The role of WNT and mTOR in human memory T cell formation

Cancer is one of the world's leading causes of death with a rising trend in incidence. These epidemiologic observations underline the need for novel treatment strategies. In this regard, a promising approach takes advantage of the adaptive effector mechanisms of the immune system, using T lymphocytes to specifically target and destroy tumour cells. However, whereas current approaches mainly depend on short-lived, terminally differentiated effector T cells, increasing evidence suggests that long lasting and maximum efficient immune responses are mediated by low differentiated memory T cells. These memory T cells should display characteristics of stem cells, such as longevity, self-renewal capacity and the ability to continuously give rise to further differentiated effectors. These stem celllike memory T (TSCM) cells are thought to be of key therapeutic value as they might not only attack differentiated tumour cells, but also eradicate the root cause of cancer, the cancer stem cells themselves. Thus, efforts are made to characterize TSCM cells and to identify the signalling pathways which mediate their induction. Recently, a human TSCM cell subset was described and the activation of the Wnt-ß-catenin signalling pathway by the drug TWS119 during naive CD8+ T (TN) cell priming was suggested to mediate their induction. However, a precise deciphering of the signalling pathways leading to TSCM cell induction and an in-depth characterization of in vitro induced and in vivo occurring TSCM cells remain to be performed. Here, evidence is presented that the induction of human and mouse CD8+ and CD4+ TSCM cells may be triggered by inhibition of mechanistic/mammalian target of rapamycin (mTOR) complex 1 with simultaneously active mTOR complex 2. This molecular mechanism arrests a fraction of activated TN cells in a stem cell-like differentiation state independently of the Wnt-ß-catenin signalling pathway. Of note, TWS119 was found to also inhibit mTORCl, thereby mediating the induction of TSCM cells. Suggesting an immunostimulatory effect, the acquired data broaden the therapeutic range of mTORCl inhibitors like rapamycin, which are, at present, exclusively used due to their immunosuppressive function. Furthermore, by performing broad metabolic analyses, a well-orchestrated interplay between intracellular signalling pathways and the T cells' metabolic programmes could be identified as important regulator of the T cells' differentiation fate. Moreover, in vitro induced CD4+ TSCM cells possess superior functional capacities and share fate-determining key factors with their naturally occurring counterparts, assessed by a first-time full transcriptome analysis of in vivo occurring CD4+ TN cell, TSCM cells and central memory (TCM) cells and in vitro induced CD4+ TSCM cells. Of interest, a group of 56 genes, with a unique expression profile in TSCM cells could be identified. Thus, a pharmacological mechanism allowing to confer sternness to activated TN cells has been found which might be highly relevant for the design of novel T cell-based cancer immunotherapies.

Impact of HLA A2 and cytomegalovirus serostatus on outcomes in patients with leukemia following matched-sibling myeloablative allogeneic hematopoietic cell transplantation.

BACKGROUND AND OBJECTIVES: Donor cytomegalovirus seropositivity was reported to improve leukemia outcomes in HLA-A2 identical hematopoietic cell transplant (HCT) recipients, due to a possible cross-reactivity of donor HLA-A2-restricted CMV-specific T cells with minor histocompatibility (H) antigen of recipient cells. This study analyzed the role of donor CMV serostatus and HLA-A2 status on leukemia outcomes in a large population of HLA-identical HCT recipients. DESIGN AND METHODS: Leukemia patients transplanted between 1992 and 2003 at the Fred Hutchinson Cancer Research Center were categorized as standard risk [leukemia first remission, chronic myeloid leukemia in chronic phase (CML-CP)] and high risk (advanced disease) patients. Time-to-event analysis was used to evaluate the risk of relapse and death associated with HLA-A2 status and donor CMV serostatus. RESULTS: In standard risk patients, acute leukemia (p<0.001) and sex mismatch (female to male, p=0.004)) independently increased the risk of death, while acute leukemia increased the risk of relapse (p<0.001). In high risk patients acute leukemia (p=0.01), recipient age > or = 40 (p=0.005) and herpes simplex virus (HSV) seropositivity (p<0.001) significantly increased the risk death; HSV seropositivity (p=0.006) increased the risk of relapse. Donor CMV serostatus had no significant effect on mortality or relapse in any HLA group. INTERPRETATION AND CONCLUSION: This epidemiological study did not confirm the previously reported effect of donor CMV serostatus on the outcomes of leukemia in HLA-A2-identical HCT recipients. Addressing the question of cross-reactivity of HLA-A2-restricted CMV-specific T cells with minor H antigens in a clinical study would require knowledge of the patient's minor H antigen genotype. However, because of the unbalanced distribution of HLA-A2-restricted minor H antigens in the population and their incomplete identification, this question might be more appropriately evaluated in in vitro experiments than in a clinical study.

IFN alpha activates dormant haematopoietic stem cells in vivo

Maintenance of the blood system is dependent on dormant haematopoietic stem cells (HSCs) with long-term self-renewal capacity. After injury these cells are induced to proliferate to quickly reestablish homeostasis(1). The signalling molecules promoting the exit of HSCs out of the dormant stage remain largely unknown. Here we show that in response to treatment of mice with interferon-alpha (IFN alpha), HSCs efficiently exit G(0) and enter an active cell cycle. HSCs respond to IFN alpha treatment by the increased phosphorylation of STAT1 and PKB/Akt (also known as AKT1), the expression of IFN alpha target genes, and the upregulation of stem cell antigen-1 (Sca-1, also known as LY6A). HSCs lacking the IFN alpha/beta receptor (IFNAR)(2), STAT1 (ref. 3) or Sca-1 (ref. 4) are insensitive to IFN alpha stimulation, demonstrating that STAT1 and Sca-1 mediate IFN alpha-induced HSC proliferation. Although dormant HSCs are resistant to the anti-proliferative chemotherapeutic agent 5-fluoro-uracil(1,5), HSCs pre-treated (primed) with IFN alpha and thus induced to proliferate are efficiently eliminated by 5-fluoro-uracil exposure in vivo. Conversely, HSCs chronically activated by IFN alpha are functionally compromised and are rapidly out-competed by non-activatable Ifnar(-/-) cells in competitive repopulation assays. Whereas chronic activation of the IFN alpha pathway in HSCs impairs their function, acute IFN alpha treatment promotes the proliferation of dormant HSCs in vivo. These data may help to clarify the so far unexplained clinical effects of IFN alpha on leukaemic cells(6,7), and raise the possibility for new applications of type I interferons to target cancer stem cells(8).

The effect of endocrine responsiveness on high-risk breast cancer treated with dose-intensive chemotherapy: results of International Breast Cancer Study Group Trial 15-95 after prolonged follow-up.

BACKGROUND: The role of adjuvant dose-intensive chemotherapy and its efficacy according to baseline features has not yet been established. PATIENTS AND METHODS: Three hundred and forty-four patients were randomized to receive seven courses of standard-dose chemotherapy (SD-CT) or three cycles of dose-intensive epirubicin and cyclophosphamide (epirubicin 200 mg/m(2) plus cyclophosphamide 4 mg/m(2) with filgrastim and progenitor cell support). All patients were assigned tamoxifen at the completion of chemotherapy. The primary end point was disease-free survival (DFS). This paper updates the results and explores patterns of recurrence according to predicting baseline features. RESULTS: At 8.3-years median follow-up, patients assigned DI-EC had a significantly better DFS compared with those assigned SD-CT [8-year DFS percent 47% and 37%, respectively, hazard ratio (HR) 0.76; 95% confidence interval 0.58-1.00; P = 0.05]. Only patients with estrogen receptor (ER)-positive disease benefited from the DI-EC (HR 0.61; 95% confidence interval 0.39, 0.95; P = 0.03). CONCLUSIONS: After prolonged follow-up, DI-EC significantly improved DFS, but the effect was observed only in patients with ER-positive disease, leading to the hypothesis that efficacy of DI-EC may relate to its endocrine effects. Further studies designed to confirm the importance of endocrine responsiveness in patients treated with dose-intensive chemotherapy are encouraged.

Ochratoxin A at nanomolar concentration perturbs the homeostasis of neural stem cells in highly differentiated but not in immature three-dimensional brain cell cultures.

Ochratoxin A (OTA), a fungal contaminant of basic food commodities, is known to be highly cytotoxic, but the pathways underlying adverse effects at subcytotoxic concentrations remain to be elucidated. Recent reports indicate that OTA affects cell cycle regulation. Therefore, 3D brain cell cultures were used to study OTA effects on mitotically active neural stem/progenitor cells, comparing highly differentiated cultures with their immature counterparts. Changes in the rate of DNA synthesis were related to early changes in the mRNA expression of neural stem/progenitor cell markers. OTA at 10nM, a concentration below the cytotoxic level, was ineffective in immature cultures, whereas in mature cultures it significantly decreased the rate of DNA synthesis together with the mRNA expression of key transcriptional regulators such as Sox2, Mash1, Hes5, and Gli1; the cell cycle activator cyclin D2; the phenotypic markers nestin, doublecortin, and PDGFRα. These effects were largely prevented by Sonic hedgehog (Shh) peptide (500ngml(-1)) administration, indicating that OTA impaired the Shh pathway and the Sox2 regulatory transcription factor critical for stem cell self-renewal. Similar adverse effects of OTA in vivo might perturb the regulation of stem cell proliferation in the adult brain and in other organs exhibiting homeostatic and/or regenerative cell proliferation.

Crosstalk of Notch with p53 and p63 in cancer growth control.

Understanding the complexity of cancer depends on an elucidation of the underlying regulatory networks, at the cellular and intercellular levels and in their temporal dimension. This Opinion article focuses on the multilevel crosstalk between the Notch pathway and the p53 and p63 pathways. These two coordinated signalling modules are at the interface of external damaging signals and control of stem cell potential and differentiation. Positive or negative reciprocal regulation of the two pathways can vary with cell type and cancer stage. Therefore, selective or combined targeting of the two pathways could improve the efficacy and reduce the toxicity of cancer therapies.

Epidemiology and outcome of fungemia in a cancer Cohort of the Infectious Diseases Group (IDG) of the European Organization for Research and Treatment of Cancer (EORTC 65031).

BACKGROUND: Anti-cancer treatment and the cancer population have evolved since the last European Organisation for Research and Treatment of Cancer (EORTC) fungemia survey, and there are few recent large epidemiological studies. METHODS: This was a prospective cohort study including 145 030 admissions of patients with cancer from 13 EORTC centers. Incidence, clinical characteristics, and outcome of fungemia were analyzed. RESULTS: Fungemia occurred in 333 (0.23%; 95% confidence interval [CI], .21-.26) patients, ranging from 0.15% in patients with solid tumors to 1.55% in hematopoietic stem cell transplantation recipients. In 297 evaluable patients age ranged from 17 to 88 years (median 56 years), 144 (48%) patients were female, 165 (56%) had solid tumors, and 140 (47%) had hematological malignancies. Fungemia including polymicrobial infection was due to: Candida spp. in 267 (90%), C. albicans in 128 (48%), and other Candida spp. in 145 (54%) patients. Favorable overall response was achieved in 113 (46.5%) patients by week 2. After 4 weeks, the survival rate was 64% (95% CI, 59%-70%) and was not significantly different between Candida spp. Multivariable logistic regression identified baseline septic shock (odds ratio [OR] 3.04, 95% CI, 1.22-7.58) and tachypnoea as poor prognostic factors (OR 2.95, 95% CI, 1.66-5.24), while antifungal prophylaxis prior to fungemia (OR 0.20, 95% CI, .06-.62) and remission of underlying cancer (OR, 0.18; 95% CI, .06-.50) were protective. CONCLUSIONS: Fungemia, mostly due to Candida spp., was rare in cancer patients from EORTC centers but was associated with substantial mortality. Antifungal prophylaxis and remission of cancer predicted better survival.

JNK controls the onset of mitosis of planarian stem cells and triggers apoptotic cell death required for regeneration and remodeling

Regeneration of lost tissues depends on the precise interpretation of molecular signals that control and coordinate the onset of proliferation, cellular differentiation and cell death. However, the nature of those molecular signals and the mechanisms that integrate the cellular responses remain largely unknown. The planarian flatworm is a unique model in which regeneration and tissue renewal can be comprehensively studied in vivo. The presence of a population of adult pluripotent stem cells combined with the ability to decode signaling after wounding enable planarians to regenerate a complete, correctly proportioned animal within a few days after any kind of amputation, and to adapt their size to nutritional changes without compromising functionality. Here, we demonstrate that the stress-activated c-jun-NH2-kinase (JNK) links wound-induced apoptosis to the stem cell response during planarian regeneration. We show that JNK modulates the expression of wound-related genes, triggers apoptosis and attenuates the onset of mitosis in stem cells specifically after tissue loss. Furthermore, in pre-existing body regions, JNK activity is required to establish a positive balance between cell death and stem cell proliferation to enable tissue renewal, remodeling and the maintenance of proportionality. During homeostatic degrowth, JNK RNAi blocks apoptosis, resulting in impaired organ remodeling and rescaling. Our findings indicate that JNK-dependent apoptotic cell death is crucial to coordinate tissue renewal and remodeling required to regenerate and to maintain a correctly proportioned animal. Hence, JNK might act as a hub, translating wound signals into apoptotic cell death, controlled stem cell proliferation and differentiation, all of which are required to coordinate regeneration and tissue renewal.

Cancer therapy-related toxicity in the immature testis

Infertility is a common late effect of childhood cancer treatment. Testicular toxicity can clinically be first detected after the onset of pubertal maturation of the patients when the testis does not grow, spermatogenesis does not initiate and serum levels of gonadotrophins rise. Improved prognosis for childhood cancer has resulted in a growing number of childhood cancer survivors with late effects. In our study, we developed novel tools for detecting cancer therapy-related testicular toxicity during development. By using these methods the effects of the tyrosine kinase inhibitor imatinib mesylate, chemotherapy agent doxorubicin and irradiation on testicular development were investigated in rat and monkey. Patients with chronic myeloid leukemia and some patients with acute lymphoblastic leukemia have fusion gene BCR-ABL which codes for abnormal tyrosine kinase protein. Imatinib mesylate (Glivec®) inhibits activity of this protein. In addition, imatinib inhibits the action of the c-kit and PDGF –receptors, which are both important for the survival and proliferation of the spermatogonial stem cell pool. Imatinib exposure during prepubertal development disturbed the development and the growth of the testis. Spermatogonial stem cells were also sensitive to the toxic effects of doxorubicin and irradiation during the initiation phase of spermatogenesis. In addition, the effect of the treatment of acute lymphoblastic leukemia on germ cell numbers and recovery of reproductive functions after sexual maturation was investigated. Therapy for childhood acute lymphoblastic leukemia seldom results in infertility. The present study gives new information on the mechanisms by which cancer treatments exert their gonadal toxicity in immature testis.

Chemical Biology Screen for Prostate Cancer Therapeutics

Prostate cancer initially responds to hormone-based therapeutics such as anti-androgen treatment or chemotherapeutics but eventually becomes resistant. Novel treatment options are therefore urgently needed. This thesis study applied a high-throughput screen of 4910 known drugs and drug-like small molecules to identify compounds that selectively inhibit growth of prostate cancer cells. In addition, the mechanisms underlying the cellular sensitivity to potent cancer selective compounds were addressed. Surprisingly, many of the compounds currently used in the clinics or studied in clinical trials were not cancer-selective. Only four drugs, aldehyde dehydrogenase inhibitor disulfiram (Antabus), antibiotic ionophore monensin, histone deacetylase inhibitor tricostatin A and fungicide thiram inhibited prostate cancer cell growth at nanomolar concentrations without major effects on non-malignant prostate epithelial cells. Disulfiram, monensin and a structurally similar compound to monensin, salinomycin, induced oxidative stress and inhibited aldehyde dehydrogenase activity. Moreover, monensin and salinomycin reduced androgen receptor signalling and steroidogenesis, enforced cell differentiation and reduced the overall levels of cancer stem cells. Taken together, novel and potentially prostate cancer-selective therapeutic agents were identified in this study, including the description of a multitude of intoxicating mechanisms such as those relating to oxidative stress. The results provide novel insights into prostate cancer biology and exemplify useful means of considering novel approaches to cancer treatment.

Health Related Quality of Life after Childhood Cancer - A Finnish Nationwide survey

The purpose of this Finnish epidemiological nationwide cross-sectional study was to evaluate the Health Related Quality of Life (HRQL) of young people that have survived childhood cancer at least four years after cancer diagnosis. The study aims were (1) to increase knowledge and understanding about the relationship between childhood cancer and its treatment and HRQL of childhood cancer survivors and (2) to identify survivors who need and could benefit from ongoing long-term follow-up, as well as (3) to identify what kind of aftercare the childhood cancer survivors will possibly need. HRQL and fatigue of currently still young survivors of extracranial childhood malignancies were evaluated with self-reports and parent proxy reports. HRQL was measured with age-appropriate generic instruments: PedsQL™, SF-36, 15D, 16D and 17D. Fatigue for children and adolescents aged below 18 years was measured with the PedsQL™ Multidimensional Fatigue Scale Finnish version. PedsQL™ parent-proxy and the PedsQL™ Multidimensional Fatigue Scale Parentproxy instruments were used to assess the perception of the parents on HRQL and fatigue of their children and adolescents. Postal-survey questionnaires were mailed to 852 childhood cancer survivors aged 11-27 years and their randomly selected gender-, age and living-place matched controls, as well as under 18-year-old children´s parents. A total of 474 survivors, 595 controls, 209 survivor’s parent and 253 control’s parent replied. The mean age of survivors at the time of the study was 18.4 years. The mean length of survival was 12.3 years, and the mean age at diagnosis 5.5 years. The most of the Finnish childhood cancer survivors evaluated that their HRQL as good. Survivors rated their HRQL equal or higher than their controls. The only dimension where the survivors scored poorer than the controls was the 15D mobility dimension. Survivors of childhood cancer did not suffer from significant fatigue. There were subgroups of childhood cancer survivors who had poorer level of HRQL, and suffered from fatigue more than the reference group. The demographic factors that associated with poorer HRQL were female gender, greater weight, living alone, need of remedial education, an additional non-cancer diagnosis, survivors with siblings, and self-reported unhappiness. Disease-related factors that associated with poorer HRQL were higher age at the time of diagnosis, the diagnosis of Wilms tumor, neuroblastoma, or osteosarcoma, and treatment with stem cell transplantation. The factors associated with more fatigue in survivors were male gender, older age at evaluation, the need of remedial education at school, lower overall average grade in the latest school marks report, length of survival more than 10 years, lower HRQL-scores, and a sarcoma diagnosis. However, all the used demographic and disease related factors explained only about one third of the variation in the HRQL scores. In open questions, the survivors were most worried about their physical health, but were also worried about their mental health, cancer inheritance, late-effects, and fertility and relapse issues. It seems that there are subgroups of survivors who need and could benefit from ongoing long-term follow-up. In the future, the survivors of childhood cancer need more information about their physical and mental health, as well as on their cancer inheritance, possible late-effects including fertility issues, and on the risk of relapse.

Pluripotency and genetic stability of human pluripotent stem cells

Pluripotent cells have the potential to differentiate into all somatic cell types. As the adult human body is unable to regenerate various tissues, pluripotent cells provide an attractive source for regenerative medicine. Human embryonic stem cells (hESCs) can be isolated from blastocyst stage embryos and cultured in the laboratory environment. However, their use in regenerative medicine is restricted due to problems with immunosuppression by the host and ethical legislation. Recently, a new source of pluripotent cells was established via the direct reprogramming of somatic cells. These human induced pluripotent stem cells (hiPSCs) enable the production of patient specific cell types. However, numerous challenges, such as efficient reprogramming, optimal culture, directed differentiation, genetic stability and tumor risk need to be solved before the launch of therapeutic applications. The main objective of this thesis was to understand the unique properties of human pluripotent stem cells. The specific aims were to identify novel factors involved in maintaining pluripotency, characterize the effects of low oxygen culture on hESCs, and determine the high resolution changes in hESCs and hiPSCs during culture and reprogramming. As a result, the previously uncharacterized protein L1TD1 was determined to be specific for pluripotent cells and essential for the maintenance of pluripotency. The low oxygen culture supported undifferentiated growth and affected expression of stem cell associated transcripts. High resolution screening of hESCs identified a number of culture induced copy number variations and loss of heterozygosity changes. Further, screening of hiPSCs revealed that reprogramming induces high resolution alterations. The results obtained in this thesis have important implications for stem cell and cancer biology and the therapeutic potential of pluripotent cells.

Regulation of self-renewal and detection of karyotypic changes of pluripotent human embryonic stem cells

Human embryonic stem cells are pluripotent cells capable of renewing themselves and differentiating to specialized cell types. Because of their unique regenerative potential, pluripotent cells offer new opportunities for disease modeling, development of regenerative therapies, and treating diseases. Before pluripotent cells can be used in any therapeutic applications, there are numerous challenges to overcome. For instance, the key regulators of pluripotency need to be clarified. In addition, long term culture of pluripotent cells is associated with the accumulation of karyotypic abnormalities, which is a concern regarding the safe use of the cells for therapeutic purposes. The goal of the work presented in this thesis was to identify new factors involved in the maintenance of pluripotency, and to further characterize molecular mechanisms of selected candidate genes. Furthermore, we aimed to set up a new method for analyzing genomic integrity of pluripotent cells. The experimental design applied in this study involved a wide range of molecular biology, genome-wide, and computational techniques to study the pluripotency of stem cells and the functions of the target genes. In collaboration with instrument and reagent company Perkin Elmer, KaryoliteTM BoBsTM was implemented for detecting karyotypic changes of pluripotent cells. Novel genes were identified that are highly and specifically expressed in hES cells. Of these genes, L1TD1 and POLR3G were chosen for further investigation. The results revealed that both of these factors are vital for the maintenance of pluripotency and self-renewal of the hESCs. KaryoliteTM BoBsTM was validated as a novel method to detect karyotypic abnormalities in pluripotent stem cells. The results presented in this thesis offer significant new information on the regulatory networks associated with pluripotency. The results will facilitate in understanding developmental and cancer biology, as well as creating stem cell based applications. KaryoliteTM BoBsTM provides rapid, high-throughput, and cost-efficient tool for screening of human pluripotent cell cultures.

Effects of gamma-radiation on cell growth, cycle arrest, death, and superoxide dismutase expression by DU 145 human prostate cancer cells

Gamma-irradiation (gamma-IR) is extensively used in the treatment of hormone-resistant prostate carcinoma. The objective of the present study was to investigate the effects of 60Co gamma-IR on the growth, cell cycle arrest and cell death of the human prostate cancer cell line DU 145. The viability of DU 145 cells was measured by the Trypan blue exclusion assay and the 3(4,5-dimethylthiazol-2-yl)-2,5,diphenyltetrazolium bromide test. Bromodeoxyuridine incorporation was used for the determination of cell proliferation. Cell cycle arrest and cell death were analyzed by flow cytometry. Superoxide dismutase (SOD), specifically CuZnSOD and MnSOD protein expression, after 10 Gy gamma-IR, was determined by Western immunoblotting analysis. gamma-IR treatment had a significant (P < 0.001) antiproliferative and cytotoxic effect on DU 145 cells. Both effects were time and dose dependent. Also, the dose of gamma-IR which inhibited DNA synthesis and cell proliferation by 50% was 9.7 Gy. Furthermore, gamma-IR induced cell cycle arrest in the G2/M phase and the percentage of cells in the G2/M phase was increased from 15% (control) to 49% (IR cells), with a nonsignificant induction of apoptosis. Treatment with 10 Gy gamma-IR for 24, 48, and 72 h stimulated CuZnSOD and MnSOD protein expression in a time-dependent manner, approximately by 3- to 3.5-fold. These data suggest that CuZnSOD and MnSOD enzymes may play an important role in the gamma-IR-induced changes in DU 145 cell growth, cell cycle arrest and cell death.

A novel alternative to cryopreservation for the short-term storage of stem cells for use in cell therapy using alginate encapsulation

Efficient transport of stem/progenitor cells without affecting their survival and function is a key factor in any practical cell-based therapy. However, the current approach using liquid nitrogen for the transfer of stem cells requires a short delivery time window is technically challenging and financially expensive. The present study aims to use semipermeable alginate hydrogels (crosslinked by strontium) to encapsulate, store, and release stem cells, to replace the conventional cryopreservation method for the transport of therapeutic cells within world-wide distribution time frame. Human mesenchymal stem cell (hMSC) and mouse embryonic stem cells (mESCs) were successfully stored inside alginate hydrogels for 5 days under ambient conditions in an air-tight environment (sealed cryovial). Cell viability, of the cells extracted from alginate gel, gave 74% (mESC) and 80% (hMSC) survival rates, which compared favorably to cryopreservation. More importantly, the subsequent proliferation rate and detection of common stem cell markers (both in mRNA and protein level) from hMSCs and mESCs retrieved from alginate hydrogels were also comparable to (if not better than) results gained following cryopreservation. In conclusion, this new and simple application of alginate hydrogel encapsulation may offer a cheap and robust alternative to cryopreservation for the transport and storage of stem cells for both clinical and research purposes.