CORRELATIONS BETWEEN A FLUORIMETRIC and MASS FRAGMENTOGRAPHIC METHOD FOR THE DETERMINATION OF 3-METHOXY-4- HYDROXYPHENYLACETIC ACID and TWO MASS FRAGMENTOGRAPHIC METHODS FOR THE DETERMINATION OF 3-METHOXY-4- HYDROXYPHENYLETHYLENE GLYCOL IN CEREBROSPINAL FLUID

One-hundred and twenty-two lumbar cerebrospinal fluid (CSF) samples were assayed for 3-methoxy-4-hydroxyphenylacetic acid (HVA) by both a fluorimetric and mass fragmentographic method. The correlation coefficient (cc) and residual standard deviation (Syx) of the results were calculated as 0.966 and 23.3 ng/ml, respectively. If only samples containing less than 100ng/ml of HVA were considered, somewhat different values for cc and Syx were found (0.854 and 10.0 ng/ml, respectively). The data obtained by the fluorimetric method were consistently 17% lower than those obtained by the mass fragmentographic method. Spiking experiments resulted in 96.5 ± 7.8% recovery for the fluorimetric method, whereas the use of a deuterated internal standard was found to compensate completely for losses in the mass fragmentographic method. In addition the correlation between two different mass fragmentographic methods for the simultaneous determination of HVA and 3-methoxy-4-hyd-roxyphenylethylene glycol (MOPEG) in CSF was studied. The measurements were performed in different laboratories and resulted in correlation coefficients of 0.941 and 0.940 and residual standard deviations of 7.6 and 1.0 ng/ml for HVA and MOPEG, respectively. From all data we conclude that mass fragmentographic methods for the determination of catecholamine metabolites in CSF are superior to fluorimetric methods because of their selectivity, reproducibility and accuracy.     

Identification of two Thormählen-positive compounds from melanotic urine by gas chromatography—mass spectrometry

The isolation of two Thormählen-positive compounds from the urine of a patient with malignant melanoma and the elucidation of their structure by gas chromatography—mass spectrometry is described. The compounds were isolated using a poly-N-vinylpyrrolidone column and separated by preparative thin-layer chromatography. After elution they were analyzed by gas chromatography and gas chromatography—mass spectrometry as their trimethylsilyl derivatives and after hydrolysis also as their tert.-butyldimethylsilyl derivatives. The results showed the main Thormählen-positive compound A to be the glucuronide of 5-hydroxy-6-methoxyindole, whereas the minor compound AX appeared to be the glucuronide of its isomer 6-hydroxy-5-methoxyindole.     

Commitment to the First Cortical Reorganization During Conjugation in Stylonychia mytilus: An Argument for Homology with Cortical Development During Binary Fission

The asexual nature of the first cortical reorganization of conjugation in Stylonychia was analyzed by comparing the effect of amputation performed at different stages of early conjugation to that performed on vegetative cells at different stages of the cell cycle. Amputation of vegetative cells delineated a point of commitment to binary fission at 0.51–0.57 of the cell cycle. Cells amputated before this point were induced to undergo the regenerative mode of asexual development, but those amputated after this point continued with binary fission. In parallel, during conjugation a similar commitment was made around the time of formation of tight mating-pairs: early conjugants amputated around this time might undergo regeneration, and those operated on after this stage continued with the first cortical reorganization as in typical conjugants. The two mates of a pair might differ in their response to amputation, suggesting that the timing of commitment to the first cortical reorganization is not related to the events of conjugation, but rather is individually determined in the vegetative cycle of the cells before they pair up in mating. These observations provide support for the notion that the first cortical reorganization of conjugants is homologous to the asexual mode of cortical development in dividers, according to the theory of developmental heterochrony in the sexual reproduction of hypotrichs. The timing of commitment to the first cortical reorganization was found to temporally correlate with the entrance of the micronuclei into meiosis. Since the first cortical reorganization can proceed without the micronucleus, this raises the possibility that initiation of micronuclear meiosis is closely coupled with, and may be determined by, the commitment to the first cortical reorganization.     

A correlation between BCL-2 modifying factor,p53 and livin gene expressions in cancer colon patients

Accumulating evidence has revealed that livin gene and BCL-2 modifying factor (BMF) gene are closely associated with the initiation and progression of colon carcinoma by activating or suppressing multiple malignant processes. Those genes that can detect colon - cancer are a promising approach for cancer screening and diagnosis. This study aimed to evaluate correlation between livin, BMF and p53 genes expression in colon cancer tissues of patients included in the study, and their relationship with clinicopathological features and survival outcome in those patients. In this study, 50 pathologically diagnosed early cancer colon patients included and their tissue biopsy with 50 matched adjacent normal tissue, and 50 adenoma tissue specimens were analyzed for livin gene and BMF gene expressions using real time PCR. The relationship of those genes expressions with clinicopathological features, tumor markers, Time to Progression and overall survival for those patients were correlated in cancer colon group. In this study, there was a significant a reciprocal relationship between over expression of livin gene and down regulation of BMF and p53 genes in colon cancer cells. Livin mRNA was significantly higher, while BMF and p53 mRNA were significantly lower in colorectal cancer tissue compared to benign and normal colon tissue specimens (P < 0.001), however, this finding was absent between colon adenomas and normal mucosa. There was a significant association between up regulation of livin and down regulation of BMF and p53 expressions with more aggressive tumor (advanced TNM stage), rapid progression with metastasis and decreased overall survival in cancer colon patients, hence these genes can serve as significant prognostic markers of poor outcome in colon cancer patients. This work highlights the role of livin, BMF and p53 genes in colorectal tumorigenesis and the applicability of using those genes as a diagnostic and prognostic markers in patients with colon carcinoma and as a good target for cancer colon treatment in the future.     

中条山油松人工林下物种对群落物种丰富度分布格局的贡献

该研究以中条山油松人工林群落为研究对象,研究林下不同大小的子群落对群落物种丰富度分布格局的贡献,并确定影响该区域群落物种丰富度分布格局的关键种,为区域物种多样性保护提供理论依据。结果表明:(1)该地区林下物种频度分布格局呈明显右偏,且不同样方物种丰富度存在明显差异。(2)常见种对群落丰富度分布格局的贡献大于稀有种。(3)最常见的物种解释了整个群落物种丰富度格局的88.4%(P0.01),而最稀有物种仅解释了24.5%(P0.05),去除最稀有物种后,最常见物种可以解释剩余物种的90.8%(P0.01),而去除最常见物种后,最稀有物种仅能解释剩余物种的48.6%(P0.01)。(4)当子群落中常见种越多时,子群落与整个群落的丰富度分布格局相关性越高。(5)连翘(Forsythia suspensa)、太平花(Philadelphus pekinensis)、鞘柄菝葜(Smilax stans)、多歧沙参(Adenophora wawreana)、金花忍冬(Lonicera chrysantha)等对群落物种丰富度分布格局的贡献最大,但并非越常见的物种对群落丰富度格局贡献越大。(6)与频度较高物种的种间关联度低的物种对于群落物种的分布格局贡献较大,但此解释并不适用于稀有种。研究发现,稀有种对中条山油松人工林群落物种丰富度分布格局存在较大的贡献,所以在油松人工林物种多样性保护过程中并不能只关注常见种而忽视稀有种。     

Aspects of reproduction and their implications for the conservation of the endangered limpet,Patella ferruginea

Summary

The protandric limpet, Patella ferruginea G., is the most endangered marine species in the Western Mediterranean and is at serious risk of extinction. Nevertheless, its biology and ecology are little known. In the present work, several reproductive aspects are studied. Recruitment take place in June and the largest individuals are the most scarce. The sex ratio is slanted towards the largest sizes, and the species seems to show sex change at sizes from 60 mm upwards, although males can be observed up to 80 mm. There is no correlation between size of oocytes and shell length; however, the larger females contribute greatly to the reproductive event, with high fecundity and GSI values. The mean diameter of oocytes was observed to be 149.78 μm, whereas the heads of spermatozoa were 3.78 μm long. The results of the present study increase the maximum length for males by double that cited in previous literature and highlight the importance of strictly protecting females (i.e., most of the largest individuals, >60 mm), since their population percentage is very low, in order to develop adequate strategies to preserve the species.     

Production of endothelial progenitor cells obtained from human Wharton's jelly using different culture conditions

Endothelial progenitor cells (EPC) participate in revascularization and angiogenesis. EPC can be cultured in vitro from mononuclear cells of peripheral blood, umbilical cord blood or bone marrow; they also can be transdifferentiated from mesenchymal stem cells (MSC). We isolated EPCs from Wharton's jelly (WJ) using two methods. The first method was by obtaining MSC from WJ and characterizing them by flow cytometry and their adipogenic and osteogenic differentiation, then applying endothelial growth differentiating media. The second method was by direct culture of cells derived from WJ into endothelial differentiating media. EPCs were characterized by morphology, Dil-LDL uptake/UEA-1 immunostaining and testing the expression of endothelial markers by flow cytometry and RT-PCR. We found that MSC derived from WJ differentiated into endothelial-like cells using simple culture conditions with endothelium induction agents in the medium.     

Assessment of essential fatty acid and omega3-fatty acid status by measurement of erythrocyte 20:3omega9 (Mead acid), 22:5omega6/20:4omega6 and 22:5omega6/22:6omega3

BACKGROUND: Early suspicion of essential fatty acid deficiency (EFAD) or omega3-deficiency may rather focus on polyunsaturated fatty acid (PUFA) or long-chain PUFA (LCP) analyses than clinical symptoms. We determined cut-off values for biochemical EFAD, omega3-and omega3/22:6omega3 [docosahexaenoic acid (DHA)]-deficiency by measurement of erythrocyte 20:3omega9 (Mead acid), 22:5omega6/20:4omega6 and 22:5omega6/22:6omega3, respectively. METHODS: Cut-off values, based on 97.5 percentiles, derived from an apparently healthy omnivorous group (six Dominica breast-fed newborns, 32 breast-fed and 27 formula+LCP-fed Dutch low-birth-weight infants, 31 Jerusalem infants, 33 Dutch 3.5-year-old infants, 69 omnivorous Dutch adults and seven Dominica mothers) and an apparently healthy group with low dietary LCP intake (81 formula-fed Dutch low-birth-weight infants, 12 Dutch vegans). Cut-off values were evaluated by their application in an EFAD suspected group of 108, mostly malnourished, Pakistani children, three pediatric patients with chronic fat-malabsorption (abetal-ipoproteinemia, congenital jejunal and biliary atresia) and one patient with a peroxisomal beta-oxidation disorder. RESULTS: Erythrocyte 20:3omega9, 22:5omega6/20:4omega6 and 22:5omega6/22:6omega3 proved age-dependent up to 0.2 years. Cut-off values for ages above 0.2 years were: 0.46mol% 20:3omega9 for EFAD, 0.068mol/mol 22:5omega6/20:4omega6 for omega3-deficiency, 0.22mol/mol 22:5omega6/22:6omega3 for omega3/DHA-marginality and 0.48mol/mol 22:5omega6/22:6omega3 for omega3/DHA-deficiency. Use of RBC 20:3omega9 and 22:5omega6/20:4omega6 cut-off values identified 20.4% of the Pakistani subjects as EFAD+omega3-deficient, 12.9% as EFAD+omega3-sufficient, 38.9% as EFA-sufficient+omega3-deficient and 27.8% as EFA-sufficient+omega3-sufficient. The patient with the peroxisomal disorder was classified as EFA-sufficient, omega3-sufficient (based on RBC 22:5omega6/20:4omega6) and omega3/DHA-deficient (based on RBC 22:5omega6/22:6omega3). The three other pediatric patients were classified as EFAD, omega3-deficient and omega3/DHA-deficient. CONCLUSION: Use of the combination of the present cut-off values for EFA, omega3 and omega3/DHA status assessment, as based on 97.5 percentiles, may serve for PUFA supplement intervention until better concepts have emerged.