| Objective: This article aims to explore the factors that influencedisease progression in acute leukemia (AL) by examining the expression ofhigh mobility group box1protein (HMGB1), vascular endothelial growthfactor (VEGF)including VEGFA and VEGFC, and their receptors VEGFR1,VEGFR2, VEGFR3, and provide a evidence for supplementing the newtreatments.Methods:(1) A total of71AL patients, hospitalized from March toOctober in2012, were recruited at the Department of Hematology, TheSecond Affiliated Hospital of Dalian Medical University, including34primary untreated patients,27complete remission cases,10relapsed orrefractory cases. Among them,42had acute non-lymphocytic leukemia (M1:1cases, M2:9cases, M3:16cases,M4:1case, M5:9cases, M6:6cases),including20patients without primary treatment,18cases with completeremission,4relapsed or refractory patients; and29had acute lymphoblasticleukemia patients (L1:12cases, L2:17cases), including14primaryuntreated patients,9complete remission cases,6relapsed or refractorypatients. All patients were diagnosed by clinical and peripheral blood, bonemarrow cell morphology and staining, in accordance with standardizeddiagnosis and treatment procesures. All cases have been eliminated suchdiseases as heart, liver and kidney dysfunctions, acute and chronicinfections, other malignancies, autoimmune diseases and pregnancy. In71cases, there were37males and34females aged14-79years old, with amedian age of46years old. The control group consisted of30healthy cases selected from the medical exam center in the same hospital, with14malesand16females aged16-75years old, with a median age of47years old.(2)For each patient, approximately1ml serum was isolated from3ml DUTfasting venous blood and stored at-20℃until further analysis.(3)Double-antibody sandwich enzyme-linked immunosorbent assay (ELISA)using antibody-coated96-well microtiter plates, is made of solid phase.Standards or samples were added into the microporous in which theantibody connects to binding antibody on the solid phase carrier, thenbiotinylated antibody was added, with unbound biotinylated antibodywashed off, and then HPR-labeled avidin was added. Undergoingthoroughly washed again, TMB was added for coloration. TMB conversesinto the blue under peroxidase catalyzing, and into yellow with the presenceof acid. The antibody was positively correlated with the depth of color andsamples. The absorbance (OD) was measured with a microplate reader at450nm wavelength, calculate the concentration of the sample.(4) The levelsof HMGB1, VEGFA, VEGFC, VEGFR1, VEGFR2and VEGFR3in serumwere determined by ELISA assay.Results:1.We found that the expressions of HMGB1in primaryuntreated group and relapsed or refractory group were significantly higherthan the normal control group in ANLL and ALL patients, the differenceswere statistically significant (P<0.05), We found in ANLL, the expressionsof HMGB1in complete remission group were significantly decreased, incomparison with primary untreated group, but compared with the normalcontrol group, the differences were not statistically significant (P>0.05), thethe expressions of HMGB1in relapsed or refractory group weresignificantly higher than normal control group and complete remissiongroup, the differences were statistically significant (P<0.05). We found inANLL group, the serum levels of HMGB1in primary untreated group were(517.17±477.49) pg/ml, the serum levels of HMGB1in complete remissiongroup were (88.66±75.74) pg/ml, the serum levels of HMGB1in relapsedor refractory group were (1016.25±978.49) pg/ml. We found in ALL group,the serum levels of HMGB1in primary untreated group were (659.98±479.55) pg/ml, the serum levels of HMGB1in complete remission group were(73.75±47.44) pg/ml, the serum levels of HMGB1in relapsed orrefractory group were (677.00±354.10) pg/ml.The serum levels of HMGB1in normal control group were (15.87±15.98) pg/ml.2. By measuring the expressions of VEGFA, VEGFC, VEGFR1,VEGFR2, VEGFR3from serum in patients with AL, we found in ANLLgroup the expressions of VEGFA, VEGFR1, VEGFR3in the primaryuntreated group were significantly higher than the normal control group, thedifferences were statistically significant (P<0.05), the expressions ofVEGFA, VEGFR1, VEGFR3in complete remission group were decreased,in comparison with the primary untreated group, compared with the normalcontrol group the differences were not statistically significant (P>0.05). theexpressions of VEGFR2in the primary untreated group and completeremission group were significantly higher than the normal control group,the differences were statistically significant (P<0.05, P=0.05), there wereno statistical differences between the levels of VEGFR2in primaryuntreated group and complete remission group (P>0.05). VEGFC were notexpressed in primary untreated group,complete remission group and normalcontrol group. ANLL group: the levels of VEGFA in the primary untreatedgroup were (559.60±542.92) pg/ml and the levels of VEGFA in completeremission were (21.24±21.47) pg/ml,the levels of VEGFA in normal controlgroup were ((92.46±42.35) pg/ml. The levels of VEGFR1in primaryuntreated group were (0.30±0.09) ng/ml, the levels of VEGFR1incomplete remission group were (0.15±0.11) ng/ml,the levels of VEGFR1innormal control group were (0.04±0.03) ng/ml. The levels of VEGFR2inprimary untreated group were (1.53±0.34) ng/ml, the levels of VEGFR2incomplete remission group were (1.30±0.10) ng/ml, the levels of VEGFR2innormal control group were (0.44±0.56) ng/ml. The levels of VEGFR3inprimary untreated group were (13.05±4.21) ng/ml, the levels of VEGFR3in complete remission group were (3.13±1.71) ng/ml, the levels ofVEGFR3in normal control group were(2.62±1.31) ng/ml.In the ALL group, we found the expressions of VEGFR2, VEGFR3levels in the primary untreated group were significantly higher than the normal control group, the differences were statistically significant (P <0.05),and the expressions of VEGFR2, VEGFR3in complete remission weresignificantly decreased, in comparison with the primary untreated group,the differences were not statistically significant (P>0.05). There were nostatistical differences between the levels of VEGFA,VEGFR1in primaryuntreated group, complete remission group and normal controlgroup(P>0.05). VEGFC were not expressed in primary untreatedgroup,complete remission group and normal control group. ALL group: thelevels of VEGFA in the primary untreated group were (473.27±469.87)pg/ml, the levels of VEGFA in complete remission group were (74.96±50.62)pg/ml, the levels of VEGFA in the normal control group were (92.46±42.35)pg/ml. The levels of VEGFR1in the primary untreated group were(0.29±0.19) ng/ml, the levels of VEGFR1in complete remission group were(0.06±0.03) ng/ml, the levels of VEGFR1in the normal control groupwere(0.04±0.03) ng/ml. The levels of VEGFR2in primary untreated groupwere (1.49±0.32) ng/ml,the levels of VEGFR2in complete remission groupwere (0.04±0.03) ng/ml, the levels of VEGFR2in normal control groupwere (0.44±0.56) ng/ml. The levels of VEGFR3in primary untreated groupwere (13.68±6.99) ng/ml, the levels of VEGFR3complete remission groupwere(5.37±2.40) ng/ml, the levels of VEGFR3in the normal control groupwere(2.62±1.31) ng/ml.3. Through analysed the serum levels of HMGB1and VEGFA,VEGFR1,VEGFR2, VEGFR3in ANLL primary untreated patients, we found thelevels of HMGB1uncorrelated with VEGFA,VEGFR1,VEGFR2,VEGFR3(r=-0.400,-0.099,-0.053,0.089; P>0.05).Through analysed the serum levels of HMGB1and VEGFA, VEGFR1,VEGFR2, VEGFR3in ALL primary untreated patients, we found that levelsof HMGB1uncorrelated with VEGFA、 VEGFR1、 VEGFR2、 VEGFR3(r=-0.029,0.005,-0.67,0.179； P>0.05).Conclusions: The expressions of HMGB1were significantly increasedin the patients with acute leukemia. HMGB1levels in primary untreatedgroup and relapsed or refractory group were significantly increased inANLL and ALL patients, the expressions of HMGB1in complete remission were significantly decreased in complete remission after treatment, thelevels of HMGB1related to the degree of disease progression. Theexpressions of HMGB1were associated with AL in different periods.HMGB1may become the reference indicator of disease outcome in thetreatment of clinical diagnosis of acute leukemia; VEGF and these receptorsfrom serum in ANLL and ALL were abnormal expressed. In ANLL group,the expressions of VEGFA, VEGFR1, VEGFR3in primary untreatedpatients were significantly higher than the remission group and normalcontrol group. In the ALL group, the expressions of VEGFR2, VEGFR3inthe primary untreated patients were significantly higher than the completeremission group and normal control group. In ANLL group and ALL group,VEGFC were not expressed in primary untreated group,complete remissiongroup and normal control group. VEGF and these receptors could be used tomonitor the development of acute leukemia and provide more valuable basisfor judging the prognosis of acute leukemia. We found the levels of HMGB1uncorrelated with VEGFA, VEGFR1, VEGFR2, VEGFR3in primaryuntreated patients of ANLL and ALL. |
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