VEGF、HIF-1α、PTEN与JAK2V617F阳性骨髓增殖性肿瘤血管新生相关性研究

发布时间：2018-06-25 20:45

本文选题：JAK2V617F + 髓增殖性肿瘤　；参考：《承德医学院》2016年硕士论文

【摘要】：目的:观察促血管新生因子:血管内皮生长因子(VEGF)、缺氧诱导因子1α(HIF-1α)及血管新生因子抑制因子:与张力蛋白同源的10号染色体缺失的磷酸酶基因(PTEN)在Janus蛋白酪氨酸激酶2(JAK2)点突变(JAK2V617F)阳性骨髓增殖性肿瘤(MPN)患者骨髓中的表达水平,探讨它们与血管新生之间的相互关系。方法:1本研究所用的52例石蜡标本均取自保定市第一医院病理科,选取自2012年1月至2014年10月保定市第一医院门诊及住院收治的42例JAK2V617F表达阳性的MPN患者及10例特发性免疫性血小板减少性紫癜(ITP)患者。MPN患者包括真性红细胞增多症(PV)10例,原发性血小板增多症(ET)17例,原发性骨髓纤维化(PMF)15例,其中男18例,女24例,年龄32~75岁,中位年龄57岁。ITP患者包括男5例,女5例,中位年龄40岁。所选取患者的诊断均符合《血液病诊断及疗效标准》,患者均知情同意及经保定市第一医院伦理委员会批准。2实验分组:JAK2V617F阳性的MPN患者中初治组(初诊未治疗组)25例;治疗组(该组患者均应用干扰素-α2b(IFN-α2b)肌肉或皮下注射,300万U/次,3~7次/周,至少应用6个月以上,联合或不联合应用羟基脲调整外周血白细胞、血红蛋白及血小板数量。)17例。10例特发性免疫性血小板减少性紫癜(ITP)患者作为对照组。3通过实时荧光定量PCR检测MPN患者JAK2V617F突变量(突变型与野生型JAK2比值)。4应用免疫组化方法检测初治组、治疗组及对照组患者骨髓病理切片p-JAK2、VEGF、HIF-1α、PTEN的蛋白表达水平及CD105标记的微血管密度(MVD)。5所有数据均采用SPSS 17.0统计分析软件,计量资料以(x±s)表示,两组均数间的比较采用t检验,多组均数比较采用方差分析,两两比较采用q检验。spearman等级相关分析各变量之间的相关性。jak2v617f突变阳性中jak2v617f突变量、vegf、hif-1α、mvd表达的比较及四指标与临床特征(血栓栓塞发生率)的关系分析采用χ2检验;p0.05为差异有统计学意义。结果:1mpn患者的jak2v617f突变量pcr结果显示:42例mpn患者在初次诊治时均经实时荧光定量pcr检测显示jak2v617f突变为阳性,经过治疗后jak2v617f突变转阴性者2例,其余40例mpn患者jak2v617f突变量在26.8%-75.4%之间。其中初治组25例,jak2v617f突变量为46.17%±19.32%,干扰素治疗组17例,jak2v617f突变量为22.69%±12.64%,前者明显高于后者(p0.05)。2免疫组织化学染色检测p-jak2、vegf、hif-1α、pten、mvd在mpn患者中的表达水平p-jak2、vegf、hif-1α及pten蛋白均表达于细胞质,其中p-jak2、vegf、hif-1α三者骨髓细胞中阳性率在初治组(82.41%±11.65%;64.72%±25.01%;45.12%±20.28%)中表达最高,其次为治疗组(60.93%±20.57%;36.58%±15.95%;32.15%±14.27%),对照组(43.05%±12.59%;25.69%±13.75%;25.07%±15.49%)最低,差异均有统计学显著性(p0.01);mvd在初治组(26.58%±5.93%)中显著高于治疗组(15.86%±4.27%)及对照组(10.76%±4.01%),pten蛋白在初治组(24.78%±14.86%)中表达最低,其次为治疗组(38.72%±18.25%),对照组表达(55.05%±21.17%)最高,差异均有统计学意义(p0.01)。3jak2v617f突变量与p-jak2、vegf、hif-1α、pten、mvd相关分析spearman等级相关分析显示mpn患者的jak2v617f突变量和p-jak2、vegf、mvd正相关(r=0.739,p0.01、r=0.589,p0.05、r=0.577,p0.05),与hif-1α无明显相关(p0.05),与pten负相关(r=-0.508,p0.05)。pten蛋白表达与mvd负相关(r=-0.584,p0.05)。以jak2v617f/jak2比值50%为界值分为50%与≥50%两组,其中50%组26例,≥50%组16例。结果显示jak2v617f/jak250%的患者其p-jak2、vegf、hif-1α及mvd(65.15%±17.59%、43.26%±18.67%、33.27%±15.26%、19.16%±5.34%)的水平均明显低于jak2v617f/jak2≥50%组(89.76%±20.56%、69.87%±27.32%、49.51%±22.65%、34.19%±6.57%),pten在jak2v617f/jak2比值50%患者的水平(34.42%±19.76%)要明显高于jak2v617f/jak2比值≥50%(22.68%±13.12%)患者(p均0.05)。初治组中(46.17%±19.32%;82.41%±11.65%;64.72%±25.01%;45.12%±20.28%;26.58%±5.93%)jak2v617f突变量及p-jak2、vegf、hif-1α、mvd的水平均明显高于ifn-α2b治疗组(22.69%±12.64%;60.93%±20.57%;64.72%±25.01%;32.15%±14.27%;15.86%±4.27%;15.86%±4.27%)及对照组(0;43.05%±12.59%;25.69%±13.75%;25.07%±15.49%;10.76%±4.01%)(p均0.05);而pten与上述相反,在初治组(24.78%±14.86%)中表达最低,其次为治疗组(15.86%±4.27%),在对照组(10.76%±4.01%)中表达最高(p均0.05)。4jak2v617f及mvd与血栓栓塞的相关性42例患者,初治组25例,血栓栓塞15例,血栓发生率为60.00%;治疗组17例,再发血栓栓塞4例(既往发生血栓者不计在内),血栓发生率为23.53%;后者明显低于前者(χ2=5.43,p0.05)。42例mpn患者中19例患者发生血栓,血栓发生率为45.24%,其中26例jak2v617f/jak2比值50%组,发生血栓栓塞者8例,血栓发生率为30.77%,16例≥50%组,发生血栓栓塞者11例,血栓发生率为68.75%,后者明显高于前者(χ2=5.77,p0.05)。42例患者总vegf为53.40%±21.97%,其中vegf53.40%患者24例,血栓栓塞7例,血栓发生率为33.33%,vegf53.40%患者18例,血栓栓塞12例,血栓发生率为66.67%,后者明显高于前者(χ2=5.84,p0.05)。42例患者总hif-1α为39.46%±18.08%,其中hif-1α39.46%患者22例,血栓栓塞6例,血栓发生率为31.82%,hif-1α39.46%患者20例,血栓栓塞13例,血栓发生率为60.00%,后者明显高于前者(χ2=6.02,p0.05)。42例患者总mvd为22.24±7.49,其中mvd22.24患者23例,血栓栓塞7例,血栓发生率为为30.43%,mvd22.24患者19例,血栓栓塞12例,血栓发生率为63.16%,后者明显高于前者(χ2=4.50,P0.05)。结果表明,经干扰素治疗后,随着JAK2V617F突变量的减低,患者血栓栓塞发生率亦减低;JAK2V617F突变量愈高,MVD愈高,VEGF表达越高,HIF-1α表达越高,患者血栓栓塞发生率愈高,呈正相关关系。结论:1 PTEN、VEGF、HIF-1α与JAK2V617F共同参与了骨髓增殖性肿瘤患者血管新生。2 JAK2V617F阳性MPN患者在应用IFN-α2b治疗后JAK2V617F突变量明显减少,部分患者转阴性,IFN-α2b能够抑制MPN患者JAK2V617F突变量。3 VEGF、HIF-1α及MVD在JAK2V617F阳性患者中表达增加,而PTEN表达减低。4 JAK2V617F基因突变量越高者VEGF、HIF-1α、MVD越高则血管新生越明显,而与PTEN表达负相关。5 JAK2V617F突变量愈高,VEGF表达越高,HIF-1α表达越高,骨髓微血管密度愈高,患者发生血栓风险则愈高,随着JAK2V617F突变量的减低,患者血栓栓塞发生率亦减低。
[Abstract]:Objective: To observe vascular endothelial growth factor: vascular endothelial growth factor (VEGF), hypoxia inducible factor 1 alpha (HIF-1 alpha) and angiogenesis factor inhibitory factor: phosphatase gene (PTEN), which is homologous to tension protein 10 chromosome deletion (PTEN) in Janus protein tyrosine kinase 2 (JAK2) point mutation (JAK2V617F) positive marrow proliferative tumor (MPN) patients' bone marrow The relationship between them and angiogenesis was discussed. Methods: 1 paraffin specimens from 1 studies were taken from the pathology department of the first hospital in Baoding. From January 2012 to October 2014, 42 cases of MPN patients with positive MPN and 10 idiopathic immune blood were treated in the outpatient and hospitalized Hospital of the first hospital of the city. .MPN patients included 10 cases of real erythrocytosis (PV), 17 cases of primary thrombocythemia (ET) and 15 cases of primary myelofibrosis (PMF), of which 18 cases were male, 24 women, age 32~75 years, and 57 year old.ITP patients, including 5 men, 5 women, and middle age 40. The diagnosis of the selected patients was in line with < blood. The diagnosis and efficacy criteria of liquid disease, the patients' informed consent and the.2 experimental group approved by the ethics committee of the first hospital of Baoding: 25 patients with JAK2V617F positive MPN patients (first diagnosed untreated group); the treatment group (all the patients were treated with interferon - alpha 2b (IFN- alpha 2b) muscle or subcutaneous injection, 3 million U/ times, 3~7 times per week, at least 6 months. Above, combined or non combined use of hydroxyurea in peripheral blood leukocytes, hemoglobin and platelet counts.) 17 cases of.10 patients with idiopathic thrombocytopenic purpura (ITP) as control group,.3 by real-time fluorescence quantitative PCR detection of MPN patients JAK2V617F process variable (mutate and wild type JAK2 ratio).4 application immunization method The p-JAK2, VEGF, HIF-1, PTEN protein expression level and the microvascular density (MVD).5 of CD105 markers in the treatment group and the control group were measured with SPSS 17 statistical analysis software, and the measurement data were (x + s). The comparison between the two groups was carried out by t test, and the multiple groups were compared with variance analysis. 22 compared the correlation analysis of the correlation.Jak2v617f mutation between the variables of.Spearman and the correlation analysis between the variables, the comparison of the VEGF, HIF-1 alpha, MVD expression and the relationship between the four indexes and the clinical characteristics (thromboembolism incidence) by the Q test. The analysis of the relationship between the four indexes and the clinical characteristics (thromboembolism incidence) was analyzed by the chi 2 test; P0.05 was statistically significant. The result: JAK2V617F of the 1mpn patients. The results of sudden variable PCR showed that 42 cases of MPN patients were detected by real-time fluorescence quantitative PCR at the first time of diagnosis and JAK2V617F mutation was positive. After treatment, 2 cases were JAK2V617F mutation negative, and the other 40 cases of MPN patients were between 26.8%-75.4%. The initial treatment group was 25 cases, JAK2V617F process variable was 46.17% + 19.32%, interferon treatment. In group 17, the JAK2V617F process variable was 22.69% + 12.64%. The former was significantly higher than that of the latter (P0.05).2 immunohistochemical staining for p-jak2, VEGF, HIF-1 alpha, PTEN, MVD in MPN patients, p-jak2, VEGF, HIF-1 alpha and protein were expressed in the cytoplasm, and the positive rate in bone marrow cells was 82.41% + 11. (82.41% + 11.). The expression of 65%, 64.72% + 25.01%, 45.12% + 20.28%) was the highest, followed by the treatment group (60.93% + 20.57%; 36.58% + 15.95%; 32.15% + 14.27%), and the control group (43.05% + 12.59%; 25.69% + 20.28%) was the lowest, and the difference was statistically significant (P0.01); MVD was significantly higher in the primary treatment group than the treatment group and the control group. .76% + 4.01%), the expression of PTEN protein was the lowest in the primary treatment group (24.78% + 14.86%), followed by the treatment group (38.72% + 18.25%) and the control group (55.05% + 21.17%), the difference was statistically significant (P0.01).3jak2v617f abrupt variable and p-jak2, VEGF, HIF-1 a, PTEN, MVD correlation analysis of Spearman grade correlation analysis showed JAK2V617F process variables of MPN patients The positive correlation with p-jak2, VEGF, MVD (r=0.739, P0.01, r=0.589, P0.05, r=0.577, P0.05) was not significantly correlated with HIF-1 alpha (P0.05), and was negatively correlated with PTEN negative correlation (r=0.739). The value of the ratio 50% was divided into 50% and more than 50% groups, including 50% groups 26 cases, and 50% groups 16 cases. The levels of p-jak2, VEGF, HIF-1 alpha and MVD (65.15% + 17.59%, 43.26% + 18.67%, 33.27% + 15.26%, 19.16% + 5.34%) were significantly lower than those of jak2v617f/jak2 > 50% group (89.76% + 20.56%, 69.87% + 27.32%, 43.26% + 33.27%). The level of PTEN at JAK2V617F /jak2 ratio was significantly higher than that of jak2v617f/. The JAK2 ratio was more than 50% (22.68% + 13.12%) (P 0.05). In the primary treatment group (46.17% + 19.32%; 82.41% + 11.65%; 64.72% + 25.01%; 45.12% + 20.28%; 26.58% + 5.93%), the level of p-jak2, VEGF, HIF-1 alpha and MVD was significantly higher than that of ifn- alpha 2b; 86% + 4.27%) and the control group (0; 43.05% + 12.59%; 25.69% + 13.75%; 25.07% + 15.49%; 10.76% + 4.01%) (P 0.05); while PTEN was the lowest in the primary treatment group (24.78% + 14.86%), followed by the treatment group, the highest (P mean).4jak2v617f and MVD and thromboembolism were expressed in the control group. In the patients, there were 25 cases in the primary treatment group, 15 cases of thromboembolism and 60% thrombus, 17 cases in the treatment group, 4 cases of recurrent thromboembolism (not included in the previous thrombus), the thrombus incidence was 23.53%, the latter was significantly lower than the former (x 2=5.43, P0.05).42 cases MPN patients with thrombosis, the incidence of thrombosis was 45.24%, of which 26 cases jak2v617f/jak2 ratio. In the 50% group, thromboembolism occurred in 8 cases, thrombus incidence was 30.77%, 16 cases were more than 50%, thromboembolism occurred in 11 cases, thrombus incidence was 68.75%, the latter was significantly higher than the former (x 2=5.77, P0.05).42 patients with total VEGF 53.40% + 21.97%, 24 cases of vegf53.40%, thromboembolism 7 cases, thrombus incidence of 33.33%, 18 cases of vegf53.40% patients, 18 cases, vegf53.40% patients, Thromboembolism in 12 cases, the incidence of thrombus was 66.67%, the latter was significantly higher than the former (x 2=5.84, P0.05).42 cases, the total HIF-1 alpha was 39.46% + 18.08%, including 22 cases of HIF-1 alpha 39.46%, thromboembolism in 6 cases, thrombus incidence of 31.82%, 20 cases of HIF-1 a 39.46%, thromboembolism in 13 cases, thrombus incidence of 60%, the latter was significantly higher than the former (x 2=6.02, P) 0.05) the total MVD of.42 patients was 22.24 + 7.49, of which 23 cases of mvd22.24, 7 thromboembolism, 30.43% thrombus, 19 cases of mvd22.24, 12 thromboembolism and 63.16% thrombus, the latter was significantly higher than that of the former (x 2=4.50, P0.05). The results showed that after interferon treatment, thrombus thrombus was decreased along with the decrease of JAK2V617F process variable. Thrombus thrombus The higher the incidence of plug is, the higher the JAK2V617F process, the higher the MVD, the higher the expression of VEGF, the higher the expression of HIF-1 a, the higher the incidence of thromboembolism in the patients. Conclusion: 1 PTEN, VEGF, HIF-1 alpha and JAK2V617F are involved in the new.2 JAK2V617F positive MPN patients in the patients with proliferative tumor of the bone marrow. IFN- alpha 2b could inhibit.3 VEGF of JAK2V617F process in MPN patients, HIF-1 A and MVD increased in JAK2V617F positive patients, while PTEN expression reduced the greater of.4 genes, the higher the higher then the higher then the more obvious, and the negative correlation with the expression of 2B. The higher the quantity, the higher the expression of VEGF, the higher the expression of HIF-1 a, the higher the density of the bone marrow microvessel, the higher the risk of thrombosis in the patients. The incidence of thromboembolism is also decreased with the decrease of the variable of JAK2V617F.
【学位授予单位】：承德医学院
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R733.3

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