The study was to explore the telomerase activity and the expression of hTERT, c-myc and bcl-2 mRNA during terminal differentiation of HL-60 cells induced by all trans-retinoic acid (ATRA) and to study the possible molecular mechanism. By use of the model of differentiated HL-60 cells induced by ATRA, the telomerase activity was determined by TRAP-PCR-ELISA and the expression of hTERT, c-myc, bcl-2 mRNA was detected by RT-PCR in differentiated HL-60 cells. The results showed that during differentiation of HL-60 cells, the telomerase activity was decreased, the expression of hTERT, c-myc and bcl-2 mRNA was downregulated, and the downregulation of hTERT occurred prior to suppression of telomerase activity. It is concluded that the telomerase activity is related to decrease expression of hTERT, c-myc and bcl-2 mRNA during HL-60 cell differentiation induced by ATRA.
Cell Differentiation ; DNA-Binding Proteins ; Gene Expression Regulation, Neoplastic ; Genes, bcl-2 ; Genes, myc ; HL-60 Cells ; Humans ; RNA, Messenger ; analysis ; Telomerase ; genetics ; metabolism ; Tretinoin ; pharmacology

OBJECTIVETo investigate the expression changes of intrinsic cytokines TGF-beta(1) and TNF-alpha, telomerase activity and bcl-2 during ongoing apoptosis of HL-60 and K562 cells induced by p53.

METHODSpN53cG (Val135), a temperature sensitive p53 mutant, which behaved like wild type p53 (wt-p53) at 32.5 degrees C, were introduced into p53-null HL-60 and K562 cells respectively by lipofectin. In the presence of G418, HL-60-pN53cG and K562 pN53cG clones expressing p53 protein were selected. The ongoing expression of intrinsic cytokines (TGF-beta(1) and TNF-alpha), bcl-2 oncogene and hTERT mRNA during the apoptosis of HL-60 and K562 cells induced by p53 and the effects of exogenous p53 gene, TGF-beta(1) and TNF-alpha antisense PS-ODNS on the apoptosis of HL-60 and K562 cells and the expression of bcl-2 were studied by RT-PCR, quantitative RT-PCR, DNA fragmentation, TdT-mediated dUTP nick end labeling (TUNEL) and flow cytometery. The levels of secreted TGF-beta(1) and telomerase activity were detected by ELISA and PCR-ELISA, respectively.

RESULTS(1) The expressions of intrinsic TGF-beta(1) and TNF-alpha mRNA were up-regulated, while that of bcl-2 and hTERT down-regulated. The levels of TGF-beta(1) in the supernatant of HL-60 and K562 cells were increased, and the level of telomerase activity decreased. (2) Antisense PS-ODNS of TGF-beta(1) and TNF-alpha could obviously inhibit the p53 inducing cell apoptosis, and restore bcl-2 mRNA and protein to pre-treated level.

CONCLUSIONSExogenous p53 induces leukemia cell apoptosis via up-regulating the expression of intrinsic TGF-beta(1) and TNF-alpha and down-regulating the expression of hTERT and bcl-2.

Apoptosis ; drug effects ; genetics ; DNA, Antisense ; pharmacology ; DNA-Binding Proteins ; Gene Expression Regulation, Neoplastic ; HL-60 Cells ; Humans ; K562 Cells ; Leukemia ; genetics ; pathology ; Mutation ; Plasmids ; genetics ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; RNA, Messenger ; drug effects ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Telomerase ; genetics ; Transfection ; methods ; Transforming Growth Factor beta ; genetics ; Transforming Growth Factor beta1 ; Tumor Necrosis Factor-alpha ; genetics ; Tumor Suppressor Protein p53 ; genetics ; physiology

To investigate the change of telomerase activity and human telomerase reverse transcriptase (hTERT) gene expression in HL-60 cells transfected with wild type p53 gene, wild type p53 gene was introduced into HL-60 cells by Lipofectin transfection. Apoptosis was analyzed by TUNEL assay. Telomerase activity and the level of hTERT mRNA were detected by telomeric repeat amplification protocol (TRAP)-ELISA and RT-PCR, respectively. The results showed that the apoptotic rate of HL-60-pN53cG cells was 8.3% and 21.0% respectively after cultured at 32.5 degrees C for 24 h and 72 h. The level of hTERT mRNA was decreased to 68.4% and 55.8% and telomerase activity to 27.3% and 8.9% of control value in HL-60-pN53cG cells at the same points. In conclusions, hTERT mRNA and telomerase activity were down-regulated in HL-60 cells transfected with p53 gene. This may be one of mechanisms of apoptosis induced by wild type p53 gene.
Apoptosis ; DNA-Binding Proteins ; Gene Expression ; Genes, p53 ; physiology ; HL-60 Cells ; Humans ; RNA, Messenger ; analysis ; Telomerase ; genetics ; metabolism

Recent research indicates that TGF-beta and type II receptor (TbetaR-II) play an important role in the pathogenesis of tumor. A high frequency of abnormalities in TbetaR-II has been demonstrated in various cancers. To identify the mutation of TbetaR-II in patients with acute leukemia, the bone marrow samples from 6 patients with acute leukemia and 11 normal individuals as control were detected by long-range RT-PCR. To detect a deletion in sequence of the TbetaR-IIgene, the PCR products were cloned to T vector and then sequenced. The results showed that there was existance of the isoform of TbetaR-II in 2 cases out of 6 patients with acute leukemia. These two patients had more poor prognosis than others. In conclusion, there was the isoform of TbetaR-II in partial patients with acute leukemia, and the isoform may be related with prognosis.
Adult ; Base Sequence ; Female ; Humans ; Leukemia, Myeloid, Acute ; metabolism ; Male ; Middle Aged ; Molecular Sequence Data ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; metabolism ; Prognosis ; Protein Isoforms ; Protein-Serine-Threonine Kinases ; classification ; metabolism ; Receptors, Transforming Growth Factor beta ; classification ; metabolism ; Signal Transduction ; Transforming Growth Factor beta ; metabolism

OBJECTIVETo identify the expression of Drosophila Eyes Absent Homologue 2 (EYA2) in non-small cell lung cancer (NSCLC) and to investigate its correlation with clinical parameters.

METHODS59 fresh specimens of lung cancer and paired normal lung tissue were obtained from 59 NSCLC cases treated in the department of thoracic surgery in our hospital from June 2006 to October 2007. Western blotting and immunohistochemistry were used to assay the specimens with goat anti-human EYA2 polyclone antibody. Clinicopathological parameters were collected and the correlation with EYA2 expression was subsequently analyzed.

RESULTSThe expression of EYA2 was detected in cytoplasm and nucleus of the cancer cells, but mostly in cytoplasm. Western blotting and immunohistochemistry showed the expression of EYA2 in NSCLC was increased and correlated with pathological type, but not with gender, age, pTNM stage, histological differentiation and lymph node metastasis. EYA2 expression was significantly up-regulated in adenocarcinoma, while not changed in lung squamous cell carcinoma.

CONCLUSIONThe results of this study suggest that expression of EYA2 in lung adenocarcinoma is augmented. EYA2 is likely participating in the development of lung adenocarcinoma as a transcriptional activator.

Adenocarcinoma ; metabolism ; pathology ; Adult ; Aged ; Carcinoma, Non-Small-Cell Lung ; metabolism ; pathology ; Carcinoma, Squamous Cell ; metabolism ; pathology ; Cytoplasm ; metabolism ; Female ; Humans ; Intracellular Signaling Peptides and Proteins ; metabolism ; Lung ; metabolism ; Lung Neoplasms ; metabolism ; pathology ; Lymphatic Metastasis ; Male ; Middle Aged ; Neoplasm Staging ; Nuclear Proteins ; metabolism ; Protein Tyrosine Phosphatases ; metabolism ; Up-Regulation

OBJECTIVETo establish the mouse model for the expression of PD-L1 by hydrodynamic injection and to study the effects of myeloablative conditioning on hydrodynamic injection-mediated PD-L1 expression.

METHODSPlasmid amplification, hydrodynamic injection, collagenase perfusion, real time PCR, ELISA and flow cytometry were applied to test the expression and function of PD-L1. Also, animal models were set up to test the effects of chemical or radiactive myeloablative conditioning on hydrodynamic injection-mediated PD-L1 expression.

RESULTSThe expression of PD-L1 mRNA and protein could be detected as early as 8 h after hyrodynamic injection and reached peak expression by 24 h, and returned to baseline level by 7 d after injection. Serum PD-L1 level reached to 100 µg/ml as early as 24 h after injection and plateaued at 7 d after injection. Serum PD-L1 persisted for 3 weeks and declined to baseline after 1 month of hydrodynamic injection. The PD-L1 function induced by hydrodynamic injection was consistent with literature reports. At each time point, the PD-L1 expression was not different significantly between the myeloablative conditioning group and control group; the mice transfected with PD-L1 showed a higher survival rate than that in control group.

CONCLUSIONMyeloablative conditioning does not affect hydrodynamic injection-mediated PD-L1 expression, indicating that the PD-L1 can be used in HSCT mouse model.

Animals ; B7-H1 Antigen ; pharmacology ; Disease Models, Animal ; Flow Cytometry ; Hydrodynamics ; Injections ; Mice ; Myeloablative Agonists ; pharmacology ; RNA, Messenger ; Transfection ; Transplantation Conditioning

Objective: To explore the efficacy and prognostic factors of hematopoietic stem cell transplantation (HSCT) for the treatment of patients with anaplastic large cell lymphoma (ALCL) . Methods: The clinical records of 33 ALCL patients after HSCT were collected and analyzed retrospectively to evaluate the rates of overall survival (OS) and recurrence after autologous (auto-HSCT) and allogeneic HSCT (allo-HSCT) and the factors influencing prognosis. Results: The median-age of this cohort of 33 ALCL cases at diagnosis was 31 (12-57) years old with a male/female ratio of 23/10, 24 cases (72.7%) were ALK(+) and 9 ones (27.3%) ALK(-). Of them, 25 patients (19 ALK(+) and 6 ALK(-)) underwent auto-HSCT and 8 cases (5 ALK(+) and 3ALK(-)) allo-HSCT with a median follow-up of 18.7 (4.0-150.0) months. Disease states before HSCT were as follows: only 6 patients achieved CR status and received auto-HSCT, 16 patients achieved PR (14 cases by auto-HSCT and 2 ones allo-HSCT) , the rest 11 cases were refractory/relapse (5 cases by auto-HSCT and 6 ones allo-HSCT) . There were 7 cases died of disease progression (5 after auto-HSCT and 2 allo-HSCT) and 5 cases treatment-related mortality (TRM) (2 after auto-HSCT and 3 allo-HSCT) , TRM of two groups were 8.0% and 37.5%, respectively. Both the median progression-free survival (PFS) and OS were 15 months after auto-HSCT, the median PFS and OS after allo-HSCT were 3.7 (1.0-90.0) and 4.6 (1.0-90.0) months, respectively. There was no statistically significant difference in terms of survival curves between the two groups (OS and PFS, P=0.247 and P=0.317) . The 2-year OS rates in auto-HSCT and allo-HSCT groups were 72% and 50%, respectively. The 5-year OS rates in auto-HSCT and allo-HSCT groups were 36% and 25%, respectively. Conclusion: ALCL treated by chemotherapy produced high rates of overall and complete responses. Chemotherapy followed by auto-HSCT remained to be good choice for patients with poor prognostic factors. High-risk patients should be considered more beneficial from allo-HSCT.
Adolescent ; Adult ; Child ; Female ; Hematopoietic Stem Cell Transplantation ; Humans ; Lymphoma, Large-Cell, Anaplastic/therapy* ; Male ; Middle Aged ; Neoplasm Recurrence, Local ; Retrospective Studies ; Transplantation, Autologous ; Transplantation, Homologous ; Treatment Outcome ; Young Adult

Objective: To evaluate clinical outcomes of autologous (auto-HSCT) and allogeneic hematopoietic stem cell transplantation (allo-HSCT) for angioimmunoblastic T-cell lymphoma (AITL) . Methods: From June 2007 to June 2017, clinical data of AITL patients who underwent HSCT in eight hospitals were assessed retrospectively. Results: Of 19 patients, 13 male and 6 female with a median age of 50 (32-60) years old, 12 auto-HSCT and 7 allo-HSCT recipients were enrolled in this study, all donors were HLA-identical siblings. Two of allo-HSCT recipients were relapsed auto-HSCT ones. There were 5 patients (5/12) in complete response (CR) status and 7 (7/12) in partial remission (PR) status before transplantation in auto-HSCT group, and 2 (2/7) in PR status and 3 (3/7) in progression disease (PD) status before transplantation in allo-HSCT group. The median follow-up for the surviving patients was 46.5 months (range, 1-100 months) for the whole series, two patients lost in auto-HSCT group. Three patients developed acute graft-versus-host disease (aGVHD) and 5 chronic graft-versus-host disease (cGVHD) after allo-HSCT. Three patients died of primary disease and 1bleeding in auto-HSCT group. One patient died of primary disease and 2 transplantation-related mortality in allo-HSCT group. The 3-year cumulative overall survival (OS) were 56% (95%CI 32%-100%) and 57% (95%CI 30%-100%) for auto-HSCT and allo-HSCT, respectively (P=0.979) . The 3-year cumulative progression-free survival (PFS) were 34% (95%CI 14%-85%) and 57% (95%CI 30%-100%) for auto-HSCT and allo-HSCT, respectively (P=0.451) . Conclusion: Both auto-HSCT and allo-HSCT were optimal choices for AITL. In clinical practice, which HSCT was better for AITL patients should be based on comprehensive factors including sensitivity to chemotherapy, risk stratification and disease status at transplantation.
Adult ; Female ; Graft vs Host Disease ; Hematopoietic Stem Cell Transplantation ; Humans ; Lymphoma, T-Cell/therapy* ; Male ; Middle Aged ; Retrospective Studies ; Transplantation, Autologous ; Transplantation, Homologous ; Treatment Outcome

BACKGROUND: There were few studies on real-world data about autologous hematopoietic stem cell transplantation (auto-HSCT) or allogeneic HSCT (allo-HSCT) in peripheral T-cell lymphoma (PTCL). This study aimed to investigate the clinical outcomes of patients who received auto-HSCT or allo-HSCT in China. METHODS: From July 2007 to June 2017, a total of 128 patients who received auto-HSCT (n  = 72) or allo-HSCT (n  = 56) at eight medical centers across China were included in this study. We retrospectively collected their demographic and clinical data and compared the clinical outcomes between groups. RESULTS: Patients receiving allo-HSCT were more likely to be diagnosed with stage III or IV disease (95% vs. 82%, P = 0.027), bone marrow involvement (42% vs. 15%, P = 0.001), chemotherapy-resistant disease (41% vs. 8%, P = 0.001), and progression disease (32% vs. 4%, P < 0.001) at transplantation than those receiving auto-HSCT. With a median follow-up of 30 (2-143) months, 3-year overall survival (OS) and progression-free survival (PFS) in the auto-HSCT group were 70%(48/63) and 59%(42/63), respectively. Three-year OS and PFS for allo-HSCT recipients were 46%(27/54) and 44%(29/54), respectively. There was no difference in relapse rate (34%[17/63] in auto-HSCT vs. 29%[15/54] in allo-HSCT, P = 0.840). Three-year non-relapse mortality rate in auto-HSCT recipients was 6%(4/63) compared with 27%(14/54) for allo-HSCT recipients (P = 0.004). Subanalyses showed that patients with lower prognostic index scores for PTCL (PIT) who received auto-HSCT in an upfront setting had a better outcome than patients with higher PIT scores (3-year OS: 85% vs. 40%, P = 0.003). Patients with complete remission (CR) undergoing auto-HSCT had better survival (3-year OS: 88% vs. 48% in allo-HSCT, P = 0.008). For patients beyond CR, the outcome of patients who received allo-HSCT was similar to that in the atuo-HSCT group (3-year OS: 51% vs. 46%, P = 0.300). CONCLUSIONS Our study provided real-world data about auto-HSCT and allo-HSCT in China. Auto-HSCT seemed to be associated with better survival for patients in good condition (lower PIT score and/or better disease control). For patients possessing unfavorable characteristics, the survival of patients receiving allo-HSCT group was similar to that in the auto-HSCT group.
China ; Hematopoietic Stem Cell Transplantation ; Humans ; Lymphoma, T-Cell, Peripheral/therapy* ; Neoplasm Recurrence, Local ; Retrospective Studies ; Transplantation, Autologous ; Transplantation, Homologous ; Treatment Outcome