Modification And Function Of Glucose And Amino Acid Metabolism In Colorectal Cancer Complicated With Diabetes Mellitus

Part I Metabolomics analysis of colorectal cancer complicated with diabetes mellitusBackgroundsColorectal cancer(CRC)is one of the most common malignant disease around the world.It is estimated that in 2012 there were more than 1.36 million new cases of CRC and a population of nearly 0.7 million were died of this kind of hindgut carcinoma.In China in 2011 the prevalence of CRC ranked No.3 among all cancers,next only to lung cancer and gastric cancer.Diabetes mellitus(DM)is now one of the most common metabolic disorders which lead to many complications.The total number of people with diabetes worldwide in 2017 has risen to 424.9 million.Epidemiology evidences proved that DM can independently increase the risk of developing and dying from CRC.DM and CRC share many risk factors,including age,sexuality,obesity,smoke,drink,and high-fat diet,but potential biologic links between the two diseases remain unclear.Due to the specific growth rate of cancer tissues,tumor cells require obviously more energy than normal cells.On the other hand,DM patients suffer from body-wide metabolism disorders like hyperglycemia and glucose metabolism dysfunction caused by insulin resistance.Thus we aimed to find out whether DM can affect biological behavior of tumor via changing metabolism and energy supply of CRC cells.Purpose1.To analyze the metabolomics changing of CRC complicated with DM,compared with normal CRC.2.Qualitative and quantitative analysis for highly diverse families of metabolites.3.To check out significantly changed metabolic process of CRC complicated with DM.Methods1.Gas chromatography-time of flight mass spectrometry(GC-TOF-MS)was used to analyze the metabolome of CRC complicated with DM.2.Principal component analysis and orthogonal partial least squares-discrimination analysis were employed to identify the differentiation of metabolome between DM and normal CRC.3.Variable importance in the projection of the first principal component and student's t-test were used to pick out highly diverse metabolites.4.Use Kyoto Encyclopedia of Genes and Genomes to check out significantly changed metabolic process of CRC complicated with DM.Results1.The metabolomics of CRC complicated with DM was obviously changed,compared with normal CRC.2.The relative concentration of glucose were significantly higher in CRC complicated with DM,but none of the other metabolites involved in glucose metabolic processes was different with that of normal CRC,neither of the glucose metabolic processes.3.Many kinds of amino acids(AAs)were markedly lower in CRC complicated with DM.Many AA metabolic processes seemed to be different.ConclusionsWe ensured that there were specific variation of metabolome of CRC complicated withDM when compared with normal CRC,indicating that metabolic characteristic modifications might make a difference in CRC promoted by DM.We found that AA metabolism but not glucose metabolism might play a potential role in this process.Part ? Modification and function of glucose metabolism and insulin sensitivity in colorectal cancer complicated with diabetes mellitusBackgroundsApparently,tumor cells require more energy than normal cells to meet the high rate of growth and proliferation.Almost all solid carcinoma cells preferred glycolysis rather than oxidation.This phenomenon is called Warburg effect.The role of tricarboxylic acid cycle(TCA cycle)changes from glucose oxidation to lipids and amino acids biosynthesis,too.Another important glucose metabolic pathway,pentose phosphate pathway(PPP)is also activated abnormally,to produce more pentose for nucleic acids replication.In general,tumor cells adjusts their way of glucose metabolism to meet the high rate of energy consumption,cell growth and proliferation.Insulin is one of the most important growth factor which can not only regulate cell metabolism,but also promote cell growth.Insulin has a definitive effect on colorectal carcinogenesis.Insulin and insulin-like growth factor receptor signal pathway are the most related signal transduction downstream of insulin,which then modify plenty biological function of cell.As a systemic metabolic disease,the main metabolic changes of DM are hyperglycemia and hyperinsulinemia.However,the real status of glucose metabolic process and insulin sensitivity in CRC complicated with DM requires further investigation.Purpose1.To observe the change of biological behavior of CRC cell lines under high glucose conditions.2.To compare the expression of glucose metabolism relative enzymes between normal CRC patients' tumor tissue and CRC combined with DM patients' tumor tissue.3.To analyze the expression of glucose metabolism relative enzymes in CRC cell lines under high glucose conditions.4.To test the activity of insulin signal pathway in CRC cell lines and to analyze the biological behavior of CRC cell lines treated with exogenous insulin under high glucose conditions.Methods1.Human CRC cell lines SW480,SW620,LoVo and HT29 were treated with normal culture medium plus high glucose concentration.Cell proliferation and migration assays were performed in these cell lines and the results were compared with that of the same cell lines under normal glucose concentration.2.Immunohistochemistry was used to detect the expression level of glucose metabolic enzymes in CRC complicated with DM and normal CRC.3.Western blot was used to analyze the expression level of glucose metabolic enzymes in CRC cell lines mentioned above.4.Human CRC cell lines were treated with normal culture medium plus high glucose concentration.After insulin stimulation western blot was used to evaluate the phosphorylation level of insulin-signal-pathway-related proteins,and cell proliferation and migration assays were performed.Results1.High glucose concentration promoted cell proliferation and migration of CRC cell lines in certain concentration range.When the glucose level was higher than the threshold,there would be no more promotion.2.The expression levels of glucose metabolism relative enzymes in tissues of CRC complicated with DM and tissues of normal CRC had no significant differences.3.The expression levels of glucose metabolism relative enzymes in CRC cell lines treated with high glucose concentration and normal CRC cell lines had no obvious differences.4.The phosphorylation levels of insulin-signal-pathway-related proteins in CRC cell lines treated with high glucose concentration were inhibited.Exogenous insulin cannot affect the proliferation and migration ability of CRC cell lines treated with high glucose any further.ConclusionsThe carcinogenesis of CRC under high glucose concentrations was limited by the unchanged expression level of glucose metabolic relative enzymes.High glucose concentration induced the insulin resistance of CRC cell lines.There should be other metabolic processes involved in CRC complicated with DMPart ? Modification and function of amino acid metabolism in colorectal cancer complicated with diabetes mellitusBackgroundsAs the smallest component of protein,the role of AAs in carcinogenesis and development of cancer has gradually been studied.It can not only be used as precursor of protein biosynthesis,but also as alternative energy producer at starvation status.Besides,some AAs molecular can take parts in the mediation of important cellular process like proliferation,apoptosis and autophagy.Imbalance of nutrient metabolism is an important feature of diabetes.The overmuch AAs might be a direct risk factor for carcinogenesis.But our metabolome analysis showed a marked decrease in many kinds of AAs.Till then there were no related reports similar to this discovery and it drew our attention to AA metabolism in CRC complicated with DM.Purpose1.To evaluate the level of AAs in tissues and cell lines of CRC complicated with DM.2.To investigate the effect of AAs on proliferation and migration ability of high glucose treated CRC cell lines.3.To investigate the influence of AAs on neoplasia on tumor-bearing mice model.4.To study the reasons and functions of AAs level change in DM combined CRC.Methods1.Human CRC cell lines SW480,SW620,HT29 and LoVo were treated with normal culture medium plus high glucose concentration.The level of AAs in these cell lines were tested.2.High glucose induced CRC cell lines were then treated with additional AAs.Cell proliferation and migration assays were performed.3.High-fat-diet(HFD)NOD/SCID mice were injected subcutaneously with CRC cell suspension and fed with compound amino acid solution.Tumor formation of the mice were observed.4.The expression of proteins related to AAs and protein metabolism were evaluated by isobaric tags for relative and absolute quantification(iTRAQ)proteomic assay.The differences were analyzed.Results1.High glucose treated CRC cell lines consumed excess AAs compared to normal CRC cell lines.2.Additional AAs didn't affect cell proliferation rate and migration ability of CRC cell lines treated with high glucose concentration.3.Compound amino acid solution didn't affect the size and mass of tumor which formed in HFD NOD/SCID mice.4.The iTRAQ proteomic assay showed that expression level of several enzymes and factors related to protein biosynthesis were up-regulated in tissues of CRC complicated with DM.ConclusionsThe level of AAs in CRC complicated with DM decreased relative to normal CRC,possibly due to increased consumption during abnormal protein biosynthesis.Changing AAs supply alone couldn't influence the biological behavior of tumor tissue and cell,which indicated that change of AAs level was not the reason for higher malignance of CRC complicated with DM.Proteomic assay showed that cellular protein biosynthetic and metabolic process but not AA metabolic process were apparently activated in tissues of CRC complicated with DM.Above all,protein metabolism should be the new point of penetration for the treatment of CRC patients with DM.