Urine metabolomics study of hepatocellular carcinoma / 中华肿瘤杂志

Objective: To investigate the urinary small molecular metabolites and their metabolic characteristics of patients with hepatocellular carcinoma (HCC). Methods: High throughput ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) was used to detect the small molecular metabolites in urine of healthy control (n=10), patients with hepatic hemangioma (n=10) and patients with HCC (n=10). The orthogonal projections to latent structures-discriminant analysis (OPLS-DA), hierarchical cluster analysis of multivariate analysis and univariate analysis were used to analyze the differential metabolites of the three groups. Results: The metabolic profiles of the three groups showed that the total of 381 differential metabolites were identified and divided into 96 up-regulated metabolites and 285 down-regulated metabolites. There were 55 urinary metabolites specifically related to HCC. Twenty-one of them were significantly up-regulated, including Acetyl-DL-Leucine, Ala Asp, HoPhe-Gly-OH, while 34 were significantly down-regulated, including Selenocystathionine, Met Trp Met Cys, Valsartan acid and so on. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that the differential metabolites were mainly enriched in glutamine/glutamate metabolism, lysine biosynthesis, tricarboxylic acid cycle and purine metabolism. Conclusions: The occurrence of HCC is accompanied by the abnormalities of multiple metabolites and metabolic pathways. The analysis of the characteristic metabolic profile of urine in patients with HCC is helpful to find metabolic markers and potential therapeutic targets for liver cancer.

Chinmedomics strategy to discover effective constituents and elucidate action mechanism of Nanshi capsule against kidney-yang deficiency syndrome / 中国中药杂志

The chinmedomics method was used to explore the effect of Nanshi capsule on endogenous metabolites of rats with kidney-yang deficiency syndrome, investigate the metabolites and metabolic pathways closely related to kidney-yang deficiency syndrome (KYDS)and identify the therapeutic basis of Nanshi capsule(NPC)as well as its action mechanism for KYDS. The routine biochemical indexes of serum were detected and histomorphology was observed. Based on the chinmedomics technology platform, discriminatory analysis in multivariate modes was conducted for rat blood and urine, thus to investigate the biomarkers of KYDS and the therapeutic effect of NPC against KYDS. Meanwhile, the main constituents of NPC in rat serum were also detected to analyze its correlation between the constituents in vivo and the biomarkers of KYDS, and determine the potential effective compounds for therapeutic effect. Eleven biomarkers of KYDS were identified in the rat models, involving steroid hormone biosynthesis, tryptophan metabolism and tyrosine metabolism. It was found that NPC could regulate steroid hormone biosynthesis, tryptophan metabolism and tyrosine metabolism; PCMS analysis showed that caffeic acid, 2-hydroxy-1-methoxy-anthraquinone, 1-hydroxy-2-methoxyanthraquinone, ferulic acid glucuronide conjugation, deacetylasperulosidic acid, cynaroside, betaine and umbelliferone were the main effective compounds of NPC for KYDS. In this study, cynaroside, betaine, umbelliferone and other compounds in NPC could integrally regulate the disturbance of metabolic profile in KYDS by improving the hormone synthesis, hormone synthesis pathway, hormone synthesis and release pathway in tyrosine metabolism and linoleic acid synthesis pathway in linoleic acid metabolism. These results indicated that the NPC had the characteristics of multi-pathway, multi-target and overall regulation in the treatment of KYDS. Chinmedomics approach can provide methodology support to discover innovative drug from traditional Chinese medicine.

Mediastinal lymph nodes micro-metastases in patients with clinical stage I-II lung cancer / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To investigate micro-metastasis in mediastinal lymph nodes (mLN) of patients with clinical stage I approximately II lung cancer and its clinical significance.</p><p><b>METHODS</b>A total of 181 mLN from 32 lung cancer patients in clinical stage I approximately II were collected during operation and their frozen sections at two different levels were examined immunohistochemically (IHC) with an anti-epithelial cell monoclonal antibody Ber-Ep4. Routine HE staining was done for comparison. The results were processed by Chi-square tests in SPSS 10.0 soft ware.</p><p><b>RESULTS</b>Fifteen of the 32 patients (46.9%) were found to have micro-metastasis in 21 of 181 mLN (11.6%) examined by immunohistochemical staining though routine histopathological examinations were negative. Of those 15 cases, micro-metastasis was detected in 9 only by IHC and in 6 both by IHC and HE stainings. The positive rate of micro-metastasis in N0, N1, and N2 stratified by routine pathology was 36.8% (7/19), 33.3% (2/6) and 85.7% (6/7), respectively (N0 vs N2, P < 0.05). When stratified according to clinical staging (cTNM), pathological staging (pTNM) and pathological staging on the basis of IHC (iTNM), the frequencies of N2 cases were 0, 18.8% and 46.9%, respectively (differences among the three groups: P < 0.01). Nine cases reported as N0(7) and N1(2) by routine histopathological examination were found to have micro-metastasis in mLN by IHC staining, therefore they were actually N2 cases.</p><p><b>CONCLUSION</b>IHC staining with a monoclonal antibody specific for epithelial cells (Ber-Ep4) is more sensitive in the detection of mediastinal micro-metastais than routine HE staining. Underestimation of the extent of mLN metastasis by cTNM and/or pTNM stagings frequently exists in patients with clinically early lung cancer.</p>

Significance of CEA, SCC and Cyfra21-1 serum test in esophageal cancer / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To study the clinical significance of serum CEA, SCC and Cyfra21-1 test in the diagnosis, prediction of prognosis and postoperative monitor of recurrence in esophageal cancer.</p><p><b>METHODS</b>The concentration of serum CEA and Cyfra21-1 was measured by electrochemiluminescence immunoassay (ECLIA) using Elecsys 2010, CEA kit and Cyfra21-1 kit. Serum SCC was measured by microparticle enzyme immunoassay (MEIA) using IMx System and SCC kit. Serum of 206 patients with esophageal cancer (203 squamous cell carcinoma, 2 small cell carcinoma and 1 adenosquamous carcinoma) was measured preoperatively, 71 of whom also measued 8 to 12 days after resection.</p><p><b>RESULTS</b>The cut-off value of CEA and Cyfra21-1 was < or = 3.25 ng/ml and < or = 2.61 ng/ml, which were determined by the data of 45 healthy Chinese measured during the same period. The positive ratios of serum CEA and Cyfra21-1 in 206 cases were 29.1% and 45.1%. The combined positive ratio of CEA and Cyfra21-1 was 57.3%. The CEA positive ratios, according to the pathological stage of 165 resectable patients, were 16.6% (stage I), 26.8% (II) and 30.8% (III). For Cyfra21-1, they were 27.8%, 37.5% and 50.5%. For CEA combined with Cyfra21-1, they were 38.9%, 50.0% and 63.7%. The mean value of CEA, SCC and Cyfra21-1 (especially SCC and Cyfra21-1) was found to be well correlated with the tumor volume, TNM stage and depth of tumor invasion. Patient with bulky tumor or advanced tumor (T4) usually had much higher mean value than those with early stage tumors. One week after radical resection, the level of the three tumor markers fell to normal level in 92.9% of 71 patients. The level of serum CEA and Cyfra21-1 varied greatly in a small part of the patients. Extremely elevated serum CEA and Cyfra21-1 usually indicated advanced lesion or tumor metastasis.</p><p><b>CONCLUSION</b>Preoperative and postoperative measurement of serum CEA, SCC and Cyfra21-1 (especially Cyfra21-1) is helpful in the diagnosis, prediction of prognosis and monitor of postoperative recurrence in patients with esophageal cancer.</p>

Cloning of human uracil N-glycosylase and its detection in cancer tissues by quantitative RT-PCR / 生物工程学报

The uracil in DNA comes from either the misincorporation of dUTP in place of dTTP or deamination of cytosine. In the latter case, it can result in a GC to AT transition mutation if the uracil is not removed before DNA replication. Base excision repair (BER) is a major pathway for removing DNA lesions arising from endogenous processes as well as those induced by exposure to exogenous chemicals or irradiation. BER is initiated by DNA glycosylases that excise aberrant bases from DNA by cleavage of the N-glycosidic bond linking to the base of its deoxyribose sugar. Uracil N-glycosylase (UNG) is the enzyme responsible for the first step in the BER pathway that specifically removes uracil from DNA. The UNG gene undergoes both temporal and spatial regulation mainly at the level of transcription. Normally cancer cells undergo over-proliferation and up-regulate their UNG during tumorigenesis. In this study we examine the correlation between UNG level and carcinogenesis, and explore the possibility of using UNG as a marker for cancer diagnosis. Human UNG gene was amplified from the total RNA of the human choriocarcinoma cell line, JEG-3, by RT-PCR. After purification, the 942bp full-length UNG cDNA coding sequence was digested with EcoR I and Sal I, and cloned into the digested pET-21 to construct a recombinant vector, pUNG. The UNG protein was expressed under the control of T7 promoter in E. coli BL21 (DE3) cells induced with IPTG. After ultrasonic treatment, the cell lysate and precipitate were analyzed by SDS-PAGE and a 39kD band was detected. The plasmid was serially diluted at appropriate concentrations and employed as standards in the subsequent quantification. Total RNAs were extracted from 18 pairs of clinical samples, each pair contains a sample of esophageal squamous cell carcinoma (ESCC) tissue and its surrounding normal esophageal epithelia. The copy numbers of UNG mRNA in these RNA samples were determined by real-time quantitative RT-PCR using a Lightcycler (Roche). UNG was present in 13 cases of ESCC (13/18, n = 18) but absent in all of the normal tissues. The results indicated that there was a correlation between high level of UNG expression and the carcinogenesis of ESCC.