One-step cell biomanufacturing platform:porous gelatin microcarrier beads promote human embryonic stem cell-derived midbrain dopaminergic progenitor cell differentiation in vitro and survival after transplantation in vivo

Numerous studies have shown that cell replacement therapy can replenish lost cells and rebuild neural circuitry in animal models of Parkinson’s disease.Transplantation of midbrain dopaminergic progenitor cells is a promising treatment for Parkinson’s disease.However,transplanted cells can be injured by mechanical damage during handling and by changes in the transplantation niche.Here,we developed a one-step biomanufacturing platform that uses small-aperture gelatin microcarriers to produce beads carrying midbrain dopaminergic progenitor cells.These beads allow midbrain dopaminergic progenitor cell differentiation and cryopreservation without digestion,effectively maintaining axonal integrity in vitro.Importantly,midbrain dopaminergic progenitor cell bead grafts showed increased survival and only mild immunoreactivity in vivo compared with suspended midbrain dopaminergic progenitor cell grafts.Overall,our findings show that these midbrain dopaminergic progenitor cell beads enhance the effectiveness of neuronal cell transplantation.

Quality-adjusted time without symptoms or toxicity analysis of haploidentical-related donor vs.identical sibling donor hematopoietic stem cell transplantation in acute myeloid leukemia

Objective:We aimed to compare the quality-adjusted time without symptoms or toxicity(Q-TWiST)in acute myeloid leukemia(AML)patients who received haploidentical-related donor(HID)and identical sibling donor(ISD)hematopoietic stem cell transplantation(HSCT).Methods:Five clinical health states were defined:toxicity(TOX),acute graft-versus-host disease(GVHD),chronic GVHD(cGVHD),time without symptoms and toxicity(TWiST)and relapse(REL).The equation used in this study was as follows:Q-TWiST=UTOX×TOX+UTWiST×TWiST+UREL×REL+UaGVHD×aGVHD+UcGVHD×cGVHD.Results:A total of 239 AML patients were enrolled.We established a mathematical model,i.e.,Q-TWiST HID HSCT>Q-TWiST ISD HSCT,to explore the range of utility coefficients satisfying the inequality.Based on the raw data,the utility coefficient is equivalent to the following inequality:10.57067UTOX-46.27733UREL+105.9374+3.388078UaGVHD-210.8198UcGVHD>0.The model showed that when UTOX,UREL,and UaGVHD were within the range of 0-1,as well as when UcGVHD was within the range of 0-0.569,the inequality Q-TWiST HID HSCT>Q-TWiST ISD HSCT was valid.According to the results of the ChiCTR1800016972 study,the median coefficients of TOX,acute GVHD(aGVHD),and cGVHD were 0.56(0.41-0.76),0.56(0.47-0.72),and 0.54(0.37-0.79),respectively.We selected a series of specific examples of the coefficients,i.e.,UTOX=0.5,UREL=0.05,UaGVHD-0.5,and UcGVHD-0.5.The Q-TWiST values of ISD and HID HSCT were 896 and 900 d,respectively(P=0.470).Conclusions:We first observed that Q-TWiST was comparable between AML patients receiving HID HSCT and those receiving ISD HSCT.

Gossypol acetic acid regulates leukemia stem cells by degrading LRPPRC via inhibiting IL-6/JAK1/STAT3 signaling or resulting mitochondrial dysfunction

BACKGROUND Leukemia stem cells(LSCs)are found to be one of the main factors contributing to poor therapeutic effects in acute myeloid leukemia(AML),as they are protected by the bone marrow microenvironment(BMM)against conventional therapies.Gossypol acetic acid(GAA),which is extracted from the seeds of cotton plants,exerts anti-tumor roles in several types of cancer and has been reported to induce apoptosis of LSCs by inhibiting Bcl2.AIM To investigate the exact roles of GAA in regulating LSCs under different microenvironments and the exact mechanism.METHODS In this study,LSCs were magnetically sorted from AML cell lines and the CD34+CD38-population was obtained.The expression of leucine-rich pentatricopeptide repeat-containing protein(LRPPRC)and forkhead box M1(FOXM1)was evaluated in LSCs,and the effects of GAA on malignancies and mitochondrial RESULTS LRPPRC was found to be upregulated,and GAA inhibited cell proliferation by degrading LRPPRC.GAA induced LRPPRC degradation and inhibited the activation of interleukin 6(IL-6)/janus kinase(JAK)1/signal transducer and activator of transcription(STAT)3 signaling,enhancing chemosensitivity in LSCs against conventional chemotherapies,including L-Asparaginase,Dexamethasone,and cytarabine.GAA was also found to downregulate FOXM1 indirectly by regulating LRPPRC.Furthermore,GAA induced reactive oxygen species accumulation,disturbed mitochondrial homeostasis,and caused mitochondrial dysfunction.By inhibiting IL-6/JAK1/STAT3 signaling via degrading LRPPRC,GAA resulted in the elimination of LSCs.Meanwhile,GAA induced oxidative stress and subsequent cell damage by causing mitochondrial damage.CONCLUSION Taken together,the results indicate that GAA might overcome the BMM protective effect and be considered as a novel and effective combination therapy for AML.

NRS2002 assesses nutritional status of leukemia patients undergoing hematopoietic stem cell transplantation

Objective： To discuss whether nutritional risk screening 2002 （NRS2002） is appropriate for nutritional risk screening for leukemia patients before and after hematopoietic stem cell transplantation （HSCT）, and whether there are risk differences in other conditions, such as age, gender and matching degree; to find the methods and indicators of nutritional risk screening for these patients before and after HSCT, in order to give timely intervention to guarantee the successful completion of the entire transplantation process. Methods： Nutritional risk of 99 leukemia patients was screened with NRS2002 before and after HSCT. The ^（2 test was applied to compare the risk differences between groups such as age, gender and matching degree, while the differences of other enumeration data, such as recent （1-3 months） weight loss, reduced food intake within one week and BMI, were compared by continuity correction. Results： Of the 99 leukemia patients, 22 cases （22.2 %） had nutritional risk before HSCT, while all patients had nutritional risk after ttSCT; there is no significant difference in nutritional risk between male and female, and patients of less than 30 years old, not-full matched, recent （1-3 months） weight loss, reduced food intake within a week or BMI 〈18.5 were more likely to have nutritional risk; and 77 cases （77.8%） had weight loss, among which 49 patients （63.6%） had more than 5% weight loss within one month. Conclusions= This study showed that leukemia patients should receive the nutritional risk screening conventionally before and after HSCT, and NRS2002 was only appropriate for nutritional risk screening before HSCT. More attention should be paid to the patients less than 30 years old or not-full matched. Weight change was one of the important nutritional indicators for patients after HSCT.

Identification and targeting leukemia stem cells: The path to the cure for acute myeloid leukemia

Accumulating evidence support the notion that acute myeloid leukemia(AML) is organized in a hierarchical system, originating from a special proportion of leukemia stem cells(LSC). Similar to their normal counterpart, hematopoietic stem cells(HSC), LSC possess selfrenewal capacity and are responsible for the continued growth and proliferation of the bulk of leukemia cells in the blood and bone marrow. It is believed that LSC are also the root cause for the treatment failure and relapse of AML because LSC are often resistant to chemotherapy. In the past decade, we have made significant advancement in identification and understanding the molecular biology of LSC, but it remains a daunting task to specifically targeting LSC, while sparing normalHSC. In this review, we will first provide a historical overview of the discovery of LSC, followed by a summary of identification and separation of LSC by either cell surface markers or functional assays. Next, the review will focus on the current, various strategies for eradicating LSC. Finally, we will highlight future directions and challenges ahead of our ultimate goal for the cure of AML by targeting LSC.

Transplantation of bone marrow-derived endothelial progenitor cells and hepatocyte stem cells from liver fibrosis rats ameliorates liver fibrosis

AIM To explore the effectiveness for treating liver fibrosisby combined transplantation of bone marrow-derived endothelial progenitor cells(BM-EPCs) and bone marrow-derived hepatocyte stem cells(BDHSCs) from the liver fibrosis environment.METHODS The liver fibrosis rat models were induced with carbon tetrachloride injections for 6 wk. BM-EPCs from rats with liver fibrosis were obtained by different rates of adherence and culture induction. BDHSCs from rats with liver fibrosis were isolated by magnetic bead cell sorting. Tracing analysis was conducted by labeling EPCs with PKH26 in vitro to show EPC location in the liver. Finally, BM-EPCs and/or BDHSCs transplantation into rats with liver fibrosis were performed to evaluate the effectiveness of BM-EPCs and/or BDHSCs on liver fibrosis.RESULTS Normal functional BM-EPCs from liver fibrosis rats were successfully obtained. The co-expression level of CD133 and VEGFR2 was 63.9% ± 2.15%. Transplanted BM-EPCs were located primarily in/near hepatic sinusoids. The combined transplantation of BM-EPCs and BDHSCs promoted hepatic neovascularization, liver regeneration and liver function, and decreased collagen formation and liver fibrosis degree. The VEGF levels were increased in the BM-EPCs(707.10 ± 54.32) and BM-EPCs/BDHSCs group(615.42 ± 42.96), compared with those in the model group and BDHSCs group(P < 0.05). Combination of BM-EPCs/BDHSCs transplantation induced maximal up-regulation of PCNA protein and HGF m RNA levels. The levels of alanine aminotransferase(AST), aspartate aminotransferase, total bilirubin(TBIL), prothrombin time(PT) and activated partial thromboplastin time in the BMEPCs/BDHSCs group were significantly improved, to be equivalent to normal levels(P > 0.05) compared with those in the BDHSC(AST, TBIL and PT, P < 0.05) and BM-EPCs(TBIL and PT, P < 0.05) groups. Transplantation of BM-EPCs/BDHSCs combination significantly reduced the degree of liver fibrosis(staging score of 1.75 ± 0.25 vs BDHSCs 2.88 ± 0.23 or BMEPCs 2.75 ± 0.16, P < 0.05).CONCLUSION The combined transplantation exhibited maximal therapeutic effect compared to that of transplantation of BM-EPCs or BDHSCs alone. Combined transplantation of autogenous BM-EPCs and BDHSCs may represent a promising strategy for the treatment of liver fibrosis, which would eventually prevent cirrhosis and liver cancer.

Comparison of phenotypic markers and neural differentiation potential of multipotent adult progenitor cells and mesenchymal stem cells

AIM: To compare the phenotypic and neural differentiation potential of human bone marrow derived multipotent adult progenitor cells (MAPC) and mesenchymal stem cells (MSC). METHODS: Cultures of MAPC and MSC were established in parallel from same samples of human bone marrow (n = 5). Both stem cell types were evaluated for expression of pluripotency markers including Oct-4 and Nanog by immunocytochemistry and reversetranscription polymerase chain reaction (RT-PCR) and expression of standard mesenchymal markers including CD14, CD34, CD44, CD45, CD73, CD90, CD105 andhuman leukocyte antigen (HLA)-ABC by flow cytometry. After treatment with neural induction medium both MAPC and MSC were evaluated for expression of neural proteins [neuronal filament-200 (NF-200) and glial fibrillar acidic protein (GFAP)] by immunocytochemistry and Western blotting and neural genes [NF-200, GFAP, Tau, microtubule-associated protein (MAP)-1B, MAP-2, neuron-specific enolase (NSE) and oligodendrocyte-1 (Olig-1)] by quantitative real-time-PCR. RESULTS: MAPC had small trigonal shaped while MSC had elongated spindle-shaped morphology. The MAPC expressed Oct-4 and Nanog both at gene and protein levels, whereas MSC were negative for these pluripotent markers. MAPC were negative for HLA-ABC while MSC had high expression of HLA-ABC. In addition, MAPC as compared to MSC had significantly lower expression of CD44 (36.56% ± 1.92% vs 98.23% ± 0.51%), CD73 (15.11% ± 2.24% vs 98.53% ± 2.22%) and CD105 (13.81% ± 3.82%vs 95.12% ± 5.65%) (P < 0.001, for all) MAPC cultures compared to MSC cultures treated with neural induction medium had significantly higher fold change expression of NF-200 (0.64), GFAP (0.52), Tau (0.59), MAP-2 (0.72), Olig-1 (0.18) and NSE (0.29) proteins (P < 0.01 for Olig-1 and P < 0.001 for rest) as well as higher fold change expression of genes of NF-200 (1.34),GFAP (1.12),Tau (1.08),MAP-1B (0.92), MAP-2 (1.14) andNSE (0.4) (P < 0.001 for all). CONCLUSION: MAPC can be differentially characterized from MSC as Oct-4 and Nanog positive stem cells with no expression of HLA-ABC and low expression of mesenchymal markers CD44, CD73 and CD105 and when compared to MSC they possess greater predilection for differentiation into neuro-ectodermal lineage.

Epigenetic therapy of cancer stem and progenitor cells by targeting DNA methylation machineries

Recent advances in stem cell biology have shed light on how normal stem and progenitor cells can evolve to acquire malignant characteristics during tumorigenesis. The cancer counterparts of normal stem and progenitor cells might be occurred through alterations of stem cell fates including an increase in self-renewal capability and a decrease in differentiation and/or apoptosis. This oncogenic evolution of cancer stem and progenitor cells, which often associates with aggressive phenotypes of the tumorigenic cells, is controlled in part by dysregulated epigenetic mechanisms including aberrant DNA methylation leading to abnormal epigenetic memory. Epigenetic therapy by targeting DNA methyltransferases(DNMT) 1, DNMT3 A and DNMT3 B via 5-Azacytidine(Aza) and 5-Aza-2'-deoxycytidine(Aza-d C) has proved to be successfultoward treatment of hematologic neoplasms especially for patients with myelodysplastic syndrome. In this review, I summarize the current knowledge of mechanisms underlying the inhibition of DNA methylation by Aza andAza-d C, and of their apoptotic- and differentiation-inducingeffects on cancer stem and progenitor cells in leukemia, medulloblastoma, glioblastoma, neuroblastoma, prostate cancer, pancreatic cancer and testicular germ cell tumors. Since cancer stem and progenitor cells are implicatedin cancer aggressiveness such as tumor formation, progression, metastasis and recurrence, I propose that effective therapeutic strategies might be achievedthrough eradication of cancer stem and progenitor cells by targeting the DNA methylation machineries to interfere their "malignant memory".

Wharton's jelly mesenchymal stem cells differentiate into retinal progenitor cells

Human Wharton＇s jelly mesenchymal stem cells were isolated from fetal umbilical cord. Cells were cultured in serumfree neural stem cellconditioned medium or neural stem cellconditioned medium supplemented with Dkk1, a Wnt/13 catenin pathway antagonist, and LeftyA, a Nodal signaling pathway antagonist to induce differentiation into retinal progenitor cells. Inverted microscopy showed that after induction, the spindleshaped or fibroblastlike Wharton＇s jelly mesenchymal stem cells changed into bulbous cells with numerous processes. Immunofluorescent cytochemical stain ing and reversetranscription PCR showed positive expression of retinal progenitor cell markers, Pax6 and Rx, as well as weakly downregulated nestin expression. These results demonstrate that Wharton＇s jelly mesenchymal stem cells are capable of differentiating into retinal progenitor cells in vitro.

Transplantation of vascular endothelial growth factor-modified neural stem/progenitor cells promotes the recovery of neurological function following hypoxic-ischemic brain damage

Neural stem/progenitor cell （NSC） transplantation has been shown to effectively improve neurological function in rats with hypoxic-isch- emic brain damage. Vascular endothelial growth factor （VEGF） is a signaling protein that stimulates angiogenesis and improves neural regeneration. We hypothesized that transplantation of VEGF-transfected NSCs would alleviate hypoxic-ischemic brain damage in neo- natal rats. We produced and transfected a recombinant lentiviral vector containing the VEGF165gene into cultured NSCs. The transfected NSCs were transplanted into the left sensorimotor cortex of rats 3 days after hypoxic-ischemic brain damage. Compared with the NSCs group, VEGF mRNA and protein expression levels were increased in the transgene NSCs group, and learning and memory abilities were significantly improved at 30 days. Furthermore, histopathological changes were alleviated in these animals. Our findings indicate that transplantation of VEGF-transfected NSCs may facilitate the recovery of neurological function, and that its therapeutic effectiveness is better than that of unmodified NSCs.

Propofol and remifentanil at moderate and high concentrations affect proliferation and differentiation of neural stem/progenitor cells

Propofol and remifentanil alter intracellular Ca^2＋ concentration （[Ca^2＋]i） in neural stem/progen-itor cells by activating γ-aminobutyric acid type A receptors and by reducing testosterone levels. However, whether this process affects neural stem/progenitor cell proliferation and differenti-ation remains unknown. In the present study, we applied propofol and remifentanil, alone or in combination, at low, moderate or high concentrations （1, 2–2.5 and 4–5 times the clinically effective blood drug concentration）, to neural stem/progenitor cells from the hippocampi of newborn rat pups. Low concentrations of propofol, remifentanil or both had no noticeable effect on cell proliferation or differentiation; however, moderate and high concentrations of propofol and/or remifentanil markedly suppressed neural stem/progenitor cell proliferation and differen-tiation, and induced a decrease in [Ca^2＋]i during the initial stage of neural stem/progenitor cell differentiation. We therefore propose that propofol and remifentanil interfere with the prolifer-ation and differentiation of neural stem/progenitor cells by altering [Ca^2＋]i. Our ifndings suggest that propofol and/or remifentanil should be used with caution in pediatric anesthesia.

Donor-Derived CD19-Targeted T Cell Infusion Eliminates B Cell Acute Lymphoblastic Leukemia Minimal Residual Disease with No Response to Donor Lymphocytes after Allogeneic Hematopoietic Stem Cell Transplantation

Leukemia relapse is still the leading cause of treatment failure after allogeneic hematopoietic stem cell transplantation (allo-HSCT) for B cell acute lymphoblastic leukemia (B-ALL). Relapsed patients with BALL after allo-HSCT have a very short median survival. Minimal residual disease (MRD) is predictive of forthcoming hematological relapse after hematopoietic stem cell transplantation (HSCT);furthermore, eliminating MRD effectively prevents relapse. Donor lymphoblastic infusion (DLI) is the main established approach to treat B-ALL with MRD after allo-HSCT. However, about one-third of patients with MRD are non-responsive to DLI and their prognosis worsens. Although donor-derived cluster of differentiation (CD)19-directed chimeric antigen receptor-modified (CAR) T cells (CART19s) can potentially cure leukemia, the efficiency and safety of infusions with these cells have not yet been investigated in patients with MRD after HSCT. Between September 2014 and February 2018, six patients each received one or more infusions of CART19s from HSCT donors. Five (83.33%) achieved MRD-negative remission, and one case was not responsive to the administration of CAR T cells. Three of the six patients are currently alive without leukemia. No patient developed acute graft-versus-host disease (aGVHD), and no patient died of cytokine release syndrome. Donor-derived CAR T cell infusions seem to be an effective and safe intervention for patients with MRD in B-ALL after allo-HSCT and for those who were not responsive to DLI.

Similarities and differences between mesenchymal stem/progenitor cells derived from various human tissues

BACKGROUND Mesenchymal stromal/stem cells (MSCs) constitute a promising tool in regenerative medicine and can be isolated from different human tissues. However, their biological properties are still not fully characterized. Whereas MSCs from different tissue exhibit many common characteristics, their biological activity and some markers are different and depend on their tissue of origin. Understanding the factors that underlie MSC biology should constitute important points for consideration for researchers interested in clinical MSC application. AIM To characterize the biological activity of MSCs during longterm culture isolated from: bone marrow (BM-MSCs), adipose tissue (AT-MSCs), skeletal muscles (SMMSCs), and skin (SK-MSCs). METHODS MSCs were isolated from the tissues, cultured for 10 passages, and assessed for: phenotype with immunofluorescence and flow cytometry, multipotency with differentiation capacity for osteo-, chondro-, and adipogenesis, stemness markers with qPCR for mRNA for Sox2 and Oct4, and genetic stability for p53 and c-Myc;27 bioactive factors were screened using the multiplex ELISA array, and spontaneous fusion involving a co-culture of SM-MSCs with BM-MSCs or AT-MSCs stained with PKH26 (red) or PKH67 (green) was performed. RESULTS All MSCs showed the basic MSC phenotype;however, their expression decreased during the follow-up period, as confirmed by fluorescence intensity. The examined MSCs express CD146 marker associated with proangiogenic properties;however their expression decreased in AT-MSCs and SM-MSCs, but was maintained in BM-MSCs. In contrast, in SK-MSCs CD146 expression increased in late passages. All MSCs, except BM-MSCs, expressed PW1, a marker associated with differentiation capacity and apoptosis. BM-MSCs and AT-MSCs expressed stemness markers Sox2 and Oct4 in long-term culture. All MSCs showed a stable p53 and c-Myc expression. BM-MSCs and AT-MSCs maintained their differentiation capacity during the follow-up period. In contrast, SK-MSCs and SM-MSCs had a limited ability to differentiate into adipocytes. BM-MSCs and AT-MSCs revealed similarities in phenotype maintenance, capacity for multilineage differentiation, and secretion of bioactive factors. Because AT-MSCs fused with SM-MSCs as effectively as BM-MSCs, AT-MSCs may constitute an alternative source for BM-MSCs. CONCLUSION Long-term culture affects the biological activity of MSCs obtained from various tissues. The source of MSCs and number of passages are important considerations in regenerative medicine.

Expression change of stem cell-derived neural stem/progenitor cell sup-porting factor gene in injured spinal cord of rats

Objective To explore the expression change of stem cell-derived neural stem/progenitor cell supporting factor （SDNSF） gene in the injuried spinal cord tissues of rats, and the relation between the expressions of SDNSF and nestin. Methods The spinal cord contusion model of rat was established according to Allen＇s falling strike method. The expression of SDNSF was studied by RT-PCR and in situ hybridization （ISH）, and the expression of nestin was detected by immunochemistry. Results RT-PCR revealed that SDNSF mRNA was upregulated on day 4 after injury, peaked on day 8-12, and decreased to the sham operation level on day 16. ISH revealed that SDNSF mRNA was mainly expressed in the gray matter cells, probably neurons, of spinal cord. The immunohistochemistry showed that accompanied with SDNSF mRNA upregulation, the nestin-positive cells showed erupted roots, migrated peripherad and proliferation on the 8-day slice. However, the distribution pattern of these new cells was different from that of SDNSF-positive cells. Conclusion （1） SDNSF is expressed in the gray matter of spinal cord. The expression of SDNSF mRNA in the spinal cord varies with injured time. （2） The nestin-positive cells proliferate accompanied with spinal cord injury repair, but do not secrete SDNSF.

Dysfunctional stem and progenitor cells impair fracture healing with age

Successful fracture healing requires the simultaneous regeneration of both the bone and vasculature;mesenchymal stem cells (MSCs) are directed to replace the bone tissue, while endothelial progenitor cells (EPCs) form the new vasculature that supplies blood to the fracture site. In the elderly, the healing process is slowed, partly due to decreased regenerative function of these stem and progenitor cells. MSCs from older individuals are impaired with regard to cell number, proliferative capacity, ability to migrate, and osteochondrogenic differentiation potential. The proliferation, migration and function of EPCs are also compromised with advanced age. Although the reasons for cellular dysfunction with age are complex and multidimensional, reduced expression of growth factors, accumulation of oxidative damage from reactive oxygen species, and altered signaling of the Sirtuin-1 pathway are contributing factors to aging at the cellular level of both MSCs and EPCs. Because of these geriatric-specific issues, effective treatment for fracture repair may require new therapeutic techniques to restore cellular function. Some suggested directions for potential treatments include cellular therapies, pharmacological agents, treatments targeting age-related molecular mechanisms, and physical therapeutics. Advanced age is the primary risk factor for a fracture, due to the low bone mass and inferior bone quality associated with aging;a better understanding of the dysfunctional behavior of the aging cell will provide a foundation for new treatments to decrease healing time and reduce the development of complications during the extended recovery from fracture healing in the elderly.

Philadelphia chromosome-positive leukemia stem cells in acute lymphoblastic leukemia and tyrosine kinase inhibitor therapy

Leukemia stem cells(LSCs),which constitute a minority of the tumor bulk,are functionally defined on the basis of their ability to transfer leukemia into an immunodeficient recipient animal.The presence of LSCs has been demonstrated in acute lymphoblastic leukemia(ALL),of which ALL with Philadelphia chromosome-positive(Ph+).The use of imatinib,a tyrosine kinase inhibitor(TKI),as part of front-line treatment and in combination with cytotoxic agents,has greatly improved the proportions of complete response and molecular remission and the overall outcome in adults with newly diagnosed Ph+ ALL.New challenges have emerged with respect to induction of resistance to imatinib via Abelson tyrosine kinase mutations.An important recent addition to the arsenal against Ph+ leukemias in general was the development of novel TKIs,such as nilotinib and dasatinib.However,in vitro experiments have suggested that TKIs have an antiproliferative but not an antiapoptotic or cytotoxic effect on the most primitive ALL stem cells.None of the TKIs in clinical use target the LSC.Second generation TKI dasatinib has been shown to have a more profound effect on the stem cell compartment but the drug was still unable to kill the most primitive LSCs.Allogeneic stem cell transplantation(SCT) remains the only curative treatment available for these patients.Several mechanisms were proposed to explain the resistance of LSCs to TKIs in addition to mutations.Hence,TKIs may be used as a bridge to SCT rather than monotherapy or combination with standard chemotherapy.Better understanding the biology of Ph+ ALL will open new avenues for effective management.In this review,we highlight recent findings relating to the question of LSCs in Ph+ ALL.

Decitabine for Relapsed Acute Lymphoblastic Leukemia after Allogeneic Hematopoietic Stem Cell Transplantation

Relapse after allogeneic hematopoietic stem cell transplantation（allo-HSCT） remains a main question on treatment failure. Current strategies for management that usually include salvage chemotherapy, donor lymphocytic infusion and second transplantation. Our study assessed the efficacy of decitabine（DAC） for treating patients with acute lymphoblastic leukemia（ALL） who relapsed after allogeneic hematopoietic stem cell transplantation（allo-HSCT）. We retrospectively analyzed the outcomes of 12 patients with relapsed ALL after allo-HSCT who received DAC therapy. Nine patients received DAC combined with chemotherapy and donor stem cell infusion, and 3 patients received single-agent DAC. Ten of the 12 patients achieved complete remission（CR）, 1 achieved a partial remission（PR）, and 1 had no response（NR） after treatment at the latest follow-up（LFU）, the median survival was 11.2 months（range, 3.8–34, 7 months）. The 1-and 2-year overall survival（OS） rates were 50%（6/12） and 25%（3/12）, respectively. Five patients were still alive; 4 had maintained CR and 1 was alive with disease. Patients with Philadelphia chromosome-positive ALL had higher survival rate than patients with Philadelphia chromosome-negative ALL（57.1% vs. 20%）. No aggravated flares of graft-versus-host disease（GVHD） were observed during DAC treatment. Therefore, DAC may be a promising therapeutic agent for ALL recurrence after allo-HSCT.

Changes in expression and secretion patterns of fibroblast growth factor 8 and Sonic Hedgehog signaling pathway molecules during murine neural stem/progenitor cell differentiation in vitro

In the present study, we investigated the dynamic expression of fibroblast growth factor 8 and Sonic Hedgehog signaling pathway related factors in the process of in vitro hippocampal neural stem/progenitor cell differentiation from embryonic Sprague-Dawley rats or embryonic Kunming species mice, using fluorescent quantitative reverse transcription-PCR and western blot analyses. Results demonstrated that the dynamic expression of fibroblast growth factor 8 was similar to fibroblast growth factor receptor 1 expression but not to other fibroblast growth factor receptors. Enzyme-linked immunosorbent assay demonstrated that fibroblast growth factor 8 and Sonic Hedgehog signaling pathway protein factors were secreted by neural cells into the intercellular niche. Our experimental findings indicate that fibroblast growth factor 8 and Sonic Hedgehog expression may be related to the differentiation of neural stem/progenitor cells.

Resveratrol-downregulated Phosphorylated Liver Kinase B1 Is Involved in Senescence of Acute Myeloid Leukemia Stem Cells

Summary： Senescence is an important obstacle to cancer development. Engaging a senescent response may be an effective way to cure acute myeloid leukemia （AML）. The aim of this study was to examine the effect of resveratrol-downregulated phosphorylated liver kinase B1 （pLKB1） on the senescence of acute myeloid leukemia （AML） stem cells. The protein expressions of pLKB 1 and Sirtuin 1 （SIRT1）, a regulator ofpLKB1, were measured in CD34＋CD38-KGla cells treated with resveratrol （40 μmol/L） or not by Western blotting. Senescence-related factors were examined, including p21 mRNA tested by real-time PCR, cell morphology by senescence-associated β-galactosidase （SA-β-gal） staining, cell pro- liferation by MTT assay and cell cycle by flow cytometry. Besides, apoptosis was flow cytometrically determined. The results showed that pLKB1 was highly expressed in CD34＋CD38- KGla cells, and resveratrol, which could downregulate pLKB1 through activation of SIRT1, induced senescence and apoptosis of CD34＋CD38- KGla cells. It was concluded that resveratrol-downregulated pLKB1 is in- volved in the senescence of AML stem cells.

Stages based molecular mechanisms for generating cholangiocytes from liver stem/progenitor cells

Except for the most organized mature hepatocytes,liver stem/progenitor cells(LSPCs)can differentiate into many other types of cells in the liver including cholangiocytes.In addition,LSPCs are demonstrated to be able to give birth to other kinds of extra-hepatic cell types such as insulin-producing cells.Even more,under some bad conditions,these LSPCs could generate liver cancer stem like cells(LCSCs)through malignant transformation.In this review,we mainly concentrate on the molecular mechanisms for controlling cell fates of LSPCs,especially differentiation of cholangiocytes,insulin-producing cells and LCSCs.First of all,to certificate the cell fates of LSPCs,the following three features need to be taken into account to perform accurate phenotyping:(1)morphological properties;(2)specific markers;and(3)functional assessment including in vivo transplantation.Secondly,to promote LSPCs differentiation,systematical attention should be paid to inductive materials(such as growth factors and chemical stimulators),progressive materials including intracellular and extracellular signaling pathways,and implementary materials(such as liver enriched transcriptive factors).Accordingly,some recommendations were proposed to standardize,optimize,and enrich the effective production of cholangiocyte-like cells out of LSPCs.At the end,the potential regulating mechanisms for generation of cholangiocytes by LSPCs were carefully analyzed.The differentiation of LSPCs is a gradually progressing process,which consists of three main steps:initiation,progression and accomplishment.It’s the unbalanced distribution of affecting materials in each step decides the cell fates of LSPCs.