941 resultados para Liver cancer stem cell


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Modern concepts for the treatment of myocardial diseases focus on novel cell therapeutic strategies involving stem cell-derived cardiomyocytes (SCMs). However, functional integration of SCMs requires similar electrophysiological properties as primary cardiomyocytes (PCMs) and the ability to establish intercellular connections with host myocytes in order to contribute to the electrical and mechanical activity of the heart. The aim of this project was to investigate the properties of cardiac conduction in a co-culture approach using SCMs and PCMs in cultured cell strands. Murine embryonic SCMs were pooled with fetal ventricular cells and seeded in predefined proportions on microelectrode arrays to form patterned strands of mixed cells. Conduction velocity (CV) was measured during steady state pacing. SCM excitability was estimated from action potentials measured in single cells using the patch clamp technique. Experiments were complemented with computer simulations of conduction using a detailed model of cellular architecture in mixed cell strands. CV was significantly lower in strands composed purely of SCMs (5.5 ± 1.5 cm/s, n = 11) as compared to PCMs (34.9 ± 2.9 cm/s, n = 21) at similar refractoriness (100% SCMs: 122 ± 25 ms, n = 9; 100% PCMs: 139 ± 67 ms, n = 14). In mixed strands combining both cell types, CV was higher than in pure SCMs strands, but always lower than in 100% PCM strands. Computer simulations demonstrated that both intercellular coupling and electrical excitability limit CV. These data provide evidence that in cultures of murine ventricular cardiomyocytes, SCMs cannot restore CV to control levels resulting in slow conduction, which may lead to reentry circuits and arrhythmias.

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One novel treatment strategy for the diseased heart focuses on the use of pluripotent stem cell-derived cardiomyocytes (SC-CMs) to overcome the heart's innate deficiency for self-repair. However, targeted application of SC-CMs requires in-depth characterization of their true cardiogenic potential in terms of excitability and intercellular coupling at cellular level and in multicellular preparations. In this study, we elucidated the electrical characteristics of single SC-CMs and intercellular coupling quality of cell pairs, and concomitantly compared them with well-characterized murine native neonatal and immortalized HL-1 cardiomyocytes. Firstly, we investigated the electrical properties and Ca2+ signaling mechanisms specific to cardiac contraction in single SC-CMs. Despite heterogeneity of the new cardiac cell population, their electrophysiological activity and Ca2+ handling were similar to native cells. Secondly, we investigated the capability of paired SC-CMs to form an adequate subunit of a functional syncytium and analyzed gap junctions and signal transmission by dye transfer in cell pairs. We discovered significantly diminished coupling in SC-CMs compared with native cells, which could not be enhanced by a coculture approach combining SC-CMs and primary CMs. Moreover, quantitative and structural analysis of gap junctions presented significantly reduced connexin expression levels compared with native CMs. Strong dependence of intercellular coupling on gap junction density was further confirmed by computational simulations. These novel findings demonstrate that despite the cardiogenic electrophysiological profile, SC-CMs present significant limitations in intercellular communication. Inadequate coupling may severely impair functional integration and signal transmission, which needs to be carefully considered for the prospective use of SC-CMs in cardiac repair.

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Deregulated expression of the MET receptor tyrosine kinase has been reported in up to 50% of patients with hepatocellular carcinoma, the most abundant form of liver cancers, and is associated with decreased survival. Consequently, MET is considered as a molecular target in this malignancy, whose progression is highly dependent on extensive angiogenesis. Here we studied the impact of MET small molecule inhibitors on angiogenesis-associated parameters and growth of xenograft liver models consisting of cells expressing MET-mutated variants M1268T and Y1248H, which exhibit constitutive kinase activity. We demonstrate that MET mutations expression is associated with significantly increased production of vascular endothelial growth factor, which is blocked by MET targeting only in cells expressing the M1268T inhibitor-sensitive but not in the Y1248H inhibitor-resistant variant. Decrease in vascular endothelial growth factor production is also associated with reduction of tyrosine phopshorylation of the vascular endothelial growth factor receptor 2 expressed on primary liver sinusoidal endothelial cells and with inhibition of vessel formation. Furthermore, MET inhibition demonstrated an efficient anti-tumor activity and considerable reduction in microvessel density only against the M1268T-derived intrahepatic tumors. Collectively, our data support the role of targeting MET-associated angiogenesis as a major biological determinant for liver tumor growth control.

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BACKGROUND Acute myeloid leukaemia mainly affects elderly people, with a median age at diagnosis of around 70 years. Although about 50-60% of patients enter first complete remission upon intensive induction chemotherapy, relapse remains high and overall outcomes are disappointing. Therefore, effective post-remission therapy is urgently needed. Although often no post-remission therapy is given to elderly patients, it might include chemotherapy or allogeneic haemopoietic stem cell transplantation (HSCT) following reduced-intensity conditioning. We aimed to assess the comparative value of allogeneic HSCT with other approaches, including no post-remission therapy, in patients with acute myeloid leukaemia aged 60 years and older. METHODS For this time-dependent analysis, we used the results from four successive prospective HOVON-SAKK acute myeloid leukaemia trials. Between May 3, 2001, and Feb 5, 2010, a total of 1155 patients aged 60 years and older were entered into these trials, of whom 640 obtained a first complete remission after induction chemotherapy and were included in the analysis. Post-remission therapy consisted of allogeneic HSCT following reduced-intensity conditioning (n=97), gemtuzumab ozogamicin (n=110), chemotherapy (n=44), autologous HSCT (n=23), or no further treatment (n=366). Reduced-intensity conditioning regimens consisted of fludarabine combined with 2 Gy of total body irradiation (n=71), fludarabine with busulfan (n=10), or other regimens (n=16). A time-dependent analysis was done, in which allogeneic HSCT was compared with other types of post-remission therapy. The primary endpoint of the study was 5-year overall survival for all treatment groups, analysed by a time-dependent analysis. FINDINGS 5-year overall survival was 35% (95% CI 25-44) for patients who received an allogeneic HSCT, 21% (17-26) for those who received no additional post-remission therapy, and 26% (19-33) for patients who received either additional chemotherapy or autologous HSCT. Overall survival at 5 years was strongly affected by the European LeukemiaNET acute myeloid leukaemia risk score, with patients in the favourable risk group (n=65) having better 5-year overall survival (56% [95% CI 43-67]) than those with intermediate-risk (n=131; 23% [19-27]) or adverse-risk (n=444; 13% [8-20]) acute myeloid leukaemia. Multivariable analysis with allogeneic HSCT as a time-dependent variable showed that allogeneic HSCT was associated with better 5-year overall survival (HR 0·71 [95% CI 0·53-0·95], p=0·017) compared with non-allogeneic HSCT post-remission therapies or no post-remission therapy, especially in patients with intermediate-risk (0·82 [0·58-1·15]) or adverse-risk (0.39 [0·21-0·73]) acute myeloid leukaemia. INTERPRETATION Collectively, the results from these four trials suggest that allogeneic HSCT might be the preferred treatment approach in patients 60 years of age and older with intermediate-risk and adverse-risk acute myeloid leukaemia in first complete remission, but the comparative value should ideally be shown in a prospective randomised study. FUNDING None.

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Vinorelbine chemotherapy with granulocyte-colony stimulating factor (G-CSF) stimulation is a widely applied non-myelosuppressive mobilization regimen in Switzerland for myeloma patients, but its neurotoxic potential limits its use in patients with bortezomib-induced polyneuropathy. In this single-center study, we alternatively evaluated safety and effectiveness of gemcitabine chemotherapy with G-CSF for mobilization of autologous stem cells. Between March 2012 and February 2013, all bortezomib-pretreated myeloma patients planned to undergo first-line high-dose melphalan chemotherapy received a single dose of 1250 mg/m(2) gemcitabine, with G-CSF started on day 4. The 24 patients in this study had received a median of four cycles of bortezomib-dexamethason-based induction. Bortezomib-related polyneuropathy was identified in 21 patients (88%) by clinical evaluation and a standardized questionnaire. Administration of gemcitabine mobilization did not induce new or aggravate pre-existing neuropathy. Stem cell mobilization was successful in all 24 patients, with a single day of apheresis being sufficient in 19 patients (78%). The median yield was 9.51 × 10(6) CD34+ cells/kg. Stem collection could be accomplished at day 8 in 67%. Our data suggest that single-dose gemcitabine together with G-CSF is an effective mobilization regimen in myeloma patients and a safe alternative non-myelosuppressive mobilization chemotherapy for myeloma patients with bortezomib-induced polyneuropathy.

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Vinorelbine chemotherapy with G-CSF stimulation is the standard mobilization regimen in Switzerland for multiple myeloma patients. However, with the increasing use of bortezomib during induction treatment, adding the neurotoxic compound vinorelbine for mobilization may aggravate bortezomib-induced polyneuropathy. In this retrospective single-center study, we aimed to explore vinorelbine mediated neuropathy in 106 consecutive bortezomib pretreated myeloma patients. We confirmed that vinorelbine with G-CSF represents a reliable and effective regimen for mobilization of autologous stem cells. However, the single administration of 35 mg/m(2) vinorelbine added significant neurotoxicity. We found that 24 patients (24%) reported vinorelbine mediated neurotoxicity: Aggravation of bortezomib-induced neuropathy was observed in 17 patients (17%), and vinorelbine mobilization induced first occurrence of polyneuropathy in additional 7 patients (7%). We observed that development of polyneuropathy was not associated with differing survival rates. Finally, affected patients reported polyneuropathy associated disease burden as "very high" in 13% and "high" in 50%. Our data indicate that a single administration of vinorelbine to mobilize autologous stem cells is associated with significant additional polyneuropathy in bortezomib pretreated myeloma patients. The efficacy of vinorelbine mobilization should be balanced against its neurotoxic potential.

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Novel strategies aiming to increase survival rates in patients with advanced-stage mantle cell lymphoma (MCL) and relapsing diffuse large B-cell lymphoma (DLBCL) are a clinical need. High-dose chemotherapy (HDCT) with autologous stem cell transplantation (ASCT) has improved progression-free (PFS) and overall survival (OS) in MCL and relapsed DLBCL. However, the role of CD34+ cell selection before ASCT in MCL and DLBCL is unclear. We retrospectively analyzed the outcome of 62 consecutive patients with advanced-stage MCL or relapsed DLBCL undergoing ASCT with (n=31) or without (n=31) prior CD34+ selection. All patients had stage III or IV disease, with 47% having DLBCL and 53% MCL. The median duration for neutrophil and platelet recovery was 12 and 16 days in CD34+ selected patients, and 11 (P<.001) and 14 days (P=.012) in the group without selection, respectively. No differences in toxicities were observed. The 5-year PFS for CD34+ selected versus not selected patients was 67% and 39% (P=.016), and the 5-year OS was 86% and 54% (P=.007). Our data suggest that using CD34+ selected autografts for ASCT in advanced stage MCL and DLBCL is associated with longer PFS and OS without increased toxicity.

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BACKGROUND & AIMS European and American guidelines have endorsed the Barcelona Clinic Liver Cancer (BCLC) staging system. The aim of this study was to assess the performance of the recently developed Hong Kong Liver Cancer (HKLC) classification as a staging system for hepatocellular carcinoma (HCC) in Europe. METHODS We used a pooled set of 1693 HCC patients combining three prospective European cohorts. Discrimination ability between the nine substages and five stages of the HKLC classification system was assessed. To evaluate the predictive power of the HKLC and BCLC staging systems on overall survival, Nagelkerke pseudo R2, Bayesian Information Criterion and Harrell's concordance index were calculated. The number of patients who would benefit from a curative therapy was assessed for both staging system. RESULTS The HKLC classification in nine substages shows suboptimal discrimination between the staging groups. The classification in five stages shows better discrimination between groups. However, the BCLC classification performs better than the HKLC classification in the ability to predict OS. The HKLC treatment algorithm tags significantly more patients to curative therapy than the BCLC. CONCLUSIONS The BCLC staging system performs better for European patients than the HKLC staging system in predicting OS. Twice more patients are eligible for a curative therapy with the HKLC algorithm, whether this translates in survival benefit remains to be investigated. This article is protected by copyright. All rights reserved.

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BACKGROUND The soluble factors secreted by mesenchymal stem cells are thought to either support or inhibit tumor growth. Herein, we investigated whether the human lung-derived mesenchymal stem cell-conditioned medium (hlMSC-CM) exerts antitumor activity in malignant pleural mesothelioma cell lines H28, H2052 and Meso4. METHODS hlMSC-CM was collected from the human lung-derived mesenchymal stem cells. Inhibition of tumor cell growth was based on the reduction of cell viability and inhibition of cell proliferation using the XTT and BrdU assays, respectively. Elimination of tumor spheroids was assessed by the anchorage-independent sphere formation assay. The cytokine profile of hlMSC-CM was determined by a chemiluminescence-based cytokine array. RESULTS Our data showed that hlMSC-CM contains a broad range of soluble factors which include: cytokines, chemokines, hormones, growth and angiogenic factors, matrix metalloproteinases, metalloproteinase inhibitors and cell-cell mediator proteins. The 48- and 72-hour hlMSC-CM treatments of H28, H2052 and Meso4 cell lines elicited significant decreases in cell viability and inhibited cell proliferation. The 72-hour hlMSC-CM incubation of H28 cells completely eliminated the drug-resistant sphere-forming cells, which is more potent than twice the half maximal inhibitory concentration of cisplatin. CONCLUSIONS Our findings indicate that the cell-free hlMSC-CM confers in vitro antitumor activities via soluble factors in the tested mesothelioma cells and, hence, may serve as a therapeutic tool to augment the current treatment strategies in malignant pleural mesothelioma.

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Abstract We explored the feasibility of unrelated donor haematopoietic stem cell transplant (HSCT) upfront without prior immunosuppressive therapy (IST) in paediatric idiopathic severe aplastic anaemia (SAA). This cohort was then compared to matched historical controls who had undergone first-line therapy with a matched sibling/family donor (MSD) HSCT (n = 87) or IST with horse antithymocyte globulin and ciclosporin (n = 58) or second-line therapy with unrelated donor HSCT post-failed IST (n = 24). The 2-year overall survival in the upfront cohort was 96 ± 4% compared to 91 ± 3% in the MSD controls (P = 0·30) and 94 ± 3% in the IST controls (P = 0·68) and 74 ± 9% in the unrelated donor HSCT post-IST failure controls (P = 0·02).The 2-year event-free survival in the upfront cohort was 92 ± 5% compared to 87 ± 4% in MSD controls (P = 0·37), 40 ± 7% in IST controls (P = 0·0001) and 74 ± 9% in the unrelated donor HSCT post-IST failure controls (n = 24) (P = 0·02). Outcomes for upfront-unrelated donor HSCT in paediatric idiopathic SAA were similar to MSD HSCT and superior to IST and unrelated donor HSCT post-IST failure. Front-line therapy with matched unrelated donor HSCT is a novel treatment approach and could be considered as first-line therapy in selected paediatric patients who lack a MSD. © 2015 John Wiley & Sons Ltd.

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Tyrosine kinase inhibitors represent today's treatment of choice in chronic myeloid leukemia (CML). Allogeneic hematopoietic stem cell transplantation (HSCT) is regarded as salvage therapy. This prospective randomized CML-study IIIA recruited 669 patients with newly diagnosed CML between July 1997 and January 2004 from 143 centers. Of these, 427 patients were considered eligible for HSCT and were randomized by availability of a matched family donor between primary HSCT (group A; N=166 patients) and best available drug treatment (group B; N=261). Primary end point was long-term survival. Survival probabilities were not different between groups A and B (10-year survival: 0.76 (95% confidence interval (CI): 0.69-0.82) vs 0.69 (95% CI: 0.61-0.76)), but influenced by disease and transplant risk. Patients with a low transplant risk showed superior survival compared with patients with high- (P<0.001) and non-high-risk disease (P=0.047) in group B; after entering blast crisis, survival was not different with or without HSCT. Significantly more patients in group A were in molecular remission (56% vs 39%; P=0.005) and free of drug treatment (56% vs 6%; P<0.001). Differences in symptoms and Karnofsky score were not significant. In the era of tyrosine kinase inhibitors, HSCT remains a valid option when both disease and transplant risk are considered.Leukemia advance online publication, 20 November 2015; doi:10.1038/leu.2015.281.

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OBJECTIVES The aetiology of hyposalivation in haematopoietic stem cell transplantation (HSCT) recipients is not fully understood. This study examined the effects of treatment-related aetiological factors, particularly medications, on stimulated salivary flow in HSCT recipients. SUBJECTS AND METHODS Adult HSCT recipients (N = 118, 66 males, 27 autologous and 91 allogeneic transplants) were examined. Stimulated whole salivary flow rates (SWSFR) were measured before HSCT and at 6 and 12 months post-HSCT. Linear regression models were used to analyse the associations of medications and transplant-related factors with salivary flow rates, which were compared to salivary flow rates of generally healthy controls (N = 247). RESULTS The SWSFR of recipients were lower pre-HSCT (mean ± standard deviation, 0.88 ± 0.56 ml/min; P < 0.001), 6 months post-HSCT (0.84 ± 0.61; P < 0.001) and 12 months post-HSCT (1.08 ± 0.67; P = 0.005) than the SWSFR of controls (1.31 ± 0.65). In addition, hyposalivation (<0.7 ml/min) was more frequent among HSCT recipients pre-HSCT (P < 0.001), 6 months post-HSCT (P < 0.001) and 12 months post-HSCT (P = 0.01) than among controls. The SWSFR was observed to improve over time being significantly higher 12 months post-HSCT compared to pre-HSCT (P < 0.001). The observed decrease of salivary flow could not be explained by the examined transplant-related factors and medications. CONCLUSIONS Decreased stimulated salivary flow rates could not be explained by the examined factors alone; these findings indicate that hyposalivation in HSCT recipients exhibits a multifactorial aetiology. CLINICAL RELEVANCE All HSCT recipients should be considered to be at high risk of hyposalivation and consequent oral diseases, and they should be treated accordingly.

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Hair cycle disturbances are common in dogs and comparable to some alopecic disorders in humans. A normal hair cycle is maintained by follicular stem cells which are predominately found in an area known as the bulge. Due to similar morphological characteristics of the bulge area in humans and dogs, the shared particularity of compound hair follicles as well as similarities in follicular biomarker expression, the dog is a promising model to study human hair cycle and stem cell disorders. To gain insight into the spatial distribution of follicular keratinocytes with stem cell potential in canine compound follicles, we microdissected hair follicles in anagen and telogen from skin samples of freshly euthanized dogs. The keratinocytes isolated from different locations were investigated for their colony forming efficiency, growth and differentiation potential as well as clonal growth. Our results indicate that i) compound and single hair follicles exhibit a comparable spatial distribution pattern with respect to cells with high growth potential and stem cell-like characteristics, ii) the lower isthmus (comprising the bulge) harbors most cells with high growth potential in both, the anagen and the telogen hair cycle stage, iii) unlike in other species, colonies with highest growth potential are rather small with an irregular perimeter and iv) the keratinocytes derived from the bulbar region exhibit characteristics of actively dividing transit amplifying cells. Our results now provide the basis to conduct comparative studies of normal dogs and those with hair cycle disorders with the possibility to extend relevant findings to human patients.

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The mammalian cerebral neocortex is a complex six-layered structure containing multiple types of neurons. Pyramidal neurons of the neocortex are formed during development in an inside-out manner, by which deep layer (DL) neurons are generated first, and upper layer (UL) neurons are generated last. Neurons within the six-layered neocortex express unique markers for their position, showing whether they are subplate, deep layer, upper layer, or Cajal-Retzius neurons. The sequential generation of cortical layers, which exists in vivo, has been partially recapitulated in vitro by differentiation of mouse embryonic stem cells (Gaspard et al., 2008) and human embryonic stem cells (hESC) (Eiraku et al., 2008). The timeline of generation of cortical neurons from hESC is still not well defined, and could be very important in the future of cell therapy. In this study we will define timeline for UL and DL neurons for our experimental paradigm as well as test the effects of fibroblast growth factors (FGF) 2 and 8 on this neuronal differentiation. Recent papers suggest that FGFs are critical for forebrain patterning (Storm et al., 2003). Neuronal differentiation after treatment with either FGF2 or FGF8 from hESCs will be examined and the proportion of specific neuronal markers will be analyzed using immunocytochemistry. Our results show that the generated pyramidal neurons will express DL and UL laminar markers in vitro as they do in vivo and that the presence of FGF8 in induction media creates a proliferative effect, while FGF2 induces hESC to differentiate at a higher rate.

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Human embryonic stem cells (hESCs) have the potential to differentiate to all adult somatic cells. This property makes hESCs a very promising area of research for the treatment of disorders in which specific cell populations need to be restored. Despite this potential, research that focuses on producing mesodermally derived cell populations from hESCs is decidedly limited, notwithstanding the prevalence of disorders involving mesodermal tissues for which treatment options are limited. Skeletal muscle myoblasts are derivatives of mesodermal cells and are characterized by the expression of the MyoD gene. These cells are difficult to obtain from hESCs in a reproducible and efficient manner. Recent developments in the field have showed some success in obtaining myogenic cells from hESCs through a mesenchymal stem cell (MSC)-like intermediate population. MSCs, which are an adult stem cell population typically derived from the bone marrow, are capable of generating multiple cell types including skeletal muscle. The aim of this study was to develop an efficient method that derives myoblasts from an MSC-like intermediate. To accomplish this goal, we first set out to isolate and expand the MSC-like intermediate from hESCs differentiated in vitro. Difficulties in reproducing published cell-differentiation methodologies, which represent a significant and familiar challenge in hESC research, are highlighted in this report.