Pharmacological Properties of White Mulberry (Morus alba L.) Leaves: Suppressing Migratory and Invasive Activities Against A549 Lung Cancer Cells.

Morus alba known as a white mulberry is a medicinal plant that has been used in food ingredients and traditional medicine. M. alba leaves contain various bioactive phenolic compounds, in particular chlorogenic acid (CGA), which is a major bioactive ingredient. Their anticancer potency of M. alba leaf extracts derived from Soxhlet extraction was evaluated based on cytotoxicity and antimigratory and antiinvasive properties. The dichloromethane extract exhibited the highest nitric oxide radical scavenging activity with a half-maximal inhibitory concentration (IC50) value of 780 µg/mL, promising cytotoxicity against HuCCA-1, MCF-7, and A-549 cells with IC50 values of 59.18, 62.20, and 103.25 µg/mL, respectively. CGA selectively inhibited the growth of MCF-7 cells with an IC50 value of 26.75 µg/mL and showed potent radical scavenging activity against DPPH radicals (IC50 = 18.85 µg/mL). An ethanolic extract derived from the gradient Soxhlet extraction suppressed A549 lung cancer cell migration and invasion more effectively than CGA with no migratory inhibition effect on noncancerous HaCaT cells. Furthermore, the ethanolic extract and CGA accelerated HaCaT wound closure at 20 µg/mL, which was the same as allantoin. Bioactive ingredients including triterpenes, steroids, phenolics, and flavonoids were mainly detected in all extracts. The highest content of CGA (52.23 g/100 g dry weight) was found in the ethanolic extract derived from the gradient Soxhlet extraction. These findings show the potency of the dichloromethane extract as a cytotoxic agent against various cancer types and the ethanolic extract as an antimetastatic agent by their antimigratory and antiinvasive activities.

PLGA-Lipid Hybrid Nanoparticles for Overcoming Paclitaxel Tolerance in Anoikis-Resistant Lung Cancer Cells.

Drug resistance and metastasis are two major obstacles to cancer chemotherapy. During metastasis, cancer cells can survive as floating cells in the blood or lymphatic circulatory system, due to the acquisition of resistance to anoikis-a programmed cell death activated by loss of extracellular matrix attachment. The anoikis-resistant lung cancer cells also develop drug resistance. In this study, paclitaxel-encapsulated PLGA-lipid hybrid nanoparticles (PLHNPs) were formulated by nanoprecipitation combined with self-assembly. The paclitaxel-PLHNPs had an average particle size of 103.0 ± 1.6 nm and a zeta potential value of -52.9 mV with the monodisperse distribution. Cytotoxicity of the nanoparticles was evaluated in A549 human lung cancer cells cultivated as floating cells under non-adherent conditions, compared with A549 attached cells. The floating cells exhibited anoikis resistance as shown by a lack of caspase-3 activation, in contrast to floating normal epithelial cells. Paclitaxel tolerance was evident in floating cells which had an IC50 value of 418.56 nM, compared to an IC50 value of 7.88 nM for attached cells. Paclitaxel-PLHNPs significantly reduced the IC50 values in both attached cells (IC50 value of 0.11 nM, 71.6-fold decrease) and floating cells (IC50 value of 1.13 nM, 370.4-fold decrease). This report demonstrated the potential of PLHNPs to improve the efficacy of the chemotherapeutic drug paclitaxel, for eradicating anoikis-resistant lung cancer cells during metastasis.

Global Analysis of Protein Expression in A549 Cells After Prolonged Nicotine Exposure by Using Label-free Quantification.

BACKGROUND/AIM: Lung cancer is the leading cause of cancer death worldwide. Cigarette smoke is the most important risk factor for cancer development. Growing evidence indicates that prolonged nicotine exposure is a potential factor associated with tumorigenesis. Here, the effect of prolonged nicotine exposure on A549 cells was investigated, using label-free quantitative proteomics. MATERIALS AND METHODS: Selection of an invasive subpopulation from the A549 cell line was performed to reveal the differential expression of proteins in relation to prolonged nicotine exposure, using Boyden chamber assays in combination with a proteomics approach. RESULTS: One hundred proteins from the NicoA549-L5 subline showed significant change in expression compared to those from the A549-L5 subline and their A549 parental cell line. Heat shock protein, protein disulfide isomerase A3, profilin-1 and legumain were expressed at higher levels in A549 cells after prolonged nicotine exposure. CONCLUSION: These aberrant proteins might serve as novel cancer biomarkers for cigarette smokers.

Effects of topical exposure to a mixture of chlorpyrifos, cypermethrin and captan on the hematological and immunological systems in male Wistar rats.

Most Thai orchid farmers heavily used pesticide mixtures, and were shown to have various hematologic/immunologic alterations. The present study investigated the effect of exposure of male Wistar rats to a mixture of three pesticides (chlorpyrifos, cypermethrin and captan) that are most often used by the farmers. Three groups of 10 rats were dermally exposed to three different doses (high, middle and low) for 28 consecutive days. The rats showed significant changes in body, liver, kidneys and adrenals weights. Significant changes were observed in various biological parameters including hematotoxicity (increased leukocyte and platelet counts, percent neutrophil, decreased RBC count, percent lymphocyte and eosinophil), hepatotoxicity (increased serum AST, decreased serum ALP, cholesterol, triglyceride, serum protein and albumin), and immunotoxicity (decrease in numbers of NK cells, decrease splenic proliferative response to LPS, and increase in serum IgG). These results confirm the potential health danger of exposure to these pesticide mixtures in orchid farmers.

Immunoaffinity chromatography in antivenomics studies: Various parameters that can affect the results.

Antivenomics is a recently developed powerful method for the study of antivenom antibody profiles when bound to homologous and heterologous snake venoms. The information obtained is useful in gaining an understanding of venom protein immunogenicity, antivenom potency and also for the improvement of antivenom potency and paraspecificity. The preferred method used in this type of study is immunoaffinity chromatography of the venom proteins on an antivenom IgG (or F(ab')2) column where the bound and unbound proteins can be separated and identified. However, there are some parameters of the immunochromatography that can significantly affect the binding of the proteins to the immunoaffinity matrix and lead to imprecise results in antivenom immunoprofiling. The present study demonstrated that the ligand density (mg IgG/ml of the matrix), the buffers used for binding and washing the venom proteins, the amount of venom loaded, the abundance of some venom protein(s) and the eluting buffers can significantly alter the binding of the proteins to the matrix and consequently the conclusions drawn from antivenomics studies. Furthermore, the immunochromatographic procedure can be extended to include the estimation of the relative affinity of venom protein-antibody interactions that can provide additional information useful to antivenomics study.

Protein A/G-based immunochromatographic test for serodiagnosis of pythiosis in human and animal subjects from Asia and Americas.

Pythiosis is a life-threatening infectious disease of both humans and animals living in Asia, Americas, Africa, and parts of Australia and New Zealand. The etiologic pathogen is the fungus-like organism Pythium insidiosum The disease has high mortality and morbidity rates. Use of antifungal drugs are ineffective against P. insidiosum, leaving radical surgery the main treatment option. Prompt treatment leads to better prognosis of affected individuals, and could be achieved by early and accurate diagnosis. Since pythiosis has been increasingly reported worldwide, there is a need for a rapid, user-friendly, and efficient test that facilitates the diagnosis of the disease. This study aims to develop an immunochromatographic test (ICT), using the bacterial protein A/G, to detect anti-P. insidiosum IgGs in humans and animals, and compare its diagnostic performance with the established ELISA. Eighty-five serum samples from 28 patients, 24 dogs, 12 horses, 12 rabbits, and 9 cattle with pythiosis, and 143 serum samples from 80 human and 63 animal subjects in a healthy condition, with thalassemia, or with other fungal infections, were recruited for assay evaluation. Detection specificities of ELISA and ICT were 100.0%. While the detection sensitivity of ELISA was 98.8%, that of ICT was 90.6%. Most pythiosis sera, that were falsely read negative by ICT, were weakly positive by ELISA. In conclusion, a protein A/G-based ICT is a rapid, user-friendly, and efficient assay for serodiagnosis of pythiosis in humans and animals. Compared to ELISA, ICT has an equivalent detection specificity and a slightly lower detection sensitivity.

A comparative study of venomics of Naja naja from India and Sri Lanka, clinical manifestations and antivenomics of an Indian polyspecific antivenom.

Naja naja (Indian cobra) from Sri Lanka and India is the WHO Category 1 medically important snakes in both countries. Some antivenom produced against Indian N. naja (NNi) were less effective against Sri Lankan N. naja (NNsl). Proteomes of NNi and NNsl venoms were studied by RP-HPLC, SDS-PAGE and LC/MS/MS. Six protein families were identified in both venoms with the most abundant were the 3 finger toxins (3FTs) where cytotoxins (CTX) subtype predominated, followed by phospholipase A2, cysteine-rich venom protein, snake venom metalloproteases, venom growth factors, and protease inhibitors. Qualitative and quantitative differences in the venomics profiles were observed. Some proteins were isolated from either NNi or NNsl venom. Postsynaptic neurotoxins (NTX) were identified for the first time in NNsl venom. Thus, there are geographic intra-specific variations of venom composition of the two N. naja. The relative abundance of CTX and NTX explained well the clinical manifestations of these venoms. Antivenomics study of an Indian antivenom (Vins) showed the antibodies effectively bound all venom toxins from both snakes but more avidly to the Indian venom proteins. The lower antibody affinity towards the 'heterologous' venom was the likely cause of poor efficacy of the Indian antivenom used to treat NNsl envenoming.

Effect of pesticide exposure on immunological, hematological and biochemical parameters in thai orchid farmers- a cross-sectional study.

Various studies have found that many Thai orchid farmers used excessive amounts of pesticides without proper protective gear, but no toxicological study has been made. This cross-sectional study aimed to evaluate the immunological, hematological and biochemical statuses of these farmers. Sixty four orchid farmers and 60 controls were studied. Plasma cholinesterase activity, the percentage and absolute number of B lymphocytes (CD19+) were significantly lower in the farmers group (3966.32±1165.48 U/L, 11.61±4.09% and 312.26±164.83 cells/mm3, respectively) as compared to those of controls (5048.85±1139.40 U/L, 14.32±4.23%, 420.34±195.18 cells/mm3, respectively). There was a statistically significant higher level of serum IgE among the orchid farmers (0.031±0.011 mg/dL vs. 0.018±0.007 mg/dL) but not IgG, IgA and IgM, levels. Serum lysozyme level, lymphocyte proliferative responses to mitogens, hematological parameters and kidney function test, were not significantly different between the two groups. The liver function profiles showed significantly lower levels of albumin and serum protein in the farmer group. Thus frequent pesticide exposure resulted in subtle changes of some biological parameters. These changes, though may not be clinically significant, strongly indicated that caution in handing pesticides by these farmers is warranted.

A defective angiogenesis in chronic kidney disease.

BACKGROUND: A progressive reduction in peritubular capillary flow is observed in chronic kidney disease (CKD) patients as the disease severity progresses. This suggests an altered vascular homeostasis in CKD patients, but such a defective mechanism needs to be verified. METHODS: To study the vascular injury as reflected by circulating endothelial cell (CEC), the balance between angiogenic factor, vascular endothelial growth factor (VEGF), and antiangiogenic factor, endostatin. RESULTS: A deficient VEGF was observed, whereas the value of endostatin and CEC were abnormally elevated in CKD patients. DISCUSSION: Enhanced CEC reflects an increased activity of vascular injury. A deficient VEGF in the presence of enhanced antiangiogenesis (endostatin) implies a defective angiogenesis. This may explain the progressive nature of renal microvascular disease observed in late stage of CKD patients.