| Objective:Myeloid Neoplasm mainly includes Acute myeloid leukemia(AML),Myelodysplastic syndrome(MDS)and Chronic myeloproliferative neoplasm(cMPN).Due to the clinical and genetic high heterogeneity of myeloid neoplasm,the specific pathogenesis is still unclear.Nowadays,with the deep study of myeloid neoplasm,more and more epigenetic abnormalities were found in tumor cells,they can regulate the gene expression by DNA methylation,chromatin remodeling,histone modifications and no-coding RNA regulations,etc.Histone modifications mainly contain methylation,acetylation,phosphorylation and ubiquitination and so on.Lysine methylation is a particularly important way in histone modifications.Lysine Specific Demethylase 1(LSD1)is the first real sense of histone lysine demethylase,a flavin adenine dinucleotide-dependent amino oxidase,which can generate demethylation lysine acid residues and formaldehyde by the amino oxidation,and then achieve the purpose of demethylation.Histone lysine methylation is mainly catalyzed by methyl transferase containing SET domain.H3K4 and H3K36 methylation are usually related to the chromatin activation,whereas,the H3K9,H4K20 and H3K27 methylation are associated with gene silencing.Histone lysine methyltransferase(enhancer of zeste homolog 2,EZH2)can catalytic the H3K27 methylation sites,then silence the tumor suppressor genes,and lead to tumorigenesis.This makes people realized that histone methylation is a dynamic process,the imbalance between histone methylation enzymes and methyltransferases can lead to the disorder of histone methylation,recent studies suggest that there is a close relationship between the imbalance of histone methylation enzymes and methyltransferases and the occurrence of myeloid neoplasm.Therefore,deeply exploring the mechanism of histone methylation enzymes and methyltransferases in the occurrence of myeloid neoplasm has great clinhical guiding significance.Methods:70 cases Myeloid Neoplasm patients' bone marrow samples from Second Hospital of Hebei Medical University were collected for the test group in this research,including AML 40 cases,MDS 15 cases,cMPN 15 cases(essential thrombocythemia,ET 7 cases;polycythemia vera,PV 5 cases;primary myelofibrosis,MF 3 cases).And 16 cases bone marrow samples of treated AML patients',17 cases bone marrow samples of patients with non-myeloid neoplasm disease were prepared to set as control group.The expression level of LSD1 mRNA in bone marrow mononuclear cells(BM-MNCs)could be detected by the application of RT-PCR,and LSD1 protein and EZH2 protein levels could be assayed by ELISA enzyme-linked immunosorbent.Results:1 The expression level of LSD1 mRNART-PCR results showed that among the untreated Myeloid Neoplasm patients,the relative expression quantity of LSD1 mRNA in AML group,MDS group and cMPN group was(0.9925± 0.1958),(0.9536± 0.1603)and(1.0062±0.1675)respectively,the relative expression level of LSD1 mRNA in treated AML group was(0.9515±0.1015),while the relative expression quantity of LSD1 mRNA in control group was(0.9621± 0.1687).These indicated that there was no statistical difference of the relative expression of LSD1 mRNA between untreated AML group and control group(P =0.587);there was no statistical difference of the relative expression of LSD1 mRNA between MDS group and control group(P =0.911);there was no statistical difference of the relative expression of LSD1 mRNA between cPMN group and control group(P =0.544);there was no statistical difference of the relative expression of LSD1 mRNA between untreated AML group and treated AML group(P =0.465),there was no statistical difference of the relative expression of LSD1 mRNA between treated AML group and control group(P =0.862).2 The expression level of LSD1 proteinELISA enzyme-linked immunosorbent results showed that among the untreated Myeloid Neoplasm patients,the relative expression quantity of LSD1 protein in AML group,MDS group and cMPN group was(18.5737± 1.8809),(12.5276± 1.3824)and(12.0236±1.1412)respectively,the relative expression level of LSD1 protein in treated AML group was(8.5756± 1.5808),while the relative expression quantity of LSD1 protein in control group was(4.6231± 1.3155).These indicated that the relative expression of LSD1 protein in untreated AML group was higher than that in control group,and the difference was statistically significant(P =0.00);the relative expression of LSD1 protein in MDS group was higher than that in control group,and the difference was statistically significant(P =0.00);the relative expression of LSD1 protein in cMPN group was higher than that in control group,and the difference was statistically significant(P =0.00).The relative expression of LSD1 protein in treated AML group was lower than that in untreated AML group,and the difference was statistically significant(P =0.00);the relative expression of LSD1 protein in treated AML group was higher than that in control group,and the difference was statistically significant(P =0.00).All the untreated AML patients were grouped as M2 15 cases,M4 13 cases,M5 12 cases by subtype,and the relative expression quantity of LSD1 protein in M2 group,M4 group and M5 group was(18.4879± 2.1489),(18.6596± 1.8493)and(8.5878±1.7078)respectively,and there was no statistical difference of the relative expression of LSD1 protein among each group(P =0.972).All the untreated AML patients were grouped as older(more than 60 years)7 cases,younger(less than 60 years)33 cases by age,and the relative expression quantity of LSD1 protein in older group and younger group was(18.3177± 2.1077)and(18.6280± 1.8605)respectively,and there was no statistical difference of the relative expression of LSD1 protein between the two groups(P =0.697).All the untreated AML patients were grouped as male 23 cases and female 17 cases by sex,and the relative expression quantity of LSD1 protein in male group and female group was(18.6377± 2.0178)and(18.4871± 1.7353)respectively,and there was no statistical difference of the relative expression of LSD1 protein between the two groups(P =0.806).All the MDS patients were grouped as high-risk group 7 cases and low-risk group 8 cases by IPSS-R,and the relative expression quantity of LSD1 protein in high-risk group and low-risk group was(13.7541± 0.7302)and(13.7541± 0.7302)respectively,and the relative expression of LSD1 protein in high-risk group was higher than that in low-risk group,and the difference was statistically significant(P =0.00).All the cMPN patients were grouped as ET 7 cases,PV 5 cases,MF 3 cases by WHO,and the relative expression quantity of LSD1 protein in ET group,PV group and MF group was(12.1291± 1.0428),(12.3386±1.3338)and(11.2523±1.0570)respectively,and there was no statistical difference of the relative expression of LSD1 protein among each group(P =0.436).3 The expression level of EZH2 proteinELISA enzyme-linked immunosorbent results showed that among the untreated Myeloid Neoplasm patients,the relative expression quantity of EZH2 protein in AML group,MDS group and cMPN group was(268.2885± 16.4276),(204.5954±17.7583)and(199.3212±10.8547)respectively,the relative expression level of EZH2 protein in treated AML group was(194.8555± 12.9244),while the relative expression quantity of EZH2 protein in control group was(175.3548± 13.9937).These indicated that the relative expression of EZH2 protein in untreated AML group was higher than that in control group,and the difference was statistically significant(P =0.00);the relative expression of EZH2 protein in MDS group was higher than that in control group,and the difference was statistically significant(P =0.00);the relative expression of EZH2 protein in cMPN group was higher than that in control group,and the difference was statistically significant(P =0.00).The relative expression of EZH2 protein in treated AML group was lower than that in untreated AML group,and the difference was statistically significant(P =0.00);the relative expression of EZH2 protein in treated AML group was higher than that in control group,and the difference was statistically significant(P =0.00).All the untreated AML patients were grouped as M2 15 cases,M4 13 cases,M5 12 cases by subtype,and the relative expression quantity of EZH2 protein in M2 group,M4 group and M5 group was(267.8381± 17.1344),(266.6426± 18.0838)and(270.63463±14.7275)respectively,and there was no statistical difference of the relative expression of EZH2 protein among each group(P =0.832).All the untreated AML patients were grouped as older(more than 60 years)7 cases,younger(less than 60 years)33 cases by age,and the relative expression quantity of EZH2 protein in older group and younger group was(267.6477± 17.4577)and(269.2461± 16.2742)respectively,and there was no statistical difference of the relative expression of EZH2 protein between the two groups(P =0.102).All the untreated AML patients were grouped as male 23 cases and female 17 cases by sex,and the relative expression quantity of EZH2 protein in male group and female group was(267.3703± 16.5247)and(269.5308±16.7175)respectively,and there was no statistical difference of the relative expression of EZH2 protein between the two groups(P =0.686).All the MDS patients were grouped as high-risk group 7 cases and low-risk group 8 cases by IPSS-R,the relative expression quantity of EZH2 protein in high-risk group and low-risk group was(221.3522± 7.7296)and(189.9333±7.2821)respectively,and the relative expression of EZH2 protein in high-risk group was higher than that in low-risk group,and the difference was statistically significant(P =0.00);All the cMPN patients were grouped as ET 7 cases,PV 5 cases,MF 3 cases by WHO,and the relative expression quantity of EZH2 protein in ET group,PV group and MF group was(202.0684± 8.5480),(201.0672±13.6585)and(190.0010±8.2533)respectively,and there was no statistical difference of the relative expression of EZH2 protein among each group(P =0.264).4 Statistical analysis Of the ratio between LSD1 protein and EZH2 proteinThe statistical results showed that among the untreated Myeloid Neoplasm patients,the ratio between LSD1 protein and EZH2 protein in AML group,MDS group and cMPN group was(0.0693± 0.0066),(0.0615±0.0073)and(0.0603±0.0035)respectively,the ratio between LSD1 protein and EZH2 protein in treated AML group was(0.0437± 0.0053),while the ratio between LSD1 protein and EZH2 protein in control group was(0.0260± 0.0056).These indicated that the ratio between LSD1 protein and EZH2 protein in untreated AML group was higher than that in control group,and the difference was statistically significant(P =0.00);the ratio between LSD1 protein and EZH2 protein in MDS group was higher than that in control group,and the difference was statistically significant(P =0.00);the ratio between LSD1 protein and EZH2 protein in cMPN group was higher than that in control group,and the difference was statistically significant(P =0.00).The ratio between LSD1 protein and EZH2 protein in treated AML group was lower than that in untreated AML group,and the difference was statistically significant(P =0.00);the ratio between LSD1 protein and EZH2 protein in treated AML group was higher than that in control group,and the difference was statistically significant(P =0.00).All the untreated AML patients were grouped as M2 15 cases,M4 13 cases,M5 12 cases by subtype,and the ratio between LSD1 protein and EZH2 protein in M2 group,M4 group and M5 group was(0.0690± 0.0070),(0.0701±0.0067)and(0.0688±0.0066)respectively,and there was no statistical difference of the ratio between LSD1 protein and EZH2 protein among each group(P =0.876).All the untreated AML patients were grouped as older(more than 60 years)7 cases,younger(less than 60 years)33 cases by age,and the ratio between LSD1 protein and EZH2 protein in older group and younger group was(0.0684± 0.0062)and(0.0692±0.0066)respectively,and there was no statistical difference of the ratio between LSD1 protein and EZH2 protein between the two groups(P =0.757).All the untreated AML patients were grouped as male 23 cases and female 17 cases by sex,and the ratio between LSD1 protein and EZH2 protein in male group and female group was(0.0698± 0.0072)and(0.0686±0.0059)respectively,and there was no statistical difference of the ratio between LSD1 protein and EZH2 protein between the two groups(P =0.601).All the MDS patients were grouped as high-risk group 7 cases and low-risk group 8 cases by IPSS-R,the ratio between LSD1 protein and EZH2 protein in high-risk group and low-risk group was(0.0621± 0.0025)and(0.0603±0.0039)respectively,and there was no statistical difference of the ratio between LSD1 protein and EZH2 protein among each group(P =0.315).All the cMPN patients were grouped as ET 7 cases,PV 5 cases,MF 3 cases by WHO,and the ratio between LSD1 protein and EZH2 protein in ET group,PV group and MF group was(0.0600± 0.0044),(0.0612±0.0027)and(0.0592±0.0031)respectively,and there was no statistical difference of the ratio between LSD1 protein and EZH2 protein among each group(P =0.738).5 Relationship between the expression of LSD1 protein and EZH2 proteinThe correlation between LSD1 protein expression and EZH2 protein expression of 103 samples were analysed.The result showed that there was a positive correlation between the two sets of data,the Spearman correlation was 0.910,which was statistically significant(P =0.00).Conclusions:1 There was no significant difference of the expression of LSD1 mRNA between the newly diagnosed Myeloid Neoplasm patients and the control group,but the LSD1 protien levels in Myeloid Neoplasm patients were higher than the control group,which indicated that LSD1 may regulate the pathogenesis of Myeloid Neoplasm by some mechanisms.2 The EZH2 protien levels in Myeloid Neoplasm patients were higher than the control group.3 From untreated AML patients to treated AML patients the expression level of LSD1 protein and EZH2 protein decreased gradually.4 There was an imbalance between LSD1 protein and EZH2 protien expressed in newly diagnosed Myeloid Neoplasm patients,and some certain correlation may be exist. |
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