Bitter gourd suppresses lipopolysaccharide-induced inflammatory responses

Bitter gourd ( Momordica charantia L.) is a popular tropical vegetable in Asian countries. Previously it was shown that bitter gourd placenta extract suppressed lipopolysaccharide (LPS)-induced TNFalpha production in RAW 264.7 macrophage-like cells. Here it is shown that the butanol-soluble fraction of bitter gourd placenta extract strongly suppresses LPS-induced TNFalpha production in RAW 264.7 cells. Gene expression analysis using a fibrous DNA microarray showed that the bitter gourd butanol fraction suppressed expression of various LPS-induced inflammatory genes, such as those for TNF, IL1alpha, IL1beta, G1p2, and Ccl5. The butanol fraction significantly suppressed NFkappaB DNA binding activity and phosphorylation of p38, JNK, and ERK MAPKs. Components in the active fraction from bitter gourd were identified as 1-alpha-linolenoyl-lysophosphatidylcholine (LPC), 2-alpha-linolenoyl-LPC, 1-lynoleoyl-LPC, and 2-linoleoyl-LPC. Purified 1-alpha-linolenoyl-LPC and 1-linoleoyl-LPC suppressed the LPS-induced TNFalpha production of RAW 264.7 cells at a concentration of 10 microg/mL.     

Alpha-eleostearic acid and its dihydroxy derivative are major apoptosis-inducing components of bitter gourd

Bitter gourd ( Momordica charantia L.) pericarp, placenta, and seed extracts were previously shown to induce apoptosis in HL60 human leukemia cells. To determine the active component that induces apoptosis in cancer cells, bitter gourd ethanol extract was fractionated by liquid-liquid partition and silica gel column chromatography. Several fractions obtained by silica gel column chromatography inhibited growth and induced apoptosis in HL60 cells. Among them, fraction 7 had the strongest activity in inhibiting growth and inducing apoptosis in HL60 cells. A component that induced apoptosis in HL60 cells was then isolated from fraction 7 by another silica gel column chromatography and high-performance liquid chromatography (HPLC) using a C18 column and was identified as (9Z,11E,13E)-15,16-dihydroxy-9,11,13-octadecatrienoic acid (15,16-dihydroxy alpha-eleostearic acid). 15,16-Dihydroxy alpha-eleostearic acid induced apoptosis in HL60 cells within 5 h at a concentration of 160 microM (50 microg/mL). (9Z,11E,13E)-9,11,13-Octadecatrienoic acid (alpha-eleostearic acid) is known to be the major conjugated linolenic acid in bitter gourd seeds. Therefore, the effect of alpha-eleostearic acid on the growth of some cancer and normal cell lines was examined. alpha-Eleostearic acid strongly inhibited the growth of some cancer and fibroblast cell lines, including those of HL60 leukemia and HT29 colon carcinoma. alpha-Eleostearic acid induced apoptosis in HL60 cells after a 24 h incubation at a concentration of 5 microM. Thus, alpha-eleostearic acid and the dihydroxy derivative from bitter gourd were suggested to be the major inducers of apoptosis in HL60 cells.     

Quantitative PCR assay for the detection of the parasitic ciliate <Emphasis Type="Italic">Cryptocaryon irritans</Emphasis>

We developed a quantitative PCR assay for detecting the parasitic ciliate Cryptocaryon irritans, which causes “white spot disease” in marine fishes, from the natural environment. A specific primer set for C. irritans was designed and its high specificity was confirmed in silico: almost all of the sequences deposited in the GenBank nucleotide database were covered, 22/23 for the forward primer and 7/7 for the reverse primer. We estimated that there were 3,415.9 rRNA gene copies per genome of C. irritans. In artificial mixture experiments to validate whether the qPCR assay is applicable to natural samples, the estimated copy numbers showed significantly positive correlations with the number of theronts added (p < 0.001). When we applied this qPCR assay to natural samples collected bimonthly from surface and bottom seawaters at an aquaculture site (water depth, 10 m) from May 2009 to March 2010, we only detected C. irritans (112.0 ± 6.3 cells/l) in the surface seawater sample in November. This qPCR assay is a useful tool for detecting C. irritans rapidly and quantitatively in natural environments; it could also help advance our understanding of the ecology of C. irritans, as well as facilitate the diagnosis of the disease.     

IL‐12p40 gene expression in lung and hilar lymph nodes of MPS‐resistant pigs

Mycoplasma pneumonia of swine (MPS) is caused by Mycoplasma hyopneumoniae (M.hp) and is a common chronic respiratory disease of pigs. Recently, a genetically selected variant of the Landrace pig (Miyagino L2) has a lower incidence of pulmonary MPS lesions. We investigated the pathological and immunological characteristics of MPS resistance in these pigs (n = 24) by comparing with the normal landrace pig (control: n = 24). The pathological MPS lung lesion score in MPS‐selected landrace pigs was significantly lower than in the control. The gene expression of interleukin (IL)‐12p40, which acts as a chemoattractant and a component of the bioactive cytokines IL‐12 and IL‐23, was significantly higher at the hilar lymph nodes, lung, and spleen in MPS‐selected landrace pigs than in control landrace pigs, and these were negatively correlated with the macroscopic MPS lung lesion score. In summary, we demonstrate that resistance against MPS in Miyagino L2 pigs is associated with IL‐12p40 up‐regulation, in comparison with normal landrace pigs without the MPS vaccine. In addition, a comparative study of macroscopic MPS lung lesions and IL‐12p40 gene expression in lung and hilar lymph nodes may lead to beneficial selection traits for the genetic selection for MPS resistance in pigs.     

The effect of high ambient temperature on Ca,P and Mg balance and bone turnover in high‐yielding dairy cows

We investigated the effect of heat stress on Ca, P and Mg balance and bone turnover in lactating cows. In a 2 × 2 crossover design, four multiparous lactating Holstein cows were kept in a chamber and subjected to a constant moderate (18°C) ambient temperature (MT) or high (28°C) ambient temperature (HT). The cows were fed total mixed ration (Ca, 0.7%; P, 0.4%; Mg, 0.2%) ad libitum. The milk yield under HT (35.4 kg/day) tended to be lower (P < 0.10) than that under MT (43.2 kg/day). The concentrations of milk P (P < 0.05) and Mg (P < 0.01) were significantly lower under HT than MT. The Ca, P and Mg intake (P < 0.10); Ca (P < 0.10), P, and Mg (P < 0.05) secretion into milk; and Ca (P < 0.05), P (P < 0.01), and Mg (P < 0.05) absorption in the intestine were lower under HT than MT. The plasma osteocalcin, a marker of bone turnover, was significantly lower (P < 0.05) under HT than MT. Heat stress did not affect plasma C‐telopeptide of collagen type I, a bone resorption marker, and plasma parathyroid hormone concentration.     

Molecular cloning of the canine c-Met/HGF receptor and its expression in normal and regenerated liver

The c-Met proto-oncogene is the receptor for hepatocyte growth factor (HGF), which is a member of the tyrosine kinase family. Activation of the HGF/c-Met signal pathway leads to cell proliferation, motility, regeneration, and morphogenesis. In this study, the complete nucleotide sequence of complementary DNA (cDNA) of canine c-Met was cloned, and its distribution was determined in tissues. The canine c-Met cDNA clone had an open reading frame of 4419 bp that encoded a putative polypeptide of 1383 amino acids. The c-Met mRNA was expressed in a variety of canine tissues including peripheral blood mononuclear cells (PBMC), bone marrow, liver, kidney, lung, stomach, uterus, testis, thymus, lymph node, small intestine, colon, adrenal gland, thyroid gland, heart, muscle, skin, pancreas, ovary, prostate, spleen, fat, cerebrum, and cerebellum. In addition, the c-Met mRNA expression in normal and regenerated liver was examined. The levels of the mRNA increased 2-fold in regenerated liver compared to that found in normal liver, indicating that c-Met is involved in various functions including remodeling of canine hepatocytes.     

Relationship between milk production and plasma concentrations of oxidative stress markers during hot season in primiparous cows

Oxidative stress markers, ascorbic acid, and sulfhydryl (SH) residue concentration in primiparous cows' plasma and their relationship to milking performance during hot seasons were investigated. The rectal temperature of cows correlated negatively with SH residue (r = −0.38, n  = 38, P  < 0.05) and ascorbic acid (r = −0.34, P  < 0.05) concentrations in the cows' plasma. The group with a higher concentration of ascorbic acid over the mean value produced significantly more milk ( P  < 0.05) than did the group with a lower ascorbic acid concentration. Although the cows' milk production showed a positive correlation with ascorbic acid concentration in plasma (r = 0.47, P  < 0.05), the relation of SH residue concentration to milk yield was not constant. The plasma SH residue concentration during the hot season correlated positively with milk protein % (r = 0.38, P  < 0.05), lactose % (r = 0.35, P  < 0.05), and solid-non-fat (SNF) % (r = 0.47, P  < 0.05), respectively, but not with fat %. On the other hand, ascorbic acid concentration in plasma showed negative correlations with milk fat % (r = −0.34, P  < 0.05) and protein % (r = −0.49, P  < 0.05), but correlated positively with lactose % (r = 0.52, P  < 0.05). The produced amount (g/day) of milk protein (r = 0.42, P  < 0.05), lactose (r = 0.61, P  < 0.05), and SNF (r = 0.56, P  < 0.05) showed positive correlations with ascorbic acid concentration in plasma.     

Localization of CO2 source by a hexapod robot equipped with an anemoscope and a gas sensor

A hexapod robot was developed to explore an agricultural field and gather agronomical information such as soil nutrients and plant growth. In addition, a sensor device for the robot to track a gas source was developed. The sensor device consisted of a CO₂ gas sensor and an anemoscope. The gas sensor detected the concentration of CO₂ in the surroundings, and the anemoscope indicated the upwind direction from the robot. An indoor test was conducted during which the hexapod robot tracked windward direction using the sensor device. According to the results, the hexapod robot with the sensor device successfully localized a CO₂ source that was in the upwind direction. The robot paused on the spot under windless conditions, and walked windward as soon as the wind started blowing again.