with lung cancer were enrolled. They were compared with 60 healthy controls, and levels of retinol, α-tocopherol, β-carotene, lycopene, β-cryptoxanthin, selenium, and zinc were lower (P < 0.05). However, peroxidase activity was higher (P = 0.002) in patients. Selenium levels were higher in early stage compared to advanced stage patients (P = 0.041). Overweight patients had higher selenium levels (0.04 mg/L) than normal BMI patients (β = 0.04, P = 0.035). Patients with SGA class C had lower selenium levels (0.03 mg/L) than those with class A (β = -0.03, P = 0.035). The poorer ECOG performance patients had significantly lower β-carotene (β = -0.192, P = 0.003) and selenium (β = -0.031, P = 0.011) levels compared with those with good ECOG performance status. CONCLUSIONS Significantly lower levels of antioxidants and selenium were found in lung cancer patients compared to healthy controls. Levels of some antioxidants and minerals differed among categories of BMI, SGA categories, or ECOG performance status. These findings may be helpful for further studies, such as the effect of nutritional supplementation on clinical outcomes.

Chitinase 3 like 1 is associated with tumor angiogenesis in cervical cancer.

Elevated serum levels of a secreted glycoprotein chitinase 3 like 1 (CHI3L1) are associated with poor prognosis and short survival time of patients with cervical cancer (CxCa). Our previous microarray data showed the increased expression of CHI3L1 in invasive CxCa compared to normal tissue, implicating a potential role of CHI3L1 in CxCa. To establish the pathological role of CHI3L1 in the development of CxCa, this study focused on its expression in CxCa and angiogenic impacts in tumor vessel formation. CHI3L1 activated angiogenesis by promoting endothelial cell migration and tube formation in vitro but failed to protect CxCa cell lines, CaSki and HeLa against apoptosis induced by γ-irradiation. In addition, the capability of CHI3L1 to induce proliferation and migration of CaSki and HeLa cells was cell type specific. In an analysis of 103 specimens from CxCa patients, increased expression levels of CHI3L1 mRNA and protein in invasive CxCa were 4-fold (P<0.05) and 2-fold (P<0.01), respectively, stronger than those in normal subjects. The immunostaining of CHI3L1 was positively correlated with VEGF expression (P=0.0019) and microvessel density (P=0.0110). Moreover, CHI3L1 expression was also positively associated with cancer metastasis (P=0.011). The data suggest the crucial role of CHI3L1 by promoting angiogenesis, which may contribute to the development and progression of CxCa. The findings help establish CHI3L1 as a prognostic biomarker and therapeutic target for CxCa patients.

Heteromtoxin (HmTx), a novel heterodimeric phospholipase A2 from Heterometrus laoticus scorpion venom

Heteromtoxin (HmTx) is a group III phospholipase A(2) produced in Heterometrus laoticus, in Thailand. In this study, HmTx was purified from venom by separation chromatography, and the PLA(2) activity of the fractions was determined by lecithin agar assay. The enzyme is an acidic protein with a pI of 5.6 and an apparent molecular weight of 14018.4 Da. The nucleotide sequence of HmTx contains 649 bp, and the mature protein is predicted to have 131 amino acid residues-104 of which make up the large subunit, and 27 of which make up the small subunit. The subunit structure of HmTx is highly similar to that of the other toxin, Pandinus imperator imperatoxin I (IpTx(i)) and to Mesobuthus tamulus phospholipase A(2) (MtPLA(2)). The 3D-structure of HmTx consists of three conserved alpha-helices: h1 (Lys24-His34), h2 (Cys59-Asp71), and h3 (Ala80-Phe89). The beta-sheet consisted of a single stranded anti-parallel beta-sheet (b1.1 at Glu43-Lys45 and b1.2 at Lys48-Asn50) that was highly similar to the conserved sequences (-CGXG-, -CCXXHDXC- and CXCEXXXXXC-) of Apis mellifera (bee) phospholipases.

Cytotoxic effects of Phytophenolics from Caesalpinia mimosoides Lamk on cervical carcinoma cell lines through an apoptotic pathway.

BACKGROUND Extracts of Caesalpinia mimosoides Lamk has been reported to possess anticancer effects, but the active ingredients and the anti-cancer mechanisms are still unknown. MATERIALS AND METHODS The effects of a C mimosoides Lamk extract on cell proliferation and apoptosis induction in human cervical carcinoma cell lines, namely HeLa, SiHa, and C33A, as well as in normal Vero cells, were investigated. RESULTS Treatment with 5 active fractions (F17-F21) of C mimosoides Lamk methanol extracts inhibited cell viability in a dose- and time- dependent manner. Neutral red assays indicated that treatment with F21 significantly decreased the viability of all cervical cancer cell lines compared to F21-treated normal cells. In addition, HPLC analysis revealed that F21 contained multiple phenolic compounds, namely gallic acid, caffeine, vanillic acid, ferulic acid and resveratrol. F21 had the lowest IC50 and, therefore, a much higher cytotoxicity than F20, F17, F19, and F18 by 20-, 25-, 46- and 47- fold, respectively. Analysis of activation of the apoptosis pathway using a caspase 3/7 activity assay revealed that F21 treatment resulted in a considerable increase in caspase activation in all cancer cell lines tested. At the same concentration of F21, HeLa cells had the highest caspase activity (6.5-fold) compared to the control. CONCLUSION C mimosoides Lamk may be of value as an alternative therapeutic agent, especially in combination with other compounds offering possible of synergy of action. Moreover, HPV- and non-HPV-related cervical cancer cells may differ in their responses to treatment regimens.

Rapid detection of the most common high-risk human papillomaviruses by loop-mediated isothermal amplification

Persistent infection with high-risk human papillomavirus (HPV) is a major risk factor for development of cervical cancer. At present, polymerase chain reaction (PCR)-based methods, the most widely molecular tools used for HPV detection, are time-consuming and require expensive instruments. In this study, loop-mediated isothermal amplification (LAMP) was established for detection of HPV types 16, 18, 45 and 58 which are frequently found in Thailand. The optimal condition for detection of these high risk HPVs was 63°C for 60min. Since a white magnesium pyrophosphate precipitate is a characteristic by product of the LAMP reaction which can be visualized directly by the naked eye, the entire assay time of LAMP is 1h compared to 6-8h of for a nested PCR detection. The detection limit of LAMP assay was shown to be equivalent to nested PCR that could amplify 10(2) copies of HPV-18 and 10(3) copies of HPV 16, 45 and 58, as determined by either turbidity detection or agarose gel electrophoresis. No cross-reaction was observed, indicating that LAMP assay has high type-specificity. The assay showed successful detection of HPV in 56 clinical specimens. Using nested PCR as the gold standard, the sensitivity, specificity, negative predictive values and positive predictive values of LAMP assay were 100%. In conclusion, LAMP assay is a high efficiency, low cost diagnostic tool, useful for rapid, accurate, direct detection of HPV for clinical diagnosis.

Gallic Acid Enhancement of Gold Nanoparticle Anticancer Activity in Cervical Cancer Cells

Cervical cancer (CxCa) is the most common cancer in women and a prominent cause of cancer mortality worldwide. The primary cause of CxCa is human papillomavirus (HPV). Radiation therapy and chemotherapy have been used as standard treatments, but they have undesirable side effects for patients. It was reported that gallic acid has antioxidant, antimicrobial, and anticancer activities. Gold nanoparticles are currently being used in medicine as biosensors and drug delivery agents. This study aimed to develop a drug delivery agent using gold nanoparticles conjugated with gallic acid. The study was performed in uninfected (C33A) cervical cancer cells, cervical cancer cells infected with HPV type 16 (CaSki) or 18 (HeLa), and normal Vero kidney cells. The results showed that GA inhibited the proliferation of cancer cells by inducing apoptosis. To enhance the efficacy of this anticancer activity, 15-nm spherical gold nanoparticles (GNPs) were used to deliver GA to cancer cells. The GNPs-GA complex had a reduced ability compared to unmodified GA to inhibit the growth of CxCa cells. It was interesting that high-concentration (150 μM) GNPs-GA was not toxic to normal cells, whereas GA alone was cytotoxic. In conclusion, GNPs-GA could inhibit CxCa cell proliferation less efficiently than GA, but it was not cytotoxic to normal cells. Thus, gold nanoparticles have the potential to be used as phytochemical delivery agents for alternative cancer treatment to reduce the side effects of radiotherapy and chemotherapy.

Bioactive compounds from crocodile (Crocodylus siamensis) white blood cells induced apoptotic cell death in hela cells.

Crocodile (Crocodylus siamensis) white blood cell extracts (WBCex) were examined for anticancer activity in HeLa cell lines using the MTT assay. The percentage viability of HeLa cells significantly deceased after treatment with WBCex in a dose‐ and time‐dependent manner. The IC50 dose was suggested to be approximately 225 μg/mL protein. Apoptotic cell death occurred in a time‐dependent manner based on investigation by flow cytometry using annexin V‐FITC and PI staining. DAPI nucleic acid staining indicated increased chromatin condensation. Caspase‐3, −8 and −9 activities also increased, suggesting the induction of the caspase‐dependent apoptotic pathway. Furthermore, the mitochondrial membrane potential (ΔΨm) of HeLa cells was lost as a result of increasing levels of Bax and reduced levels of Bcl‐2, Bcl‐XL, Bcl‐Xs, and XIAP. The decreased ΔΨm led to the release of cytochrome c and the activation of caspase‐9 and −3. Apoptosis‐inducing factor translocated into the nuclei, and endonuclease G (Endo G) was released from the mitochondria. These results suggest that anticancer agents in WBCex can induce apoptosis in HeLa cells via both caspase‐dependent and ‐independent pathways.

Genetic analysis of the merozoite surface protein-1 block 2 allelic types in Plasmodium falciparum clinical isolates from Lao PDR

BackgroundMSP-1 is one of the potential malarial vaccine candidate antigens. However, extensive genetic polymorphism of this antigen in the field isolates of Plasmodium falciparum represents a major hindrance for the development of an effective vaccine. Therefore, this study aimed to establish the prevalence and genetic polymorphisms of K1, MAD20 and RO33 allelic types of msp-1 block 2 among P. falciparum clinical isolates from Lao PDR.MethodsPlasmodium falciparum isolates were collected from 230 P. falciparum-infected blood samples from three regions of Lao PDR. K1, MAD20 and RO33 were detected by nested PCR; SSCP was used for polymorphism screening. The nested PCR products of each K1, MAD20 and RO33 allelic types that had different banding patterns by SSCP, were sequenced.ResultsThe overall prevalence of K1, MAD20 and RO33 allelic types in P. falciparum isolates from Lao PDR were 66.95%, 46.52% and 31.30%, respectively, of samples under study. Single infections with K1, MAD20 and RO33 allelic types were 27.83%, 11.74% and 5.22%, respectively; the remainders were multiple clonal infections. Neither parasite density nor age was related to MOI. Sequence analysis revealed that there were 11 different types of K1, eight different types of MAD20, and 7 different types of RO33. Most of them were regional specific, except type 1 of each allelic type was common found in 3 regions under study.ConclusionsGenetic polymorphism with diverse allele types was identified in msp-1 block 2 among P. falciparum clinical isolates in Lao PDR. A rather high level of multiple clonal infections was also observed but the multiplicity of infection was rather low as not exceed 2.0. This basic data are useful for treatment and malaria control program in Lao PDR.

High sensitivity, loop-mediated isothermal amplification combined with colorimetric gold-nanoparticle probes for visual detection of high risk human papillomavirus genotypes 16 and 18.

High-risk human papillomavirus (HR-HPV) causes cervical cancer. HPV16 and HPV18 are the most prevalent strains of the virus reported in women worldwide. Loop-mediated isothermal amplification (LAMP) is an alternative method for DNA detection under isothermal conditions. However, it results in a turbid amplified product which is not easily detected by the naked eye. This study aimed to develop an improved technique by using gold nanoparticles (AuNPs) attached to a single-stranded DNA probe for the detection of HPV16 and HPV18. Detection of the LAMP product by AuNP color change was compared with detection by visual turbidity. The optimal conditions for this new LAMP-AuNP assay were an incubation time of 20min and a temperature of 65°C. After LAMP amplification was complete, its products were hybridized with the AuNP probe for 5min and then detected by the addition of magnesium salt. The color changed from red to blue as a result of aggregation of the AuNP probe under high ionic strength conditions produced by the addition of the salt. The sensitivity of the LAMP-AuNP assay was greater than the LAMP turbidity assay by up to 10-fold for both HPV genotypes. The LAMP-AuNP assay showed higher sensitivity and ease of visualization than did the LAMP turbidity for the detection of HPV16 and HPV18. Additionally, AuNP-HPV16 and AuNP-HPV18 probes were stable for over 1year. The combination of LAMP and the AuNP-probe colorimetric assay offers a simple, rapid and highly sensitive alternative diagnostic tool for the detection of HPV16 and HPV18 in district hospitals or field studies.

Targeted delivery of 5-fluorouracil to cholangiocarcinoma cells using folic acid as a targeting agent

There are limits to the standard treatment for cholangiocarcinoma (CCA) including drug resistance and side effects. The objective of this study was to develop a new technique for carrying drugs by conjugation with gold nanoparticles and using folic acid as a targeting agent in order to increase drug sensitivity. Gold nanoparticles (AuNPs) were functionalized with 5-fluorouracil (5FU) and folic acid (FA) using polyethylene glycol (PEG) shell as a linker (AuNPs-PEG-5FU-FA). Its cytotoxicity was tested in CCA cell lines (M139 and M213) which express folic acid receptor (FA receptor). The results showed that AuNPs-PEG-5FU-FA increased the cytotoxic effects in the M139 and M213 cells by 4.76% and 7.95%, respectively compared to those treated with free 5FU+FA. It is found that the cytotoxicity of the AuNPs-PEG-5FU-FA correlates with FA receptor expression suggested the use of FA as a targeted therapy. The mechanism of cytotoxicity was mediated via mitochondrial apoptotic pathway as determined by apoptosis array. In conclusion, our findings shed some light on the use of gold nanoparticles for conjugation with potential compounds and FA as targeted therapy which contribute to the improvement of anti-cancer drug efficacy. In vivo study should be warranted for its effectiveness of stability, biosafety and side effect reduction.