Haibo Cai

East China University of Science and Technology, Shanghai, Shanghai Shi, China

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Publications (22)46.74 Total impact

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    ABSTRACT: Ex-vivo expansion technologies were developed for application of hematopoietic stem cells (HSCs) derived from cord blood (CB). The cytokine combination was essential to expand HSCs ex vivo and maintain the function of expanded HSCs. However the optimal cytokine combination was not determined. In this study, two combinations of cytokines were applied in ex-vivo expansion of HSCs to investigate the effect on the hematopoietic repopulating ability of expanded HSCs. CB CD34(+) cells were expanded with SCF + TPO + FL (STF) or SCF + TPO + FL + IL-3 + IL-6 (STF36) for 7 days and got 30.3 ± 6.4 and 39.8 ± 7.3 folds of total cells, respectively. The cells cultured by both STF and STF36 could engraft and repopulate in non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice effectively; however, the STF group achieved higher level of engraftment. These result demonstrated that the cytokine combination of STF36 favored the expansion of cells, while the cytokine combination of STF facilitated the engraftment and multi-lineage repopulating in vivo. These findings may have important implications for the cell therapy.
    Artificial cells, nanomedicine, and biotechnology (Print). 03/2014;
  • J Ge, H Cai, Q Li, Z Du, W S Tan
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    ABSTRACT: OBJECTIVE: Ex vivo expansion of CD34(+) cells has become critically important in order to obtain sufficient haematopoietic stem cells for clinical application. Among major regulators involved in ex vivo expansion, telomerase activity and apoptosis have been revealed to be closely linked to cell cycle progression. However, all exact roles remain to be elucidated. Here, change in telomerase activity and level of apoptosis in cord blood (CB) CD34(+) cells were evaluated together with specific cell population growth rate during ex vivo culture. MATERIALS AND METHODS: CD34(+) cells isolated from human CB were expanded ex vivo over a 28-day period. Besides monitoring cell proliferation kinetics of the CD34(+) cells, changes in telomerase activity and apoptotic levels were investigated. Several relevant genes were quantified by qRT-PCR during the culture period. RESULTS: Significant elevation of telomerase activity had close relationship to activation of CB CD34(+) cell expansion. Peak apoptotic level was accompanied by a remarkable decline in cell-specific growth rate, and apoptotic level of differentiated CD34(-) population was significantly higher than that of the CD34(+) population. CONCLUSION: Although telomerase activity was activated during the culture, expansion of CB CD34(+) cells seemed to be more susceptible to apoptotic suppression when cultured ex vivo, which implied that apoptosis may serve as a rate-limiting factor involved in controlling expansion efficiency.
    Cell Proliferation 12/2012; · 2.27 Impact Factor
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    ABSTRACT: To investigate the relationship between the expressions of B-cell-specific monoclonal leukemia virus insert site 1 (Bmi1) and human telomerase reverse transcriptase (hTERT) genes and the proliferative capacity of stem cells. Cord blood CD34+ cells were cultured in IMDM medium containing fetal bovine serum, stem cell growth factor, thrombopoietin, and Fms-like tyrosine kinase 3 ligand during a 28-day ex vivo expansion process, while the expansion fold, specific growth rate, and the colony-forming efficiency were monitored. Then the Bmi1 and hTERT mRNA expressions in CD34+ cells were detected by fluorescence quantitative PCR, and the relations between the expressions and the cell proliferative capacity were analyzed. CD34+ cells expanded (20.1 +/- 3.5) folds during the 28-day culture, while the proportion declined from 95.5% +/- 2.6% before culture to 2.1% +/- 0.4%. Both the specific growth rate and colony-forming efficiency of CD34+ cells declined significantly after 7 days, while the expression levels of Bmi1 and hTERT mRNA in CD34+ cells reached top at 7 days and declined gradually thereafter. The expressions of Bmi1 and hTERT genes may have a close relation to the proliferative capacity of CD34+ cells.
    Zhongguo xiu fu chong jian wai ke za zhi = Zhongguo xiufu chongjian waike zazhi = Chinese journal of reparative and reconstructive surgery 09/2012; 26(9):1112-6.
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    ABSTRACT: Stem cell factor (SCF) plays important roles in ex vivo expansion of hematopoietic stem cells (HSCs). In this study, the effects of dose and feeding time of SCF on ex vivo expansion of CD34(+) cells were investigated in serum-free medium supplemented with a cytokine cocktail composed of SCF, thrombopoietin (TPO) and flt3-ligand (FL). Among the four tested doses (0, 5, 50 and 500ng/mL), a SCF dose of 50ng/mL was demonstrated to be most favorable for ex vivo expansion of CD34(+) cells, which resulted in 34.22±10.80 and 8.89±1.25 folds of expansion regarding total cells and CD34(+) cells, respectively. Meanwhile, the specific growth rate of cells, the consumption rate of SCF and the percentage of CD34(+)c-kit(+) cells during the 21-day culture process were analyzed. The results indicated that initial 4-day period was a critical stage for SCF functioning on CD34(+) cells during ex vivo expansion. Based on this, a modified SCF feeding regimen was proposed, in which SCF (50ng/mL) was only supplemented on day 0 in the cytokine cocktail and cells were then fed with TPO and FL till the end of culture. It was found that this SCF feeding regimen could expand CD34(+) cells efficiently, thus providing a cost-effect expansion protocol for HSCs.
    Journal of Biotechnology 08/2012; 164(2):211-219. · 3.18 Impact Factor
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    ABSTRACT: The mononuclear cells were cultivated in stirred flasks at different agitation speeds of 30 rpm, 45 rpm, 60 rpm and 80 rpm. At the agitation speed of 30 rpm, total cells achieved higher expansion folds and the CFC density increased. When at higher agitation speed of 60 rpm or 80 rpm, the number of cells dropped rapidly and characteristics of hematopoietic stem/progenitor cells (HSPCs) were not maintained. Moreover, the culture duration of 6-9 days was better for HSPCs ex vivo expansion. These data indicated that HSPCs should be cultured at relatively low agitation speed and for a short-term period when cultured in stirred suspension system.
    Artificial Cells Blood Substitutes and Biotechnology (formerly known as Artificial Cells Blood Substitutes and Immobilization Bi 08/2012; · 0.94 Impact Factor
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    ABSTRACT: Oxygen tension regulates the biological characteristics of hematopoietic stem and progenitor cells (HSPCs) by modulating intracellular reactive oxygen species (ROS). To better understand oxygen tension mechanism on HSPCs culture, gene expression analysis of human CD34(+)CD38(-) HSPCs was performed using microarrays. The CD34(+)CD38(-) HSPCs cultured under normoxia, hypoxia, or with N-acetyl cysteine (NAC, an ROS scavenger) were isolated for transcriptional profilings. Compared to normoxia group, 1 gene was up-regulated and 22 genes were down-regulated in hypoxia group, while 1 gene was up-regulated and 29 genes were down-regulated in NAC group. These differently expressed genes were involved in cell surface markers, blood activation and differentiation. The common down-regulated genes related to dendritic cells (DCs) maturation (CD80, CD86, and JAG1) were confirmed by real-time RT-PCR. Furthermore, the analysis of the phenotypes of DCs, including the DC-characteristic surface molecule CD1a, the costimulatory molecules CD80 and CD86, and HLA-DR, associated with the capacity of DCs to stimulate allogeneic T cells, showed that hypoxia-mediating ROS inhibited the potential of CD34(+)CD38(-) HSPCs differentiating to mature DCs. All these results demonstrated that hypoxia-reducing ROS down-regulated the genes driving CD34(+)CD38(-) HSPCs differentiation, which provides an interesting molecular hint to direct their development to DCs during cultures.
    Journal of Biotechnology 01/2012; 158(3):104-11. · 3.18 Impact Factor
  • J Ge, H Cai, W S Tan
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    ABSTRACT:   Ex vivo expansion is a feasible strategy, which may overcome limitation of the very low frequency of haematopoietic stem/progenitor cells, in umbilical cord blood (UCB). However, both quality of cells and safety of expanded population are critical issues to be addressed for their clinical application. Hence, in this study, we evaluated genetic stability of UCB-derived CD34(+) cells during ex vivo culture, based on karyotype analysis, as well as its effect on cell proliferation characteristics.   CD34(+) cells were isolated from human UCB samples by immunomagnetic separation and were expanded ex vivo over a 28-day period. Expansion of total nucleate cells, CD34(+) cells and CD34(+) CD38(-) cells was investigated. Karyotype analysis of the expanded cells from six randomly selected UCB samples was performed to evaluate their genetic stability.   Chromosomal abnormality of expanded cells mainly appeared by day 14, but was seldom sustained until day 28. None of the chromosomal abnormal samples displayed neoplastic proliferation, and expanded cells with altered chromosomes did not show obvious transformation phenomena according to soft agar assay.   Ex vivo expansion could lead to occurrence of chromosomal abnormality, although here it did not produce excessive proliferative advantage of the expended cells. Importantly, chromosomal alteration seemed not to be inheritable and unlikely to result in malignant transformation. However, further in-depth evaluation of potential clinical risks of chromosomal abnormality is warranted.
    Cell Proliferation 12/2011; 44(6):550-7. · 2.27 Impact Factor
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    ABSTRACT: Heart failure remains the leading cause of morbidity and mortality. Recently, it was reported that the adult heart has intrinsic regenerative capabilities, prompting a great wave of research into applying cell-based therapies, especially with skeletal myoblasts and bone marrow-derived cells, to regenerate heart tissues. While the mechanism of action for the observed beneficial effects of bone marrow-derived cells remains unclear, new cell candidates are emerging, including embryonic stem (ES) and introduced pluripotent stem (iPS) cells, as well as cardiac stem cells (CSCs) from adult hearts. However, the very low engraftment efficiency and survival of implanted cells prevent cell therapy from turning into a clinical reality. Injectable hydrogel biomaterials based on hydrophilic, biocompatible polymers and peptides have great potential for addressing many of these issues by serving as cell/drug delivery vehicles and as a platform for cardiac tissue engineering. In this review, we will discuss the application of stem cells and hydrogels in myocardial regeneration.
    Advanced drug delivery reviews 02/2011; 63(8):688-97. · 11.96 Impact Factor
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    ABSTRACT: Interferon beta (IFN-beta) and TNF-related apoptosis-inducing ligand (TRAIL) are effective anticancer agents. Adeno-associated virus (AAV) is one of the current most promising gene delivery vectors. Previously, we constructed tumor-targeting AAV-hTERT-IFN-beta and AAV-hTERT-TRAIL by inserting IFN-beta or TRAIL gene into AAV controlled by hTERT promoter. The studies showed that either single IFN-beta or TRAIL gene therapy exhibited a certain extent anticancer effect. Here, we report their inhibitory effects on A549 lung cancer cell growth in vitro and in vivo by combined AAV-hTERT-IFN-beta and AAV-hTERT-TRAIL. Expression of secreted IFN-beta in lung cancer A549 cells infected by AAV-hTERT-IFN-beta was detected by enzyme-linked immunosorbent assay (ELISA). The growth-suppressing effect of AAV-hTERT-IFN-beta in combination with AAV-hTERT-TRAIL on several cancer cell lines was assessed by MTT assay. Apoptosis of A549 cancer cells infected by AAV-hTERT-IFN-beta alone, AAV-hTERT-TRAIL alone, and their combination was evaluated by apoptotic cell staining and flow cytometry (FCM), respectively. The antitumor effect of the combination of AAV-hTERT-IFN-beta with AAV-hTERT-TRAIL in vivo was further evaluated through A549 lung cancer xenograft in nude mice. The results showed that the combinational treatment was superior to any alone and presented intensified tumor cytotoxic and apoptotic effect on A549 cancer cells. Most importantly, the combination of AAV-hTERT-IFN-beta with AAV-hTERT-TRAIL exhibited significant antitumor effect and eliminated all tumor masses in nude mice, which lay a foundation for exploring the molecular mechanisms of combined IFN-beta and TRAIL anti-tumor activity.
    Sheng wu gong cheng xue bao = Chinese journal of biotechnology 06/2010; 26(6):780-8.
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    ABSTRACT: TNF-related apoptosis-inducing ligand (TRAIL) functions as a soluble cytokine and has been demonstrated significant antitumor activity against a variety of cancer cell lines without toxicity to most normal cells. Cisplatin is a potent anticancer agent and is widely used in the clinical for treatment of human cancers. Adeno-associated virus (AAV2) is a promising gene delivery vehicle for its advantage of low pathogenicity and long-term gene expression. However, lack of tissue specificity caused low efficiency of AAV transfer to target cells. The promoter of human telomerase reverse transcriptase (hTERT) is a good candidate to enhance targeting efficiency of AAV in cancer cells. Although AAV-mediated TRAIL controlled by hTERT promoter (AAV-hTERT-TRAIL) has obvious antitumor activity, the tumor cannot be completely eradicated. In this study, we first examined the effectiveness of combination therapy of cisplatin and AAV-hTERT-TRAIL on human hepatocellular carcinoma (HCC) in vitro and in vivo. For in vitro experiments, tumor cell lines were treated with cisplatin, virus, or both. The transgene TRAIL expression controlled by hTERT promoter was evaluated in BEL7404 HCC cell line. Cytotoxicity was performed by MTT analysis. Cell apoptosis was detected by flow cytometry analysis. The in vivo antitumor efficacy of combination treatment with cisplatin and AAV-hTERT-TRAIL was assessed in human hepatocellular carcinoma xenografts mouse model. The enhanced TRAIL expression was observed in BEL7404 cells treated with AAV-hTERT-TRAIL plus cisplatin. Treatment with both AAV-hTERT-TRAIL and cisplatin exhibited stronger cytotoxicity and induced more significant apoptosis in cancer cells compared with AAV-hTERT-TRAIL or cisplatin alone, respectively. Moreover, in animal experiments, the combined treatment greatly suppressed tumor growth and resulted in tumor cell death. AAV-mediated therapeutic gene expression in combination with chemotherapy provides a promising therapeutic strategy for human cancers. These data suggest that combined use of AAV-hTERT-TRAIL and cisplatin may have potential clinical application.
    Journal of Cancer Research and Clinical Oncology 03/2010; 136(12):1827-37. · 2.91 Impact Factor
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    ABSTRACT: To investigate the effect of hepatocyte-like cells induced by CD34+ cells in vitro on the repair of the injured hepatic tissues of mice in vivo. Mononuclear cells were isolated from umbilical blood by density gradient centrifugation and enriched CD34+ cells were obtained. The cells were (1 x 10(5) cells/mL) cultured in serum-free medium containing stem cell factor (SCF), hepatocyte growth factor (HGF), EGF, oncostatin M (OSM), bFGF (the concentration were 50, 20, 20, 10, 10 ng/mL respectively) in vitro for 10 days. Forty-eight 6-week-old female ICR mice were chosen to prepare liver injury model by injecting carbon tetrachloride and 2-acetylamionoflu-orene. The mice were randomly divided into two groups (n = 24 per group): the experimental group, the cultured cells were injected into the mice through the tail vein; the control group, the equivalent serum-free medium was injected. Six mice from each group were killed at 7, 14, 21, and 28 days after operation to receive HE staining, PCR gel electrophoresis, immunohistochemistry staining, and hepatic function detection. HE staining: the morphology of injured hepatic tissues in the control group recovered to normal 28 days after operation, while in the experimental group, it recovered to normal 14 days after operation. PCR gel electrophoresis and immunohistochemistry staining: the cells expressing human serum albumin were detected in the hepatic tissue of the experimental group at each time point after operation; while in the control group, no such cells were detected within 28 days after operation. Hepatic function detection: the activity of alanine aminotransferase in the control group recovered to normal 14 days after operation; the mean activity of aspartate aminotransferase of two groups failed to recover within 28 days. The hepatocyte-like cells induced by CD34+ cells in vitro can promote the morphological and functional recovery of the injured hepatic tissue in mice. Moreover, it can be transformed into human-derived hepatic cells in liver-injured mice.
    Zhongguo xiu fu chong jian wai ke za zhi = Zhongguo xiufu chongjian waike zazhi = Chinese journal of reparative and reconstructive surgery 10/2009; 23(10):1235-40.
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    ABSTRACT: We have synthesized heparin-immobilized copolymers of L-lactide (LA) and 5-methyl-5-benzyloxycarbonate-1,3-dioxan-2-one (MBC) as non-inflammatory and non-thrombogenic biodegradable coating materials. These copolymers were used in fabricating arsenic trioxide (As(2)O(3))-eluting stents to reduce the late-stage adverse events, such as thrombosis, localized hypersensitivity and inflammation, that occur when applying stents to treat coronary artery diseases. Heparinized copolymers effectively reduced platelet adhesion and protein adsorption while increasing the plasma recalcification time and thromboplastin time in vitro. Histological analysis of the polymer-coated stents in a porcine coronary artery injury model indicated that one heparinized copolymer (Hep-Co90, LA:MBC=90:10), with the highest LA content of 90% and the lowest degradation rate, induced the least foreign body reactions and inflammation, which were as small as those induced by bare metal stents. Consequently, Hep-Co90 was used as the stent coating material for local As(2)O(3) delivery. Histomorphometric evaluations suggested no significant difference between bare metal stents and As(2)O(3)-eluting stents at 1 and 3 months post-implantation. At 6 months, the lumen area in the porcine coronary arteries treated with As(2)O(3)-eluting stents is 32.4% higher than those treated with bare metal stents while the neointimal area, neointimal thickness and stenosis rate decreased by 25.8%, 32.5% and 31.2%, respectively. The As(2)O(3)-eluting stent using Hep-Co90 as the drug carrier and stent coating material presented in this study represents a novel promising device in preventing in-stent restenosis.
    Acta biomaterialia 08/2009; 6(2):534-46. · 5.68 Impact Factor
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    ABSTRACT: Ex vivo expansion of hematopoietic stem cells (HSCs) is very important for clinical applications of cord blood (CB). With the aim to find proper culture duration for ex vivo expansion, mononuclear cells (MNC) was applied as starting culture cells to expand HSCs and the repopulating potential of seven-day and fourteen-day cultured CD34+ cells were compared. The average expansion of total cells and CD34+ cells cultured for 7 days were higher than those cultured for 14 days. The results of phenotypic analysis of fresh and cultured cells showed that the percentage of CD3+ cells declined and the percentage of CD33+ cells increased during culture. The engraftment levels of fourteen-day cultured CD34+ cells were higher than those of fresh and seven-day cultured CD34+ cells. Fourteen-day cultured CD34+ cells also showed better multilineage reconstitution ability than fresh and seven-day cultured CD34+ cells. The results of the present study demonstrated that prolonged culture could preserve the hematopoietic reconstitution ability of ex vivo cultured CB cells and improve the engraftment level in NOD/SCID mice. Keywords cord blood - mononuclear cells - CD34 + cells -ex vivo expansion - NOD/SCID mice
    Biotechnology and Bioprocess Engineering 01/2009; 14(4):429-435. · 1.28 Impact Factor
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    ABSTRACT: The hematopoietic repopulating ability of fresh and cultured CD34+ cells and CD34- cells derived from cord blood were compared by nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mouse model. Fresh CD34+ cells and CD34- cells were isolated from fresh cord blood. Cultured CD34+ cells and CD34- cells were separated from cultured mononuclear cells (MNC). We transplanted these cells into sublethally irradiated NOD/SCID mice via the tail vein and sacrificed surviving mice after 6 weeks. The peripheral blood, spleen and bone marrow from each mouse were harvested for flow cytometry, colony-forming cells and human Alu sequences analyses. The proportions of CD45+ cells and human multilineage hematopoietic cells in NOD/SCID mice received CD34+ cells were close to that in the mice received both CD34+ cells and CD34- cells, while it was significantly higher than that in the mice received CD34- cells. Six weeks after transplantation, all the mice injected with cultured CD34- cells dead. The survival rate of mice injected with cultured CD34+ cells was 66.7%. All of the mice injected with both cultured CD34- and CD34+ cells survived. Moreover, CD45+ cells could be detected in all surviving mice, and human CD34, CD3, CD19, CD33 and CD71 antigen also could be detected on these CD45+ cells. The results showed that both fresh and cultured CD34+ cells had the capability of engraftment and hematopoiesis reconstitution, but CD34- cells hadn't the ability. However, CD34- cells had assistant effect on the hematopoietic repopulating ability of CD34+ cells.
    Sheng wu gong cheng xue bao = Chinese journal of biotechnology 10/2008; 24(9):1588-94.
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    ABSTRACT: Hypoxia maintained biological characteristics of CD34(+) cells through keeping lower intracellular reactive oxygen specials (ROS) levels. The effects of normoxia and hypoxia on antioxidant enzymes and glutathione redox state were compared in this study. Hypoxia decreased the mRNA expression of both catalase (CAT) and glutathione peroxidase (GPX), but not affected mRNAs expression of superoxide dismutase (SOD). While the cellular GPX activities under hypoxia were apparently less than those under normoxia, neither SOD activities nor CAT activities were affected by hypoxia. The analysis of glutathione redox status and ROS products showed the lower oxidized glutathione (GSSG) levels, the higher reduced glutathione (GSH) levels, the higher GSH/GSSG ratios, and the less O(2)- and H(2)O(2) generation under hypoxia (versus normoxia). Meanwhile more primary CD34(+)CD38(-) cells were obtained when cultivation was performed under hypoxia or with N-acetyl cysteine (the precursor of GSH) under normoxia. These results demonstrated the different responses of anti-oxidative mechanism between normoxia and hypoxia. Additionally, the present study suggested that the GSH-GPX antioxidant system played an important role in HSPCs preservation by reducing peroxidation.
    Comparative Biochemistry and Physiology Part B Biochemistry and Molecular Biology 07/2008; 151(2):153-8. · 2.07 Impact Factor
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    ABSTRACT: Adeno-associated virus (AAV) has rapidly become a promising gene delivery vehicle for its excellent advantages of low pathogenicity and long-term gene expression. However, lack of tissue specificity caused low efficiency of AAV transfer to target cells. The promoter of human telomerase reverse transcriptase (hTERT) has been implicated in mediating gene expression in cancer cells as hTERT is transcriptionally upregulated in most cancer cells. Thereby, the hTERT promoter becomes a good candidate to enhance the targeting efficiency of AAV in cancer cells. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) functions as a soluble cytokine to selectively kill various cancer cells without toxicity to most normal cells. It remains to be determined whether the hTERT promoter can efficiently mediate TRAIL gene therapy in cancer cells using AAV vector. A novel AAV vector containing the TRAIL gene under the control of the hTERT promoter (AAV-hTERT-TRAIL) was generated. The specific expression of hTERT-controlled genes was evaluated in cell lines. The antitumor efficacy of AAV-hTERT-TRAIL was assessed in tumor cell lines and human hepatocellular carcinoma xenograft mouse model. TRAIL expression was observed in tumor cells infected with AAV-hTERT-TRAIL at both the protein and mRNA level. AAV-hTERT-TRAIL displayed cancer-specific cytotoxicity and induced tumor cell apoptosis. Moreover, in animal experiments, intratumoral administration of AAV-hTERT-TRAIL significantly suppressed the growth of xenograft tumors and resulted in tumor cell death. AAVs in combination with hTERT-mediated therapeutic gene expression provide a promising targeting approach for developing effective therapy for human cancers. These data suggest that AAV-hTERT-TRAIL is a potent therapeutic agent for cancer therapy.
    The Journal of Gene Medicine 06/2008; 10(5):518-26. · 2.16 Impact Factor
  • Meiqin Zhou, Haibo Cai, Wensong Tan
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    ABSTRACT: Based on the requirement of culture conditions for hematopoietic stem and progenitor cells (HSPCs) ex vive expansion, we developed a new-type bioreactor by combining superiorities of static and stirred culture models. Stem cell factor (SCF), thrombopoietin (TPO), FLT-3 ligand(Flt-3) were used as the cytokines cocktails. The effects of the static and cyclic culture on the expansion characteristics of CD34+ selected cells were compared in the new-type bioreactor. After 7 d cultures, the expansion of total cells in the static culture was 13.86 +/- 4.26 fold, higher than that in the cyclic culture (7.23 +/- 2.67 fold). The analysis of the fold expansion and the proportion of CD34+ cells showed that there was no marked difference between the static culture and the cyclic culture. However, the fold expansion and the proportion of CD34+CD38- cells were higher in the cyclic culture than those in the static culture (3.90 +/- 0.85 versus 1.82 +/- 0.58), which reflected more primary CD34+CD38- cells were obtained in the cyclic culture. The above results demonstrated that both the static culture and the cyclic culture could be used in ex vive expansion of CD34+ cells with the new-type bioreactor. In static culture hematopoietic stem cells differentiated into progenitor cells, whilst the cyclic culture favored the expansion of primary HSPCs.
    Sheng wu gong cheng xue bao = Chinese journal of biotechnology 06/2008; 24(5):786-92.
  • Yiqi Chen, Haibo Cai, Wensong Tan
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    ABSTRACT: To observe the effect of cytokines and combinations in the inducement of human umbilical cord blood-derived CD34+ cells into hepatocyte-like cells. The mononuclear cells (MNCs) were derived by density gradient centrifugation and the CD34+ cells were separated from MNCs. The human umbilical cord blood-derived CD34+ cells were cultivated through 49 different combinations of cytokines including leukemia inhibitor factor (LIF), oncostatin M, bFGF, aFGF, hepatocyte growth factor, EGF and stem cell factor for 28 days, and the concentrations of the cytokines were 10, 10, 10, 10, 20, 20 and 50 ng/mL, respectively. The mRNAs of cytokeratin 19 (CK-19), CK-18, glutamine synthetase (GS), human albumin (ALB) and alpha-fetoprotein (AFP) were detected every seven days. The ALB secretion ability, detoxification ability and hepatic synthesis ability of the induced cells were detected by immunofluorescence assay, indocyanine green (ICG) and periodic acid-schiff assay, respectively. The fresh umbilical cord blood-derived CD34+ cells were detected at the same time as a control. The mRNAs of CK-19, CK-18 and GS could be transcribed in all the induced cells, but the transcription of the mRNAs of ALB and AFP which was the special mark of mature hepatocyte and liver stem cell, respectively, was not found. All the mRNAs could not be found in freshly isolated umbilical cord blood-derived CD34+ cells. All the cells induced in vitro could not release ALB, and not help the detoxification of ICG which was the fundamental function of mature liver cells. These results were the same in the control group. The hepatic synthesis ability of all the induced cells increased by comparison to the fresh ones. Though some mRNAs of proteins which are transcribed in hepatocytes can be found in the induced cells, umbilical cord blood-derived CD34+ cells could not be transdifferentiated into hepatocyte-like cells through cytokines in vitro.
    Zhongguo xiu fu chong jian wai ke za zhi = Zhongguo xiufu chongjian waike zazhi = Chinese journal of reparative and reconstructive surgery 06/2008; 22(6):747-52.
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    ABSTRACT: The hematopoietic reconstitution of cord blood (CB) CD34(+)cells grown in static and stirred system was studied. Static cultures were better than stirred cultures for cell expansion. Engraftment of stirred-culture hematopoietic stem cells (HSCs) was higher than static-culture HSCs. Stirred-culture HSCs had better multilineage reconstitution ability and colony-forming ability than static-culture HSCs. Static cultures thus favor the expansion of HSCs and stirred cultures are more effective in preserving functional HSCs.
    Biotechnology Letters 02/2008; 30(1):61-5. · 1.85 Impact Factor
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    Jinli Fan, Haibo Cai, Wen-Song Tan
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    ABSTRACT: Hypoxia favored the preservation of progenitor characteristics of hematopoietic stem and progenitor cells (HSPCs) in bone marrow. This work aimed at studying the role of reactive oxygen species (ROS)-generating NADPH oxidase system regulated by hypoxia in ex vivo cultures of cord blood CD34+ cells. The results showed that NADPH oxidase activity and ROS generation were reduced in hypoxia with respect to normal oxygen tension. Meanwhile the ROS generation was found to be inhibited by diphenyleneiodonium (the NADPH oxidase inhibitor), or N-acetylcysteine (the ROS scavenger). Accordingly NADPH oxidase mRNA and p67 protein levels decreased in hypoxia. The analysis of progenitor characteristics, including the proportion of cultured cells expressing the HSPCs marker CD34+CD38-, colony production ability of the colony-forming cells (CFCs), and the re-expansion capability of the cultured CD34+ cells, showed that either 5% pO(2) or reduced ROS favored preserving the characteristics of CD34+ progenitors, and promoted the expansion of CD34+CD38- cells as well. The above results demonstrated that hypoxia effectively maintained biological characteristics of CD34+ cells through keeping lower intracellular ROS levels by regulating NADPH oxidase.
    Journal of Biotechnology 08/2007; 130(4):455-62. · 3.18 Impact Factor