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This study was purposed to investigate the effect of aminopeptidase N/CD13 on bestatin enhancing all-trans-retinoic acid (ATRA)-inducing differentiation in NB4 cells. The nitroblue-tetrazolium (NBT) reduction assay was performed to determine the differentiation of NB4 cells, MR2 cells and primary APL blasts. The expression of P38 MAPK protein and the phosphorylation of P38 MAPK protein in NB4, MR2 and K562 cells were detected by Western blot. The results showed that pre-incubation with 5 µg/ml WM-15 blocked the enhancement effect of bestatin on differentiation of NB4 cells induced by ATRA. 5 µg/ml CD13 antibody WM-15 partly blocked the inhibition of bestatin on the phosphorylation of P38 MAPK in NB4 cells. 100 µg/ml bestatin inhibited the phosphorylation of P38 MAPK in NB4 cells and MR2 cells in a time-dependent manner. 100 µg/ml bestatin had no effect on the phosphorylation of P38 MAPK in K562 cells with low level of CD13. Bestatin could not restore the sensitivity to ATRA in ATRA-resistant primary APL blasts and MR2 cells. It is concluded that aminopeptidase N/CD13 inhibitor bestatin may enhance the differentiation-inducing activity of ATRA through inhibiting the phosphorylation of P38 MAPK in NB4 cells mediated by the cell surface APN/CD13.

The aim of this study was to investigate the combined effects of bortezomib (Bor) and daunorubicin (DNR) or each drug alone on proliferation of human multiple myeloma cell line KM3. KM3 cells were cultured with different concentrations of Bor and DNR, Bor or DNR alone for different times. The cell proliferation was analyzed by MTT assay, and the concentration of 50% growth inhibition (IC(50)) was calculated. The results indicated that both of Bor and DNR inhibited KM3 cell proliferation in dose dependent manner. The IC(50) of both drugs were 0.27 micromol/L and 0.16 micromol/L respectively. The inhibiting rate of Bor plus DNR on KM3 cells was much higher than that of Bor (p < 0.05). It is concluded that the Bor has synergistic inhibitory effect with DNR on the growth of KM3 cell in vitro.

Candida arthritis in patient with hematological malignancy is rare. A case of Candida tropicalis arthritis of knee occurred in a patient with acute monocytic leukemia was reported during the recovery phase of post chemotherapy myelosuppression and agranulocytosis. The patient was diagnosed as Candida tropicalis arthritis of knee according to the Candida tropicalis isolated from the synovial fluid. Itraconazole and amphotericin B were intravenously injected for therapy for 4 - 5 weeks based on the susceptibility test in vitro, which showed better efficacy. But the arthritis relapsed at 4 - 6 weeks after the drug withdrawal. The curative effect was found in patient after treatment with fluconazole injection and articular cavity douching with amphotericin B for 8 weeks. In conclusion, although Candida arthritis in patient with hematological malignancy is rare, it still occurred in the patient with hypoimmunity. The treatment emphasis showed be placed on the full dosage and full treatment course of antifungal agent.

The study was aimed to investigate the expression level of TRF1 protein in human acute leukemia and relationship between expression level of TRF1 protein and activity of telomerase. A quantitative Western blot technique was developed using anti-TRF1(33 - 277) monoclonal antibody and GST-TRF1 fusion protein as a standard to further determine the expression level of TRF1 protein in total proteins extracted from clinical specimens. 20 cases of acute leukemias were studied when 11 normal volunteer's bone marrow was used as control. The results showed that the expression level of TRF1 protein in normal bone marrow (2.217 +/- 0.461 microg/microl) was significantly higher than that in bone marrow of acute leukemia patients (0.754 +/- 0.343 microg/microl) (P < 0.01). There was no remarkable difference of expression level of TRF1 protein between ALL and ANLL (0.628 +/- 0.281 microg/microl vs 0.844 +/- 0.360 microg/microl, P > 0.05). After chemotherapy, TRF1 expression level of patients with complete remission raised (0.772 +/- 0.307 microg/microl vs 1.683 +/- 0.344 microg/microl, P < 0.01), but lower than that of normal (2.217 +/- 0.461 microg/microl, P < 0.01). TRF1 expression level of patients without complete remission was not remarkable different after chemotherapy (0.726 +/- 0.443 microg/microl vs 0.894 +/- 0.338 microg/microl, P > 0.05). TRF1 expression level of patients with complete remission was higher than that in patients without complete remession (1.683 +/- 0.344 microg/microl vs 0.894 +/- 0.338 microg/microl, P < 0.01). For all sample the telomerase activity was determined. It was confirmed that the activity of telomerase in normal bone marrow was lower than that in bone marrow of acute leukemia patients (0.125 +/- 0.078 microg/microl vs 0.765 +/- 0.284 microg/microl, P < 0.01). There was no significantly difference of expression level of TRF1 protein between ALL and ANLL (0.897 +/- 0.290 microg/microl vs 0.677 +/- 0.268 microg/microl, P > 0.05). After chemotherapy, telomerase activity of patients with complete remission reduced (0.393 +/- 0.125 microg/microl), but higher than that of normal (0.125 +/- 0.078 microg/microl, P < 0.01). It is concluded that expression level of TRF1 protein in AL patients is significantly decrese and associated with therapeutic efficaciousness and the activity of telomerase (P < 0.001).

The objective of this study was to investigate the effect of cytomegalovirus (CMV) infection on actin and microfilament in human embryo fibroblast cells (HF) and to explore the possible relationship with CMV replication. The cell shape was observed by microscopy after the infection of CMV, RT-PCR assay was used to detect the mRNA expression of beta-actin gene, while Westen-blot was used to measure the level of beta-actin protein. CMV immediately early antigen (IE) in HF cells was analyzed by indirect immunofluorescence assay. Microfilament alteration was determined by cytoskeleton fluorescence probe. The results showed that CMV IE was observed in more than 95% of HF cells after infection, primarily located in nucleus. HF cells infected by CMV changed from thin shuttle shape to round and thick ball shape, even detached from wall. Beta-actin got a significant and gradual decreasing of mRNA level in time-dependent manner (P < 0.05). Compared with uninfected group, the expression of beta-actin protein decreased to (74.2 +/- 13.4)% at 96 hours after infection (P < 0.05). In infected HF cells, microfilaments were ruptured, arranged turbulently, as well as cells merged and fluorescence density of microfilament obviously reduced. It is concluded that cytomegalovirus can induce alteration of actin and microfilament, which may be helpful for CMV to infect, replicate and reactivate in host cells.

This study was purposed to investigate whether aminopeptidase inhibitor, bestatin, can potentiate all-trans retinoic acid (ATRA)-inducing differentiation in NB4 cells, and to explore its mechanism. The NB4 cells were exposed to either bestatin and ATRA alone or in combination, the morphological changes of NB4 cells were observed by optical microscopy, the CD11b expression was measured by flow cytometry, the function of defferentiation cells was analyzed by nitroblue-tetrazolium (NBT) reduction assay, the mRNA expressions of c-myc and c-EBPepsilon in NB4 cells were detected by RT-PCR, the c-Myc protein expression was determined by Western blot. The results showed that treatment with bestatin alone induced no significant changes in morphology, NBT reduction activity and CD11b expression in NB4 cells. NB4 cells incubated with 10 nmol/L ATRA plus 100 microg/ml bestatin showed more morphologic feature of metamyelocyte and band neutrophil than ATRA alone treated cells. 100 microg/ml bestatin enhanced the NBT reduction activity in NB4 cells induced by various concentrations of ATRA (10, 20, 40 nmol/L). The effects of various concentrations of ATRA in combination with 100 microg/ml bestatin were statistically different from the effect of ATRA alone (P < 0.01). From 48 to 96 hours, 100 microg/ml bestatin time-dependently increased NBT reduction in NB4 cells induced by 10 nmol/L ATRA (P < 0.01). 10 nmol/L ATRA plus 100 microg/ml bestatin for 72 hours prominently elevated CD11b expression in NB4 cells as compared with ATRA alone treated NB4 cells (P < 0.01). There was a substantial decrease in c-myc mRNA levels when 100 microg/ml bestatin was added to 10 nmol/L ATRA (P < 0.05). Various concentrations (50, 75, 100 microg/ml) of bestatin combined with 10 nmol/L ATRA down-regulated the expression of c-Myc protein, which was negatively correlated with the NBT reduction activity of NB4 cells induced by 10 nmol/L ATRA alone or plus bestatin at various concentrations (r = -0.940, P = 0.017). However, 100 microg/ml bestatin plus 10 nmol/L ATRA could not induce any significant changes in the levels of c-EBPepsilon mRNA as compared with ATRA alone treated NB4 cells. It is concluded that an aminopeptidase inhibitor bestatin can potentiate ATRA-inducing differentiation of NB4 cells, possibly by down-regulating c-myc expression in synergy with ATRA.

The study was aimed to detect expression rate of survivin gene in APL cell and to explore the relationship between its expression and clinical manifestation. PML/RARalpha and survivin mRNA expression were analyzed by using reverse transcriptase polymerase chain reaction (RT-PCR) technique. The results showed: (1) the survivin gene expression was detected in NB4 cell line. By treatment with ATRA, survivin mRNA expression in NB4 cell gradually decreased along with time delay and almost could not be detected at the 72th hour. (2) the positive and negative rate of survivin mRNA expression was 67% and 33% respectively, while in all 36 cases of de novo and relapse APL patients, the PML/RAR(alpha) fusion gene expression was positive. In 22 cases at remission stage, the PML/RARalpha fusion gene expression was negative, and the positive and negative rate of survivin mRNA expression was 36% and 64% respectively. The survivin mRNA expression positive rates in the de novo group, relapse group and PML/RARalpha fusion gene L-type positive group were obviously higher than those in remission period group (P < 0.05) and were significantly lower than those in acute leukemia group (P < 0.05, < 0.001). (3) whether the survivin mRNA expression was positive or negative in 36 cases of de novo and relapse APL patients, all the 36 cases could obtain complete remission. 4 APL patients with positive expression of survivin mRNA had DIC and serious infection (one patient died). The clinical symptom showed slight skin or mucosa bleeding, fever and asthenic in the patients with negative expression of survivin mRNA. When 2 APL patients with positive expression of survivin mRNA had been treated with ATRA, induction differentiation sign in their peripheral blood and bone marrow figures was not obvious. It is concluded that the survivin gene positive expression rate is lower in acute promyelocytic leukemia than that in any other types of leukemia and is related to clinical manifestation.

To explore the regulation mechanism of survivin gene, the NB4 and HL-60 cells were used in experiments, the cell culture in vitro and cell morphological observation were performed and survivin mRNA expression was detected by semi-quantitative RT-PCR. The results showed that the survivin expression in NB4 cell was positive. By treatment of 1 micromol/L ATRA, cell differentiation antigen CD11b was gradually increased ([chi = 47.002, P = 0.000) and CD33 was gradually decreased (chi = 1.614, P = 0.806) with time. Simultaneously, survivin mRNA expression was down-regulated and the cell cycle was arrested at G(0)-G(1) phase (chi = 58.566, P = 0.000). ATRA could down-regulate the survivin mRNA expression of HL-60 cell, but G-CSF, GM-CSF and PHA could up-regulate the survivin expression of HL-60 cell. The cytokine could regulate survivin expression in gene transcription level. The up-regulation of survivin expression was observed while HL-60 cell was stimulated by PHA. The survivin gene expression could be blocked by the survivin antisense oligonucleotide. The survivin mRNA expression of NB4 cell was inhibited by 100 nmol/L-1000 nmol/L survivin antisense oligonucleotide in a dose-dependent manner. The survivin mRNA expression in the NB4 cell was obviously inhibited in 600 nmol/L survivin AS-ODN groups (38%) while the AS-ODN dose increases, the inhibition rate does not descend, but was not inhibited in the control groups, liposomes groups and ODN groups. After NB4 cell was treated by survivin AS-ODN, the typical morphological changes for the apoptosis emerged in NB4 cell. These changes were not found in control groups. It is concluded that PHA, GM-CSF and G-CSF can up-regulate the survivin gene expression, but survivin AS-ODN and ATRA can down-regulate survivin gene expression. The cell cycle arrest at G(0)-G(1) phase while the survivin gene expression was down-regulated by ATRA. It suggested that the survivin gene expression is very related to cell cycle. The morphological changes of cell apoptosis can be observed when the survivin gene expression of NB4 cell was suppressed.

The purpose of this study was to evaluate the effecacy and safety of CHOEP mobilization regimen, and the effect and tolerance of sequential chemotherapy combined with tandem autotransplants of peripheral blood stem cells for aggressive lymphoma. The clinical data of 5 patients with recurrent, aggressive lymphoma treated with of sequential chemotherapy combined with tandem autotransplants were analyzed retrospectively. The patients included 1 HD and 4 NHL. Mobilization regimen was CHOEP combined with G-CSF 5 microg/(kg x d). The conditioning regimen for the tandem transplantation was high-dose CHOEP. The interval of the tandem autotransplantation was 9 (5 - 31) weeks. In tandem autotransplant, the cell number of MNC transfused was 3.05 (1.91 - 4.14) x 10(8)/kg and 3.55 (2.23 - 6.0) x 10(8)/kg; CD34(+) cells were 4.11 (2.59 - 4.94) x 10(6)/kg and 5.70 (2.77 - 10.6) x 10(6)/kg; CFU-GM was 2.96 (2.01 - 4.54) x 10(5)/kg and 2.44 (1.78 - 2.9) x 10(5)/kg respectively (P > 0.05). The results showed that all patients gained prompt and sustained hemotopoietic reconstitution. The interval of ANC >or= 0.5 x 10(9)/L was 10 (8 - 12) days and 10.5 (9 - 12) days; Pt >or= 2.0 x 10(9)/L was 11 (10 - 14) days and 12.5 (10 - 15) days respectively (P > 0.05). Four patients survived, three patients among them were alive in disease-free for median of 46 (9 - 88) months. The overall survival was 80%, and the disease-free survival was 60%. In conclusion, the method of sequential high-dose CHOEP chemotherapy combined with autotransplants of peripheral blood stem cells in tandem for aggressive lymphoma is probably safe and effective.

To investigate the mechanisms of the telomerase regulations during the apoptosis of the human MDS-RAEB cell line MUTZ-1 cells induced by arsenic trioxide (As(2)O(3)), telomerase activity was detected by TRAP-ELISA and the expressions of mRNAs of hTERT, TRF1 (TTAGGG repeat binding factor 1), TRF2 (TTAGGG repeat binding factor 2), bcl-2, and bax genes were detected by RT-PCR. Apoptosis was detected by translocation of phosphatidylserine (PS) by flow cytometry. The results showed that 1 - 8 micromol/L of As(2)O(3) induced typical apoptosis of MUIZ-1 cells in the dose-and time-dependent manners, the telomerase activity could be down-regulated at this concentration and negatively correlated with increased apoptosis (r = -0.938, P = 0.018). The expression of telomerase activity was positively related to the expression of hTERT (r = 0.783, P = 0.022), but As(2)O(3) had no effect on the mRNA expression of TRF1 and TRF2 genes. The inhibition of telomerase activity by As(2)O(3) on MUTZ-1 cells was accompanied with the low expression of bcl-2 gene and the decrease of bcl-2/bax ratio. It is concluded that the apoptosis of MUTZ-1cells induced by As(2)O(3) may occur via the inhibition of telomerase activity and down-regulation of the expression of hTERT mRNA, and this may be one of the mechanisms inducing apoptosis in MUTZ-1 cells treated by As(2)O(3).

To elucidate the antileukemi effects of berbamine and the possible molecular mechanisms in vitro and in vivo, MTT method was used to examine the effect of berbamine on K562 cell growth. The apoptosis rate was measured by flow cytometry. The mRNA expression level of BCR/ABL gene (semiquantity value) was determined by RT-PCR and the BCR/ABL protein (P210) level was detected by Western blot. The K562-bearing mice were used to reveal the therapeutic effect in vivo. The results showed that a significant time- and concentration-dependent inhibition of cell growth was found in the cells treated with berbamine. After the cells were exposed to 8.0 microg/ml berbamine for 24, 48 and 72 hours, the percentage of growth inhibition of K562 cells progressively increased by (26.63 +/- 3.57)%, (61.84 +/- 4.74)%, (75.32 +/- 1.95)%, respectively (compared with control, P < 0.01). The IC(50) (72 hours) value was 5.227 +/- 1.307 microg/ml. The apoptosis rate of K562 cells treated with 8.0 microg/ml berbamine for 24 and 72 hours increased from (29.20 +/- 3.82)% to (61.77 +/- 4.35)% (P < 0.01). Berbamine down-regulated the expression levels of bcr/abl gene and P210 in K562 cells in a time- and concentration-dependent manner. The bcr/abl expression decreased from (1.38 +/- 0.02) to (0.97 +/- 0.01) after exposure of the cells to 8.0 microg/ml berbamine for 0 and 72 hours (P < 0.01). When the cells were treated with 4.0 - 16.0 microg/ml berbamine for 24 hours, the level of P210 decreased from (0.95 +/- 0.03) to (0.63 +/- 0.01) (P < 0.01). In vivo, after treatment for 4 weeks, the tumor weight of berbamine-treated group was also lower than that of untreated group [(1.46 +/- 0.43) g vs (2.90 +/- 0.94) g, P < 0.01] and the inhibition rate was 49.66%, moreover, berbamine down-regulated the expression level of bcr/abl gene of tumor cells. It is concluded that berbamine can obviously inhibit the cell proliferation and induce apoptosis in K562 cell lines in a time- and concentration-dependent manner in vitro. The mechanisms of berbamine-induced apoptosis may be involved in down-regulation of bcr/abl gene expression and P210 level. In vivo, berbamine can aslo display a better antileukemic effect and down-regulate expression of bcr/abl gene. Berbamine extracted from Chinese herb may be a promising candidate of new drug for clinical anticancer treatment, especially for bcr-abl(+) diseases.

To investigate the influence and mechanisms of CD47 engagement by its soluble mAb B6H12 on the maturation and function of cultured dendritic cells (DCs), monocyte-derived DCs were propagated in granulocyte-macrophage colony stimulating factor (GM-CSF) combined with lipopolysaccharide (LPS) and interleukin (IL)-4, in the presence or absence of soluble anti-CD47 monoclonal antibodies (anti-CD47 mAbs, B6H12). The characteristic morphology of DCs was identified by using the transmission electron microscopy. Flow cytometry was used to detect the cell surface phenotypes. The concentration of IL-12 P70 in supernatant was measured by ELISA. The antigen-presenting functions of DCs were determined in one-way mixed leukocyte reaction by BrdU-ELISA. Electrophoretic mobility shift assay (EMSA) was applied to examine the activity of NF-kappaB. The results indicated that the anti-CD47 mAbs markedly suppressed the expression of CD80, CD86, CD83, CD1a and HLA-DR on the surface of DCs (P < 0.05). The data of the mixed leukocyte reaction and IL-12 P70 production were consistent with the results by flow cytometry (P < 0.01). Pre-exposure to B6H12 mAb during the development of DCs resulted in a dramatic depletion of the DNA binding activity of NF-kappaB toward nucleus protein. Moreover, such an inhibition effect seemed to be dose dependent. In conclusion, the soluble mAb B6H12 inhibits the expression of the costimulatory molecules and MHCII molecules on the DCs. The antigen-presenting function of DCs was also impaired by B6H12. And these modulations are closely related with the depletive DNA binding activity of NF-kappaB. It is suggested that the soluble B6H12 exerts a negative effect on the maturation and function of in vitro cultured DCs due to inhibition of NF-kappaB binding activity.

To explore the correlation between the cellular inhibitors of apoptosis proteins (cIAPs) and the apoptosis of myelodysplastic syndrome (MDS) cell line (RAEB type) cells induced by aclacinomycin (ACM), the apoptosis of MDS cell line MUTZ-1 cells induced by ACM was analyzed with terminal deoxyribonucleotidy transferase mediated dUTP-biotin nick end labeling (TUNEL) technique. By using semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR), the expression levels of cIAP-1 and cIAP-2 mRNA in MUTZ-1 cells were assayed. The results showed as follow: (1) Using 0.5 micromol/L, 1.0 micromol/L ACM treated cells for 24 hours, the relative expression level of cIAP-1 mRNA (cIAP-1/GAPDH) was lower than those in the untreated cell group (P=0.002, 0.0002, respectively). (2) Using 0.5 micromol/L ACM treated for 6, 12, 24 hours, the relative expression level of cIAP-1 mRNA was 0.95 +/- 0.04, 0.73 +/- 0.05, 0.38 +/- 0.07, respectively and the relative expression level of cIAP-1 mRNA was correlated negatively with the time treated by ACM (r=-0.996, P <0.01). (3) Using 0.5 micromol/L ACM treated for 3, 6, 12, 24 hours, the relative expression level of cIAP-2 mRNA was 1.17 +/- 0.06, 0.91 +/- 0.03, 0.69 +/- 0.07 and 0.00 +/- 0.00, respectively and relative expression level of CIAP-2 mRNA was correlated negatively with the time treated by ACM (r=-0.091, P <0.01). (4) The percentages of TUNEL positive cells were correlated negatively with the relative expression level of CIAP1 and CIAP2 mRNA (r=-0.984, -0.959 and P=0.002, 0.013 respectively) when treated with increasing concentration of ACM. In conclusion, ACM can induce significantly MUTZ-1 cell apoptosis via suppressing expression of anti-apoptotic gene cIAP-1 and cIAP-2 mRNA.

To investigate the effect of 5-aza-2'-deoxycytidine (5-Aza-CdR) on cell of high-risk patients with myelodysplastic syndrome (MDS) in vitro, the growth inhibition of MUTZ-1 cell induced by 5-Aza-CdR was detected by MTT method; apoptosis was detected by morphological observation and translocation of phosphatidylserine (PS) was examined by flow cytometry assay; the expressions of P15INK4B, DNA methyltransferases (DNMT)(1), DNMT(3A) and DNMT(3B) gene on mRNA level were detected by RT-PCR; methylation of p15INK4B gene in MUTZ-1 cells was detected by PCR using methylation specific primer (MSP). The results showed that 5-Aza-CdR inhibited the growth of MUTZ-1 cells. The IC(50) values of 24, 48 and 72 hours were 6.75, 2.82 and 5.45 mmol/L respectively. Characteristic changes of apoptosis emerged in MUTZ-1 cells after being exposed to 5-Aza-CdR in the different concentration from 0.8 mmol/L to 3.2 mmol/L, and the positive cells of annexin V on the membrane of MUTZ-1 cells were analyzed by flow cytometry. 5-Aza-CdR could activate the p15INK4B gene expression in MUTZ-1 cells by demethylation of the p15INK4B gene in a dose-dependent manner after the cells were treated for 48 hours. Furthermore, 5-Aza-CdR could significantly down-regulate the expressions of DNA methyltransferase genes DNMT(3A) at mRNA level in a dose dependent manner. However, it had no effects on DNMT(1) gene and DNMT(3B) gene. It is concluded that 5-Aza-CdR can inhibit the growth of MUTZ-1 cells and induce the apoptosis of these cells within the range of concentration from 0.8 mmol/L to 3.2 mmol/L, which may be one of the mechanisms of antitumor effects of 5-Aza-CdR. The drug can activate the expression of p15INK4B gene in MUTZ-1 cells by demethylation of the p15INK4B gene through inhibiting the expression of DNMT(3A) gene. It may be the mechanism of 5-Aza-CdR in the treatments of MDS.

In order to explore a new way to study allogeneic reactive T lymphocytes, detection of cytokine-secreting T lymphocytes after allogeneic peripheral blood mononuclear cells (PBMNCs) stimulation and investigation of its clinical significance were performed. A novel cytokine secretion assay (CKSA) was first applied to detect T lymphocytes secreting cytokine including IFN-gamma, IL-4 and IL-10 at single cell level in human mixed lymphocytes reaction. IFN-gamma-secreting T cells from PBMNCs were then evaluated in 2 patients with acute graft versus host disease (aGVHD) after allogeneic bone marrow transplantation. The results showed that compared with IL-4 and IL-10 (which were 0.12 +/- 0.03% and 0.10 +/- 0.03% respectively), a sizable proportion of IFN-gamma-secreting T lymphocytes could be detected (1.12 +/- 0.13)% after allogeneic PBMNCs stimulation. Preliminary results indicated that frequency of IFN-gamma-secreting T lymphocytes correlated with the onset and severity of clinical aGVHD. In conclusion, it is feasible to detect IFN-gamma secreting T lymphocytes after allogeneic PBMNCs stimulation and to apply the CKSA technique for clinical identification of aGVHD.

To explore the hematopoietic reconstitution and transplantation-related complications of two units of unrelated umbilical cord blood combined transplantation for the treatment of adult hematologic malignancies, one adult patient with chronic myelogenous leukemia received two units of unrelated umbilical cord blood combined transplantation. The conditioning regimen was busulfan and cyclophosphamide (Bu-Cy). GVHD prophylaxis regimen consisted of mycophenolate mofetil (MMF), cyclosporine A (CsA) and methotrexate (MTX). The patient received total nucleated cells 4.63 x 10(7)/kg with CD34+ cells 8.34 x 10(5)/kg. Engraftment was documented by the analysis of short tandem repeat with polymerase chain reaction (STR-PCR). The results showed that the STR-PCR analysis for peripheral blood at day 31, 46 and 71 after transplantation suggested that one of two units of cord blood were completely engrafted. The ANC > 0.5 x 10(9)/L in the patient occurred at day 23, blood platelet counts > 20 x 10(9)/L at day 33 and > 50 x 10(9)/L at day 47. The Philadelphia chromosome and bcr/abl fusion gene of the patient also turned to negative after engraftment. Acute GVHD grade II occurred at day 13 and cured after treatment. It is concluded that umbilical cord blood can be used in adult hematopoietic stem cell transplantation. Two or more units umbilical cord blood combined transplantation might be the way to solve the problem of the low counts of nucleated cells when be used for adult.

To investigate the mechanism and its significance of the depression of telomerase activity in HL-60 cells exposed to homoharringtonine (HHT), the semi-quantitative RT-PCR and PCR-ELISA were used to detect the expression of hTERT mRNA and telomerase activity in HL-60 cells, the apoptotic rate of HL-60 cells was assayed with TUNEL. The results indicated that compared with the control, the level of hTERT mRNA synthesis in HL-60 cells treated with HHT at 0.005 - 0.03 micro g/ml for 12 hours did not change significantly, but it reduced obviously at 0.04 micro g/ml and was undetectable at 0.05 micro g/ml. When HL-60 cells incubated with HHT at 0.02 micro g/ml for various hours, the expression of hTERT mRNA did not decrease after 6 - 18 hours and reduced significantly after 24 hours and was undetectable at 30 hours. The tendency for suppression of telomerase activity was consistent with decrease of the expression of hTERT mRNA in HHT-treated HL-60 cells. The apoptotic rate of HL-60 cells was apparently increased with the depression of hTERT mRNA and telomerase activity. In conclusion, the transcription of hTERT mRNA in HL-60 cells can be inhibited by HHT. The relationship between regulation of hTERT mRNA expression and HHT-induced apoptosis is worth investigating furtherly.

To investigate the expression of hTERT mRNA in bone marrow mononuclear cells (MNCs) from acute leukemia patients, the method of semi-quntitative RT-PCR was used to examine the expression of hTERT mRNA in marrow MNCs, and the telomerase activity of marrow MNCs was determined with the method of TRAP-PCR-ELISA by using a commercial kit. The results indicated that the expression of hTERT mRNA of marrow MNCs in 30 untreated AL patients was markedly higher than that in 12 CR cases (0.71 +/- 0.34 vs 0.43 +/- 0.25, P < 0.05) and 6 normal volunteers (0.71 +/- 0.34 vs 0.22 +/- 0.21, P < 0.01), respectively. Telomerase activity of marrow MNCs in 30 untreated AL patients was significantly higher than that in 12 CR cases (0.235 +/- 0.395 vs 0.012 +/- 0.015, P = 0.007). Moreover, there was a positive correlation between the hTERT mRNA synthesis and telomerase activity in AL cells (r = 0.421, P < 0.01). The pencentage of blast cells in marrow smear of the untreated AL patients was positively correlated with both the expression of hTERT mRNA and the telomerase activity of bone marrow MNCs (r = 0.457, P < 0.05 and r = 0.411, P < 0.05), respectively. It is concluded that the expression of hTERT mRNA in bone marrow MNCs from untreated AL patients was correlated with their telomerase activity. It is suggested that the expression of hTERT mRNA leukemic cells indicates their higher proliferation ability.