Alteration of fecal tryptophan metabolism correlates with shifted microbiota and may be involved in pathogenesis of colorectal cancer

BACKGROUNDGut tryptophan (Trp) metabolites are produced by microbiota and/or host metabolism. Some of them have been proven to promote or inhibit colorectal cancer (CRC) in vitro and animal models. We hypothesized that there is an alteration of gut Trp metabolism mediated by microbiota and that it might be involved in the pathogenesis of cancer in patients with CRC.AIMTo investigate the features of Trp metabolism in CRC and the correlation between fecal Trp metabolites and gut microbiota.METHODSSeventy-nine patients with colorectal neoplastic lesions (33 with colon adenoma and 46 with sporadic CRC) and 38 healthy controls (HCs) meeting the inclusion and exclusion criteria were included in the study. Their demographic and clinical features were collected. Fecal Trp, kynurenine (KYN), and indoles (metabolites of Trp metabolized by gut microbiota) were examined by ultraperformance liquid chromatography coupled to tandem mass spectrometry. Gut barrier marker and indoleamine 2,3-dioxygenase 1 (IDO1) mRNA were analyzed by quantitative real-time polymerase chain reaction. Zonula occludens-1 (ZO-1) protein expression was analyzed by immunohistochemistry. The gut microbiota was detected by 16S ribosomal RNA gene sequencing. Correlations between fecal metabolites and other parameters were examined in all patients.RESULTSThe absolute concentration of KYN [1.51 (0.70, 3.46) nmol/g vs 0.81 (0.64, 1.57) nmol/g, P = 0.036] and the ratio of KYN to Trp [7.39 (4.12, 11.72) × 10^-3 vs 5.23 (1.86, 7.99) × 10^-3, P = 0.032] were increased in the feces of patients with CRC compared to HCs, while the indoles to Trp ratio was decreased [1.34 (0.70, 2.63) vs 2.46 (1.25, 4.10), P = 0.029]. The relative ZO-1 mRNA levels in patients with CRC (0.27 ± 0.24) were significantly lower than those in HCs (1.00 ± 0.31) (P < 0.001), and the relative IDO1 mRNA levels in patients with CRC [1.65 (0.47-2.46)] were increased (P = 0.035). IDO1 mRNA levels were positively associated with the KYN/Trp ratio (r = 0.327, P = 0.003). ZO-1 mRNA and protein levels were positively correlated with the indoles/Trp ratio (P = 0.035 and P = 0.009, respectively). In addition, the genera Asaccharobacter (Actinobacteria) and Parabacteroides (Bacteroidetes), and members of the phylum Firmicutes (Clostridium XlVb, Fusicatenibacter, Anaerofilum, and Anaerostipes) decreased in CRC and exhibited a positive correlation with indoles in all subjects.CONCLUSIONAlteration of fecal Trp metabolism mediated by microbiota is associated with intestinal barrier function and tissue Trp metabolism, and may be involved in the pathogenesis of CRC.     

替尼达普对慢性颞叶癫(癎)大鼠海马内向整流钾通道2.3亚单位表达的影响

目的 观察慢性颞叶癫(癎)(TEE)大鼠致(癎)后不同时点海马组织中内向整流钾通道(Kir)2.3亚单位mRNA和蛋白的表达变化规律及通道特异性开放剂替尼达普对其表达的影响,探讨Kir2.3通道与TLE发病机制的关系,并为临床应用钾通道开放剂作为抗癫(癎)药物提供依据.方法 匹罗卡品诱导大鼠产生癫(癎)持续状态(SE),持续观察2周,成模为慢性TLE大鼠.用逆转录PCR(RT-PCR)和Western blot方法检测对照组及致(癎)组大鼠SE终止后0、6、72 h和2周后海马组织中Kit2.3通道mRNA和蛋白表达变化趋势;随后以慢性期2周作为观察时间点,检测替尼达普对大鼠海马Kir2.3通道mRNA及蛋白表达变化的影响.结果 对照组、致(癎)组SE终止后0、6、72 h和2周时Kir2.3 mRNA/β-actin比值分别为0.080±0.030、0.103±0.045、0.164±0.026、0.132±0.024和0.011±0.008(F:23.684,P<0.01);各时间点Kir2.3蛋白/甘油醛-3磷酸脱氢酶(GAPDH)比值分别为0.305±0.030、0.263±0.028、0.767±0.167、0.498±0.077和0.176±0.026(F=44.183,P<0.05),其表达呈现急性期增高、慢性期明显降低的动态变化趋势.在慢性期,替尼达普给药组大鼠Kir2.3 mRNA/β-actin与Kir2.3蛋白/GAPDH比值分别为0.021±0.006和0.636±0.140,与未给药组相比表达增高,差异有统计学意义(F=25.216、47.355,P<0.05、0.01).结论 Kir2.3通道mRNA和蛋白在慢性期表达下调,可能是难治性癫(癎)发病的分子生物学机制之一.替尼达普通过增加Kir2.3通道的表达,最终可能减少(癎)样放电的产生.     

In vivo determination of endogenous biogenic amines in rat brain using HPLC and push-pull cannula

Combining the methods of push-pull cannulation with those of high performance liquid chromatography (HPLC), we have measured the content of a number of biogenic amines in the perfusate of freely moving rats. In an initial study, the lateral hypothalamus (LH) was chronically implanted with a push-pull cannula and was perfused with 0.9% NaCl. Fifteen minute samples were collected through the push-pull cannula (flow rate: 25 μl/min) and aliquots of 200 μl were injected into the HPLC without any extraction or prepurification procedure. Simultaneous determination of the levels of 5-hydroxytryptophan (5-HTP), 5-hydroxytryptamine (5-HT), 5-hydroxyindoleacetic acid (5-HIAA), norepinephrine (NE) and dopamine (DA) in the perfusate was accomplished by means of HPLC with electrochemical detection. The HPLC system utilized a C-18 reverse phase column coupled with a glassy carbon detector. Results indicate that this combination of push-pull perfusions and HPLC assay can provide a simple, rapid, and sensitive technique for the in vivo simultaneous determination of the compounds released in discrete brain areas. In preliminary studies in which these methods were used, 50 mg/kg D,L-5-HTP was injected subcutaneously (SC) into rats implanted with push-pull cannulae and working on a variable interval (VI 1) schedule of reinforcement. Increases in 5-HTP, 5-HT and 5-HIAA were measured during the period of behavioral depression following 5-HTP administration. This technique could provide a useful tool in the assessment of neurochemical changes in brain during ongoing steady-state behaviors or during the disruption of behavior following administration of drugs, precursors, or other perturbations.     

Synthesis and antioxidant activity of new tetrahydro-naphthalene-indole derivatives as retinoid and melatonin analogs

A number of retinoid-related compounds represent classes of antioxidative and proapoptotic agents with promising potential in the treatment of neoplastic diseases. Indeed, the synthetic retinoid amide fenretinide [N-(4-hydroxyphenyl)retinamide] induces apoptosis of cancer cells and acts as a chemotherapeutic drug in cancer therapy. In the present work, and as a continuation of our studies on retinoid-type compounds, the synthesis of melatonin retinamide derivatives was studied as a novel series of melatonin retinoids, using the condensation reaction sequence involving tetrahydrotetramethylnaphthalene carboxylic acid and appropriate melatonin-type moieties. Despite of the weak DPPH inhibition activity pattern of the synthesized compounds, some of them showed a strong inhibition on lipid peroxidation (IVa-b, Va, and VIIa-c, 88, 96, 90, 94, 93, and 86%, respectively at 10(-4) M concentration) when melatonin (85% at 10(-4) M concentration) was used as a reference compound.     

HPLC determination of biogenic amines in discrete brain areas in food deprived rats.

Norepinephrine (NE), dopamine (DA), 5-hydroxytryptophan (5-HTP), 5-hydroxytryptamine (5-HT), and 5-hydroxyindole acetic acid (5-HIAA) levels in the lateral hypothalamus (LH), ventromedial hypothalamus (VMH), median raphe (MR) and dorsal raphe (DR) were determined in nondeprived and 48 hr food deprived rats. Simultaneous determination of these compounds was accomplished by means of high performance liquid chromatography (HPLC) with electrochemical detection. When compared with controls, food deprived animals showed significant increases in 5-HT and 5-HIAA levels in the raphe nuclei, significant increases in 5-HIAA in the LH, but no changes in either 5-HT or 5-HIAA levels in the VMH. No changes in catecholamine levels were found in any of the brain areas studied. These results show that indoles in the raphe nuclei, as well as in the LH, are affected by food deprivation. The lack of change in indole levels in the VMH indicates that specific nuclei within the hypothalamus are differentially affected by food deprivation.     

卡他莫拉菌快速鉴定的方法学及耐药性研究

目的　建立快速鉴定卡他莫拉菌 (MC)的方法 ,对不同血源、营养配方的琼脂培养基进行了MC的生长指数 (GI)的测定 ,了解MC的耐药状况。方法　使用含 5 0 μg/ml万古霉素的兔血巧克力琼脂培养基分离MC ,以 3 乙酰吲哚纸片法检测MC产生的乙酰酯酶 ,与DNA酶试验为对照 ;将MC接种于 9种培养基上 ,计算和比较MC在 9种培养基上的平均GI;同时用Nitrocefin纸片检测MC的 β内酰胺酶发生率 ,以琼脂平皿二倍稀释法测定最低抑菌浓度 (MIC)。结果　检测 2 2 8株MC的乙酰酯酶结果 ,其敏感性为 10 0 % ,与 2 0 3株近缘菌比较特异性为 10 0 % ,与DNA酶试验有很高的相关性 ;MC在巧克力琼脂培养基上生长较血琼脂培养基好 ,但与GI值比较 ,差异无明显意义 (P >0 0 5 ) ;MC产 β内酰胺酶的阳性率为 93 0 % ;哌拉西林 /舒巴坦和哌拉西林 /他唑巴坦的MIC50 和MIC90 分别较单哌拉西林 ,降低了 8倍 ;氨苄西林 /舒巴坦的MIC50 和MIC90 较单氨苄西林降低了 32倍和 4倍。结论　用 3 乙酰吲哚纸片法检测MC产生的乙酰酯酶 ,是临床实验室快速鉴定MC的良好方法。MC产 β内酰胺酶阳性率之高 ,应引起临床医生的重视。哌拉西林 /舒巴坦、氨苄西林 /舒巴坦、头孢哌酮 /舒巴坦、哌拉西林 /他唑巴坦及亚胺培南 /西司他丁对 β内酰胺酶阳性MC有很     

丙戊茶碱对大鼠大脑皮层星形胶质细胞NGF和IL-1β释放的影响

目的 探讨丙戊茶碱对大鼠大脑皮层星形胶质细胞神经生长因子(NGF)和IL-1β释放的影响.方法 出生1～3 d的SD大鼠,体重6～8 g,原代培养大脑皮层星形胶质细胞,传代培养到第4代时以1×105个/ml的密度接种到24孔培养板中,随机分为8组,每组6孔:正常对照组(C组):无血清培养基继续培养;不同浓度丙戊茶碱组(p1～3组):分别加入含有丙戊茶碱10、100和1000μmol/L的培养基中;LPS组:加入含有LPS 1 μg/ml的培养基中;P1+LPS组、P2+LPS组和P3+LPS组:分别加入含有丙戊茶碱10、100和1000μmol/L和LPS 1μg/ml的培养基中.于孵育1、3 d时检测上清液IL-1β和NGF的浓度,以反映其释放量.结果 与C组比较,P1组、P2组和P3组星形胶质细胞NGF释放量升高,IL-1β释放量降低,LPS组NGF释放量降低,IL-1β释放量升高(P＜0.05),P1+LPS组、P2+LPS组和P3+LPS组NGF和IL-1β释放量差异无统计学意义(P＞0.05);P1组、P2组和P3组星形胶质细胞NGF释放量依次升高(P＜0.05),3组IL-1β释放量比较差异无统计学意义(P＞0.05);与LPS组比较,P1+LPS组、P2+LPS组和P3+LPS组星形胶质细胞NGF释放量升高,P2+LPS组和P3+LPS组IL-1β释放量降低(P＜0.05).结论 丙戊茶碱可促进大鼠大脑皮质星形胶质细胞释放NGF,抑制其释放IL-1β.     

神经外科开颅术后患者呕吐及预防用药的临床回顾

目的：评价高选择性5-羟色胺（5-HT3）受体拮抗剂托烷司琼对神经外科术后患者出现的恶心、呕吐的预防作用。方法：回顾调查了我科过去6年收治的神经外科术后患者,按照应用药物的与否分为两组：未用药组（n=3 332）,托烷司琼组（n=3519）。分别统计入室后6、12、24 h的如下指标：（1）呕吐发生率,（2）其它不良反应发生情况。结果：与未应用呕吐药物组相比,托烷司琼组恶心呕吐的发生率均明显降低,差异有统计学意义（P〈0.01）。结论：本研究结果显示应用选择性5-HT3受体拮抗剂后,神经外科开颅术后患者术后呕吐的发生率明显降低。     

他汀类药物对急性冠状动脉综合征患者vWF因子的影响

目的：探讨洛伐他汀和氟伐他汀对急性冠脉综合征（ACS）患者血浆血管性血友病因子（von Willebrand factor，vWF）及血脂水平的影响。方法：88例ACS患者随机单盲分为3组，其中对照组（A组）18例；洛伐他汀组（B组）34例，20mg／d；氟伐他汀组（C组）36例，20mg／d；检测患者治疗前后30d血清vWF浓度和血脂浓度变化。结果：B、C组能显著降低血总胆固醇（TC）、甘油三酯（TG）及低密度脂蛋白（LDL）水平、升高血高密度脂蛋白（HDL）；两组间疗效无明显差异；B、C组能显著降低vWF的浓度，两组间疗效无明显差异；B、C组患者的血清vWF水平降低与血清TC、TG、LDL的水平降低、HDL的水平升高不相关。结论：洛伐他汀和氟伐他汀治疗急性冠脉综合征患者30天就可明显改善血脂，降低vWF水平，具有完善内皮功能、抑制血小板黏附聚集等作用。     

Identification of three indolic compounds in a pigmented-melanoma cell-culture supernatant by gas chromatography-mass spectrometry

Summary In the supernatant of melanoma cell culture SK mel 25 three indolic compounds-5-hydroxy-6-methoxyindole, 5-hydroxy-6-methoxyindolyl-2-carboxylic, and 6-hydroxy-5-methoxyindolyl-2-carboxylic acids-have been identified. The supernatants were extracted with ethyl acetate, derivatized, and analyzed by gas chromatography-mass spectrometry. The significance of this finding is discussed.Abbreviations 5,6DHI 5,6-dihydroxyindole - 5H6MI 5-hydroxy-6-methoxyindole - 6H5MI 6-hydroxy-5-methoxyindole - 5,6DHI2C 5,6-dihydroxyindolyl-2-carboxylic acid - 5H6MI2C 5-hydroxy-6-methoxyindolyl-2-carboxylic acid - 6H5MI2C 6-hydroxy-5-methoxyindolyl-2-carboxylic acid - PFPA pentafluoropropionic anhydride - HFIP 1,1,1,3,3,3-hexafluoroisopropanol - DOPA L-3,4-dihydroxyphenylalanine - COMT catechol-O-methyltransferase (EC 2.1.1.6) - GC-MS gas chromatography-mass spectrometry This work was supported by grants from Koningin Wilhelmina Fonds (Netherlands Cancer Foundation) Nos.82-10 and 82-6 GUKC