The Role Of Long Non-coding RNA In Metastatic Spine Tumor And HCC And Its Clinical Significance

With the environmental pollution and food safety problems become increasingly serious,smoking,drinking and other bad habits more and more to the younger development,resulting in increased incidence of cancer year after year.Currently the tumor has become the major disease threatening human health,regardless of incidence or mortality tops in the world,to national health caused great harm.The study of tumor and its related complications has been paid more and more attention.In recent years,tumor diagnosis methods and treatment has made considerable progress.However,the treatment effect,prognosis and long-term survival rate of lung cancer,liver cancer,pancreatic cancer and other tumors are still poor.The main reason is that the mechanism of tumor development and development is not deep enough.The spine is the most common site of tumor bone metastasis,up to 70% of cancer patients will appear spinal metastases.metastatic spine tumor can lead to spinal cord compression and pathological fractures,so that patients with paralysis,complete loss of self-care ability,seriously affecting the quality of life of patients,thus accelerating the progress of the disease.The current mechanism of spinal metastases is not clear,resulting in the lack of effective treatment.For the treatment of lung cancer spinal metastases is still a major problem plagued the medical profession.Although the operation can effectively promote the recovery of patients,but complications,trauma and short easy to relapse,some patients die within a month after surgery.Therefore,to explore the key target molecules of lung cancer metastasis,it is to improve the clinical efficacy of patients with spinal metastases and improve the prognosis and quality of life in new directions.Some tumors are easily metastasis to bone,such as liver cancer,lung cancer,breast cancer and so on.Liver cancer bone metastasis is the most common site of the spine,the occurrence of spinal metastases in patients with survival rate of less than3%.Hepatocellular carcinoma(HCC)is a common malignancy within a global range,it accounts for more than 90% of liver cancer.Patients are often accompanied by intrahepatic and extrahepatic metastases.Early clinical symptoms and signs of HCCatypical,it is difficult to early detection,a clear diagnosis of most of the course has entered the late,loss of surgical opportunities.Because HCC has a high degree of malignancy,early detection is difficult,rapid progress,treatment complex,postoperative recurrence and metastasis rate is high,so that poor treatment of liver cancer patients,the prognosis is poor.In the past,HCC was mainly based on microRNA and mRNAs,long non-coding RNA was less studied.At present,the molecular mechanism of HCC at home and abroad is not deep enough,so we need to further study the molecular mechanism of HCC in the process of occurrence,development and metastasis,to find molecular markers that can predict the occurrence of HCC.The human genome is made up of large and complex nucleotide sequences.It is known that genes that encode proteins contain only 2% of the human genome sequence,and the remaining 98% of sequences that do not have the ability to encode proteins are considered to be useless "noises" in the genome.Such RNA molecules that are transcribed from the genome and that do not encode proteins are called non-coding RNAs(ncRNAs).Previously,the researchers only focused on coding RNA,but in recent years the researchers gradually attracted by non-coding RNA.With the deepening of research,it has been found that lncRNA is closely related to the occurrence and development of various human diseases,such as Alzheimer’s disease,cancer,Huntington’s disease.Long non-coding RNAs(lncRNAs)are defined as transcripts greater than 200nucleotides(nt)in length with limited or no protein-coding potential.However,the latest research shows that lncRNA plays an important role in the process of life activity and can influence biological behavior by regulating mRNA and protein expression levels.With the deepening of research,more and more tumor-related lncRNA were found,which through a variety of ways to regulate the tumor growth,metastasis,invasion and apoptosis,become a new tumor markers and drug targets,significantly advanced Advances in tumor research.Ji found by sequencing the metastasis-associated lung adenocarcinoma transcript1(MALAT 1),225 patients with stage Ⅲ non-small cell lung cancer(NSCLC)patients were screened,found that patients with metastatic specimens of 70 patients MALAT 1 over-expression,and has course of disease and tissue-specific,suggesting MALAT 1 can be used as stage Ⅰ NSCLC patients with potential markers.The genewas also found in other tumors,but was most pronounced in NSCLC patients.MALAT 1 promotes tumor metastasis by activating the relevant gene.It can also be involved in variable cleavage of mRNA precursors and phosphorylation of serine /arginine to influence gene function and expression.With the deepening of tumor genomics and transcriptomics research,the researchers found that a variety of different lncRNAs could be present in the same tumor,and that the same lncRNA could also be expressed abnormally in different types of tumors.HULC was first found in liver cancer cells overexpressed,can be used as primary liver cancer is more sensitive molecular markers,its in prostate cancer,gastric cancer also appears abnormal expression.H19 as a proto-oncogene in a variety of digestive system tumors increased expression.The above studies have shown that lncRNA plays an important role in oncology research,but at present both domestic and foreign research on lncRNA in metastatic spine tumor is still blank,the study of HCC is still less,and the vast majority of lncRNA Regulatory mechanism is not clear.Prompting us to screen and verify by high-throughput sequencing technology to find the new lncRNA as a tumor diagnosis and treatment target,for the metastatic spine tumor and liver cancer clinical diagnosis and treatment to provide a new strategy.Based on this we have carried out the following three parts of the study,are summarized below.Part 1: Analysis of lncRNA expression profile in spinal metastases of lung cancerObjective: Establish spinal metastases of lung cancer lncRNA and mRNA expression profiles,clearly differentially expressed mRNA molecular function,biological processes and information related to disease pathways.Method: First,we extracted RNA from tumor tissues and adjacent tissues of metastatic spine tumor,construction of lncRNA and mRNA Expression Profiles in metastatic spine tumor by High-throughput Sequencing Technique.And then the molecular function of differentially expressed m RNA,the biological process and disease pathways involved in the disease pathway analysis and GO analysis.Finally,the length of lncRNA and the number of exons were predicted.The expression of lncRNA was differentiated by coding the gene position.Result: We constructed lncRNA and mRNA expression profile of metastaticspine tumor and found 171 differentially expressed lncRNAs and 2929 differentially expressed mRNAs,of which 66 were new lncRNAs.The differentially expressed mRNA was analyzed by pathway analysis and GO analysis.The results showed that differentially expressed mRNAs in metastatic spine tumor were enriched in 31 diseases,79 pathways and 1535 molecular functions,thus further clarifying the molecular function of differentially expressed m RNA,participating in biological processes and pathogenesis adjust.Finally,we found that most of the differences in the expression of lncRNA exon number less than 5,the length of less than 10000 nt,are less than mRNA.Antisense total of 50,known 19,unknown 31;Sence total of 53,known 51,unknown 2;Bidirectionsl a total of 6,known 4,unknown 2;Intergenic total 40,known 20,unknown 20;Intronic a total of 22,known 8,unknown 14.Conclution: Construction of metastatic spine tumor lncRNA,mRNA expression profiles.Found 171 differentially expressed lncRNA,of which 66 are new.Found lncRNA 66 complete new unreported in the 171 differentially expressed in lncRNA.The 2929 differentially expressed mRNAs were identified for molecular function,biological processes and regulation of disease pathways.Part 2: Expression of lnc000489 in plasma of patients with metastatic spine tumor and its correlation with clinical featuresObjective: Find the key lncRNA mediated metastatic spine tumor,to understand it molecular functions,biological processes and disease pathways related information,to determine the diagnostic value of lncRNA in spinal metastases of lung cancer.Method: Combined with literature research,from the metastatic spine tumor lncRNA expression profile selected 20 new lncRNA.The plasma of lung cancer patients with spinal metastases was set as the experimental group and the healthy volunteers as the control group.The expression of lncRNA was significantly differentiated by qRT-PCR.The sensitivity and specificity were calculated by the ROC curve.And then differential expression of lncRNA co-expression analysis,to find co-expression of mRNA,pathway analysis and GO analysis of co-expressed mRNA were performed.The correlation between lncRNA and clinical features was analyzed statistically.Result: It was found that lnc00489 was significantly up-regulated in patients with metastatic tumor(P<0.05),and ROC curve analysis showed that lnc00489 haddefinite diagnostic value for metastatic spine tumor.There were 51 mRNAs that were co-expressed with lnc00489.Tumor-related diseases can be enriched in solid tumors,oral cancer,ovarian cancer,prostate cancer,etc,and Wnt,Hh,Hedgehog signaling pathway and Basal cell carcinoma.Further illustrates lnc00489 might have correlated with tumorigenesis,development and metastasis.The association of lnc00489 expression with various clinicopathological factors indicated that lnc00489 expression in plasma of patients were dramatically associated with the number of vertebral metastasis and visceral metastasis.The number of metastatic vertebral body increased,the more serious the disease.The main pathway of tumor metastasis is blood transfer,visceral and vertebral blood supply is relatively rich.In summary,lnc00489 plays an important role in the process of spinal metastasis,which is expected to be a potential plasma marker,which provides a new idea for the diagnosis and treatment of spinal metastases.Conclution: Lnc00489 is highly expressed in plasma of patients with metastatic spine tumor and has diagnostic value for spinal metastases.Lnc00489 co-expressed mRNA can be linked to a variety of tumor-related diseases and pathways.The expression level of lnc00489 in plasma was correlated with the number of metastatic vertebral body and visceral metastasis.It is expected to become a new molecular marker for the diagnosis of metastatic spine tumor.Part 3: Expression of lnc34822 in plasma of patients with HCC and its correlation with clinical featuresObjective: To find a different expression lncRNA in the HCC from expression profile of the metastatic spine tumor,clear its diagnostic value in the HCC.Method: Combined with related literature,10 highly expressed lncRNA were selected from the lncRNA expression profile of metastatic spine tumor.Large sample validation clearly indicates whether there are lncRNAs that are highly expressed in HCC.The sensitivity and specificity were calculated using ROC.Use qRT-PCR technology to detect candidate lncRNA expression in a tissue,plasma and cell lines(Hep3B、HepG-2、SMMC-7721、Huh7.5、LO2).QRT-PCR was used to verify the stability of differentially expressed lncRNA in plasma and its relationship with blood cells.Verify the differential expression of lncRNA in patients with liver cancer before,during and after the recurrence of plasma changes in the expression of the situation.The correlation between lncRNA and clinical features was analyzed statistically.Result: It has been reported that the same lncRNA can be regulated in different tumors.The same tumor can also be affected by a variety of different lncRNA.Ten lncRNAs were screened from the lncRNA database of the metastatic spine tumor.It was found that lnc34822 was significantly up-regulated in patients with HCC(P<0.01),and it can be expressed stably in plasma.The expression level of lnc34822 in hepatocellular carcinoma cell line was significantly higher than that in normal liver cell line,and was not correlated with peripheral blood cell.ROC curve analysis showed that lnc34822 had definite diagnostic value for HCC.Lnc34822 in HCC patients with postoperative plasma expression was significantly reduced,and the recurrence of rapid rise.It is concluded that lnc34822 expected to be a dynamic marker for monitoring the progression of HCC patients.The expression of lncRNA34822 in plasma was positively correlated with AFP and tumor number by correlation analysis with clinical information.The level of AFP in blood of patients with liver cancer was significantly increased,and it has become a widely used tumor marker.Increased number of liver tumors is more severe.Indicating that lnc34822 is expected to be a potential plasma marker for HCC.Conclution: Lnc34822 was highly expressed in plasma of HCC patients and was not associated with peripheral blood cells.It has a certain diagnostic value of HCC.The expression of lnc34822 in plasma can dynamically detect the progression of HCC patients.It is expected to be a new plasma molecule marker for the diagnosis of HCC.Through the above research,we successfully constructed a new lncRNA database,which provides data resources for oncology research.we found the role of regulation of lncRNA in the development of metastatic spine tumor and HCC.For the spinal metastases and HCC clinical treatment and treatment provides a new idea.