Limited Number Of Stem Cells Research Articles

Regenerative Potential of Granulation Tissue in Periodontitis: A Systematic Review and Meta-analysis.

Electronic searches were conducted in five databases including CENTRAL, MEDLINE, EMBASE, Web of Science, and Dentistry & Oral Sciences Source using a combination of MeSH terms and keywords up to 21 June 2022. Human studies including patients aged over 18 years with all forms of periodontitis were included. Following the risk of bias assessment, both qualitative and quantitative analyses were performed. A total of twelve studies were included in qualitative analysis and six of them in quantitative analyses. The evidence suggested that cells derived from periodontitis granulation tissue have osteogenic, adipogenic, chondrogenic, neurogenic, and angiogenic differentiation abilities as well as immunoregulatory properties. In particular, CD44+, CD73+, CD90+, CD105+, and CD146+ cells were found widely in granulation tissue whilst the only meta-analysis confirmed that CD90+ cells were present in lower numbers within the granulation tissue when compared with healthy periodontal tissue (WMD = -23.43%, 95% CI -30.43 to -16.44, p < 0.00001). This review provided further evidence that granulation tissue from patients with periodontitis can be a potential stem cell source for regenerative therapy.

Harvest of Stem Cells from Umbilical Cord Blood: Relevance of Perinatal Factors for the Quality of Umbilical Cord Transplant Units

ZusammenfassungEinleitung Nabelschnurblut (NSB) enthält hämatopoetische Stammzellen mit therapeutischem Potenzial und einzigartigen zellulären Eigenschaften. Aufgrund der begrenzten Anzahl an Stammzellen im NSB (Surrogatmarker total nucleated cells, TNC) eignet sich nur jede fünfte Spende für eine Transplantation. Ziel dieser Studie war es, zu untersuchen, ob prädiktive Faktoren für eine TNC-Zahl über der 99. Perzentile existieren. Material und Methodik Retrospektive Datenanalyse der 100 größten NSB-Spenden (Top100-Kohorte) aus 2299 registrierten Einheiten. Unterschiede zwischen maternalen, fetalen und geburtshilflichen Faktoren wurden analysiert und mit einer standardisierten Kohorte von 731 NSB-Spenden verglichen. Ergebnisse Das mütterliche Alter und der BMI in der Top100-Kohorte waren höher als in der Vergleichskohorte (32 vs. 31 Jahre, p=0,007; 30 kg/m2 vs. 29 kg/m2, p=0,024). Es gab mehr Erstgebärende (76,0 vs. 62,8%, p=0,013) und Gestationsdiabetikerinnen (5,00 vs. 1,65%, p=0,044). Die Schwangerschaftswoche, das Geburtsgewicht, der Anteil vaginal-operativer Geburten und sekundärer Sectiones war in der Top100-Kohorte höher (40+4 vs. 40+1 SSW, p=0,002), (3700 vs. 3450 g, p<0,001), (53,0 vs. 22,7%, p<0,001) (10 vs. 6,2%, p=0,014). Fazit Für eine erfolgreiche Transplantation ist die Höhe der TNC-Zahl entscheidend. Vaginal-operative Entbindungen, sekundäre Sectiones und ein Geburtsgewicht über 3700 g sind günstige Faktoren. Gerade bei Geburten mit einem pathologischen Verlauf sollte nach sicherer Versorgung von Mutter und Kind nicht auf eine Entnahme verzichtet werden.

Towards manufactured red blood cells for the treatment of inherited anemia.

Patients with inherited anemia and hemoglobinopathies (such as sickle cell disease and β-thalassemia) are treated with red blood cell (RBC) transfusions to alleviate their symptoms. Some of these patients may have rare blood group types or go on to develop alloimmune reactions, which can make it difficult to source compatible blood in the donor population. Laboratory-grown RBC represent a particularly attractive alternative which could satisfy an unmet clinical need. The challenge, however, is to produce - from a limited number of stem cells - the 2x1012 RBC required for a standard adult therapeutic dose. Encouraging progress has been made in RBC production from adult stem cells under good manufacturing practice. In 2011, the Douay group conducted a successful proof-of-principle mini-transfusion of autologous manufactured RBC in a single volunteer. In the UK, a trial is planned to assess whether manufactured RBC are equivalent to RBC produced naturally in donors, by testing an allogeneic mini-dose of laboratory-grown manufactured RBC in multiple volunteers. This review discusses recent progress in the erythroid culture field as well as opportunities for further scaling up of manufactured RBC production for transfusion practice.

Does the Global Outbreak of COVID-19 or Other Viral Diseases Threaten the Stem Cell Reservoir Inside the Body?

The COVID-19 pandemic has profoundly influenced public health and contributed to global economic divergences of unprecedented dimensions. Due to the high prevalence and mortality rates, it is then expected that the consequence and public health challenges will last for long periods. The rapid global spread of COVID-19 and lack of enough data regarding the virus pathogenicity multiplies the complexity and forced governments to react quickly against this pandemic. Stem cells represent a small fraction of cells located in different tissues. These cells play a critical role in the regeneration and restoration of injured sites. Because of their specific niche and a limited number of stem cells, the key question is whether there are different anti-viral mechanisms against viral infection notably COVID-19. Here, we aimed to highlight the intrinsic antiviral resistance in different stem cells against viral infection. These data could help us to understand the possible viral infections in different stem cells and the activation of specific molecular mechanisms upon viral entrance.Graphical

Preparing Chromatin and RNA from Rare Cell Types with Fluorescence-Activated Nuclear Sorting (FANS).

The application of fluorescent tags to generate cell type-specific translational and transcriptional reporter lines is routine in plants, but separation of different cell types for downstream analyses is hampered by the presence of cell walls and tight connections between cells. Enzymatic removal of cell walls induces a wound response, dedifferentiation, or reprogramming of the resulting protoplasts. Their osmotic and mechanical instability and their large size range are challenging for FACS, a flow -sorting procedure based on differential expression of fluorescent tags. In contrast, plant nuclei are relatively robust and easy to isolate. Here, we describe a protocol for fluorescence-activated nuclear sorting (FANS) that allows efficient purification of very few fluorescence-tagged nuclei from a large background of non-labeled tissue. Purified nuclei are suitable for genome, epigenome, transcriptome, or proteome analyses. We describe in detail how to analyze nuclear RNA and DNA methylation from sorted nuclei representing the limited number of stem cells in the shoot apical meristem of Arabidopsis.

CD24 and CD49f expressions of E14.5 mouse mammary anlagen cells define putative distribution of earlier embryonic mammary stem cell activities.

Stem cell biology offers promise for understanding the origins of the mammary gland. However, the distribution of mammary stem cell (MaSC) activities at earlier embryonic stages has not been fully identified. The markers for sorting adult MaSC, CD24, CD29, and CD49f have been applied to analyze fetal MaSCs. Here we explored mammary anlagen MaSCs by investigating the expression of CD24 and CD49f. According to the comparative analysis between adult mammary gland and fetal mammary anlagen, we found that fetal mouse mammary anlagen may possess a high percentage of potential MaSCs. Flow cytometry analysis revealed 2 distinct mammary anlagen populations: Lin-CD24med and Lin-CD24high. Sphere-forming and mammary repopulating assays confirmed that the stem cell activity of E14.5 mouse mammary anlagen was restricted to the Lin-CD24med cell population. Furthermore, CD24med mammary anlagen cells were separated into Lin-CD24medCD49f+ and Lin-CD24medCD49f- populations and identified, respectively. The results proved that the mammary anlagen Lin-CD24medCD49f+ cell population possesses more stem cell activities than the Lin-CD24medCD49f- cell population. However, a limited numbers of stem cells and large numbers of stromal cells were identified in mammary anlagen in the Lin-CD24med cell population.

Concentrated Bone Marrow Aspirate for the Treatment of Chondral Injuries and Osteoarthritis of the Knee: A Systematic Review of Outcomes.

Background:Bone marrow aspirate concentrate (BMAC) has emerged as a novel treatment for pathology of the knee. Despite containing a limited number of stem cells, BMAC serves as a source of growth factors that are thought to play an important role as a result of their anabolic and anti-inflammatory effects. To our knowledge, there is no systematic review regarding the outcomes of bone marrow aspirate concentrate used for the treatment of chondral defects and osteoarthritis of the knee.Purpose:To perform a systematic review on the outcomes of bone marrow aspirate concentrate for the treatment of chondral defects and osteoarthritis of the knee.Study Design:Systematic review; Level of evidence, 4.Methods:A systematic review of the literature was performed using the Cochrane Database of Systematic Reviews, the Cochrane Central Register of Controlled Trials, PubMed, and MEDLINE from 1980 to present. Inclusion criteria were as follows: use of BMAC for treatment of chondral defects and osteoarthritis of the knee, English language, and human studies. We excluded cadaveric studies, animal studies, basic science articles, editorial articles, surveys, and studies that did not include the knee. After applying inclusion and exclusion criteria, studies were evaluated for efficacy and safety of BMAC for treatment of articular cartilage knee pathologies.Results:Eleven studies were considered. Of these, 5 were prospective studies, 1 was a retrospective study, 2 were case series, and 3 were case reports. Three comparative studies (2 with level 2 evidence, 1 with level 3 evidence) were found in our search; none of them were randomized. Three studies investigated the clinical efficacy of BMAC in the treatment of osteoarthritis, and 8 studies evaluated the efficacy of BMAC on focal cartilage injuries. All 3 studies regarding osteoarthritis and all 8 studies regarding focal chondral defects reported good to excellent overall outcomes with the use of BMAC.Conclusion:Although a growing interest for biological alternatives of treating knee pathology has been observed in the past few years, there still remains a paucity of high-quality studies. The studies included in this systematic review reported varying degrees of beneficial results with the use of BMAC with and without an additional procedure for the treatment of chondral defects and early stages of osteoarthritis. Most articles present the use of BMAC as a safe procedure and report good results.

The effects of commonly used intravitreal steroids on proliferation index of ciliary body-derived mesenchymal stem cells: an in vitro study

Aim: To investigate the effects of commonly used intravitreal steroids on survival and proliferation (namely, proliferation index) of ciliary body-derived mesenchymal stem cells (CB-MSC).Methods: CB-MSCs were isolated from newborn rats’ eye, and they were expanded in the medium. Commonly used intravitreal steroids such as dexamethasone (Dex) and triamcinolone acetonide (TA) were added into the medium at commonly used concentration in clinical practice (0.1 mg/mL) and at lower concentration (0.01 mg/mL). Proliferation indexes of CB-MSCs were analyzed with the xCELLigence system at nine consecutive times (at 3rd, 6th, 21th, 30th, 45th, 60th, 75th, 90th and 100th h).Results: Both TA and Dex at both 0.01 mg/mL and 0.1 mg/mL concentrations had negative effect on proliferation indexes of CB-MSC. Although negative effect of TA on proliferation index of CB-MSC at both concentrations was not statistically significant, statistically significant negative effect of Dex at 0.01 mg/mL concentration started 60th h (p = 0.017) and 0.1 mg/mL concentration started 30th h (p = 0.014).Discussion: Even therapeutic doses of intravitreal corticosteroid agents might have negative effects on limited numbers of stem cells. Especially, Dex caused statistically significant toxic effects on CB-MSCs even at lower concentrations of those used clinically. These novel findings deserve further in vivo investigations.

Neuroprotective effects of intravitreally transplanted adipose tissue and bone marrow–derived mesenchymal stem cells in an experimental ocular hypertension model

Background aimsThe purpose of this study was to investigate the neuroprotective effects of bone marrow bone marrow–derived and adipose tissue–derived mesenchymal stromal cells (MSCs) that were intravitreally transplanted in an experimental ocular hypertension (OHT) model. MethodsAn OHT rat model was generated by means of intracameral injection of hyaluronic acid into the anterior chamber. MSCs labeled with green fluorescence protein were transplanted intravitreally 1 week after OHT induction. At the end of the second and fourth weeks, retinal ganglion cells were visualized with the use of a flat-mount retina method and were evaluated by means of immunofluorescence staining against green fluorescence protein, vimentin, CD105, and cytokines (interleukin [IL]-1Ra, prostaglandin E2 receptor, IL-6, transforming growth factor-β1, interferon-γ and tumor necrosis factor-α). ResultsThe retinal ganglion cell numbers per area were significantly improved in stem cell–treated OHT groups compared with that in the non-treated OHT group (P < 0.05). The results of immunohistochemical analyses indicated that a limited number of stem cells had integrated into the ganglion cell layer and the inner nuclear layer. The number of cells expressing proinflammatory cytokines (interferon-γ and tumor necrosis factor-α) decreased in the MSC-transferred group compared with that in the OHT group after 4 weeks (P < 0.01). On the other hand, IL-1Ra and prostaglandin E2 receptor expressions were increased in the rat bone marrow–derived MSC group but were more significant in the rat adipose tissue–derived MSC group (P < 0.01). ConclusionsAfter intravitreal transplantation, MSCs showed a neuroprotective effect in the rat OHT model. Therefore, MSCs promise an alternative therapy approach for functional recovery in the treatment of glaucoma.

Abstract SY31-02: Building highly effective anti-tumor T cells

Abstract Many tumors contain relatively undifferentiated transformed progenitors termed ‘cancer stem cells’ (CSC). Stem cells are defined by the ability to self-renew and generate differentiated progeny, qualities maintained by evolutionarily conserved pathways that can lead to cancer when deregulated. The existence of stem cells has been documented across multiple tissues both during development on into adulthood including the hematopoietic and central nervous systems, intestine, skin, cardiac muscle and lung. All of these sites employ the presence of limited numbers of stem cells in tissues enable a few daughter cells to retain a regenerative potential while the majority of daughter cells enter an irreversible process of differentiation culminating in the generation of specialized cell types. Recently, we have found evidence supporting the existence and function of stem cells within the adaptive immune system in both the CD8+ and CD4+ T cell lineages. This stem cell-like population has been termed ‘T memory stem cells’ (TSCM). In preclinical models using both mouse and human cells, TSCM can overcome many limitations of currently employed T cell-based immunotherapies, especially in the setting of adoptive T cell transfer, including inefficient T cell engraftment, persistence, and ability to mediate prolonged immune attack. Conferring stemness to anti-tumour T cells might unleash the full potential of cellular therapies. Our goal is to trigger stem cell-like pathways in tumor-reactive T cells or to reprogram terminally differentiated tumor-infiltrating lymphocytes (TILs) to confer stem cell-like properties to augment immunotherapies against cancer and CSC, akin to fighting ‘fire with fire.’ In new work from our laboratory (1) as well as unpublished experiments, we have identified several candidate transcription factors that are differentially expressed in stem cell-like T cells. In addition, new experiments have shed significant light on the metabolic properties of highly effective stem cell-like T cell subsets. Reference 1. Roychoudhuri, R, et al, Nature, 2013. Citation Format: Nicholas P. Restifo. Building highly effective anti-tumor T cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr SY31-02. doi:10.1158/1538-7445.AM2014-SY31-02

Angiopoietin-like protein 3 promotes preservation of stemness during ex vivo expansion of murine hematopoietic stem cells.

Allogeneic hematopoietic stem cell (HSC) transplantations from umbilical cord blood or autologous HSCs for gene therapy purposes are hampered by limited number of stem cells. To test the ability to expand HSCs in vitro prior to transplantation, two growth factor cocktails containing stem cell factor, thrombopoietin, fms-related tyrosine kinase-3 ligand (STF) or stem cell factor, thrombopoietin, insulin-like growth factor-2, fibroblast growth factor-1 (STIF) either with or without the addition of angiopoietin-like protein-3 (Angptl3) were used. Culturing HSCs in STF and STIF media for 7 days expanded long-term repopulating stem cells content in vivo by ∼6-fold and ∼10-fold compared to freshly isolated stem cells. Addition of Angptl3 resulted in increased expansion of these populations by ∼17-fold and ∼32-fold, respectively, and was further supported by enforced expression of Angptl3 in HSCs through lentiviral transduction that also promoted HSC expansion. As expansion of highly purified lineage-negative, Sca-1+, c-Kit+ HSCs was less efficient than less pure lineage-negative HSCs, Angptl3 may have a direct effect on HCS but also an indirect effect on accessory cells that support HSC expansion. No evidence for leukemia or toxicity was found during long-term follow up of mice transplanted with ex vivo expanded HSCs or manipulated HSC populations that expressed Angptl3. We conclude that the cytokine combinations used in this study to expand HSCs ex vivo enhances the engraftment in vivo. This has important implications for allogeneic umbilical cord-blood derived HSC transplantations and autologous HSC applications including gene therapy.

Comparing the cultivated cochlear cells derived from neonatal and adult mouse

BackgroundPrevious reports showed the presence of limited numbers of stem cells in neonatal murine cochlear sensory epithelia and these cells are progressively lost during the postnatal development. The goal of this study was to investigate whether stem cells can be derived from mature mouse cochleae under suspension culture conditions, and to analyze the expression of the stem cell and inner ear progenitor cell markers in cells dissociated from neonatal and adult mouse organs of Corti.MethodsOrgans of Corti were dissected from postnatal day 1 (P1) or postnatal day 60 (P60) mouse. The dissociated cells were cultivated under suspension cultures conditions. Reverse transcription-polymerase chain reaction (RT-PCR) and immunocytochemistry were conducted for phenotype characterization.ResultsThe number of cochlear stem cells (otospheres) yielded from P1 organ of Corti was significantly higher than that of the P60 organ of Corti. RT-PCR analyses showed that the stem markers, such as nanog, sox2, klf4, and nestin can be found to be distributed similarly in the cells derived from both of organisms, but the inner ear developmental/progenitor cell markers showed lower expression in P60 organ of Corti compared to P1. Immunocytochemistry results also revealed the evidence that P60 otospheres lacking of differentiation potential in vitro, which opposed to the strong differentiation potential of otospheres at P1 stage.ConclusionsOur findings suggest that the loss of numbers and features of stem cells in the adult organ of Corti is associated with the substantial down-regulation of inner ear progenitor key-markers during maturation of the cells in organ of Corti.

GSK‐3β inhibition promotes early engraftment of ex vivo‐expanded haematopoietic stem cells

Umbilical cord blood (UCB) is a source of stem cells used for allogeneic transplantation, in addition to bone marrow and peripheral blood. Limited numbers of stem cells in a single UCB unit is associated with slow haematopoietic recovery and high risk of graft failure, particularly in adult patients. UCB stem cells can be expanded ex vivo; however, rapid differentiation reduces their regenerative potential. We have recently shown that Wnt/β-catenin signalling is down-regulated in ex vivo-expanded stem cells; therefore, we propose that re-activation of Wnt signalling using GSK-3β inhibition may act to improve regenerative potential of these ex vivo-expanded stem cells. Immunocompromised mice were employed in transplantation studies to determine stem-cell engraftment. Flow cytometry was used to phenotype the engrafted human cells. Retroviral gene transfer was used to examine the role of Myc gene up-regulated by GSK-3β inhibition, in ex vivo-expanded stem cells. Treatment with GSK-3β inhibitor, 6-bromoindirubin 3'-oxime (BIO) improved early human cell engraftment in the mice and elevated the numbers of myeloid progenitor cells in cytokine-stimulated culture. BIO up-regulated β-catenin and c-myc in ex vivo-expanded stem cells. Ectopic expression of Myc acted to increase clonogenic potential and to delay differentiation of haematopoietic progenitor cells, suggesting the potential mechanism to improve regenerative potential of ex vivo-expanded grafts. Pharmacological inhibition of GSK-3β provided a novel approach to improve early engraftment of ex vivo-expanded haematopoietic progenitor cells.

Bioreactor design for clinical‐grade expansion of stem cells

The many clinical trials currently in progress will likely lead to the widespread use of stem cell-based therapies for an extensive variety of diseases, either in autologous or allogeneic settings. With the current pace of progress, in a few years' time, the field of stem cell-based therapy should be able to respond to the market demand for safe, robust and clinically efficient stem cell-based therapeutics. Due to the limited number of stem cells that can be obtained from a single donor, one of the major challenges on the roadmap for regulatory approval of such medicinal products is the expansion of stem cells using Good Manufacturing Practices (GMP)-compliant culture systems. In fact, manufacturing costs, which include production and quality control procedures, may be the main hurdle for developing cost-effective stem cell therapies. Bioreactors provide a viable alternative to the traditional static culture systems in that bioreactors provide the required scalability, incorporate monitoring and control tools, and possess the operational flexibility to be adapted to the differing requirements imposed by various clinical applications. Bioreactor systems face a number of issues when incorporated into stem cell expansion protocols, both during development at the research level and when bioreactors are used in on-going clinical trials. This review provides an overview of the issues that must be confronted during the development of GMP-compliant bioreactors systems used to support the various clinical applications employing stem cells.

Umbilical Cord Blood: Medical Waste or Important Source of Stem Cells?

The identification of new sources of stem cells may provide significant clinical benefits in the regenerative medicine. Although sometimes still regarded as a medical waste, the blood remained in the umbilical vein after birth (Umbilical Cord Blood, UCB) has become a valuable alternative source of haematopoietic stem cells for the treatment of various disorders. Using UCB is advantageous because it is obtained by a simple, safe and painless procedure when the baby is delivered. The immaturity of UCB cells resulted in a reduced graft-versus-host disease when compared to bone marrow grafts. Furthermore, there may be particular utility in using UCB in the context of HLA mismatch between available donor and recipient. However, due to the limited number of stem cells, the progress in the field has been largely restricted to children. Nevertheless, evidence supporting the efficacy of a double transplant of UCB in adults has significantly increased over the past years, as an alternative to bone marrow transplantation in those adult patients where no compatible donors are available. Today, new parents may choose to have the UCB stored in a stem cell bank. These banks can be public (non-profit) or private (for-profit). The public banks store UCB from donors and provide it when transplantation is prescribed to an unrelated patient. Unfortunately, donation to a public bank is not possible everywhere, although their number is growing. On contrary, the private banks offer a commercial service to parents in order to preserve the UCB for future needs of their child. Storing UCB in such private banks is controversial and recommended only in the case of historical existence of a genetic disease; otherwise the likelihood of stored UCB being used in autologous transplant is negligible. This and other ethical aspects of UCB banking are discussed in this paper.

Phase II Trial of Parathyroid Hormone after Double Umbilical Cord Blood Transplantation

Transplantation of 1 or 2 umbilical cord blood products is a useful alternative stem cell source. However, the limited number of stem cells in each infusion results in slow engraftment. In mouse models, administration of parathyroid hormone (PTH) is an effective way to enhance the ability of limited numbers of hematopoietic stem cells to support hematopoiesis. In this study, patients received either a myeloablative or a reduced-intensity double umbilical cord blood transplantation, followed by PTH at 100 μg/day for 28 days. Thirteen patients (median age, 42 years) were enrolled. All patients engrafted; the median time to neutrophil and platelet engraftment of >20 × 10(9) cells/L was 30 days and 61 days, respectively. The incidence of grade II-IV acute GVHD was 38.5% at day 100. Four deaths occurred before day 100, prompting early study closure. No patient who received a myeloablative regimen relapsed. Overall survival at 6 months after transplantation was 62%, and disease-free survival at 2 years was 39%. At the dose and schedule studied, there was no evidence that PTH influenced blood count recovery.

Haplotype-Based Banking of Human Pluripotent Stem Cells for Transplantation: Potential and Limitations

High expectations surround the area of stem cells therapeutics. However, the cells' source-adult or embryonic-and the cells' origin-patient-derived autologous or healthy donor genetically unrelated-remain subjects of debate. Autologous origins have the advantage of a theoretical absence of immune rejection by the recipient. However, this approach has several limitations with regard to the disease of the recipient and to potential problems with the generation, expansion, and manipulation of autologous induced pluripotent stem cells (iPS cells) preparation. An alternative to using autologous cells is the establishment of a bank of well-characterized adult cells that would be used to generate iPS cells and their derivatives. In the context of transplantation, such cells would come from genetically unrelated donors and the immune system of the recipient would reject the graft without immunosuppressive therapy. To minimize the risk of rejection, human leukocyte antigen (HLA) compatibility is certainly the best option, and the establishment of an HLA-organized bank would mean having a limited number of stem cells that would be sufficient for a large number of recipients. The concept of haplobanking with HLA homozygous cell lines would also limit the number of HLA mismatches, but such an approach will not necessarily be less immunogenic in terms of selection criteria, because of the limited number of HLA-compatible loci and the level of HLA typing resolution.

Sparc Is Dispensable for Murine Hematopoiesis, Despite Its Suspected Role in 5q- Myelodysplastic Syndrome

Abstract 4822Hematopoiesis is a complex process where a limited number of stem cells give rise to all mature blood cells. It involves interplay of several factors, many of which are yet to be identified. In a search for novel regulators of hematopoiesis, we chose to study SPARC (Secreted Protein Acidic and Rich in Cysteine, also known as Osteonection and BM40) because it is downregulated upon hematopoietic differentiation (Bruno et al., Mol Cell Biol, 2004) and might therefore play a role in the regulation of hematopoietic stem cells (HSC). SPARC is a matricellular protein that forms a major component of bone and is ubiquitously expressed in a variety of tissues. It is the founding member of a family of SPARC-like proteins. Several publications have indicated an important role for SPARC in hematopoiesis. In particular – knockdown of SPARC in zebrafish embryos resulted in an altered number of circulating blood cells, and a knockout mouse model showed thrombocytopenia and reduced erythroid colony formation.We carried out an in depth phenotypic and functional analysis of the hematopoietic system of SPARC knockout mice; using it as a model to gain insight into the role of SPARC in hematopoiesis. These mice are viable and fertile but show severe osteopenia and age-onset cataract at about six months of age. They also show an altered response to tumour growth and wound healing. We used mice (129SVJ background) (Gilmour et al. EMBO, 1998) that were less than six months old. These mice had normal peripheral blood counts and the bone marrow and spleen showed no alterations in morphology or cellularity. A detailed phenotypic analysis of precursors within the bone marrow showed no significant differences in myelo-erythroid precursors as compared to wild types (n=6). Though in vitro, the precursors showed lower ability to form BFU-E (n=5, p=0.048).In transplantations of lethally irradiated recipient mice, SPARC knockout cells gave rise to multi-lineage long-term reconstitution. Also, when competed with wild type cells, they provided reconstitution as well as their wild type counterparts. When SPARC knockout mice (n=8) were transplanted with wild type cells, there was normal reconstitution, indicating that a SPARC deficient niche can fully support normal hematopoiesis.We also tested if SPARC deficient mice respond differently to hematopoietic stress. We subjected mice (n=7) to sub lethal dose of irradiation and to experimentally induced anemia (n=7) and followed recovery by analyzing peripheral blood counts. In both SPARC knockouts and wild type mice, the blood counts recovered in a similar fashion.In conclusion, we find that SPARC is dispensable for murine hematopoiesis. It is possible that there are compensatory mechanisms involving other members of the SPARC family that ultimately lead to normal hematopoiesis in the murine model.In humans, SPARC maps to the deleted region in 5q MDS and has been reported to be 71 % down regulated in patient samples (Lehmann et al. Leukemia, 2007). It is the most prominent gene that is up regulated in response to lenalidomide, a drug that inhibits the malignant clone (Pellagatti et al. PNAS, 2007). SPARC is thus increasingly speculated to be involved in the pathophysiology of this hematopoetic disease. We analysed the expression levels of SPARC mRNA in the hematopoietic stem/progenitor cell compartment and found high expression levels in the CD34+ fraction of human cord blood cells. In contrast, there is very low level of SPARC expression in all compartments of murine HSCs. Therefore SPARC function may play a more important role in human hematopoiesis than in murine blood cell regulation. Disclosures:No relevant conflicts of interest to declare.

In vitro expansion of human adipose-derived stem cells in a spinner culture system using human extracellular matrix powders

Stem cell therapy requires large numbers of stem cells to replace damaged tissues, but only limited numbers of stem cells can be harvested from a single patient. To obtain large quantities of stem cells with differentiation potential, we explored a spinner culture system using human extracellular matrix (hECM) powders. The hECM was extracted from adipose tissue and fabricated into powders. Human adipose-derived stem cells (hASCs) were isolated, seeded on hECM powders, and cultivated in a spinner flask. The 3-D culture system, using hECM powders, was highly effective for promoting cell proliferation. The number of hASCs in the 3-D culture system significantly increased for 10days, resulting in an approximately 10-fold expansion, whereas a traditional 2-D culture system showed just a 2.8-fold expansion. Surface markers, transcriptional factors, and differentiation potential of hASCs were assayed to identify the characteristics of proliferated cells in 3-D culture system. The hASCs expressed the pluripotency markers, Oct-4 and Sox-2 during 3-D culture and retained their capacity to differentiate into adipogenic, osteogenic, and chondrogenic lineages. These findings demonstrate that the 3-D culture systems using hECM powders provide an efficient in vitro environment for stem cell proliferation, and could act as stem cell delivery carriers for autologous tissue engineering and cell therapy.

Cx3Dp: A Parallel Framework for Modeling the Growth and Development of Neural Tissue

Event Abstract Back to Event Cx3Dp: A Parallel Framework for Modeling the Growth and Development of Neural Tissue Andreas Hauri1* and Rodney Douglas1 1 UZH / ETHZ, Institute of Neuroinformatics, Switzerland The development of neural tissues such as the neocortex unfolds from a limited number of stem cells by successive mitosis, migration, differentiation and adaptation of cells that finally come to take up specific functional roles. Unlike self-assembly in which the necessary components are given, and assembly depends largely on complementary structure and forces of interaction. The self-construction of development makes explicit use of structural and organizational information encoded in the genome. We have been exploring this complex process by modeling and simulation. In a previous publication, we described a software framework (Cx3D), that enables the simulation of development in physical 3D environment that respects the physical of interaction of objects as well as diffusion of morphogens. (Zubler and Douglas, 2009). However, the success of that system in simulating development soon brought us to the limits of single-threaded, single-computer applications. To open the route to simulating larger cortical areas and their inter-areal connections, we require a more general version of Cx3D that can exploit multiprocessor/multicomputer systems. To this end we have designed a parallel version of Cx3D (Cx3Dp) that addresses these improvements and can also be run cross-platform. The system scales well with addition of processors and computers. Computers can be dynamically added to the simulation if the need for more computational power or more memory arises. The load balancing will acquire the new resources and balance the system in a way that each computer takes the same amount of time to complete one time step. The simulation can be saved after any time step, over multiple computers, and can be reloaded and continued again if necessary. The diffusion framework has also been redesigned. The diffusion is now implemented on an octree grid system that can be adapted to a desired resolution, according to the resolution necessary for the diffusion. Compared to the previous version of Cx3D. This improvement allows higher flexibility in assigning the computational resources to the diffusion. Currently we are running simulations of up to 200’000 cells and 400’000 cell compartments on 3 machines with 64 GB of memory and 24 cores each, using only 25% of the memory available. The system scales well in speed when adding cores, and in speed and memory on adding machines. The new Cx3Dp thus opens the way to very large scale simulation of developing systems on simple networks of consumer computers. Keywords: Large scale modeling Conference: 4th INCF Congress of Neuroinformatics, Boston, United States, 4 Sep - 6 Sep, 2011. Presentation Type: Demo Presentation Topic: Large scale modeling Citation: Hauri A and Douglas R (2011). Cx3Dp: A Parallel Framework for Modeling the Growth and Development of Neural Tissue. Front. Neuroinform. Conference Abstract: 4th INCF Congress of Neuroinformatics. doi: 10.3389/conf.fninf.2011.08.00106 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 17 Oct 2011; Published Online: 19 Oct 2011. * Correspondence: Dr. Andreas Hauri, UZH / ETHZ, Institute of Neuroinformatics, Zurich, Switzerland, haurian@ini.ch Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Andreas Hauri Rodney Douglas Google Andreas Hauri Rodney Douglas Google Scholar Andreas Hauri Rodney Douglas PubMed Andreas Hauri Rodney Douglas Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.