Cadherin-1, -2 and -4 expression in the cranial ganglia and lateral line system of developing zebrafish

Cadherins are cell adhesion molecules that play important roles in development of a variety of tissues and organs including the nervous system. In this study we analyzed expression patterns of three zebrafish classical (type I) cadherins (cadherin-1, -2, and -4) in the embryonic zebrafish cranial ganglia and lateral line system using in situ hybridization and immunohistochemical methods. All three cadherins exhibit distinct spatiotemporal patterns of expression during cranial ganglia and lateral line system development. cadherin-1 message was detected in the trigeminal and facial ganglia, in the lateral line ganglia, and in most of neuromasts in the lateral lines. Cadherin-2 mRNA and protein were expressed by the majority of the cranial ganglia and lateral line system. Both cadherins were found in embryos younger than 24 hours post fertilization as well as in 2-3-day old embryos and larvae. In contrast, cadherin-4 mRNA and protein expression was detected in embryos older than 30 hours post fertilization and limited to the trigeminal, statoacoustic, and vagal cranial ganglia, and the lateral line ganglia of older embryos and larvae.     

Application of high-performance liquid chromatography-mass spectrometry to detection of diuretics in human urine

A rapid, sensitive and reliable high-performance liquid chromatographic-mass spectrometric method for the detection of 25 diuretics in human urine has been developed. Atmosphere pressure chemical ionization (APCI) and electrospray ionization (ESI) modes were evaluated. A 2-ml volume of urine was extracted under basic conditions and separated on an Agilent Zorbax SB-C(18) column (150 x 2.1 mm, 5 microm). The mobile phase consisted of formic ammonium-formic acid buffer (pH 3.5) and acetonitrile. The effects of capillary temperature, sheath gas pressure and compositions of mobile phase on the sensitivity were studied. The recoveries of most of the diuretics were 75-95%. In the full scan mode, the limits of detection of the 25 diuretics were 0.25-25 ng/ml for APCI and 0.6-250 ng/ml for ESI. Under the optimal conditions, 14 diuretics from authentic urine samples were detected successfully by LC-APCI-MS. To obtain more fragmentation information on the chemical structure for positive confirmation, tandem mass analysis was also investigated.     

Preferential expression of cystein-rich secretory protein-3 (CRISP-3) in chronic pancreatitis

BACKGROUND: Chronic pancreatitis (CP) is a progressive inflammatory process resulting in exocrine and endocrine pancreatic insufficiency in advanced stages. Cysteine-rich secretory protein (CRISP-3) has been identified as a defense-associated molecule with predominant expression in the salivary gland, pancreas and prostate. AIMS: In this study, we investigated CRISP-3 expression in normal pancreatic tissues, chronic pancreatitis tissues, pancreatic cancer tissues and pancreatic cancer cell lines, as well as in other gastrointestinal organs. MATERIALS AND METHODS: 15 normal pancreatic tissues, 14 chronic pancreatitis tissues and 14 pancreatic cancer tissues as well as three pancreatic cancer cell lines were analyzed. Moreover, hepatocellular carcinoma and esophageal, stomach and colon cancers were also analyzed together with the corresponding normal controls. RESULTS: CRISP-3 was expressed at moderate to high levels in chronic pancreatitis tissues and at moderate levels in pancreatic cancer tissues but at low levels in normal pancreatic tissues, and was absent in three pancreatic cancer cell lines. CRISP-3 expression was below the level of detection in all cancerous gastrointestinal tissues and in all normal tissues except 2 of 16 colon tissue samples. CRISP-3 mRNA signals and immunoreactivity were strongly present in the cytoplasm of degenerating acinar cells and in small proliferating ductal cells in CP tissues and CP-like lesions in pancreatic cancer tissues. In contrast, CRISP-3 expression was weak to absent in the cytoplasm of cancer cells as well as in acinar cells and ductal cells in pancreatic cancer tissues and normal pancreatic tissues. CONCLUSION: These results reveal that the distribution of CRISP-3 in gastrointestinal tissues is predominantly in the pancreas. High levels of CRISP-3 in acinar cells dedifferentiating into small proliferating ductal cells in CP and CP-like lesions in pancreatic cancer suggests a role of this molecule in the pathophysiology of CP.     

Induction of interleukin-6 by coal containing bioavailable iron is through both hydroxyl radical and ferryl species

Coal mining causes health problems, such as pneumoconiosis. We have previously shown that prevalence of pneumoconiosis in workers from various coalmine regions positively correlates with levels of bioavailable iron (BAI) in the coals from that region. In the present study, the nature of reactive oxygen species formed by BAI in the coals and its mechanisms of the induction of biological responses were investigated. Human lung epithelial cell line, A549 cells, were used to examine the induction of interleukin-6 (IL-6), a pro-inflammatory cytokine, which is known to play a crucial role in the development of pneumoconiosis. We found that levels of IL-6 protein as well as its mRNA were significantly increased in the cells treated for 24 h with 20 microg/cm2 of the BAI-containing Pennsylvania (PA) coal; for example we observed 6.7-fold increase in IL-6 protein. Levels of IL-6 protein in cells treated with the Utah (UT) coal containing low-BAI were only 1.9-fold of the control levels. The enhancing effect on the IL-6 by the PA coal was similar to that caused by hydrogen peroxide. Superoxide dismutase (SOD), catalase (CAT), and N-acetyl-L-cysteine (NAC) all had inhibitory effects on the PA coal-induced IL-6 formation. However, CAT had the least protective effect as compared to SOD and NAC. Our results indicate that BAI in the PA coal may induce IL-6 through both ferryl species (via iron autoxidation) and hydroxyl radicals (via the Fenton/Haber Weiss reactions).     

Production of gamma-linolenic acid by disrupted mycelia of Mortierella isabellina

AIMS: To optimize the production of linolenic acid by disrupted mycelia of Mortierella isabellina. METHODS AND RESULTS: Effects of incubation conditions such as incubation time, pH of reaction mixture, concentration of Mg2+ or malate and incubation temperature on production of linolenic acid were studied. The production of gamma-linolenic acid reached 224 mg g-1 dry cells when the reaction mixture was composed of 1.0 g (dry mycelial mass) of disrupted mycelia of M. isabellina, 50 ml (50 mmol l(-1)) potassium phosphate buffer supplemented with 0.312 mmol l(-1) of Mg2+ and 10 mmol l(-1) of malate, pH 7.0 and incubated at 5 degrees C for 1 day. CONCLUSIONS: Incubation temperature, concentration of Mg2+ and malate showed major effects on the increased linolenic acid production. SIGNIFICANCE AND IMPACT OF THE STUDY: This study highlights conditions for increasing gamma-linolenic acid production by cell-free mycelia of M. isabellina and an insight into rapidly gaining high production of polyunsaturated fatty acids.     

Fasting differentially regulates expression of agouti-related peptide,pro-opiomelanocortin,prepro-orexin,and vasoactive intestinal polypeptide mRNAs in the hypothalamus of Japanese quail

Research in mammals has established the existence of a neuronal network that lies within the hypothalamus and that regulates energy homeostasis. However, it is unknown whether this system has been evolutionarily conserved. The objective of the present study was therefore to examine the influence of the agouti-related peptide (AGRP), pro-opiomelanocortin (POMC), prepro-orexin, and vasoactive intestinal polypeptide (VIP) genes on energy balance in birds by quantifying the effect of a 24-h fast on their expression in the hypothalamus of the Japanese quail. In situ hybridization revealed strong signals for AGRP and POMC mRNAs in the infundibular nucleus (IN), for prepro-orexin in the lateral hypothalamic area (LHy) and periventricular hypothalamic nucleus, and for VIP in the LHy. POMC mRNA was co-localized with -melanocyte-stimulating hormone-like immunoreactivity in individual IN neurons. Compared with the ad-libitum-fed state, a 24-h fast resulted in a 2.2-fold increased expression of AGRP mRNA in the IN. However, fasting did not induce changes in POMC, prepro-orexin, or VIP mRNAs. The results suggest an involvement of the central melanocortin system in the regulation of energy balance in birds, as in mammals. In contrast, orexins in birds may be primarily involved in the control of physiological functions other than energy homeostasis.This research was supported by a Commonwealth Fellowship to D.P.-S. and a BBSRC Fellowship to T.B.