ABSTRACT
The aim of study was to investigate the effect of acute lymphoblastic leukemia (ALL) children bone marrow mesenchymal stem cells (MSC) on resistance of K562/A02 cells and its mechanism. MSC obtained from bone marrow of AL children were cultured and identified. The co-culture of MSC and K562/A02 and the culture of K562/A02 cell suspension alone was performed, of which 2 kinds of cells were treated with same concentration of adriamycin (ADM), and the rate of apoptosis was detected by flow cytometry, bcl-2 and bax of K562/A02 were detected by RT-PCR, while mdr1 gene level was detected by FQ-PCR. The results indicated that the MSC separation and proliferation were viable and steady. The apoptosis rate of the K562/A02 cells co-cultured with MSC was 1.97 ± 0.11%, while apoptosis rate of the K562/A02 cells cultured alone was 8.38 ± 0.29%, there was significant difference (p < 0.05). As compared with the K562/A02 cells cultured alone, the bcl-2 gene expression in K562/A02 cells co-cultured with MSC obviously increased; ratio of bcl-2/bax was obviously enhanced. The mdr1 gene level in K562/A02 co-cultured with MSC was no statistical different from K562/A02 cultured alone (p > 0.05), which suggested that adhesion co-cultured with MSC did not induce mdr1 expression higher than the culture of suspension. It is concluded that the MSC of ALL children can escape the leukemia cells from proapoptotic effect of drugs, the resistance of K562/A02 to ADM may be involved in enhancement of bcl-2 gene expression of K562/A02 cells co-cultured with MSC, but not in relation to mdr1 gene in K562/A02 cells themselves.
Subject(s)
Child , Child, Preschool , Female , Humans , Male , ATP Binding Cassette Transporter, Subfamily B , ATP Binding Cassette Transporter, Subfamily B, Member 1 , Genetics , Bone Marrow Cells , Doxorubicin , Pharmacology , Drug Resistance, Multiple , Genetics , Drug Resistance, Neoplasm , Genetics , Gene Expression Regulation, Leukemic , K562 Cells , Mesenchymal Stem Cells , Precursor Cell Lymphoblastic Leukemia-Lymphoma , Genetics , Proto-Oncogene Proteins c-bcl-2 , Genetics , bcl-2-Associated X Protein , GeneticsABSTRACT
<p><b>OBJECTIVE</b>To investigate the safety and therapeutic effect of low dose (1000 U/m(2)) L-asparaginase (L-Asp) in the treatment of children with acute lymphoblastic leukemia (ALL).</p><p><b>METHODS</b>Six patients were treated with low dose L-Asp after previously suffered severe side effects from standard dose L-Asp (5000 - 10,000 U/m(2)). Twenty-eight blood samples were obtained randomly from 5 of them. Plasma asparagine concentration was detected by reverse phase-high performance liquid chromatography (RP-HPLC).</p><p><b>RESULTS</b>All the patients treated with low dose L-Asp showed no any toxic symptoms. The plasma asparagine levels in the patients were all above 5 micromol/L except case 4 (4.91 micromol/L) before receiving L-Asp, and were all decreased below 0.5 micromol/L five days after receiving low dose L-Asp, except case 3 (3.70 micromol/L), the results being like that of receiving standard dose L-Asp.</p><p><b>CONCLUSION</b>Low dose L-Asp has definite efficacy for childhood ALL, while avoids serious side effects from standard dose L-Asp.</p>
Subject(s)
Adolescent , Child , Child, Preschool , Female , Humans , Male , Antineoplastic Agents , Blood , Asparaginase , Blood , Precursor Cell Lymphoblastic Leukemia-Lymphoma , Drug Therapy , Treatment OutcomeABSTRACT
Primer Express 2.0 software was used to design the primers and the MGB probe. With the plasmid pHaMDR1/A containing mdr1 cDNA as the template, we established a real-time fluorescent quantitative polymerase chain reaction system, which, at the template concentration of 3.061 x 10(3) to 3.061 x 10(9) cps/ml, had a correlation coefficient of 0.988243 between template concentration and threshold cycle value. This PCR method allows sensitive, specific and quantitative detection of human mdr1 gene.
Subject(s)
Female , Humans , Male , ATP Binding Cassette Transporter, Subfamily B, Member 1 , Genetics , DNA Primers , Fluorescent Dyes , Fluorometry , Methods , Genes, MDR , Genetics , Polymerase Chain Reaction , Methods , Taq PolymeraseABSTRACT
<p><b>OBJECTIVE</b>To investigate the changes in the activity of Escherichia coli asparaginase (L-asp) and the concentration of asparagines (ASN) in the plasma of the acute lymphoblastic leukemia (ALL) children receiving L-asp containing chemotherapeutic protocol to explore more reasonable usage of L-asp in the treatment of childhood ALL.</p><p><b>METHODS</b>L-asp containing hemotherapy regimen of VDLP was used, in which L-asp (10,000 U/m(2)) was administered intravenously every other day for 10 doses in 15 children with ALL. A total of 340 peripheral blood samples were collected at scheduled time points during the therapy and plasma L-asp activity (by spectrophotometric assay) and asparagines concentration (by RP-HPLC) were measured.</p><p><b>RESULTS</b>During the administration of L-asp, the plasma L-asp activity was increasing gradually peaked after eight doses and then decreased gradually, while the plasma concentration of asparagines maintained in complete or nearly complete depletion status. After the therapy courses finished, a plasma L-asp activity above 100 U/L with asparagines almost complete depletion status was lasting for about seven days.</p><p><b>CONCLUSION</b>The current L-asp containing chemotherapeutic protocols in which L-asp was administered in a dose of 10 000/m(2) intravenously every other day, are efficient enough for the depletion of plasma ASN.</p>