Mesenchymal stem cell transplantation for myocardial reparation of rat experimental heart failure

Mesenchymal stem cells (MSC) are resident pluripotent cells of bone marrow stroma. MSC have the ability to differentiate into osteoblasts, chondroblasts and adipocytes, neurons, glia and also into cardiomyocytes. The problem of MSC use in cell therapy of various diseases and in myocardial infarction therapy is widely discussed at present. The experiments were carried out on the inbred line Wistar--Kyoto rats. Myocardial experimental infarction (EI) was induced by left descending coronary artery ligation. MSC were isolated from bone marrow, cultivated in vitro and injected into the tail vein on the day of experimental infarction operation. It was shown that the structure of injured myocardium in experimental group significantly differed from that in control group. MSC transplantation led to inflammatory process acceleration and to increased angiogenesis in the damaged myocardium; also, live cardiomyocyte layers were detected in the scar. As a result, ventricular dilatation and overload of the border zone of infarct region decreased, no features of infarction relapse were shown in the border zone.     

Mesenchymal stem cell therapy: A promising cell‐based therapy for treatment of myocardial infarction

For decades, mesenchymal stem (MSCs) cells have been used for cardiovascular diseases as regenerative therapy. This review is an attempt to summarize the types of MSCs involved in myocardial infarction (MI) therapy, as well as its possible mechanisms effects, especially the paracrine one in MI focusing on the studies (human and animal) conducted within the last 10 years. Recently, reports showed that MSC therapy could have infarct‐limiting effects after MI in both experimental and clinical trials. In this context, various types of MSCs can help cardiac regeneration by either revitalizing the cardiac stem cells or revascularizing the arteries and veins of the heart. Furthermore, MSCs could produce paracrine growth factors that increase the survival of nearby cardiomyocytes, as well as increase angiogenesis through recruitment of stem cell from bone marrow or inducing vessel growth from existing capillaries. Recent research suggests that the paracrine effects of MSCs could be mediated by extracellular vesicles including exosomes. Exosomal microRNAs (miRNAs) released by MSCs are promising therapeutic hotspot target for MI. This could be attributed to the role of miRNA in cardiac biology, including cardiac regeneration, stem cell differentiation, apoptosis, neovascularization, cardiac contractility and cardiac remodeling. Furthermore, gene‐modified MSCs could be a recent promising therapy for MI to enhance the paracrine effects of MSCs, including better homing and effective cell targeted tissue regeneration. Although MSC therapy has achieved considerable attention and progress, there are critical challenges that remains to be overcome to achieve the most effective successful cell‐based therapy in MI.     

A Notch positive feedback in the intestinal stem cell niche is essential for stem cell self‐renewal

The intestinal epithelium is the fastest regenerative tissue in the body, fueled by fast‐cycling stem cells. The number and identity of these dividing and migrating stem cells are maintained by a mosaic pattern at the base of the crypt. How the underlying regulatory scheme manages this dynamic stem cell niche is not entirely clear. We stimulated intestinal organoids with Notch ligands and inhibitors and discovered that intestinal stem cells employ a positive feedback mechanism via direct Notch binding to the second intron of the Notch1 gene. Inactivation of the positive feedback by CRISPR/Cas9 mutation of the binding sequence alters the mosaic stem cell niche pattern and hinders regeneration in organoids. Dynamical system analysis and agent‐based multiscale stochastic modeling suggest that the positive feedback enhances the robustness of Notch‐mediated niche patterning. This study highlights the importance of feedback mechanisms in spatiotemporal control of the stem cell niche.     

Mesenchymal stem cell transplantation for the infarcted heart: a role in minimizing abnormalities in cardiac-specific energy metabolism

Intense interest has been focused on cell-based therapy for the infarcted heart given that stem cells have exhibited the ability to reduce infarct size and mitigate cardiac dysfunction. Despite this, it is unknown whether mesenchymal stem cell (MSC) therapy can prevent metabolic remodeling following a myocardial infarction (MI). This study examines the ability of MSCs to rescue the infarcted heart from perturbed substrate uptake in vivo. C57BL/6 mice underwent chronic ligation of the left anterior descending coronary artery to induce a MI. Echocardiography was performed on conscious mice at baseline as well as 7 and 23 days post-MI. Twenty-eight days following the ligation procedure, hyperinsulinemic euglycemic clamps assessed in vivo insulin sensitivity. Isotopic tracer administration evaluated whole body, peripheral tissue, and cardiac-specific glucose and fatty acid utilization. To gain insight into the mechanisms by which MSCs modulate metabolism, mitochondrial function was assessed by high-resolution respirometry using permeabilized cardiac fibers. Data show that MSC transplantation preserves insulin-stimulated fatty acid uptake in the peri-infarct region (4.25 ± 0.64 vs. 2.57 ± 0.34 vs. 3.89 ± 0.54 μmol·100 g(-1)·min(-1), SHAM vs. MI + PBS vs. MI + MSC; P < 0.05) and prevents increases in glucose uptake in the remote left ventricle (3.11 ± 0.43 vs. 3.81 ± 0.79 vs. 6.36 ± 1.08 μmol·100 g(-1)·min(-1), SHAM vs. MI + PBS vs. MI + MSC; P < 0.05). This was associated with an enhanced efficiency of mitochondrial oxidative phosphorylation with a respiratory control ratio of 3.36 ± 0.18 in MSC-treated cardiac fibers vs. 2.57 ± 0.14 in the infarct-only fibers (P < 0.05). In conclusion, MSC therapy exhibits the potential to rescue the heart from metabolic aberrations following a MI. Restoration of metabolic flexibility is important given the metabolic demands of the heart and the role of energetics in the progression to heart failure.     

Endometrial stem cell transplantation in MPTP‐ exposed primates: an alternative cell source for treatment of Parkinson's disease

Parkinson's disease (PD) is a neurodegenerative disease caused by the loss of dopaminergic neurons in the substantia nigra. Cell‐replacement therapies have emerged as a promising strategy to slow down or replace neuronal loss. Compared to other stem cell types, endometrium‐derived stem cells (EDSCs) are an attractive source of stem cells for cellular therapies because of their ease of collection and vast differentiation potential. Here we demonstrate that endometrium‐derived stem cells may be transplanted into an MPTP exposed monkey model of PD. After injection into the striatum, endometrium‐derived stem cells engrafted, exhibited neuron‐like morphology, expressed tyrosine hydroxylase (TH) and increased the numbers of TH positive cells on the transplanted side and dopamine metabolite concentrations in vivo. Our results suggest that endometrium‐derived stem cells may provide a therapeutic benefit in the primate model of PD and may be used in stem cell based therapies.     

Mesenchymal stem cell-conditioned media: A novel alternative of stem cell therapy for quality wound healing

Mesenchymal stem cells-conditioned media (MSCs-CM) contains several growth factors and cytokines, thus may be used as a better alternative to stem cell therapy, which needs to be elucidated. The present study was conducted to evaluate the therapeutic potential of caprine, canine, and guinea pig bone marrow-derived MSCs-CM in excision wound healing in a guinea pig model. MSCs were obtained from bone marrow, expanded ex vivo and characterized as per ISCT criteria. CM was collected assayed by western blot to ascertain the presence of important secretory biomolecules. Quantitative estimation by enzyme-linked immunosorbent assay was done for a vascular epidermal growth factor (VEGF) and interleukin-6 (IL-6) in caprine MSCs-CM and optimum time for collection of CM was decided as 72 hr. CM from all the species was lyophilized by freeze-drying method. Full-thickness (2 × 2 cm²) excision skin wounds were created in guinea pigs (six animals in each group) and respective lyophilized CM mixed with laminin gel was applied topically at weekly interval. On Day 28, histopathological examinations of healed skin were done by hemotoxylin and eosin staining. MSCs were found to secrete important growth factors and cytokines (i.e., VEGF, transforming growth factor-β1, fibroblast growth factor-2, insulin-like growth factor-1, stem cell factor, and IL-6) as demonstrated by immunohistochemistry and western blot assay. It was found that allogenic and xenogenic application of CM significantly improved quality wound healing with minimal scar formation. Thus, MSCs-CM can be used allogenically as well as xenogenically for quality wound healing.     

Non-myeloablative stem cell transplantation and donor lymphocyte infusion for the treatment of cancer and life-threatening non-malignant disorders

Allogeneic bone marrow or blood stem call transplantation (BMT) represents an important therapeutic tool for the treatment of otherwise incurable malignant and non-malignant diseases. Until recently, autologous and allogeneic bone marrow and mobilized blood stem cell transplantations were used primarily to replace malignant, genetically abnormal or deficient immunohematopoietic compartments, and therefore highly toxic myeloablative regimens were considered to be mandatory for the effective eradication of all undesirable host-derived hematopoietic elements. Our preclinical and ongoing clinical studies have indicated that much more effective eradication of the host immunohematopoietic system cells can be achieved by adoptive allogeneic cell therapy with donor lymphocyte infusion following BMT. Thus, eradication of blood cancer cells, especially in patients with chronic myeloid leukemia and, less frequently, in patients with other hematologic malignancies, can frequently be accomplished despite the complete resistance of such tumor cells to maximally tolerated doses of chemoradiotherapy. Our cumulative experience has suggested that graft-vs.-leukemia (GVL) effects might be a useful tool for the eradication of otherwise resistant tumor cells of host origin. Based on the cumulative clinical experience and experimental data in animal models of human diseases, it appears that the induction of host-vs.-graft tolerance as an initial step may allow the durable engraftment of donor immunocompetent lymphocytes, which may be used for the induction of effective biologic warfare against host-type immunohematopoietic cells that need to be replaced, including malignant, genetically abnormal or self-reactive cells. Based on the aforementioned rationale, we speculated that the therapeutic benefit of BMT may be improved by using safer conditioning as part of the transplant procedure, with the goal being to induce host-vs.-graft tolerance to enable subsequent induction of GVL, possibly graft-vs.-tumor or even graft-vs.-autoimmunity effects, rather than attempting to eliminate host cells with hazardous myeloablative chemoradiotherapy. This hypothesis suggested that effective BMT procedures could be accomplished without lethal conditioning of the host, using new well-tolerated non-myeloablative regimens, thus possibly minimizing immediate and late side-effects related to the myeloablative procedures until recently considered to be mandatory for the conditioning of BMT recipients. Recent clinical data presented in this review suggest that effective BMT procedures may be accomplished with well-tolerated non-myeloablative stem cell transplantation (NST) regimens, with no major toxicity. Thus, new NST approaches may offer the feasibility of safer BMT procedures for a large spectrum of clinical indications in children and elderly individuals, without lower or upper age limits, while minimizing procedure-related toxicity and mortality. Taken together, our data suggest that high-dose chemotherapy and radiation therapy may be successfully replaced by a more effective biologic tool, alloreactive donor lymphocytes, thus setting the stage for innovative therapeutic procedures for safer and more effective treatment of patients in need of BMT.     

Three‐dimensional cell culture microarray for high‐throughput studies of stem cell fate

We have developed a novel three‐dimensional (3D) cellular microarray platform to enable the rapid and efficient tracking of stem cell fate and quantification of specific stem cell markers. This platform consists of a miniaturized 3D cell culture array on a functionalized glass slide for spatially addressable high‐throughput screening. A microarray spotter was used to deposit cells onto a modified glass surface to yield an array consisting of cells encapsulated in alginate gel spots with volumes as low as 60 nL. A method based on an immunofluorescence technique scaled down to function on a cellular microarray was also used to quantify specific cell marker protein levels in situ. Our results revealed that this platform is suitable for studying the expansion of mouse embryonic stem (ES) cells as they retain their pluripotent and undifferentiated state. We also examined neural commitment of mouse ES cells on the microarray and observed the generation of neuroectodermal precursor cells characterized by expression of the neural marker Sox‐1, whose levels were also measured in situ using a GFP reporter system. In addition, the high‐throughput capacity of the platform was tested using a dual‐slide system that allowed rapid screening of the effects of tretinoin and fibroblast growth factor‐4 (FGF‐4) on the pluripotency of mouse ES cells. This high‐throughput platform is a powerful new tool for investigating cellular mechanisms involved in stem cell expansion and differentiation and provides the basis for rapid identification of signals and conditions that can be used to direct cellular responses. Biotechnol. Bioeng. 2010; 106: 106–118. © 2010 Wiley Periodicals, Inc.     

Pooled analysis of clinical outcome for EGFR TKI‐treated patients with EGFR mutation‐positive NSCLC

Patients with non‐small‐cell lung cancer (NSCLC) appear to gain particular benefit from treatment with epidermal growth factor receptor (EGFR) tyrosine‐kinase inhibitors (TKI) if their disease tests positive for EGFR activating mutations. Recently, several large, controlled, phase III studies have been published in NSCLC patients with EGFR mutation‐positive tumours. Given the increased patient dataset now available, a comprehensive literature search for EGFR TKIs or chemotherapy in EGFR mutation‐positive NSCLC was undertaken to update the results of a previously published pooled analysis. Pooling eligible progression‐free survival (PFS) data from 27 erlotinib studies (n = 731), 54 gefitinib studies (n = 1802) and 20 chemotherapy studies (n = 984) provided median PFS values for each treatment. The pooled median PFS was: 12.4 months (95% accuracy intervals [AI] 11.6–13.4) for erlotinib‐treated patients; 9.4 months (95% AI 9.0–9.8) for gefitinib‐treated patients; and 5.6 months (95% AI 5.3–6.0) for chemotherapy. Both erlotinib and gefitinib resulted in significantly longer PFS than chemotherapy (permutation testing; P = 0.000 and P = 0.000, respectively). Data on more recent TKIs (afatinib, dacomitinib and icotinib) were insufficient at this time‐point to carry out a pooled PFS analysis on these compounds. The results of this updated pooled analysis suggest a substantial clear PFS benefit of treating patients with EGFR mutation‐positive NSCLC with erlotinib or gefitinib compared with chemotherapy.     

The effects of mesenchymal stem cell mitochondrial transplantation on doxorubicin‐mediated nephrotoxicity in rats

The effect of dysfunctional mitochondria in several cell pathologies has been reported in renal diseases, including diabetic nephropathy and acute kidney injury. Previous studies have reported that mitochondrial transplantation provided surprising results in myocardial and liver ischemia, as well as in Parkinson's disease. We aimed to investigate the beneficial effects of isolated mitochondria transplantation from mesenchymal stem cells (MSCs) in vivo, to mitigate renal damage that arises from doxorubicin‐mediated nephrotoxicity and its action mechanism. In this study, a kidney model of doxorubicin‐mediated nephrotoxicity was used and isolated mitochondria from MSCs were transferred to the renal cortex of rats. The findings showed that the rate of isolated mitochondria from MSCs maintains sufficient membrane integrity, and was associated with a beneficial renal therapeutic effect. Following doxorubicin‐mediated renal injury, isolated mitochondria or vehicle infused into the renal cortex and rats were monitored for five days. This study found that mitochondrial transplantation decreased cellular oxidative stress and promoted regeneration of tubular cells after renal injury (P < .001, P = .009). Moreover, mitochondrial transplantation reduced protein accumulation of tubular cells and reversed renal deficits (P = .01, P < .001). Mitochondrial transplantation increased Bcl‐2 levels, and caspase‐3 levels decreased in injured renal cells (P < .015, P < .001). Our results provide a direct link between mitochondria dysfunction and doxorubicin‐mediated nephrotoxicity and suggest a therapeutic effect of transferring isolated mitochondria obtained from MSCs against renal injury. To our knowledge, this study is the first study in the literature that showed good therapeutic effects of mitochondrial transplantation in a nephrotoxicity model, which is under‐researched.     

Amyotrophic lateral sclerosis in pregnancy: clinical outcome during the post-partum period after stem cell transplantation into the frontal motor cortex

Background aimsAmyotrophic lateral sclerosis (ALS) is rare in pregnant patients. Stem cell therapy has been proposed as a potential therapeutic strategy for ALS.MethodsWe describe a young woman with sporadic ALS that started during the second trimester of pregnancy with a rapid progression after delivery and severe motor impairment. Several drugs and stem cell injection by lumbar puncture were performed without changes before the patient was referred to our institution.ResultsAfter bilateral autologous stem cell transplantation into the frontal motor cortices, we observed stabilization in ALS functional rating scale, significant delay of ALS progression and an extension in her life expectancy.ConclusionsStem cell transplantation may alter the clinical course of ALS and improve quality of life in pre-menopausal women.     

Mesenchymal stromal cell‐derived extracellular vesicles as cell‐free biologics for the ex vivo expansion of hematopoietic stem cells

Hematopoietic stem cell transplantation (HSCT) is the ultimate choice of treatment for patients with hematological diseases and cancer. The success of HSCT is critically dependent on the number and engraftment efficiency of the transplanted donor hematopoietic stem cells (HSCs). Various studies show that bone marrow‐derived mesenchymal stromal cells (MSCs) support hematopoiesis and also promote ex vivo expansion of HSCs. MSCs exert their therapeutic effect through paracrine activity, partially mediated through extracellular vesicles (EVs). Although the physiological function of EVs is not fully understood, inspiring findings indicate that MSC‐derived EVs can reiterate the hematopoiesis, supporting the ability of MSCs by transferring their cargo containing proteins, lipids, and nucleic acids to the HSCs. The activation state of the MSCs or the signaling mechanism that prevails in them also defines the composition of their EVs, thereby influencing the fate of HSCs. Modulating or preconditioning MSCs to achieve a specific composition of the EV cargo for the ex vivo expansion of HSCs is, therefore, a promising strategy that can overcome several challenges associated with the use of naïve/unprimed MSCs. This review aims to speculate upon the potential role of preconditioned/primed MSC‐derived EVs as “cell‐free biologics,” as a novel strategy for expanding HSCs in vitro.     

Human embryonic stem cell differentiation into insulin secreting β‐cells for diabetes

hESC (human embryonic stem cells), when differentiated into pancreatic β ILC (islet‐like clusters), have enormous potential for the cell transplantation therapy for Type 1 diabetes. We have developed a five‐step protocol in which the EBs (embryoid bodies) were first differentiated into definitive endoderm and subsequently into pancreatic lineage followed by formation of functional endocrine β islets, which were finally matured efficiently under 3D conditions. The conventional cytokines activin A and RA (retinoic acid) were used initially to obtain definitive endoderm. In the last step, ILC were further matured under 3D conditions using amino acid rich media (CMRL media) supplemented with anti‐hyperglycaemic hormone‐Glp1 (glucagon‐like peptide 1) analogue Liraglutide with prolonged t_½ and Exendin 4. The differentiated islet‐like 3D clusters expressed bonafide mature and functional β‐cell markers‐PDX1 (pancreatic and duodenal homoeobox‐1), C‐peptide, insulin and MafA. Insulin synthesis de novo was confirmed by C‐peptide ELISA of culture supernatant in response to varying concentrations of glucose as well as agonist and antagonist of functional 3D β islet cells in vitro. Our results indicate the presence of almost 65% of insulin producing cells in 3D clusters. The cells were also found to ameliorate hyperglycaemia in STZ (streptozotocin) induced diabetic NOD/SCID (non‐obese diabetic/severe combined immunodeficiency) mouse up to 96 days of transplantation. This protocol provides a basis for 3D in vitro generation of long‐term in vivo functionally viable islets from hESC.     

17β‐Oestradiol promotes differentiation of human embryonic stem cells into dopamine neurons via cross‐talk between insulin‐like growth factors‐1 and oestrogen receptor β

Human embryonic stem cells (hESCs) can self‐renew and differentiate into all cell lineages. E2 is known to exhibit positive effects on embryo development. Although the importance of E2 in many physiological processes has been reported, to date few researchers have investigated the effects of E2 on hESCs differentiation. We studied the effects of E2 on dopamine (DA) neuron induction of hESCs and its related signalling pathways using the three‐stage protocol. In our study, 0.1 μM E2 were applied to hESCs‐derived human embryoid bodies (hEBs) and effects of E2 on neural cells differentiation were investigated. Protein and mRNA level assay indicated that E2 up‐regulated the expression of insulin‐like growth factors (IGF)‐1, ectoderm, neural precursor cells (NPC) and DA neuron markers, respectively. The population of hESC‐derived NPCs and DA neurons was increased to 92% and 93% to that of DMSO group, respectively. Furthermore, yield of DA neuron‐secreted tyrosine hydroxylase (TH) and dopamine was also increased. E2‐caused promotion was relieved in single inhibitor (ICI or JB1) group partly, and E2 effects were repressed more stronger in inhibitors combination (ICI plus JB1) group than in single inhibitor group at hEBs, hNPCs and hDA neurons stages. Owing to oestrogen receptors regulate multiple brain functions, when single or two inhibitors were used to treat neural differentiation stage, we found that oestrogen receptor (ER)β but not ERα is strongly repressed at the hNPCs and hDA neurons stage. These findings, for the first time, demonstrate the molecular cascade and related cell biology events involved in E2‐improved hNPC and hDA neuron differentiation through cross‐talk between IGF‐1 and ERβ in vitro.