Simultaneously quantifying hundreds of acylcarnitines in multiple biological matrices within ten minutes using ultrahigh-performance liquid-chromatography and tandem mass spectrometry.

Acylcarnitines are metabolic intermediates of fatty acids and branched-chain amino acids having vital biofunctions and pathophysiological significances. Here, we developed a high-throughput method for quantifying hundreds of acylcarnitines in one run using ultrahigh performance liquid chromatography and tandem mass spectrometry (UPLC-MS/MS). This enabled simultaneous quantification of 1136 acylcarnitines (C0-C26) within 10-min with good sensitivity (limit of detection < 0.7 fmol), linearity (correlation coefficient > 0.992), accuracy (relative error < 20%), precision (coefficient of variation (CV), CV < 15%), stability (CV < 15%), and inter-technician consistency (CV < 20%, n = 6). We also established a quantitative structure-retention relationship (goodness of fit > 0.998) for predicting retention time (tR) of acylcarnitines with no standards and built a database of their multiple reaction monitoring parameters (tR, ion-pairs, and collision energy). Furthermore, we quantified 514 acylcarnitines in human plasma and urine, mouse kidney, liver, heart, lung, and muscle. This provides a rapid method for quantifying acylcarnitines in multiple biological matrices.

Rapid quantification of 50 fatty acids in small amounts of biological samples for population molecular phenotyping.

Efficient quantification of fatty-acid (FA) composition (fatty-acidome) in biological samples is crucial for understanding physiology and pathophysiology in large population cohorts. Here, we report a rapid GC-FID/MS method for simultaneous quantification of all FAs in numerous biological matrices. Within eight minutes, this method enabled simultaneous quantification of 50 FAs as fatty-acid methyl esters (FAMEs) in femtomole levels following the efficient transformation of FAs in all lipids including FFAs, cholesterol-esters, glycerides, phospholipids and sphingolipids. The method showed satisfactory inter-day and intra-day precision, stability and linearity (R2 > 0.994) within a concentration range of 2-3 orders of magnitude. FAs were then quantified in typical multiple biological matrices including human biofluids (urine, plasma) and cells, animal intestinal content and tissue samples. We also established a quantitative structure-retention relationship (QSRR) for analytes to accurately predict their retention time and aid their reliable identification. We further developed a novel no-additive retention index (NARI) with endogenous FAMEs reducing inter-batch variations to 15 seconds; such NARI performed better than the alkanes-based classical RI, making meta-analysis possible for data obtained from different batches and platforms. Collectively, this provides an inexpensive high-throughput analytical system for quantitative phenotyping of all FAs in 8-minutes multiple biological matrices in large cohort studies of pathophysiological effects.

Rab5 regulates the proliferation, migration and invasion of glioma cells via cyclin E.

Glioma is the most common and lethal type of primary brain tumor, with a high mortality and recurrence rate. Rab5, which serves as a classic ontogenetic gene, is highly expressed in various types of tumor, including lung cancer, hepatocellular carcinoma and ovarian cancer. However, the exact role and the underlying mechanism of Rab5 in glioma remain unknown. Herein, the role of Rab5 in the tumorigenesis and metastasis of glioma cells was investigated. The upregulation of Rab5 in glioma tissues and cells was observed. The expression of Rab5 was positively associated with proliferation, migration and invasion of glioma cells. Moreover, Rab5 was involved in the cell cycle of glioma cells via the regulation of cyclin E. Data presented in the present study suggest Rab5 as a potential novel diagnostic and prognosis marker of glioma.

Transmembrane protein 45A regulates the proliferation, migration, and invasion of glioma cells through nuclear factor kappa-B.

Gliomas are the most common and aggressive type of primary brain cancer in adults. The expression of transmembrane protein 45A (TMEM45A) in glioma patients and glioma cell lines was analyzed by quantitative real-time PCR. The influence of TMEM45A on the survival of glioma patients was also explored in this study. To verify the interaction between TMEM45A and key genes, correlation analysis of expression levels and the siRNA knock down method were performed. TMEM45A was upregulated in glioma tissues, and its overexpression was strongly correlated with the poor survival of glioma patients. Experiments using the overexpression and knock down of TMEM45A were carried out to demonstrate its correlation with enhanced proliferation, migration, and invasion in glioma cells. Nuclear factor kappa-B (NFκB) expression was shown to be a downstream factor of TMEM45A in glioma cells. In conclusion, TMEM45A is an oncogenic gene in glioma. The proliferation, migration, and invasion of gliomas could be effectively impeded by inhibition of TMEM45A, and the cancer-promoting effect of TMEM45A on gliomas was involved with the NFκB pathway.

LncRNATCF7 promotes the growth and self-renewal of glioma cells via suppressing the miR-200c-EpCAM axis.

Accumulating evidence has demonstrated that long non-coding RNAs (lncRNAs) are involved in tumor initiation and development. Recent studies illustrated that lncRNATCF7 was highly expressed in lung cancer and liver cancer, however, the expression pattern and function of lncRNATCF7 in glioma remains to be elucidated. Here, we found that lncTCF7 was highly expressed in glioma tissues and cell lines. Overexpression of lncTCF7 promoted the proliferation and migration of glioma cells. Down-regulation of lncTCF7 significantly suppressed the tumorigenesis of glioma. Mechanistically, lncTCF7 enhanced the self-renewal of glioma cells via up-regulating the expression of epithelial cell adhesion molecule (EpCAM). The detailed molecular mechanism uncovered that lncTCF7 bound to miR-200c and decreased the abundance of miR-200c, which consequently attenuated the negative regulation of miR-200c on EpCAM. Collectively, these data provide evidence to demonstrate the critical role of lncTCF7 in the tumorigeneis of glioma, which suggested that lncTCF7 might be a promising target in the treatment of glioma.