Induction of apoptosis by the Lactuca indica L. in human leukemia cell line and its active components

Lactuca indica L. (Compositae family) is used as a folk medicine in anti-inflammatory, antibacterial, antidiabetic, and other medications in Asia. The objectives of this study were to evaluate the antiproliferative effect of ethanol extracts of Lactuca indica L. (EEL) on human leukemic HL-60 cell lines and its active components. The results showed that EEL exhibited strong cytotoxic effects against HL-60 cells; the IC50 value was 313 microg/mL. Flow cytometric analysis of the externalization of phosphatidylserine (PS) using the annexin V/PI method on EEL-treated HL-60 cells showed a concentration-dependent increase of apoptosis. Moreover, EEL could induce typical DNA fragmentation in a concentration- and time-dependent manner as determined by electrophoresis and TUNEL assays. The treatment of HL-60 cells with EEL induced significant accumulation of cells in the G0/G1 phase, indicating that EEL is a cell-cycle-dependent anticancer agent. Our results also indicate that EEL-induced apoptosis in HL-60 cells is associated with the loss of mitochondrial membrane potential (delta psi m). EEL contains 5% phenolic compounds, such as quercetin, caffeic acid, rutin, and chlorogenic acid. Among the four active phenolic compounds, quercetin was found to be the most effective in inhibition against cell viability and in alteration of mitochondrial function. Our results suggest that the induction of apoptosis by EEL might offer a pivotal mechanism for its chemopreventive action.     

Altering the substrate specificity of Candida rugosa LIP4 by engineering the substrate-binding sites

Candida rugosa (formerly Candida cylindracea) lipase (CRL) is an important industrial enzyme that is widely used in biotechnological applications such as the production of fatty acids and the synthesis of various esters. CRL comprises at least seven isozymes (LIP1-LIP7), which share a similar amino acid sequence but with different specificities for substrates. Previously, LIP4 was reported to have higher esterase activity toward long acyl-chain ester and lower lipase activity toward triglycerides. A296 and V344 of LIP4 were predicted to play decisive roles in its substrate specificity. In this study, site-specific saturation mutagenesis has been employed to study the substrate specificity of LIP4. Point mutations were separately introduced into A296 and V344 positions using degenerate primer sets containing 32 codons to generate two libraries of variants. LIP4 variants were heterologously expressed in the yeast Pichia pastoris. A specific plate assay was used to identify lipase-producing P. pastoris clones in a medium containing tributyrin. LIP4 variants with high activity toward short fatty acyl-chain triglyceride (tributyrin) were screened. Specificity analysis and biochemical characterization indicated that the recombinant variants A296I, V344Q, and V344H had properties remarkably different from those of wild-type LIP4. All three variant enzymes had significantly higher specific activities toward tributyrin than LIP4. In addition to short-chain triglyceride, A296I and V344Q also improved hydrolytic activities of triglycerides toward medium- and long-chain triglycerides tested. The results suggested that A296 played an important role in lipase activity and high-temperature dependence of LIP4, whereas it had no effect on the chain-length specificity in lipolytic reaction. The V344 residue had a significant effect on the substrate chain-length specificity of LIP4.     

Gallic acid induces apoptosis in 3T3-L1 pre-adipocytes via a Fas- and mitochondrial-mediated pathway

Gallic acid (3,4,5-trihydroxybenzoic acid) is a naturally abundant plant phenolic compound. Our previous studies have shown that some phenolic acids such as gallic acid inhibit cell growth and induce apoptosis in 3T3-L1 pre-adipocytes. However, the molecular mechanism of gallic acid in the induction of cell apoptosis is still unclear. In this study, we investigated the effect of gallic acid on the apoptotic pathway in 3T3-L1 pre-adipocytes. Western blot data revealed that gallic acid stimulated an increase in the protein expression of Fas, FasL, and p53. The ratio of expression levels of pro- and anti-apoptotic Bcl-2 family members was changed by gallic acid treatment. Gallic acid released mitochondrial cytochrome c into the cytosol and subsequently induced the activation of caspase-9 and caspase-3, which were followed by the cleavage of poly(ADP-ribose) polymerase. Pretreatment with a general caspase-9 inhibitor (Z-LEHD-FMK) and caspase-3 inhibitor (Z-DEVD-FMK) prevented gallic acid from inhibiting cell viability in 3T3-L1 pre-adipocytes. The data also indicated that treatment with gallic acid inhibited histone deacetylase activity in 3T3-L1 pre-adipocytes. These results demonstrate that gallic acid induces apoptosis in 3T3-L1 pre-adipocytes through the Fas and mitochondrial pathway. The induction of cell apoptosis by gallic acid may prove to be a pivotal mechanism for decreased pre-adipocyte proliferation.     

Effects of flavonoids and phenolic acids on the inhibition of adipogenesis in 3T3-L1 adipocytes

Obesity has become a global epidemic in both developed and developing countries, and it is a significant risk factor for various diseases such as diabetes, cancer, heart disease, and hypertension. In the present study, the effect of naturally occurring antioxidants (flavonoids and phenolic acids) on the inhibition of adipogenesis in 3T3-L1 adipocytes was investigated. The results showed that o-coumaric acid and rutin had the highest inhibition on intracellular triglyceride (61.3 and 83.0%, respectively) among 15 phenolic acids and 6 flavonoids tested. However, the oil red o stained material (OROSM) showed that cell number in 3T3-L1 adipocytes was not influenced by those compounds. For glycerol-3-phosphate dehydrogenase (GPDH) activity, the data indicated that o-coumaric acid and rutin had the highest inhibition on GPDH activity (54.2 and 66.8%, respectively) among the compounds tested. o-Coumaric acid and rutin also inhibited the expression of PPARgamma, C/EBPalpha and leptin and then up-regulated expression of adiponectin at the protein level. Some naturally occurring antioxidants efficiently suppressed adipogenesis in 3T3-L1 adipocytes. These results suggest that o-coumaric acid and rutin targeted for adipocyte functions could be effective in improving the symptoms of metabolic syndrome.     

Comparison of larval performance between Thai and Vietnamese freshwater giant prawn, Macrobrachium rosenbergii (de Man): a preliminary study

This study compared the performance of larvae of giant freshwater prawn (Macrobrachium rosenbergii) from Thailand and Vietnam. Thai broods were collected from a farm and Vietnamese broods were collected from a river. Larvae from these two sources were raised in identical environments, and their rates of development and survival as well as their ability to withstand environmental stresses were assessed. The first postlarval stage was observed in both strains on day 22. All larvae (100%) from Thai broods completed metamorphosis by day 36, but only 89% of the Vietnam strain completed metamorphosis by day 45. The stages of development were more uniform in larvae from the Thai stock than the Vietnamese stock. Survival differences were observed within the first week (92% in the Thai group vs. 55% in the Vietnamese group). These differences were most pronounced beginning in week 5 (52% and 16% respectively) and week 7 (33% and 2% respectively). No clear difference was found in brood size and fecundity between the two groups. No difference in survival was observed between the two groups of larvae challenged with salinity, oxygen or formalin. Differential development rates of larvae in the same batch would naturally create conditions favourable for cannibalism and impaired ability to capture food. The earlier and more uniform rate of development, and subsequent higher rate of survival of Thai stock compared with the Vietnamese wild stock is suggested to be due to inadvertent selection in the domesticated stock.     

Effect of ractopamine on growth, carcass traits, and fasting heat production of U.S. contemporary crossbred and Chinese Meishan pure- and crossbred pigs.

Twenty-four U.S. crossbred (Duroc x White composite; D x Wc; 83.9 kg), 24 purebred Meishan (M; 59.4 kg), and 24 Meishan x White composite crossbred (M x Wc; 83.4 kg) barrows were allotted within genotype to a 16% CP corn-soybean meal diet or this basal diet + 20 ppm of ractopamine and allowed ad libitum access to feed for 52 d. No genotype x ractopamine interactions were detected (P greater than .05) in pigs for growth, lean cuts, ham and loin characteristics, leaf fat and visceral organ weights, fasting whole-animal heat production, and carcass traits except longissimus muscle area (LMA). The LMA was increased by ractopamine in D x Wc and M x Wc pigs (P less than .05) but not in M pigs. Compared with D x Wc and M x Wc pigs, M pigs had lower ADG, ADFI, and gain to feed ratio (G/F), shorter carcasses, and lower dressing percentage, LMA, predicted amount of muscle, weights of trimmed picnic, loin, and ham cuts, percentage of ham lean, and CP in ham lean, but heavier liver, kidneys, pancreas, and entire gastrointestinal tract with greater percentage of ham fat and ham bone (P less than .05). The M x Wc pigs had lower ADG, G/F, dressing percentage, LMA, amount of muscle, weights of trimmed cuts, and percentage of ham lean but heavier lungs, pancreas, stomach, and large intestine than did D x Wc pigs (P less than .05). Supplemental ractopamine increased ADG, G/F, dressing percentage, amount of muscle, trimmed loin weight, percentage of ham lean, and CP in ham lean and decreased weights of heart, lungs, kidneys, and pancreas in pigs (P less than .05). Neither genotype nor ractopamine had any effect on 4- to 24-h postprandial whole-animal heat production of pigs (P greater than .05). These results indicate that ractopamine will improve growth performance and carcass leanness in pure- and crossbred Meishan pigs.     

Crossability percentages of bread wheat landraces from Sichuan Province,China with rye

Summary The crossabilities of 177 landraces of wheat (Triticum aestivum L.) from Sichuan Basin and its adjacent mountain ranges with rye (Secale cereale L.) have been tested. 16 landraces possess a higher crossability than Chinese Spring, 34 landraces have a similar and 127 landraces have a lower crossability than Chinese Spring or are non-crossable with rye. Most landraces with high crossability occur in Qinling Mountain and Dabashan Mountain Ranges in north of Sichuan and the valleys of Minjiang River, Fujiang River and Jialinjiang River in Sichuan Basin.     

Genomic Relationships of Triticum sharonense with Other S-Genome Diploid Triticum Species

Triticum sharonense was hybridized with autotetra-ploid T. speltoides, T. longissimum and T. bicorne. Meiotic analysis of these hybrids showed that T. sharonense is almost equally related to both T. speltoidcs and T. longissimum, while it is comparatively distant from T. bicorne. Therefore, this study does not support treating T. sharonense as a subspeies or variety of T. lottgissimum.     

Sources of resistance to head scab in Triticum

Multi-floret and single-floret injection inoculation methods were used to test 1076 accessions of Triticum for resistance to initial infection and resistance to pathogen spread within spike tissue respectively. The data obtained between years or inoculation dates were comparatively similar and were little influenced by climatic factors, especially humid condition. All the tested materials uniformly exhibited susceptibility to initial infection, but there were great differences in resistance to spread among species or cultivars. The diploid and tetraploid wheats were severely susceptible. Only 30 genotypes of common wheat of T. aestivum concv. vulgare showed high resistance to spread. The highest frequency of high spread resistance existed in these landraces which were distributed in Zhejiang, Hubei, Hunan, Jiangsu provinces and Shanghai City, Guizhou, Sichuan and Yunnan provinces ranked second. The majority of wheat landraces from Hebei, Shanxi, Shaanxi Province and Beijing City showed susceptibility or high susceptibility. No resistance was found in Tibetian wheats. This revised version was published online in July 2006 with corrections to the Cover Date.     

Genetic variability for resistance to sorghum downy mildew (Peronosclerospora sorghi) and Rajasthan downy mildew (P. heteropogoni) in the tropical/sub-tropical Asian maize germplasm

Downy mildews cause considerable damage to maize (Zea mays L.) worldwide, particularly in the tropical Asia. We have evaluated a set of 42 tropical/sub-tropical maize inbred lines developed in different countries in Asia (India, Thailand and Philippines), and Mexico, for analysing the genetic variability for resistance to sorghum downy mildew [Peronosclerospora sorghi; SDM] and Rajasthan downy mildew [P. heteropogoni; RDM]. Experiments were carried out in replicated trials under artificial infection in field conditions against SDM and RDM at Mandya in Karnataka, India, and Udaipur in Rajasthan, India, respectively, during 1999 and 2000. The study resulted in identification of five inbred lines offering consistent and strong resistance to both SDM and RDM, while several inbred lines revealed resistance only to RDM. It was also revealed that the SDM-resistant inbreds are invariably resistant to RDM, while the RDM-resistant inbreds might show differential responses to the SDM. The maize inbred lines identified in this study with broad-spectrum resistance to downy mildews could be potentially useful for basic and applied research work on downy mildews in tropical Asia. This revised version was published online in August 2006 with corrections to the Cover Date.