Exploration of Preservation Methods for Utilizing Porcine Fetal-Organ-Derived Cells in Regenerative Medicine Research.

Human pluripotent stem cells have been employed in generating organoids, yet their immaturity compared to fetal organs and the limited induction of all constituent cell types remain challenges. Porcine fetal progenitor cells have emerged as promising candidates for co-culturing with human progenitor cells in regeneration and xenotransplantation research. This study focused on identifying proper preservation methods for porcine fetal kidneys, hearts, and livers, aiming to optimize their potential as cell sources. Extracted from fetal microminiature pigs, these organs were dissociated before and after cryopreservation-thawing, with subsequent cell quality evaluations. Kidney cells, dissociated and aggregated after vitrification in a whole-organ form, were successfully differentiated into glomeruli and tubules in vivo. In contrast, freezing hearts and livers before dissociation yielded suboptimal results. Heart cells, frozen after dissociation, exhibited pulsating heart muscle cells similar to non-frozen hearts. As for liver cells, we developed a direct tissue perfusion technique and successfully obtained highly viable liver parenchymal cells. Freezing dissociated liver cells, although inferior to their non-frozen counterparts, maintained the ability for colony formation. The findings of this study provide valuable insights into suitable preservation methods for porcine fetal cells from kidneys, hearts, and livers, contributing to the advancement of regeneration and xenotransplantation research.

The Characteristics and Function of Small Extracellular Vesicles Derived from Human Bone Marrow and Umbilical Cord Mesenchymal Stromal Cells Are Influenced by Cell Culture Conditions.

Extracellular vesicles (EVs) derived from mesenchymal stromal cells (MSC-EVs) have been proposed as a novel therapeutic tool with numerous clinically related advantages. However, their characteristics and functionality are dependent on the source of MSCs and their cell culture conditions. Fetal bovine serum (FBS) provides a source of nutrients and growth factors to the cultured cells. However, certain pitfalls are associated with its supplementation to the culture media, including introduction of exogenous FBS-derived EVs to the cultured cells. Thus, recent practices recommend utilization of serum-free (SF) media or EV-depleted FBS. On the contrary, evidence suggests that the immunomodulatory ability of MSC-EVs can be improved by exposing MSCs to an inflammatory (IF) environment. The objective of this study was to (1) compare EVs isolated from two tissue sources of MSCs that were exposed to various cell culture conditions and (2) to evaluate their anti-inflammatory effects. Bone marrow-derived mesenchymal stromal cells (BM-MSCs) and umbilical cord-derived mesenchymal stromal cells (UC-MSCs) were exposed to either a SF media environment, an IF environment, or media supplemented with 5% EV-depleted FBS. Following isolation of MSC-EVs, the isolates were quantified and evaluated for particle size, phenotypic changes, and their immunomodulatory potential. A statistically significant difference was not identified on the yield and protein concentration of different isolates of EVs from BM-MSCs and UC-MSCs, and all isolates had a circular appearance as evaluated via electron microscopy. A significant difference was identified on the phenotype of different EVs isolates; however, all isolates expressed classical markers such as CD9, CD63, and CD81. The addition of BM-derived MSC-EVs from FBS environment or UC-derived MSC-EVs from IF environment resulted in statistically significant downregulation of IL-6 messenger RNA (mRNA) in stimulated leukocytes. This study confirms that EVs produced by different MSC sources and cell culture conditions affect their phenotype and their immunomodulatory capacities.

Mesenchymal Stem Cells Isolated from Equine Hair Follicles Using a Method of Air-Liquid Interface.

Equine mesenchymal stem cells (MSC) of various origins have been identified in horses, including MSCs from the bone marrow and adipose tissue. However, these stem cell sources are highly invasive in sampling, which thereby limits their clinical application in equine veterinary medicine. This study presents a novel method using an air-liquid interface to isolate stem cells from the hair follicle outer root sheath of the equine forehead skin. These stem cells cultured herewith showed high proliferation and asumed MSC phenotype by expressing MSC positive biomarkers (CD29, CD44 CD90) while not expressing negative markers (CD14, CD34 and CD45). They were capable of differentiating towards chondrogenic, osteogenic and adipogenic lineages, which was comparable with MSCs from adipose tissue. Due to their proliferative phenotype in vitro, MSC-like profile and differentiation capacities, we named them equine mesenchymal stem cells from the hair follicle outer root sheath (eMSCORS). eMSCORS present a promising alternative stem cell source for the equine veterinary medicine.

A comparative study on various cell sources for constructing tissue-engineered meniscus.

Injury to the meniscus is a common occurrence in the knee joint and its management remains a significant challenge in the clinic. Appropriate cell source is essential to cell-based tissue regeneration and cell therapy. Herein, three commonly used cell sources, namely, bone marrow mesenchymal stem cell (BMSC), adipose-derived stem cell (ADSC), and articular chondrocyte, were comparatively evaluated to determine their potential for engineered meniscus tissue in the absence of growth factor stimulus. Cells were seeded on electrospun nanofiber yarn scaffolds that share similar aligned fibrous configurations with native meniscus tissue for constructing meniscus tissue in vitro. Our results show that cells proliferated robustly along nanofiber yarns to form organized cell-scaffold constructs, which recapitulate the typical circumferential fiber bundles of native meniscus. Chondrocytes exhibited different proliferative characteristics and formed engineered tissues with distinct biochemical and biomechanical properties compared to BMSC and ADSC. Chondrocytes maintained good chondrogenesis gene expression profiles and produced significantly increased chondrogenic matrix and form mature cartilage-like tissue as revealed by typical cartilage lacunae. In contrast, stem cells underwent predominately fibroblastic differentiation and generated greater collagen, which contributes to improved tensile strengths of cell-scaffold constructs in comparison to the chondrocyte. ADSC showed greater proliferative activity and increased collagen production than BMSC. These findings indicate that chondrocytes are superior to stem cells for constructing chondrogenic tissues while the latter is feasible to form fibroblastic tissue. Combination of chondrocytes and stem cells might be a possible solution to construct fibrocartilage tissue and meniscus repair and regeneration.

Cartilage Tissue Engineering: An Introduction.

Once damaged, cartilage has limited healing capability. This has led to a huge body of research that aims to repair or regenerate this important tissue. Despite the progress made, significant hurdles still need to be overcome. This chapter highlights some of the progress made, while elaborating on areas that need further research. The concept of translation and the route to clinical translation must be kept in mind if some of the promising preclinical research is to make it to routine clinical application.

Advances in single-cell nanoencapsulation and applications in diseases.

Single-cell nanoencapsulation is a method of coating the surface of single cell with nanomaterials. In the early 20th century, with the introduction of various types of organic or inorganic nano-polymer materials, the selection of cell types, and the functional modification of the outer coating, this technology has gradually matured. Typical preparation methods include interfacial polycondensation, complex condensation, spray drying, microdroplet ejection, and layer-by-layer (LbL) self-assembly. The LbL assembly technology utilises nanomaterials with opposite charges deposited on cells by strong interaction (electrostatic interaction) or weak interaction (hydrogen bonding, hydrophobic interaction), which drives compounds to spontaneously form films with complete structure, stable performance and unique functions on cells. According to the needs of the disease, choosing appropriate cell types and biocompatible and biodegradable nanomaterials could achieve the purpose of promoting cell proliferation, immune isolation, reducing phagocytosis of the reticuloendothelial system, prolonging the circulation time in vivo, and avoiding repeated administration. Therefore, encapsulated cells could be utilised in various biomedical fields, such as cell catalysis, biotherapy, vaccine manufacturing and antitumor therapy. This article reviews cell nanoencapsulation therapies for diseases, including the various cell sources used, nanoencapsulation technology and the latest advances in preclinical and clinical research.

Extracellular Vesicles as Drug Delivery System for Treatment of Neurodegenerative Disorders: Optimization of the Cell Source.

Extracellular vesicles (EVs) represent a next generation drug delivery system that combines nanoparticle size with extraordinary ability to cross biological barriers, reduced immunogenicity, and low offsite toxicity profiles. A successful application of this natural way of delivering biological compounds requires deep understanding EVs intrinsic properties inherited from their parent cells. Herein, we evaluated EVs released by cells of different origin, with respect to drug delivery to the brain for treatment of neurodegenerative disorders. The morphology, size, and zeta potential of EVs secreted by primary macrophages (mEVs), neurons (nEVs), and astrocytes (aEVs) were examined by nanoparticle NTA, DLS, cryoTEM, and AFM. Spherical nanoparticles with average size 110-130 nm and zeta potential around -20 mV were identified for all EVs types. mEVs showed the highest levels of tetraspanins and integrins compared to nEVs and aEVs, suggesting superior adhesion and targeting to the inflamed tissues by mEVs. Strikingly, aEVs were preferentially taken up by neuronal cells in vitro, followed by mEVs and nEVs. Nevertheless, the brain accumulation levels of mEVs in a transgenic mouse model of Parkinson's disease were significantly higher than those of nEVs or aEVs. Therefore, mEVs were suggested as the most promising nanocarrier system for drug delivery to the brain.

The study on the function and cell source of interleukin-6 in interstitial cystitis/bladder painful syndrome rat model.

OBJECTIVE: The elevated expression of interleukin-6 (IL-6) in patients with interstitial cystitis/bladder painful syndrome (IC/BPS) has been demonstrated, but the role of IL-6 in IC/BPS and its source remain to be explored. METHODS: IC/BPS rat model was created in female rats by using long-term intermittent intravesical hyaluronidase (0.5 ml, 4 mg/ml). After modeling, IL-6 stimulation group, and anti-IL-6R group were treated with recombinant rat IL-6 and tocilizumab, respectively. Symptomatic changes were detected by Vonfrey pain score and urodynamics, and hematoxylin-eosin (HE) staining, mast cell staining and Masson staining were used to evaluate the changes of inflammation in the bladder tissue of rats. Cell sources of IL-6 was explored through enzyme linked immunosorbent assay (ELISA) test, reverse transcription polymerase chain reaction (RT-PCR), and western-blot test on the supernatant of coculturing rat bladder epithelial cells and rat macrophages. RESULTS: The Vonfrey pain scores of the model group and IL-6 stimulation group were significantly higher than those of the control group, while the anti-IL-6R group were significantly lower (p < .05). Compared with the blank control group, urodynamic results showed that the urination interval of the model group and IL-6 stimulation group was significantly shortened, and the maximum bladder capacity was significantly reduced (p < .05), and anti-IL-6R treatment significantly alleviated the inflammatory response of bladder tissue. The results of HE, Mast cell staining, and Masson staining showed that the inflammatory response of bladder tissue after anti-IL-6R treatment was significantly reduced. Through cells coculture, the relative expression of IL-6 from model group was found significantly higher than blank control group by RT-PCR, ELISA, and western blot test (p < .05). CONCLUSIONS: IL-6 played an essential role in the development of IC/BPS rat model as a proinflammation cytokine. Further evidence from coculture proved that macrophages are the cell resource of IL-6 in IC/BPS.

Osteogenic Differentiation Factors of Multipotent Mesenchymal Stromal Cells in the Current Understanding.

BACKGROUND: Molecular genetic mechanisms, signaling pathways, conditions, factors, and markers of the osteogenic differentiation of mesenchymal stem cells (MSCs) are being actively studied and are among the most studied areas in the field of cellular technology. This attention is largely due to the mounting contradictions in the seemingly classical knowledge and the constant updating of results in the analyzed areas. In this regard, we focus on the main classical concepts and some new factors and mechanisms that have a noticeable regulatory effect on the differentiation potential of postnatal MSCs. RESULTS: This review considers the importance of the sources of MSCs for the realization of their differentiation potential, molecular genetic factors and signaling pathways of MSC differentiation, the role of inflammatory cytokines and chemokines in osteogenesis, biomechanical signals, and the effect of conformational changes in the cellular cytoskeleton on MSC differentiation. CONCLUSION: It is concluded that it is necessary to move from studies focused on the effects of local genes to those taking multiple measurements of the gene-regulatory profile and the biomolecules critical for the implementation of numerous, incompletely studied osteogenic factors of endogenous and exogenous origin. Among the cornerstones of future (epi)genetic studies, whether osteomodulatory effects are realized through specific signaling pathways and/or whether cross-signaling with known genes drives the osteogenic differentiation of MSCs remains to be determined.

On the Quest for In Vitro Platelet Production by Re-Tailoring the Concepts of Megakaryocyte Differentiation.

The demand of platelet transfusions is steadily growing worldwide, inter-donor variation, donor dependency, or storability/viability being the main contributing factors to the current global, donor-dependent platelet concentrate shortage concern. In vitro platelet production has been proposed as a plausible alternative to cover, at least partially, the increasing demand. However, in practice, such a logical production strategy does not lack complexity, and hence, efforts are focused internationally on developing large scale industrial methods and technologies to provide efficient, viable, and functional platelet production. This would allow obtaining not only sufficient numbers of platelets but also functional ones fit for all clinical purposes and civil scenarios. In this review, we cover the evolution around the in vitro culture and differentiation of megakaryocytes into platelets, the progress made thus far to bring the culture concept from basic research towards good manufacturing practices certified production, and subsequent clinical trial studies. However, little is known about how these in vitro products should be stored or whether any safety measure should be implemented (e.g., pathogen reduction technology), as well as their quality assessment (how to isolate platelets from the rest of the culture cells, debris, microvesicles, or what their molecular and functional profile is). Importantly, we highlight how the scientific community has overcome the old dogmas and how the new perspectives influence the future of platelet-based therapy for transfusion purposes.

Exosomes from different cells: Characteristics, modifications, and therapeutic applications.

Exosomes are cystic vesicles secreted by living cells with a phospholipid bilayer membrane. Importantly, these vesicles could serve to carry lipids, proteins, genetic materials, and transmit biological information in vivo. The cell-specific proteins and genetic materials in exosomes are capable of reflecting their cell origin and physiological status. Based on the different tissues and cells (macrophage, dendritic cells, tumor cells, mesenchymal stem cells, various body fluids, and so on), exosomes exhibit different characteristics and functions. Furthermore, owing to their high delivery efficiency, biocompatibility, and multifunctional properties, exosomes are expected to become a new means of drug delivery, disease diagnosis, immunotherapy, and precise treatment. At the same time, in order to supplement or enhance the therapeutic applicability of exosomes, chemical or biological modifications can be used to broaden, change or improve their therapeutic capabilities. This review focuses on three aspects: the characteristics and original functions of exosomes secreted by different cells, the modification and transformation of exosomes, and the application of exosomes in different diseases.

Mesenchymal stem cells - a promising strategy for treating knee osteoarthritis.

AIMS: The purpose of our study was to determine whether mesenchymal stem cells (MSCs) are an effective and safe therapeutic agent for the treatment of knee osteoarthritis (OA), owing to their cartilage regeneration potential. METHODS: We searched PubMed, Embase, and the Cochrane Library, with keywords including "knee osteoarthritis" and "mesenchymal stem cells", up to June 2019. We selected randomized controlled trials (RCTs) that explored the use of MSCs to treat knee OA. The visual analogue scale (VAS), Western Ontario and McMaster University Osteoarthritis Index (WOMAC), adverse events, and the whole-organ MRI score (WORMS) were used as the primary evaluation tools in the studies. Our meta-analysis included a subgroup analysis of cell dose and cell source. RESULTS: Seven trials evaluating 256 patients were included in the meta-analysis. MSC treatment significantly improved the VAS (mean difference (MD), -13.24; 95% confidence intervals (CIs) -23.28 to -3.20, p = 0.010) and WOMAC (MD, -7.22; 95% CI -12.97 to -1.47, p = 0.010). The low-dose group with less than 30 million cells showed lower p-values for both the VAS and WOMAC. Adipose and umbilical cord-derived stem cells also had lower p-values for pain scores than those derived from bone marrow. CONCLUSION: Overall, MSC-based cell therapy is a relatively safe treatment that holds great potential for OA, evidenced by a positive effect on pain and knee function. Using low-dose (25 million) and adipose-derived stem cells is likely to achieve better results, but further research is needed. Cite this article: Bone Joint Res 2020;9(10):719-728.

Effect of Stem Cell Source and Dose on Allogeneic Hematopoietic Stem Cell Transplantation in Adult Patients with Idiopathic Aplastic Anemia: Data from the Korean Aplastic Anemia Trials.

OBJECTIVE: We aimed to evaluate the effect of stem cell source and dose on the survival of various donor subgroups, such as matched sibling donor (MSDs) and alternative donors (ADs), upon bone marrow (BM) or peripheral blood stem cell (PBSC) infusion in aplastic anemia (AA). METHODS: We retrospectively investigated the effects of stem cell source and dose on allogeneic hematopoietic stem cell transplantation (alloHSCT) in AA. RESULTS: A total of 267 patients were included in this analysis. The BM-treated group showed an association with low incidence of any-grade acute graft versus host disease (GvHD) (p < 0.001). A higher stem cell dose was related with a low incidence of extensive chronic GvHD in MSDs (p = 0.025). Multivariate analysis for overall survival (OS) revealed that only age at alloHSCT <31 years (p = 0.010) and prior platelet transfusion <86 U (p = 0.046) in MSDs and higher stem cell dose (hazard ratio = 2.596, p = 0.045) in ADs were favorable prognostic factors. CONCLUSION: PBSCs could be preferred in AD because high stem cell dose may be easily achieved to improve the OS at the expense of acute GvHD. However, BM stem cells are preferred in MSDs.

Influence of donor type, stem cell source and conditioning on outcomes after haploidentical transplant for lymphoma - a LWP-EBMT study.

Haploidentical stem cell transplantation (haploSCT) is becoming a major transplant modality for lymphoma. To assess the effects of donor characteristics, stem cell source and conditioning on outcomes, we identified 474 adults with Hodgkin (HL; 240), peripheral T-cell (PTCL; 88), diffuse large B-cell (77), mantle cell (40) or follicular lymphoma (FL; 29), who received haploSCT with post-transplant cyclophosphamide. Median follow-up of alive patients was 32 months. On multivariate analysis, acute graft-versus-host disease (GVHD) grade 2-4 was lower with offspring donors or bone marrow cells, whereas extensive chronic GVHD was higher in partial response at haploSCT or when using sisters, haploidentical donors beyond first degree, or female donors in male patients. Progression-free survival (PFS) was better for FL, HL and PTCL, whereas overall survival (OS) was better for HL and PTCL. Complete remission at haploSCT improved PFS and OS whereas these were negatively affected by cytomegalovirus donor positive/recipient positive status. No other donor characteristics (age, gender, human leucocyte antigen mismatch, ABO incompatibility) affected PFS or OS except use of haploidentical donors beyond first degree, which negatively affected OS. PFS and OS are mostly influenced by disease status and lymphoma subtype, supporting the use of any first degree haploidentical family member as a donor.

Methods to generate tissue-derived constructs for regenerative medicine applications.

The shortage of donor organs for transplantation remains a continued problem for patients with irreversible end-stage organ failure. Tissue engineering and regenerative medicine aims to develop therapies to provide viable solutions for these patients. Use of decellularized tissue scaffolds has emerged as an attractive approach to generate tissue constructs that mimic native tissue architecture and vascular networks. The process of decellularization which involves the removal of resident cellular components from donor tissues has been successfully translated to the clinic for applications in patients. However, transplantation of bioengineered solid organs using this approach remains a challenge as the process requires repopulating target cells to achieve functioning organs. This article presents a comprehensive overview of the methods used to achieve decellularization, the types of decellularizing agents, and the potential cell sources that could be used to achieve tissue function. Understanding the mechanism of action of the decellularizing agent and the processing methods will provide the optimal results for applications.

Pre-transplantation Risks and Transplant-Techniques in Haematopoietic Stem Cell Transplantation for Acute Leukaemia.

BACKGROUND: The role of conditioning intensity and stem cell source on modifying pre-transplantation risk in allogeneic haematopoietic stem cell transplantation (HSCT) is a matter of debate, but crucial when benchmarking centres. METHODS: This Retrospective, multicenter exploratory-validation analysis of 9103 patients, (55.5% male, median age 50 years; 1-75 years range) with an allogeneic HSCT between 2010 and 2016 from a matched sibling (N = 8641; 95%) or matched unrelated donor (N = 462; 5%) for acute myeloid (N = 6432; 71%) or acute lymphoblastic (N = 2671; 29%) leukaemia in first complete remission, and reported by 240 centres in 30 countries to the benchmark database of the European Society for Blood and Marrow Transplantation (EBMT) searched for factors associated with use of transplant techniques (standard N = 6375;70% or reduced intensity conditioning N = 2728;30%, respectively bone marrow N = 1945;21% or peripheral blood N = 7158;79% as stem cell source), and their impact on outcome. FINDINGS: Treatment groups differed significantly from baseline population (p < 0.001), and within groups regarding patient-, disease-, donor-, and centre-related pre-transplantation risk factors (p < 0.001); choice of technique did depend on pre-transplantation risk factors and centre (p < 0.001). Probability of overall survival at 5 years decreased systematically and significantly with increasing pre-transplantation risk score (score 2 vs 0/1 HR: 1·2, 95% c.i. [1·1-1·.3], p = 0.002; score 3 vs 0/1 HR: 1·5, 95% c.i. [1·3-1·7], p < 0.001; score 4/5/6 vs 0/1 HR: 1·9, 95% c.i. [1·6-2·2], p < 0.001) with no significant differences between treatment groups (likelihood ratio test on interaction: p = 0.40). Overall survival was significantly associated with selection steps and completeness of information (p < 0.001). INTERPRETATION: Patients' pre-transplantation risk factors determine survival, independent of transplant techniques. Transplant techniques should be regarded as centre policy, not stratification factor in benchmarking. Selection criteria and completeness of data bias outcome. Outcomes may be improved more effectively through better identifying pre-transplantation factors as opposed to refinement of transplant techniques. FUNDING: The study was funded by EBMT.

Immune reconstitution following hematopoietic stem cell transplantation using different stem cell sources.

Introduction: Adequate immune reconstitution post-HSCT is crucial for the success of transplantation, and can be affected by both patient- and transplant-related factors. Areas covered: A systematic literature search in PubMed, Scopus, and abstracts of international congresses is performed to investigate immune recovery posttransplant. In this review, we discuss the pattern of immune recovery in the post-transplant period focusing on the impact of stem cell source (bone marrow, peripheral blood stem cells, and cord blood) on immune recovery and HSCT outcome. We examine the impact of serotherapy on immune reconstitution and the need to tailor dosing of serotherapy agents when using different stem cell sources. We discuss new techniques being used particularly with cord blood and haploidentical grafts to improve immune recovery in each scenario. Expert opinion: Cord blood T cells provide a unique CD4+ biased immune reconstitution. Initial studies using targeted serotherapy with cord grafts showed improved immune recovery with limited alloreactivity. Two competing haploidentical approaches have developed in recent years including TCRαß/CD19 depleted grafts and post-cyclophosphamide haplo-HSCT. Both approaches have comparable survival rates with limited alloreactivity. However, delayed immune reconstitution is still an ongoing problem and could be improved by modified donor lymphocyte infusions from the same haploidentical donor.

Functional surfaces for efficient differentiation of neural stem/progenitor cells into dopaminergic neurons.

Various techniques and systems have been reported for the efficient differentiation of neural stem/progenitor cells into dopaminergic neurons. Although a comparatively high percentage of dopaminergic neurons can be obtained using these techniques, the differentiated cells display varied cellular phenotypes such as astrocytes and oligodendrocytes. Generation of highly pure dopaminergic neurons is important for cell-based therapy and in vitro evaluation of dopaminergic neuron function. In this study, we developed a culture surface anchored with several neurotrophic factors and a neuronal cell-adhesive protein for efficient differentiation of neural stem/progenitor cells into dopaminergic neurons. Oligohistidine-fused brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor, synthesized using genetic engineering, were co-immobilized on the surface via metal chelation. To facilitate cell adhesion, a cell-adhesive chimeric protein derived from laminin (LN-G) was also immobilized on the surface. Approximately 40% of the cells cultured for 14 days with these protein-immobilized substrates expressed tyrosine hydroxylase, a marker of dopaminergic neurons, with a three-fold increase in differentiation efficiency than that reported previously. In addition, the number of tyrosine hydroxylase-positive cells increased to approximately 80% of the culture after 30 days. These cells secreted dopamine and expressed dopaminergic neuron-specific genes. Interestingly, cell types (glial cells and oligodendrocytes) other than neuronal cells (immature and mature dopaminergic neurons) were not detected on the protein-anchored surface. Our results demonstrate that highly pure dopaminergic neurons can be exclusively obtained using the novel substrate without extra purification steps such as cell sorting. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 107A: 860-871, 2019.

Peripheral Blood or Bone Marrow Stem Cells? Practical Considerations in Hematopoietic Stem Cell Transplantation.

Although peripheral blood stem cells (PBSC) have worldwide become the predominant source of progenitor cells for hematopoietic stem cell transplantation (HSCT), debate about their role compared with bone marrow (BM) has recently intensified, in large part based on the results of a multicenter Clinical Trials Network study which showed lower incidence of chronic graft-versus-host disease (cGVHD) and improved quality of life in recipients of myeloablative HLA-matched unrelated BM compared with PBSC transplants. However, in certain patient populations, PBSC may lead to improved clinical outcomes due to faster hematologic recovery, a lower risk of graft failure, and possibly a lower probability of relapse. This review will provide a comprehensive summary of studies comparing PBSC with BM as the graft source in terms of acute and chronic GVHD incidence, time to engraftment, and disease-free and overall survival probabilities after HLA-matched related and unrelated donor transplantation and haploidentical donor transplantation. Recommendations based on these studies regarding the use of PBSC versus BM for HSCT are offered.

Explant culture of oral mucosal epithelial cells for fabricating transplantable epithelial cell sheet.

INTRODUCTION: Carrier-free autologous mucosal epithelial cell sheets have been clinically utilized as a cell therapy for various epithelial disorders. Fabrication of a transplantable oral mucosal epithelial cell sheet without mouse feeder layers requires a higher seeding density than that of a sheet with mouse feeder layer culture; therefore, a large amount of donor mucosal tissue is needed. However, cell grafts co-cultured with mouse feeder layers are classified by the US Food and Drug Administration (FDA) as xenogeneic products. The goal of this study was to evaluate the utility of oral mucosal epithelial cells expanded by primary explant culture for the fabrication of an adequate number of transplantable epithelial cell sheets without mouse feeder layers. METHODS: Small fragments derived from minced oral mucosal tissue were placed into culture dishes for primary explant culture in keratinocyte culture medium. After primary explant culture, the outgrown cells were treated with trypsin-EDTA and were seeded on a temperature-responsive cell culture insert. After subculture, the cultured cells were harvested as a confluent cell sheet from the culture vessel by temperature reduction. RESULTS: Carrier-free human oral mucosal epithelial cell sheets were fabricated in all human cases, and autologous transplantation of the harvested cell sheets showed rapid epithelial regeneration to cover epithelial defects in a rabbit model. The explant culture method, involving the use of small fragments for primary culture, was sufficient for preparing a large number of mucosal epithelial cells without mouse feeder layers. Moreover, oral mucosal epithelial cells derived from the primary explant culture after cryopreservation allowed for the fabrication of cell sheets. CONCLUSIONS: This method for fabricating transplantable oral mucosal epithelial cell sheets is an attractive technique for regenerative medicine. It offers a patient-friendly manufacturing method in which a small amount of biopsy material from the patient represents a sufficient epithelial cell source, and a manufacturing plan for preparing cell grafts can be easily tailored.