Transplantation of Autologous Adipose Stem Cells Lacks Therapeutic Efficacy in the Experimental Autoimmune Encephalomyelitis Model

Multiple sclerosis (MS), characterized by chronic inflammation, demyelination, and axonal damage, is a complicated neurological disease of the human central nervous system. Recent interest in adipose stromal/stem cell (ASCs) for the treatment of CNS diseases has promoted further investigation in order to identify the most suitable ASCs. To investigate whether MS affects the biologic properties of ASCs and whether autologous ASCs from MS-affected sources could serve as an effective source for stem cell therapy, cells were isolated from subcutaneous inguinal fat pads of mice with established experimental autoimmune encephalomyelitis (EAE), a murine model of MS. ASCs from EAE mice and their syngeneic wildtype mice were cultured, expanded, and characterized for their cell morphology, surface antigen expression, osteogenic and adipogenic differentiation, colony forming units, and inflammatory cytokine and chemokine levels in vitro. Furthermore, the therapeutic efficacy of the cells was assessed in vivo by transplantation into EAE mice. The results indicated that the ASCs from EAE mice displayed a normal phenotype, typical MSC surface antigen expression, and in vitro osteogenic and adipogenic differentiation capacity, while their osteogenic differentiation capacity was reduced in comparison with their unafflicted control mice. The ASCs from EAE mice also demonstrated increased expression of pro-inflammatory cytokines and chemokines, specifically an elevation in the expression of monocyte chemoattractant protein-1 and keratin chemoattractant. In vivo, infusion of wild type ASCs significantly ameliorate the disease course, autoimmune mediated demyelination and cell infiltration through the regulation of the inflammatory responses, however, mice treated with autologous ASCs showed no therapeutic improvement on the disease progression

Age of the Donor Reduces the Ability of Human Adipose-Derived Stem Cells to Alleviate Symptoms in the Experimental Autoimmune Encephalomyelitis Mouse Model

There is a significant clinical need for effective therapies for primary progressive multiple sclerosis, which presents later in life (i.e., older than 50 years) and has symptoms that increase in severity without remission. With autologous mesenchymal stem cell therapy now in the early phases of clinical trials for all forms of multiple sclerosis (MS), it is necessary to determine whether autologous stem cells from older donors have therapeutic effectiveness. In this study, the therapeutic efficacy of human adipose-derived mesenchymal stem cells (ASCs) from older donors was directly compared with that of cells from younger donors for disease prevention. Mice were induced with chronic experimental autoimmune encephalomyelitis (EAE) using the myelin oligodendrocyte glycoprotein35-55 peptide and treated before disease onset with ASCs derived from younger ( \u3c 35 years) or older ( \u3e 60 years) donors. ASCs from older donors failed to ameliorate the neurodegeneration associated with EAE, and mice treated with older donor cells had increased central nervous system inflammation, demyelination, and splenocyte proliferation in vitro compared with the mice receiving cells from younger donors. Therefore, the results of this study demonstrated that donor age significantly affects the ability of human ASCs to provide neuroprotection, immunomodulation, and/or remyelination in EAE mice. The age-related therapeutic differences corroborate recent findings that biologic aging occurs in stem cells, and the differences are supported by evidence in this study that older ASCs, compared with younger donor cells, secrete less hepatocyte growth factor and other bioactive molecules when stimulated in vitro. These results highlight the need for evaluation of autologous ASCs derived from older patients when used as therapy for MS

Obesity inhibits the osteogenic differentiation of human adipose-derived stem cells

Additional file 3: Figure S3. No observable differences in lnASCs and obASCs during early bone regeneration. Critical size calvarial defects were created in the parietal bone of nude mice and assessed after 2 weeks. (A) Representative images of microCT scanning. (B) Quantification of microCT. Scale bar represents 1 mm. Bars, Âą SEM

Barriers to effective discharge planning: a qualitative study investigating the perspectives of frontline healthcare professionals

<p>Abstract</p> <p>Background</p> <p>Studies have shown that effective discharge planning is one of the key factors related to the quality of inpatient care and unnecessary hospital readmission. The perception and understanding of hospital discharge by health professionals is important in developing effective discharge planning. The aims of this present study were to explore the perceived quality of current hospital discharge from the perspective of health service providers and to identify barriers to effective discharge planning in Hong Kong.</p> <p>Methods</p> <p>Focus groups interviews were conducted with different healthcare professionals who were currently responsible for coordinating the discharge planning process in the public hospitals. The discussion covered three main areas: current practice on hospital discharge, barriers to effective hospital discharge, and suggested structures and process for an effective discharge planning system.</p> <p>Results</p> <p>Participants highlighted that there was no standardized hospital-wide discharge planning and policy-driven approach in public health sector in Hong Kong. Potential barriers included lack of standardized policy-driven discharge planning program, and lack of communication and coordination among different health service providers and patients in both acute and sub-acute care provisions which were identified as mainly systemic issues. Improving the quality of hospital discharge was suggested, including a multidisciplinary approach with clearly identified roles among healthcare professionals. Enhancement of health professionals' communication skills and knowledge of patient psychosocial needs were also suggested.</p> <p>Conclusions</p> <p>A systematic approach to develop the structure and key processes of the discharge planning system is critical in ensuring the quality of care and maximizing organization effectiveness. In this study, important views on barriers experienced in hospital discharge were provided. Suggestions for building a comprehensive, system-wide, and policy-driven discharge planning process with clearly identified staff roles were raised. Communication and coordination across various healthcare parties and provisions were also suggested to be a key focus.</p

Characterization and response to inflammatory stimulation of human endometrial-derived mesenchymal stem/stromal cells

The human endometrium has emerged as an attractive source of endometrial-derived mesenchymal stem/stromal cells (eMSCs) that can be easily isolated by non-invasive procedures. The prominent capacity of the endometrium for efficient and scarless regeneration each menstrual cycle indicates the increased eMSC immunomodulatory and pro-angiogenic properties. Herein the authors investigated the molecular responses of eMSCs to an inflammatory environment and whether those intrinsic responses affected their functional attributes. Human eMSCs immunophenotypic, transcriptional and secretory profiles were evaluated at passage three (P3) and passage eight (P8) to determine culture effects. Functionally, P3 and P8 non-induced and TNF-α/IFN-γ-induced eMSCs were interrogated for their capacity to suppress stimulated peripheral blood mononuclear cell (PBMC) proliferation, whereas non-induced eMSCs were assessed for their support to vascular network formation in co-cultures with human umbilical vein endothelial cells in vitro. Non-induced P3 and P8 eMSCs exhibited similar spindle-shaped morphology and clonogenic capacity. Nevertheless, P8 eMSCs showed reduced growth rate capacity and telomere length. The eMSCs displayed the typical MSC-related immunophenotypic profile, with P3 and P8 eMSCs expressing high levels (>98%) of CD140β, intermediate levels (35–60%) of CD146 and SUSD2 and low levels (∼8%) of NG2 pericytic markers. Non-induced P3 and P8 showed similar transcriptional and secretory profiles, though the expression of immunomodulatory HLA-G and IL-8 genes was significantly downregulated in P8 compared with P3 eMSCs. Upon TNF-α/IFN-γ induction, eMSCs showed an immunophenotypic profile similar to that of non-induced eMSCs, except for significant upregulation of HLA-DR protein expression in both induced P3 and P8 eMSCs. However, induced P3 and P8 eMSCs showed significant upregulation of CD10, HLA-G, IDO, IL-6, IL-8, LIF and TSG gene expression compared with non-induced cultures. TNF-α/IFN-γ induction strongly increased the secretion of inflammatory-/angiogenesis-related molecules, whereas growth factor secretion was similar to the non-induced eMSCs. Functionally, P3 and P8 eMSCs showed a strong inhibitory effect on stimulated PBMC proliferation and the capacity to support neovascularization in vitro. The authors’ study suggests that serial expansion does not affect eMSC immunophenotypic, transcriptional and secretory profiles. This is directly reflected by the functional immunomodulatory and pro-angiogenic properties of eMSCs, which remain unaltered until P8 in vitro. However, exposure of eMSCs to inflammatory environments enhances their immunomodulatory transcriptional and inflammatory-/angiogenesis-related secretory profiles. Therefore, the resulting evidence of eMSCs serial expansion and exposure to inflammation could serve as a foundation for improved eMSCs manufacturing and potential clinical translation efforts. Experimental design illustrating endometrial tissue obtained from the second day of menstrual cycle, endometrial-derived mesenchymal stem/stromal cell (eMSC) isolation and expansion, eMSC inflammatory stimulation and eMSC profiling and functionality assessments in vitro. [Display omitted

Reliable Reference Genes for Gene Expression Assessment in Tendon-Derived Cells under Inflammatory and Pro-Fibrotic/Healing Stimuli

Tendon cells (TCs) are important for homeostatic maintenance in the healthy tendon and to promote tissue healing after injury. Further, resident and rare populations of tendon stem/progenitor cells, located at various sites within the tendon, contribute to tendon recovery by differentiating into repairing TCs. Gene expression analysis, through quantitative reverse-transcription polymerase chain reaction (qRT-PCR), constitutes a useful tool to study cellular responses, including the transition from initial inflammation to healing processes. A critical step required for data normalization is the choice of reliable reference genes (RGs), a process highly underestimated in tendon biology. In this study, the suitability of five commonly used RGs ( , , , , and ) was evaluated using TCs samples cultured in both standard and progenitor-enriching conditions, as well as under either inflammatory (IFNγ + TNFα) or pro-fibrotic/healing (CTGF) stimulation. The stability of the candidate RGs was computationally determined using NormFinder, geNorm, BestKeeper, and DeltaCt applets. Overall, resulted as the most stable RG on the basis of the integration of each gene weight, whereas and performed poorly. To further validate 's optimal performance, we evaluated the expression of , coding for an immune-related cell surface glycoprotein, and , encoding collagen type I that is the main component of the tendon extracellular matrix (ECM), both known to be modulated by inflammation. The expression of both genes was heavily affected by the RGs used. Consequently, when analyzing gene expression in tendon-derived cells subjected to various stimulatory protocols, the use of a suitable RG should be considered carefully. On the basis of our results, can be reliably used when analyzing different TC types exposed to pathological conditions

CD10/Neprilysin Enrichment in Infrapatellar Fat Pad–Derived Mesenchymal Stem Cells Under Regulatory-Compliant Conditions: Implications for Efficient Synovitis and Fat Pad Fibrosis Reversal

Background:Synovitis and infrapatellar fat pad (IFP) fibrosis participate in various conditions of the knee. Substance P (SP), a neurotransmitter secreted within those structures and historically associated with nociception, also modulates local neurogenic inflammatory and fibrotic responses. Exposure of IFP mesenchymal stem cells (IFP-MSCs) to a proinflammatory/profibrotic environment (ex vivo priming with TNFα, IFNγ, and CTGF) induces their expression of CD10/neprilysin, effectively degrading SP in vitro and in vivo.Purpose/Hypothesis:The purpose was to test the therapeutic effects of IFP-MSCs processed under regulatory-compliant protocols, comparing them side-by-side with standard fetal bovine serum (FBS)–grown cells. The hypothesis was that when processed under such protocols, IFP-MSCs do not require ex vivo priming to acquire a CD10-rich phenotype efficiently degrading SP and reversing synovitis and IFP fibrosis.Study Design:Controlled laboratory study.Methods:Human IFP-MSCs were processed in FBS or either of 2 alternative conditions—regulatory-compliant pooled human platelet lysate (hPL) and chemically reinforced medium (Ch-R)—and then subjected to proinflammatory/profibrotic priming with TNFα, IFNγ, and CTGF. Cells were assessed for in vitro proliferation, stemness, immunophenotype, differentiation potential, transcriptional and secretory profiles, and SP degradation. Based on a rat model of acute synovitis and IFP fibrosis, the in vivo efficacy of cells degrading SP plus reversing structural signs of inflammation and fibrosis was assessed.Results:When compared with FBS, IFP-MSCs processed with either hPL or Ch-R exhibited a CD10Highphenotype and showed enhanced proliferation, differentiation, and immunomodulatory transcriptional and secretory profiles (amplified by priming). Both methods recapitulated and augmented the secretion of growth factors seen with FBS plus priming, with some differences between them. Functionally, in vitro SP degradation was more efficient in hPL and Ch-R, confirmed upon intra-articular injection in vivo where CD10-rich IFP-MSCs also dramatically reversed signs of synovitis and IFP fibrosis even without priming or at significantly lower cell doses.Conclusion:hPL and Ch-R formulations can effectively replace FBS plus priming to induce specific therapeutic attributes in IFP-MSCs. The resulting fine-tuned, regulatory-compliant, cell-based product has potential future utilization as a novel minimally invasive cell therapy for the treatment of synovitis and IFP fibrosis.Clinical Relevance:The therapeutic enhancement of IFP-MSCs manufactured under regulatory-compliant conditions suggests that such a strategy could accelerate the time from preclinical to clinical phases. The therapeutic efficacy obtained at lower MSC numbers than currently needed and the avoidance of cell priming for efficient results could have a significant effect on the design of clinical protocols to potentially treat conditions involving synovitis and IFP fibrosis

Evaluating Vascularization of Heterotopic Islet Constructs for Type 1 Diabetes Using an In Vitro Platform

Type 1 diabetes (T1D) results from the autoimmune destruction of β-cells within the pancreatic islets of Langerhans. Clinical islet transplantation from healthy donors is proposed to ameliorate symptoms, improve quality of life, and enhance the life span of afflicted T1D patients. However, post-transplant outcomes are dependent on the survival of the transplanted islets, which relies on the engraftment of the islets with the recipient's vasculature among other factors. Treatment strategies to improve engraftment include combining islets with supporting cells including endothelial cells (EC) and mesenchymal stem cells (MSC), dynamic cells capable of robust immunomodulatory and vasculogenic effects. In this study, we developed an in vitro model of transplantation to investigate the cellular mechanisms that enhance rapid vascularization of heterotopic islet constructs. Self-assembled vascular beds of fluorescently stained EC served as reproducible in vitro transplantation sites. Heterotopic islet constructs composed of islets, EC, and MSC were transferred to vascular beds for modeling transplantation. Time-lapsed imaging was performed for analysis of the vascular bed remodeling for parameters of neo-vascularization. Moreover, sampling of media following modeled transplantation showed secretory profiles that were correlated with imaging analyses as well as with islet function using glucose-stimulated insulin secretion. Together, evidence revealed that heterotopic constructs consisting of islets, EC, and MSC exhibited the most rapid recruitment and robust branching of cells from the vascular beds suggesting enhanced neo-vascularization compared to islets alone and control constructs. Together, this evidence supports a promising cell transplantation strategy for T1D and also demonstrates a valuable tool for rapidly investigating candidate cellular therapies for transplantation

Signature quality attributes of CD146 + mesenchymal stem/stromal cells correlate with high therapeutic and secretory potency

CD146 bone marrow-derived mesenchymal stem/stromal cells (BM-MSCs) play key roles in the perivascular niche, skeletogenesis, and hematopoietic support; however, comprehensive evaluation of therapeutic potency has yet to be determined. In this study, in vitro inflammatory priming to crude human BM-MSCs (n = 8) captured a baseline of signature responses, including enriched CD146 with coexpression of CD107a , CXCR4 , and LepR , transcriptional profile, enhanced secretory capacity, and robust immunomodulatory secretome and function, including immunopotency assays (IPAs) with stimulated immune cells. These signatures were significantly more pronounced in CD146 (POS)-sorted subpopulation than in the CD146 (NEG). Mechanistically, POS BM-MSCs showed a markedly higher secretory capacity with significantly greater immunomodulatory and anti-inflammatory protein production upon inflammatory priming compared with the NEG BM-MSCs. Moreover, IPAs with stimulated peripheral blood mononuclear cells and T lymphocytes demonstrated robust immunosuppression mediated by POS BM-MSC while inducing significant frequencies of regulatory T cells. in vivo evidence showed that POS BM-MSC treatment promoted pronounced M1-to-M2 macrophage polarization, ameliorating inflammation/fibrosis of knee synovium and fat pad, unlike treatment with NEG BM-MSCs. These data correlate the expression of CD146 with innately higher immunomodulatory and secretory capacity, and thus therapeutic potency. This high-content, reproducible evidence suggests that the CD146 (POS) MSC subpopulation are the mediators of the beneficial effects achieved using crude BM-MSCs, leading to translational implications for improving cell therapy and manufacturing