p53 acetylation enhances Taxol-induced apoptosis in human cancer cells

Microtubule inhibitors (MTIs) such as Taxol have been used for treating various malignant tumors. Although MTIs have been known to induce cell death through mitotic arrest, other mechanisms can operate in MTI-induced cell death. Especially, the role of p53 in this process has been controversial for a long time. Here we investigated the function of p53 in Taxol-induced apoptosis using p53 wild type and p53 null cancer cell lines. p53 was upregulated upon Taxol treatment in p53 wild type cells and deletion of p53 diminished Taxol-induced apoptosis. p53 target proteins including MDM2, p21, BAX, and β-isoform of PUMA were also upregulated by Taxol in p53 wild type cells. Conversely, when the wild type p53 was re-introduced into two different p53 null cancer cell lines, Taxol-induced apoptosis was enhanced. Among post-translational modifications that affect p53 stability and function, p53 acetylation, rather than phosphorylation, increased significantly in Taxol-treated cells. When acetylation was enhanced by anti-Sirt1 siRNA or an HDAC inhibitor, Taxol-induced apoptosis was enhanced, which was not observed in p53 null cells. When an acetylation-defective mutant of p53 was re-introduced to p53 null cells, apoptosis was partially reduced compared to the re-introduction of the wild type p53. Thus, p53 plays a pro-apoptotic role in Taxol-induced apoptosis and acetylation of p53 contributes to this pro-apoptotic function in response to Taxol in several human cancer cell lines, suggesting that enhancing acetylation of p53 could have potential implication for increasing the sensitivity of cancer cells to Taxol.     

Mycobacterium kansasii-induced death of murine macrophages involves endoplasmic reticulum stress responses mediated by reactive oxygen species generation or calpain activation

Although pathogenic mechanisms of tuberculosis have been extensively studied, little is known about the pathogenic mechanisms of Mycobacterium kansasii. In this work the influence of virulence and ER-stress mediated apoptosis of macrophages during two different strains of M. kansasii infection was investigated. We show that M. kansasii infection is associated with ER stress-mediated apoptosis in the murine macrophage cell line RAW 264.7. Infection of RAW 264.7 cells in vitro with apoptosis-inducing a clinical isolate of M. kansasii SM-1 (SM-1) resulted in strong induction of ER stress responses compared with M. kansasii type strain (ATCC 12478)-infected RAW 264.7 cells. Interestingly, inhibition of calpain prevented the induction of CHOP and Bip in ATCC 12478-infected RAW 264.7 cells but not in RAW 264.7 cells infected with SM-1. In contrast, reactive oxygen species (ROS) were significantly increased only in RAW 264.7 cells infected with SM-1. We propose that ROS generation is important for triggering ER stress-mediated apoptosis during SM-1 infection, whereas ATCC 12478-induced, ER stress-mediated apoptosis is associated with calpain activation. Our results demonstrate that the ER stress pathway plays important roles in the pathogenesis of M. kansasii infections, and that different strains of M. kansasii induce different patterns of ER stress-mediated apoptosis.     

Delphinidin prevents hypoxia-induced mouse embryonic stem cell apoptosis through reduction of intracellular reactive oxygen species-mediated activation of JNK and NF-κB, and Akt inhibition

Delphinidin, gallic acid, betulinic acid, and ursolic acid, which are bio-active ingredients in a variety of fruits, vegetables, and herbs, have potent antioxidant activity and various biological activities. However, it is not clear whether these bio-active ingredients can significantly contribute to the protection of embryonic stem (ES) cells from hypoxia-induced apoptosis. In the present study, hypoxia-induced ES cells apoptosis with time, which were abrogated by pretreatment with all ingredients. Hypoxia-induced ROS generation was blocked by pretreatment with all ingredients in a dose-dependent manner, with the maximum ROS scavenging effect observed for delphinidin. Hypoxia increased phosphorylation of JNK and NF-κB were blocked by pretreatment of delphinidin as well as NAC. Hypoxia decreased phosphorylation of Akt^thr308 and ^ser473; these decreases were reversed by pretreatment with delphinidin or NAC. However, Akt inhibition did not affect NF-κB phosphorylation. Delphinidin attenuated the hypoxia-induced increase in Bax, cleaved caspase-9, cleaved caspase-3, and decrease in Bcl-2, which were diminished by pretreatment of Akt inhibitor. Hypoxia induced Bax translocation from the cytosol to mitochondria. Furthermore, hypoxia induced mitochondria membrane potential loss and cytochrome c release in cytosol, which were blocked by delphinidin pretreatment. Hypoxia induced cleavage of procaspase-9 and procaspase-3 which were blocked by delphinidin or SP600125, but Akt inhibitor abolished the protection effect of delphinidin. Moreover, inhibition of JNK and NF-κB abolished hypoxia-induced ES cell apoptosis and inhibition of Akt attenuated delphinidin-induced blockage of apoptosis. The results indicate that delphinidin can prevent hypoxia-induced apoptosis of ES cells through the inhibition of JNK and NF-κB phosphorylation, and restoration of Akt phosphorylation.     

Molecular cloning,characterization and comparison of bile salt hydrolases from Lactobacillus johnsonii PF01

Aims

To clone, characterize and compare the bile salt hydrolase (BSH) genes of Lactobacillus johnsonii PF01.

Methods and Results

The BSH genes were amplified by polymerase chain reaction (PCR) using specific oligonucleotide primers, and the products were inserted into the pET21b expression vector. Escherichia coli BLR (DE3) cells were transformed with pET21b vectors containing the BSH genes and induced using 0·1 mmol l^?1 isopropylthiolgalactopyranoside. The overexpressed BSH enzymes were purified using a nickel–nitrilotriacetic acid (Ni²⁺‐NTA) agarose column and their activities characterized. BSH A hydrolysed tauro‐conjugated bile salts optimally at pH 5·0 and 55°C, whereas BSH C hydrolysed glyco‐conjugated bile salts optimally at pH 5·0 and 70°C. The enzymes had no preferential activities towards a specific cholyl moiety.

Conclusions

BSH enzymes vary in their substrate specificities and characteristics to broaden its activity. Despite the lack of conservation in their putative substrate‐binding sites, these remain functional through motif conservation.

Significance and Impact of the Study

This is to our knowledge the first report of isolation of BSH enzymes from a single strain, showing hydrolase activity towards either glyco‐conjugated or tauro‐conjugated bile salts. Future structural homology studies and site‐directed mutagenesis of sites associated with substrate specificity may elucidate specificities of BSH enzymes.     

Metabolic profiling and enhanced production of phytosterols by elicitation with methyl jasmonate and silver nitrate in whole plant cultures of Lemna paucicostata

In this study, the effects of methyl jasmonate (MJ) and silver nitrate (SN) treatment on metabolic profiles and yields of phytosterols such as campesterol, stigmasterol, and β-sitosterol in whole plant cultures of Lemna paucicostata were investigated using gas chromatography–mass spectrometry coupled with multivariate statistical analysis. The MJ and SN treatments retarded the growth of L. paucicostata plants, while they enhanced the yields of three phytosterols, compared to control. Higher yields of phytosterols were attained at day 28 compared to day 42. Moreover, stigmasterol yield was the highest at 0.85 mg/g from day 28 plants grown under MJ + SN co-treated culture. Among the various metabolites, the levels of palmitic and stearic acids, which might participate in a defense mechanism, were higher in the MJ + SN condition than in control. To determine the optimal timing of MJ + SN addition, MJ + SN was added on days 21, 28, and 35 after inoculation. The total yield and productivity of phytosterol reached maximum levels when the MJ + SN was added at day 35. The highest productivity of stigmasterol (6.08 mg/L) was also achieved when MJ + SN was added on day 35.     

Mobile image sensors for object detection using color segmentation

The intelligent mobile robot with sensors and image processing embedded system combines the suction and aerodynamic attraction to achieve good balance between strong adhesion force and high mobility. Experimental results showed that the robot can move upward on the wall at the speed of 2.9 m/min and carry 5 kg payload in addition to 2.5 kg self-weight, which record the highest payload capacity among climbing robots of similar size. It also implements object detection ability using effective color transform and segmentation technique for the exact target detection on the wall using a embedded camera system, communication module and several active sensors.     

A frequency monitoring system development for wide-area power grid protection

There have been recent research activities on GPS-based FNET to prevent wide-area blackouts by monitoring frequency deviation. This paper introduces a system for monitoring regional frequencies in power grid developed as an advanced research project for intelligent wide-area protective relaying in South Korea. The system is implemented by modeling an actual 345 kV transmission system of South Korea using EMTP-RV and by measuring voltages and currents at five regions. The frequencies are estimated using our previous proposed method, a frequency estimation algorithm with gain compensation. The monitoring system implemented is used to examine frequency propagation effects and various fault conditions. Simulation results showed that the proposed system could achieve good performance regarding the frequency monitoring under both steady states and dynamic fault conditions.     

Effectiveness of autologous serum as an alternative to fetal bovine serum in adipose-derived stem cell engineering

In cell culture, medium supplemented with fetal bovine serum is commonly used, and it is widely known that fetal bovine serum supplies an adequate environment for culture and differentiation of stem cells. Nevertheless, the use of xenogeneic serum can cause several problems. We compared the effects of four different concentrations of autologous serum (1, 2, 5, and 10 %) on expansion and adipogenic differentiation of adipose-derived stem cells using 10 % fetal bovine serum as a control. The stem cells were grafted on nude mice and the in vivo differentiation capacity was evaluated. The isolation of adipose-derived stem cells was successful irrespective of the culture medium. The proliferation potential was statistically significant at passage 2, as follows: 10 % autologous serum >10 % fetal bovine serum = 5 % autologous serum >2 % autologous serum = 1 % autologous serum. The differentiation capacity appeared statistically significant at passage 4, as follows: 10 % fetal bovine serum >10 % autologous serum = 5 % autologous serum >2 % autologous serum = 1 % autologous serum. Ten percent autologous serum and 10 % fetal bovine serum had greater differentiation capacity than 1 and 2 % autologous serum in vivo, and no significant difference was observed between the groups at ≥5 % concentration at 14 weeks. In conclusion, 10 % autologous serum was at least as effective as 10 % fetal bovine serum with respect to the number of adipose-derived stem cells at the end of both isolation and expansion, whereas 1 and 2 % autologous serum was inferior.     

Immunostimulating Effects of Extract of Acanthopanax sessiliflorus

As malfunction/absence of immune cells causes a variety of immunosuppressive disorders and chemical synthetic drugs for curing these diseases have many adverse effects, vigorous studies are being conducted. The Acanthopanax family has been used as traditional medicines for gastric ulcer, diabetes, etc. and culinary materials in East-South Asia. In this study, the immunostimulating properties of A. sessiliflorus were evaluated. A. sessiliflorus increased not only the splenocyte number but also immune-related cytokines such as TNF-α. However, it could not upregulate the expressions of IFN-γ and IL-2. A. sessiliflorus increased the swimming time, and comparison of organ weights relative to body weights for immune-related organs such as the spleen and thymus after a forced swim test showed that it could recover the spleen and thymus weights. It also increased the expression of TNF-α and slightly increased the concentration of IFN-γ but not IL-2. From the results, we concluded that as A. sessiliflorus has not only a host defense effect but also a stress-ameliorating property, further study it will be a promising material of immunostimulating material.