高效液相色谱直接衍生化法测定血清游离脂肪酸组成

目的脂肪酸摄入和代谢在高血压的发病和进展中发挥作用,但血清游离脂肪酸（FFA）组成测定方法尚未统一,本研究探讨高效液相色谱直接衍生化法测定空腹血清游离脂肪酸组成的可靠性。方法社区筛选出原发性高血压患者103例,其中肥胖患者65例,非肥胖患者38例,和血压正常个体128例,其中肥胖个体70例,非肥胖者58例,年龄35～59岁,各组饮食习惯相近。采空腹静脉血离心取上清分成两个1．5ml,用75％的乙醇60℃下除去血清中与FFA结合的白蛋白,无需常规的分离步骤,应用2-硝基苯肼盐酸盐（2-NPH·HCI）作为衍生试剂,在1-乙基-3-（3-二甲基氨丙基碳化二亚胺）盐酸盐（1-EDC·HCI）的催化下,直接衍生血清中的FFA,形成不挥发的能在紫外-可见光区产生吸收的脂肪酸的肼盐衍生化物,再用磷酸缓冲液酸化增加各种脂肪酸的分离度,衍生化的脂肪酸用正己烷萃取法与其它的酰肼和干扰成分分离出来,氮气吹干后甲醇定容,应用C18反向色谱柱进样30min内洗脱分析测定。结果共测定血清中主要的9种脂肪酸,从C16：0到C22：6,未鉴定的峰少于总量的1％。脂肪酸组成以个体脂肪酸浓度表示,浓度单位μmol／L。应用C17：0作内标,各脂肪酸测定的线性范围为2．6～200．0μmol／L,回收率在87．8％～120．5％之间,重复性检验变异系数〈10％,相关系数为0．973～0．999,批内RSD范围为2．78％～12．90％,批间RSD范围为为0．95％～15．4％。与血压正常个体比较,原发性高血压患者血清游离多不饱和脂肪酸（PUFA）水平降低（P=0．004）,饱和脂肪酸（SFA）水平有增高趋势（P=0．06）,P／S比值降低（P=0．000）。结论高效液相色谱直接衍生化法测定空腹血清FFA组成重复性、准确性良好,可作为血清游离脂肪酸组成常规检测的有用的工具。     

乳腺癌超声征象与淋巴结转移关系的单因素及多因素分析

目的 探讨乳腺癌原发肿块、腋窝淋巴结超声征象与淋巴结转移的关系.方法 回顾性分析146例乳腺癌患者151个原发肿块(66个伴淋巴结转移,85个未转移)及腋窝淋巴结声像图特征,与病理结果对照,用单因素(X~2检验)和多因素(Logistic回归)分析各声像图特征与淋巴结转移的关系,对超声征象和病理的吻合度进行k系数检验.对乳腺癌中提示淋巴结转移的超声征象与有转移的腋窝淋巴结征象进行相关分析.结果 单因素和多因素分析显示肿块最大直径、血流分级及淋巴结最大皮质厚度与淋巴结转移关系密切(X~2=37.939、13.153、69.128,OR=6.632、3.714、35.442,P均<0.01),且与病理结果有较好的吻合度(k=0.501、0.690、0.673),能较准确地判断腋窝淋巴结转移.肿块最大直径、血流分级与淋巴结最大皮质厚度有较好相关性(X2=40.132、33.128,P均<0.01).肿块距体表距离以及间质纤维结构改变等征象对判断乳腺癌淋巴结转移有一定提示作用.结论 采用单因素及多因素分析乳腺癌超声征象能为术前准确判断腋窝淋巴结转移提供重要的依据.     

Identification of the dominant hydrogeochemical processes and characterization of potential contaminants in groundwater in Qingyuan,China, by multivariate statistical analysis

In karst areas, groundwater is an important water source for drinking and irrigation purposes; however, karst aquifers are vulnerable and recovery from damage is difficult. We collected surface water (pond and river water) and groundwater (hand-pump well, dug well, and borehole water) samples in Qingyuan city, China, to determine the major chemicals in the water with the primary goals of evaluating the geochemical composition, identifying the geochemical processes governing the water chemistry, and identifying the probable sources of potential contaminants in shallow and deep groundwater in the study area. The results revealed marked differences in water chemistry between shallow and deep groundwater. The groundwater composition was largely controlled by rock–water interactions, particularly the dissolution of evaporite minerals (e.g., calcite, gypsum, and anhydrite), and ion exchange processes were important drivers of the chemical compositions of groundwater in the study area. Moreover, in shallow and deep groundwater, Mg²⁺ and SO₄²⁻ concentrations were increased due to the long residence time of deep groundwater, while K⁺ and Na⁺ concentrations were decreased due to anthropogenic input. Finally, factor analysis of the major and trace elements differentiated between anthropogenic and geogenic sources of potential contaminants in karst aquifers.

In karst areas, groundwater is an important water source for drinking and irrigation purposes; however, karst aquifers are vulnerable and recovery from damage is difficult.     

Role of free fatty acids and insulin in determining free fatty acid and lipid oxidation in man.

Plasma FFA oxidation (measured by infusion of 14C-palmitate) and net lipid oxidation (indirect calorimetry) are both inhibited by insulin. The present study was designed to examine whether these insulin-mediated effects on lipid metabolism resulted from a decline in circulating FFA levels or from a direct action of the hormone on FFA/lipid oxidation. Nine subjects participated in two euglycemic insulin clamps, performed with and without heparin. During each insulin clamp study insulin was infused at two rates, 4 and 20 mU/m2.min for 120 min. The studies were performed with indirect calorimetry and 3-3H-glucose and 14C-palmitate infusion. During the control study plasma FFA fell from 610 +/- 46 to 232 +/- 42 to 154 +/- 27 mumol/liter, respectively. When heparin was infused basal plasma FFA concentration remained constant. During the control study, FFA/lipid oxidation rates decreased in parallel with the fall in the plasma FFA concentration. During the insulin/heparin study, plasma 14C-FFA oxidation remained unchanged while net lipid oxidation decreased. In conclusion, when the plasma FFA concentration is maintained unchanged by heparin infusion, insulin has no direct effect on FFA turnover and disposal. These results thus suggest that plasma FFA oxidation is primarily determined by the plasma FFA concentration, while net lipid oxidation is regulated by both the plasma FFA and the insulin level.     

In vitro and in vivo effects of increased concentrations of free fatty acids on free thyroxin measurements as determined by five assays

To compare in vitro and in vivo effects of increased concentrations of free fatty acids (FFA) on free thyroxin (FT4) values, we measured FT4 in three pooled sera supplemented with oleate and in serum from 18 euthyroid patients before and after an infusion of fat emulsion (Intralipid). We used five FT4 RIA kits: two two-step methods [Gammacoat, Baxter (GC); Ria-gnost, Behring (RG)], two analog RIAs [Amerlex-M, Amersham (AM); Coat-Ria, BioMérieux (CR)], and one kit with labeled antibodies [Amerlex-MAB*, Amersham (AA)]. In vitro, at the maximum oleate addition of 5 mmol/L, FT4 increased when measured by the GC and RG kits, decreased by the AM kit, and showed no significant change by the CR and AA kits. In vivo, post-Intralipid, FFA concentrations rose significantly and the FT4 changes agreed with the results of the in vitro experiments, except for the RG kit, for which FT4 increased in only nine patients. We conclude that in vitro oleate addition is useful to predict the in vivo effect of increased FFA on FT4 values; moreover, in serum from euthyroid subjects with high concentrations of FFA, FT4 analyzed with the CR or AA kits should better agree with normal results for thyrotropin than FT4 values measured with the other kits.     

Identification of potential serum biomarkers for gastric cancer by a novel computational method, multiple normal tissues corrected differential analysis

BackgroundGenes specifically expressed in one or a few tissues and upregulated in tumors are potentially good serum biomarkers.MethodsBy applying a recently developed computational method, called multiple normal tissues corrected differential analysis (MNTDA), we identified genes that are likely to be upregulated in the blood of gastric cancer patients as compared to normal controls.ResultsWe identified four genes (MMP-1, MMP-3, MMP-12, and CXCL5) as potential serum biomarkers for gastric cancer. Of these four genes, only MMP-1 was significantly upregulated in the sera of 40 gastric cancer patients, as compared to 40 control sera. The same pattern was observed in the second cohort of 80 gastric cancer patients and 80 controls. In a combined analysis, the level of serum MMP-1 in gastric cancer patients was significantly higher than the level in control samples (P < 0.0001). The use of MMP-1 was 62.5% sensitive and 62.5% specific in detecting gastric cancer patients. Patients with high serum levels of MMP-1 had a significantly worse outcome than patients with low serum MMP-1 levels. Finally, we determined that preoperative serum MMP-1 levels were prognostic, independent of tumor stage.ConclusionsMMP-1 is a potential prognostic marker for gastric cancer patients after gastrectomy.     

Neoadjuvant chemotherapy for breast cancer determined by chemosensitivity assay achieves better tumor response.

BACKGROUND: Neoadjuvant chemotherapy can potentially reduce tumor size and help downstage the tumor before definitive operation was performed. However, it was not possible to tell whether the patient would respond to the regimen until given. This difficulty can be overcome by testing the susceptibility of a sample of cancer cells in vitro: a "patient-tailored approach". In this pilot study, we attempt to demonstrate an improved response by this "patient-tailored" approach over standard regimen. MATERIALS AND METHODS: The study included 36 women with moderately advanced local breast cancer larger than 2 cm in diameter. Twelve were allocated to receive a standard regimen of 5-fluorouracil, epirubicin and cyclophosphamide (FEC) preoperatively as controls, and 24 were given the most suitable regimen according to testing; the options were FEC, cyclophosphamide, methotrexate and 5-fluorouracil (CMF), 5-fluorouracil, adriamycin and mitomycin C (FAM) and paclitaxel alone. The cell activities of drug-treated solid tumors were compared to controls with a highly sensitive ATP bioluminescence assay. Patients received chemotherapy according to sensitivity results and the tumor area clinically measured before and after chemotherapy. RESULTS: Sensitivity-directed treatment helped patients achieve a higher rate of complete clinical response (10/24 vs. 0/12), larger mean reduction in tumor area (75% vs. 26%), and 25% pathological complete response (pCR). The paclitaxel subgroup achieved 80% (pCR). CONCLUSION: It is a useful in vitro assay to provide a reference of the particular patient who received treatment according to her sensitivity result. It may improve pathologic complete response, clinical tumor response and lead to less extensive surgery.     

Comparison of fatty acid profiles in the serum of patients with prostate cancer and benign prostatic hyperplasia

OBJECTIVE: The role of dietary fatty acids (FAs) in benign and malignant prostatic diseases was investigated by comparing the composition value of serum fatty acids in the normal controls, and patients with prostate cancer (PC) and benign prostatic hyperplasia (BPH). Also, to estimate a possible association between PC risk and PUFAs, omega-3, omega-6 and omega-3/omega-6 FA composition ratios were compared among these groups. METHODS: Serum samples were obtained from 24 BPH and 19 PC patients, and from 21 age-matched normal male subjects. The serum concentration of 21 fatty acids was determined using gas chromatography/mass spectrometry. RESULT: The proportional values of saturated fatty acids (SFAs) groups demonstrated no specific difference between the control subjects and the patients. In the polyunsaturated fatty acids (PUFAs), we found that the omega-3 PUFAs level was significantly decreased in patient with BPH and PC and that the omega-6 PUFAs level was increased in PC only. The ratio of omega-3/omega-6 PUFAs decreased in the following order of normal, BPH, and PC. CONCLUSION: It was proposed that the changed composition level of PUFAs including omega-3 and omega-6 PUFAs have certain relationship with both prostatic diseases. Therefore, the ratio of omega-3/omega-6 PUFAs also may have an important association with the benign and malignant status of prostatic disease.     

Integrated bioinformatics analysis of potential biomarkers for pancreatic cancer

BackgroundPancreatic cancer, particularly pancreatic ductal adenocarcinoma (PDA), is an aggressive malignancy associated with a low 5‐year survival rate. Poor outcomes associated with PDA are attributable to late detection and inoperability. Most patients with PDA are diagnosed with locally advanced and metastatic disease. Such cases are primarily treated with chemotherapy and radiotherapy. Because of the lack of effective molecular targets, early diagnosis and successful therapies are limited. The purpose of this study was to screen key candidate genes for PDA using a bioinformatic approach and to research their potential functional, pathway mechanisms associated with PDA progression. It may help to understand the role of associated genes in the development and progression of PDA and identify relevant molecular markers with value for early diagnosis and targeted therapy.Materials and methodsTo identify novel genes associated with carcinogenesis and progression of PDA, we analyzed the microarray datasets {"type":"entrez-geo","attrs":{"text":"GSE62165","term_id":"62165"}}GSE62165, {"type":"entrez-geo","attrs":{"text":"GSE125158","term_id":"125158"}}GSE125158, and {"type":"entrez-geo","attrs":{"text":"GSE71989","term_id":"71989"}}GSE71989 from the Gene Expression Omnibus (GEO) database. Differentially expressed genes (DEGs) were identified, and the Database for Annotation, Visualization, and Integrated Discovery (DAVID) was used for Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. A protein‐protein interaction (PPI) network was constructed using STRING, and module analysis was performed using Cytoscape. Gene Expression Profiling Interactive Analysis (GEPIA) was used to evaluate the differential expression of hub genes in patients with PDA. In addition, we verified the expression of these genes in PDA cell lines and normal pancreatic epithelial cells.ResultsA total of 202 DEGs were identified and these were found to be enriched for various functions and pathways, including cell adhesion, leukocyte migration, extracellular matrix organization, extracellular region, collagen trimer, membrane raft, fibronectin‐binding, integrin binding, protein digestion, and absorption, and focal adhesion. Among these DEGs, 12 hub genes with high degrees of connectivity were selected. Survival analysis showed that the hub genes (HMMR, CEP55, CDK1, UHRF1, ASPM, RAD51AP1, DLGAP5, KIF11, SHCBP1, PBK, and HMGB2) may be involved in the tumorigenesis and development of PDA, highlighting their potential as diagnostic and therapeutic factors in PDA.ConclusionsIn summary, the DEGs and hub genes identified in the present study not only contribute to a better understanding of the molecular mechanisms underlying the carcinogenesis and progression of PDA but may also serve as potential new biomarkers and targets for PDA.     

Proteomics and bioinformatics approaches for identification of serum biomarkers to detect breast cancer

BACKGROUND: Surface-enhanced laser desorption/ionization (SELDI) is an affinity-based mass spectrometric method in which proteins of interest are selectively adsorbed to a chemically modified surface on a biochip, whereas impurities are removed by washing with buffer. This technology allows sensitive and high-throughput protein profiling of complex biological specimens. METHODS: We screened for potential tumor biomarkers in 169 serum samples, including samples from a cancer group of 103 breast cancer patients at different clinical stages [stage 0 (n = 4), stage I (n = 38), stage II (n = 37), and stage III (n = 24)], from a control group of 41 healthy women, and from 25 patients with benign breast diseases. Diluted serum samples were applied to immobilized metal affinity capture Ciphergen ProteinChip Arrays previously activated with Ni2+. Proteins bound to the chelated metal were analyzed on a ProteinChip Reader Model PBS II. Complex protein profiles of different diagnostic groups were compared and analyzed using the ProPeak software package. RESULTS: A panel of three biomarkers was selected based on their collective contribution to the optimal separation between stage 0-I breast cancer patients and noncancer controls. The same separation was observed using independent test data from stage II-III breast cancer patients. Bootstrap cross-validation demonstrated that a sensitivity of 93% for all cancer patients and a specificity of 91% for all controls were achieved by a composite index derived by multivariate logistic regression using the three selected biomarkers. CONCLUSIONS: Proteomics approaches such as SELDI mass spectrometry, in conjunction with bioinformatics tools, could greatly facilitate the discovery of new and better biomarkers. The high sensitivity and specificity achieved by the combined use of the selected biomarkers show great potential for the early detection of breast cancer.     

Species identification of corynebacteria by cellular fatty acid analysis

We evaluated the usefulness of cellular fatty acid analysis for the identification of corynebacteria. Therefore, 219 well-characterized strains belonging to 21 Corynebacterium species were analyzed with the Sherlock System of MIDI (Newark, DE). Most Corynebacterium species have a qualitative different fatty acid profile. Corynebacterium coyleae (subgroup 1), Corynebacterium riegelii, Corynebacterium simulans, and Corynebacterium imitans differ only quantitatively. Corynebacterium afermentans afermentans and C. coyleae (subgroup 2) have both a similar qualitative and quantitative profile. The commercially available database (CLIN 40, MIDI) identified only one third of the 219 strains correctly at the species level. We created a new database with these 219 strains. This new database was tested with 34 clinical isolates and could identify 29 strains correctly. Strains that remained unidentified were 2 Corynebacterium aurimucosum (not included in our database), 1 C. afermentans afermentans, and 2 Corynebacterium pseudodiphtheriticum. Cellular fatty acid analysis with a self-created database can be used for the identification and differentiation of corynebacteria.     

应用生物信息学分析确定PLK4是乳腺癌的关键预后基因

目的 应用生物信息学分析探讨Polo样激酶4(PLK4)在乳腺癌中的生物学作用、预后价值和对局部免疫微环境的影响.方法 通过Oncomine数据库挖掘PLK4在多种肿瘤中的表达水平;使用GEPIA数据库及HPA数据库检索PLK4在乳腺癌组织与正常组织中的表达差异;分别通过UALCAN数据库及TIMER数据库分析PLK4表达水平与乳腺癌患者临床特点及免疫浸润的相关性;采用K-M Plotter数据库评估PLK4在乳腺癌中的预后价值.应用Coexpedia数据库构建PLK4在乳腺癌中的共表达基因网络,并通过DAVID数据库对PLK4及其关键基因进行功能分析.结果 乳腺癌组织中,PLK4 mRNA和蛋白表达较正常乳腺组织均上调,特别是在P53突变和三阴性乳腺癌中差异更为显著(P＜0.05),且预示着更差的预后.PLK4在乳腺癌免疫微环境中与免疫细胞的表达呈正相关.筛选出PLK4的25个关键基因,即TPX2、CCNB2、CCNB1、BIRC5、CDC20和FOXM1等基因,其生物学功能主要与细胞分裂、细胞周期、细胞增殖和泛素化修饰等相关.KEGG通路富集分析表明,PLK4与P53信号通路、FoxO信号通路、MAPK信号通路和PI3 K-Akt信号通路等相关.结论 PLK4在乳腺癌组织中呈高表达可提示患者不良预后,其影响乳腺癌局部免疫微环境,在一定程度上表现为癌基因的特性,推测其可作为乳腺癌治疗疗效的预测指标,有望成为乳腺癌临床诊疗的新靶点.     

Plasma free fatty acid metabolism during storage of platelet concentrates for transfusion

New containers allow storage of platelet concentrates (PC) at 22 degrees C for up to 7 days, during which glycolytic and oxidative metabolism is vigorous. Recent evidence suggests that 85 percent of adenosine triphosphate regeneration is based on oxidative metabolism and that substrates other than glucose may be used. Because platelets can oxidize free fatty acids (FFA) as a possible source of energy during storage, the authors studied their availability, distribution, and turnover. Plasma FFA concentration was unchanged after 1 day of PC storage but significantly increased on Days 3, 5, and 7. Platelet-free plasma (PFP) stored under the same conditions as PC demonstrated a progressive increase in FFA, suggesting that some of the FFA accumulating in PC were derived from plasma rather than platelets. Indeed, during PC storage, plasma triglycerides decreased significantly, suggesting that they are a possible source of the increased levels of FFA found on Day 3 and thereafter. Thus, PC have a plasma FFA pool available continuously for oxidation during storage. Studies with radiolabeled palmitate suggested that FFA oxidation by platelets occurs during storage. The current findings show that plasma FFA could be a significant substrate for oxidative metabolism during storage of PC and that the oxidized FFA are replenished at least in part from plasma. These results may allow platelet storage to be improved, particularly in synthetic media.     

Mechanisms for inhibition of free fatty acid mobilization by nicotinic acid and sodium salicylate in canine subcutaneous adipose tissue in situ

Mechanisms for reduced free fatty acids (FFA) mobilization effected by nicotinic acid (NA) and sodium salicylate (SS) were studied in canine adipose tissue in situ. Both drugs inhibited adipose tissue lipolysis as evidenced by reduced release of glycerol. In addition, although the total amount of FFA re-esterified was not significantly changed, the amount of FFA re-esterified relative to the amount of FFA liberated intracellularly was significantly increased by both drugs. These effects were most pronounced during isoprenaline-stimulated lipolysis. Thus NA and SS reduced mobilization of FFA from canine adipose tissue through a combined effect on re-esterification and lipolysis.     

Tumor-associated macrophages in breast cancer as potential biomarkers for new treatments and diagnostics

While several inflammatory cell types participate in cancer development, macrophages specifically play a key role in breast cancer, where they appear to be part of the pathogenesis of high-grade tumors. Tumor-associated macrophages (TAMs) produce factors that promote angiogenesis, remodel tissue and dampen the immune response to tumors. Specific macrophage types contribute to increased metastases in animal models, while human studies show an association between TAMs and tumors with poor prognostic features. Macrophages display a spectrum of phenotypic states, with the tumor microenvironment skewing TAMs towards a 'nonclassical' activation state, known as the M2, or wound healing/regulatory state. These TAMs are found in high-risk breast cancers, making them an important therapeutic target to explore. Improved techniques for identifying TAMs should translate into clinical applications for prognosis and treatment.