Can a Scaffold Enriched with Mesenchymal Stem Cells Be a Good Treatment for Spinal Cord Injury?

Spinal cord injury (SCI) is a worldwide highly crippling disease that can lead to the loss of motor and sensory neurons. Among the most promising therapies, there are new techniques of tissue engineering based on stem cells that promote neuronal regeneration. Among the different types of stem cells, mesenchymal stem cells (MSCs) seem the most promising. Indeed, MSCs are able to release trophic factors and to differentiate into the cell types that can be found in the spinal cord. Currently, the most common procedure to insert cells in the lesion site is infusion. However, this causes a low rate of survival and engraftment in the lesion site. For these reasons, tissue engineering is focusing on bioresorbable scaffolds to help the cells to stay in situ. Scaffolds do not only have a passive role but become fundamental for the trophic support of cells and the promotion of neuroregeneration. More and more types of materials are being studied as scaffolds to decrease inflammation and increase the engraftment as well as the survival of the cells. Our review aims to highlight how the use of scaffolds made from biomaterials enriched with MSCs gives positive results in in vivo SCI models as well as the first evidence obtained in clinical trials.     

Combinational Therapy of Cardiac Atrial Appendage Stem Cells and Pyridoxamine: The Road to Cardiac Repair?

Myocardial infarction (MI) occurs when the coronary blood supply is interrupted. As a consequence, cardiomyocytes are irreversibly damaged and lost. Unfortunately, current therapies for MI are unable to prevent progression towards heart failure. As the renewal rate of cardiomyocytes is minimal, the optimal treatment should achieve effective cardiac regeneration, possibly with stem cells transplantation. In that context, our research group identified the cardiac atrial appendage stem cells (CASCs) as a new cellular therapy. However, CASCs are transplanted into a hostile environment, with elevated levels of advanced glycation end products (AGEs), which may affect their regenerative potential. In this study, we hypothesize that pyridoxamine (PM), a vitamin B6 derivative, could further enhance the regenerative capacities of CASCs transplanted after MI by reducing AGEs’ formation. Methods and Results: MI was induced in rats by ligation of the left anterior descending artery. Animals were assigned to either no therapy (MI), CASCs transplantation (MI + CASCs), or CASCs transplantation supplemented with PM treatment (MI + CASCs + PM). Four weeks post-surgery, global cardiac function and infarct size were improved upon CASCs transplantation. Interstitial collagen deposition, evaluated on cryosections, was decreased in the MI animals transplanted with CASCs. Contractile properties of resident left ventricular cardiomyocytes were assessed by unloaded cell shortening. CASCs transplantation prevented cardiomyocyte shortening deterioration. Even if PM significantly reduced cardiac levels of AGEs, cardiac outcome was not further improved. Conclusion: Limiting AGEs’ formation with PM during an ischemic injury in vivo did not further enhance the improved cardiac phenotype obtained with CASCs transplantation. Whether AGEs play an important deleterious role in the setting of stem cell therapy after MI warrants further examination.     

Can a Large Number of Transplanted Mesenchymal Stem Cells Have an Optimal Therapeutic Effect on Improving Ovarian Function?

Mesenchymal stem cells (MSCs) are next-generation treatment in degenerative diseases. For the application of mesenchymal stem cell therapy to degenerative disease, transplantation conditions (e.g., optimized dose, delivery route and regenerating efficacy) should be considered. Recently, researchers have studied the mode of action of MSC in the treatment of ovarian degenerative disease. However, the evidence for the optimal number of cells for the developing stem cell therapeutics is insufficient. The objective of this study was to evaluate the efficacy in ovarian dysfunction, depends on cell dose. By intraovarian transplantation of low (1 × 10⁵) and high (5 × 10⁵) doses of placenta-derived mesenchymal stem cells (PD-MSCs) into thioacetamide (TAA)-injured rats, we compared the levels of apoptosis and oxidative stress that depend on different cell doses. Apoptosis and oxidative stress were significantly decreased in the transplanted (Tx) group compared to the non-transplanted (NTx) group in ovarian tissues from TAA-injured rats (* p < 0.05). In addition, we confirmed that follicular development was significantly increased in the Tx groups compared to the NTx group (* p < 0.05). However, there were no significant differences in the apoptosis, antioxidant or follicular development of injured ovarian tissues between the low and high doses PD-MSCs group. These findings provide new insights into the understanding and evidence obtained from clinical trials for stem cell therapy in reproductive systems.     

Do Drug-likeness Rules Apply to Oral Prodrugs?

This paper describes a comparative analysis of the physicochemical and structural properties of prodrugs and their corresponding drugs with regard to drug-likeness rules. The dataset used in this work was obtained from the DrugBank. Sixty-five pairs of prodrugs/drugs were retrieved and divided into the following categories: carrier-linked to increase hydrophilic character, carrier-linked to increase absorption, and bioprecursors. We compared the physicochemical properties related to drug-likeness between prodrugs and drugs. Our results show that prodrugs do not always follow Lipinski's Rule of 5, especially as we observed 15 prodrugs with more than 10 hydrogen bond acceptors and 18 with a molecular weight greater than 500 Da. This fact highlights the importance of extending Lipinski's rules to encompass other parameters as both strategies (filtering of drug-like chemical libraries and prodrug design) aim to improve the bioavailability of compounds. Therefore, critical reasoning is fundamental to determine whether a structure has drug-like properties or could be considered a potential orally active compound in the drug-design pipeline.     

Do Extracellular Vesicles Derived from Mesenchymal Stem Cells Contain Functional Mitochondria?

Extracellular vesicles (EV) derived from stem cells have become an effective complement to the use in cell therapy of stem cells themselves, which has led to an explosion of research into the mechanisms of vesicle formation and their action. There is evidence demonstrating the presence of mitochondrial components in EV, but a definitive conclusion about whether EV contains fully functional mitochondria has not yet been made. In this study, two EV fractions derived from mesenchymal stromal stem cells (MSC) and separated by their size were examined. Flow cytometry revealed the presence of mitochondrial lipid components capable of interacting with mitochondrial dyes MitoTracker Green and 10-nonylacridine orange; however, the EV response to the probe for mitochondrial membrane potential was negative. Detailed analysis revealed components from all mitochondria compartments, including house-keeping mitochondria proteins and DNA as well as energy-related proteins such as membrane-localized proteins of complexes I, IV, and V, and soluble proteins from the Krebs cycle. When assessing the functional activity of mitochondria, high variability in oxygen consumption was noted, which was only partially attributed to mitochondrial respiratory activity. Our findings demonstrate that the EV contain all parts of mitochondria; however, their independent functionality inside EV has not been confirmed, which may be due either to the absence of necessary cofactors and/or the EV formation process and, probably the methodology of obtaining EV.     

Photoactivable Elimination of Tumorigenic Human Induced Pluripotent Stem Cells by Using a Lectin–Doxorubicin Prodrug Conjugate

Human pluripotent stem cells (hPSCs) are attractive resources for regenerative medicine, but medical applications are hindered by their tumorigenic potential. Previously, a hPSC-specific lectin probe, rBC2LCN, was identified through comprehensive glycome analysis by using high-density lectin microarrays. Herein, a lectin–doxorubicin (DOX) prodrug conjugate, with controllable photolysis activation for the elimination of tumorigenic human induced pluripotent stem cells, has been developed. rBC2LCN was fused with a biotin-binding protein, tamavidin (BC2Tama), and the fusion protein was expressed in Escherichia coli and purified by means of affinity chromatography. BC2Tama was then conjugated with doxorubicin-photocleavable biotin (DOXPCB). The BC2Tama–DOXPCB conjugates were observed to bind to hPSCs followed by internalization. Upon exposure to ultraviolet light, DOX was released inside the cells, which allowed specific killing of the hPSCs. Thus, BC2Tama–DOXPCB should be useful for the targeted elimination of hPSCs contained in hPSC-derived cell therapy products. This is the first report of the generation of lectin–prodrug conjugates. BC2Tama should be applicable for the targeted delivery of various types of biotinylated compounds into hPSCs.     

Perivascular Mesenchymal Stem/Stromal Cells,an Immune Privileged Niche for Viruses?

Mesenchymal stem cells (MSCs) play a critical role in response to stress such as infection. They initiate the removal of cell debris, exert major immunoregulatory activities, control pathogens, and lead to a remodeling/scarring phase. Thus, host-derived ‘danger’ factors released from damaged/infected cells (called alarmins, e.g., HMGB1, ATP, DNA) as well as pathogen-associated molecular patterns (LPS, single strand RNA) can activate MSCs located in the parenchyma and around vessels to upregulate the expression of growth factors and chemoattractant molecules that influence immune cell recruitment and stem cell mobilization. MSC, in an ultimate contribution to tissue repair, may also directly trans- or de-differentiate into specific cellular phenotypes such as osteoblasts, chondrocytes, lipofibroblasts, myofibroblasts, Schwann cells, and they may somehow recapitulate their neural crest embryonic origin. Failure to terminate such repair processes induces pathological scarring, termed fibrosis, or vascular calcification. Interestingly, many viruses and particularly those associated to chronic infection and inflammation may hijack and polarize MSC’s immune regulatory activities. Several reports argue that MSC may constitute immune privileged sanctuaries for viruses and contributing to long-lasting effects posing infectious challenges, such as viruses rebounding in immunocompromised patients or following regenerative medicine therapies using MSC. We will herein review the capacity of several viruses not only to infect but also to polarize directly or indirectly the functions of MSC (immunoregulation, differentiation potential, and tissue repair) in clinical settings.     

Human Amnion Epithelial Cells: A Potential Cell Source for Pulp Regeneration?

The aim of this study was to analyze the suitability of pluripotent stem cells derived from the amnion (hAECs) as a potential cell source for revitalization in vitro. hAECs were isolated from human placentas, and dental pulp stem cells (hDPSCs) and dentin matrix proteins (eDMPs) were obtained from human teeth. Both hAECs and hDPSCs were cultured with 10% FBS, eDMPs and an osteogenic differentiation medium (StemPro). Viability was assessed by MTT and cell adherence to dentin was evaluated by scanning electron microscopy. Furthermore, the expression of mineralization-, odontogenic differentiation- and epithelial–mesenchymal transition-associated genes was analyzed by quantitative real-time PCR, and mineralization was evaluated through Alizarin Red staining. The viability of hAECs was significantly lower compared with hDPSCs in all groups and at all time points. Both hAECs and hDPSCs adhered to dentin and were homogeneously distributed. The regulation of odontoblast differentiation- and mineralization-associated genes showed the lack of transition of hAECs into an odontoblastic phenotype; however, genes associated with epithelial–mesenchymal transition were significantly upregulated in hAECs. hAECs showed small amounts of calcium deposition after osteogenic differentiation with StemPro. Pluripotent hAECs adhere on dentin and possess the capacity to mineralize. However, they presented an unfavorable proliferation behavior and failed to undergo odontoblastic transition.     

Endometrial Cancer Stem Cells: Where Do We Stand and Where Should We Go?

Endometrial cancer is one of the most common malignant diseases in women worldwide, with an incidence of 5.9%. Thus, it is the most frequent cancer of the female genital tract, with more than 34,000 women dying, in Europe and North America alone. Endometrial Cancer Stem Cells (CSC) might be drivers of carcinogenesis as well as metastatic and recurrent disease. Therefore, targeting CSCs is of high interest to improve prognosis of patients suffering of advanced or recurrent endometrial cancer. This review describes the current evidence of molecular mechanisms in endometrial CSCs with special emphasis on MYC and NF-κB signaling as well as mitochondrial metabolism. Furthermore, the current status of immunotherapy targeting PD-1 and PD-L1 in endometrial cancer cells and CSCs is elucidated. The outlined findings encourage novel therapies that target signaling pathways in endometrial CSCs as well as immunotherapy as a promising therapeutic approach in the treatment of endometrial cancer to impede cancer progression and prevent recurrence.     

Can organic farming help to reduce N-losses?

This study is in two parts. In the first part, nitrogen (N) losses per unit of milk and meat in Danish conventional and organic pig and dairy farming were compared on the basis of farm data. In the second part, organic and conventional dairy farming were compared in detail, using modelling. N-surpluses at different livestock densities, fodder intensities, and soil types were simulated. Finally, simulated N-surpluses were used in national scenarios for conversion to organic dairy farming in Denmark. In Part one, pig farming was found to have a higher N-efficiency than dairy farming. Organic pig production had a lower N-efficiency and a higher N-surplus per kg meat than conventional pig production. The possibilities to reduce N-loss by conversion to organic pig production therefore appear to be poor. Organic dairy farming had a higher N-efficiency and a lower N-surplus per kg milk than conventional dairy farming. Conversion from conventional to organic dairy farming may therefore reduce N-losses. In Part two, a positive correlation between livestock density and N-surplus ha^-1 was found for dairy farming. For all simulated livestock densities, fodder feeding intensities and soil types, organic systems showed a lower N-surplus per unit of milk produced than conventional systems. National scenarios for dairy farming showed that the present Danish milk production could be achieved with a 24% lower total N-surplus if converted from intensive conventional farming to extensive organic farming. At the same time, N-surplus ha^-1 and N-surplus (t milk)^-1 would be lowered by 50% and 25% respectively. Changing from intensive to extensive conventional dairy farming with a livestock density equal to that in the organic scenario resulted in a reduction in N-surplus ha^-1 of 15%. It was concluded that a reduction in total N-loss from agriculture is possible by converting from conventional to organic dairy farming but at the cost of either lower production on the present dairy farm area, or the current production on a substantially larger area.     

Wnt16 Signaling Is Required for IL-1β-Induced Matrix Metalloproteinase-13-Regulated Proliferation of Human Stem Cell-Derived Osteoblastic Cells

We established a differentiation method for homogeneous α7 integrin-positive human skeletal muscle stem cell (α7⁺hSMSC)-derived osteoblast-like (α7⁺hSMSC-OB) cells, and found that interleukin (IL)-1β induces matrix metalloproteinase (MMP)-13-regulated proliferation of these cells. These data suggest that MMP-13 plays a potentially unique physiological role in the regeneration of osteoblast-like cells. Here, we examined whether up-regulation of MMP-13 activity by IL-1β was mediated by Wingless/int1 (Wnt) signaling and increased the proliferation of osteoblast-like cells. IL-1β increased the mRNA and protein levels of Wnt16 and the Wnt receptor Lrp5/Fzd2. Exogenous Wnt16 was found to increase MMP-13 mRNA, protein and activity, and interestingly, the proliferation rate of these cells. Treatment with small interfering RNAs against Wnt16 and Lrp5 suppressed the IL-1β-induced increase in cell proliferation. We revealed that a unique signaling cascade IL-1β→Wnt16→Lrp5→MMP-13, was intimately involved in the proliferation of osteoblast-like cells, and suggest that IL-1β-induced MMP-13 expression and changes in cell proliferation are regulated by Wnt16.     

BMP3 Alone and Together with TGF-β Promote the Differentiation of Human Mesenchymal Stem Cells into a Nucleus Pulposus-Like Phenotype

Human mesenchymal stem cells (MSCs) have the potential to differentiate into nucleus pulposus (NP)-like cells under specific stimulatory conditions. Thus far, the effects of bone morphogenetic protein 3 (BMP3) and the cocktail effects of BMP3 and transforming growth factor (TGF)-β on MSC proliferation and differentiation remain obscure. Therefore, this study was designed to clarify these unknowns. MSCs were cultured with various gradients of BMP3 and BMP3/TGF-β, and compared with cultures in basal and TGF-β media. Cell proliferation, glycosaminoglycan (GAG) content, gene expression, and signaling proteins were measured to assess the effects of BMP3 and BMP3/TGF-β on MSCs. Cell number and GAG content increased upon the addition of BMP3 in a dose-dependent manner. The expression of COL2A1, ACAN, SOX9, and KRT19 increased following induction with BMP3 and TGF-β, in contrast to that of COL1A1, ALP, OPN, and COMP. Smad3 phosphorylation was upregulated by BMP3 and TGF-β, but BMP3 did not affect the phosphorylation of extracellular-signal regulated kinase (ERK) 1/2 or c-Jun N-terminal kinase (JNK). Our results reveal that BMP3 enhances MSC proliferation and differentiation into NP-like cells, as indicated by increased cell numbers and specific gene expressions, and may also cooperate with TGF-β induced positive effects. These actions are likely related to the activation of TGF-β signaling pathway.     

What Molecular Recognition Systems Do Mesenchymal Stem Cells/Medicinal Signaling Cells (MSC) Use to Facilitate Cell-Cell and Cell Matrix Interactions? A Review of Evidence and Options

Mesenchymal stem cells, also called medicinal signaling cells (MSC), have been studied regarding their potential to facilitate tissue repair for >30 years. Such cells, derived from multiple tissues and species, are capable of differentiation to a number of lineages (chondrocytes, adipocytes, bone cells). However, MSC are believed to be quite heterogeneous with regard to several characteristics, and the large number of studies performed thus far have met with limited or restricted success. Thus, there is more to understand about these cells, including the molecular recognition systems that are used by these cells to perform their functions, to enhance the realization of their potential to effect tissue repair. This perspective article reviews what is known regarding the recognition systems available to MSC, the possible systems that could be looked for, and alternatives to enhance their localization to specific injury sites and increase their subsequent facilitation of tissue repair. MSC are reported to express recognition molecules of the integrin family. However, there are a number of other recognition molecules that also could be involved such as lectins, inducible lectins, or even a MSC-specific family of molecules unique to these cells. Finally, it may be possible to engineer expression of recognition molecules on the surface of MSC to enhance their function in vivo artificially. Thus, improved understanding of recognition molecules on MSC could further their success in fostering tissue repair.     

A Novel Human TGF-β1 Fusion Protein in Combination with rhBMP-2 Increases Chondro-Osteogenic Differentiation of Bone Marrow Mesenchymal Stem Cells

Transforming growth factor-beta (TGF-β) is involved in processes related to the differentiation and maturation of osteoprogenitor cells into osteoblasts. Rat bone marrow (BM) cells were cultured in a collagen-gel containing 0.5% fetal bovine serum (FBS) for 10 days in the presence of rhTGF (recombinant human TGF)-β1-F2, a fusion protein engineered to include a high-affinity collagen-binding decapeptide derived from von Willebrand factor. Subsequently, cells were moderately expanded in medium with 10% FBS for 4 days and treated with a short pulse of rhBMP (recombinant human bone morphogenetic protein)-2 for 4 h. During the last 2 days, dexamethasone and β-glycerophosphate were added to potentiate osteoinduction. Concomitant with an up-regulation of cell proliferation, DNA synthesis levels were determined. Polymerase chain reaction was performed to reveal the possible stemness of these cells. Osteogenic differentiation was evaluated in terms of alkaline phosphatase activity and mineralized matrix formation as well as by mRNA expression of osteogenic marker genes. Moreover, cells were placed inside diffusion chambers and implanted subcutaneously into the backs of adult rats for 4 weeks. Histological study provided evidence of cartilage and bone-like tissue formation. This experimental procedure is capable of selecting cell populations from BM that, in the presence of rhTGF-β1-F2 and rhBMP-2, achieve skeletogenic potential in vitro and in vivo.     

Temporary Increase of PPAR-γ and Transient Expression of UCP-1 in Stromal Vascular Fraction Isolated Human Adipocyte Derived Stem Cells During Adipogenesis

In this study, cells from the stromal vascular fraction of human subcutaneous tissues were induced to differentiate toward adipose cells in vitro for 2 weeks. During adipogenic differentiation, we followed the chronological changes in their morphology with Coherent anti-Stokes Raman scattering (CARS) microscopy and checked the PPAR-γ and UCP-1 expression with RT-PCR. On day 4 after inducing adipogenic differentiation, CARS imaging showed multiple small lipid droplets (LD) distributed peripherally along the cellular membrane. PPAR-γ began to express at this time and increased until day 14 at a steady rate. On day 7, the cells appeared as brown adipocytes with numerous small LD throughout the cytoplasm, and the mRNA level of UCP-1 rose abruptly by 6- to 7-fold. After an additional 7 days, CARS imaging showed the development of a large LD, which is characteristic of white adipocytes, and the mRNA level of UCP-1 slumped significantly. These results demonstrate the possibility that ADSC pass through a brown adipocyte-like stage while differentiating into white adipocytes.     

Should Degenerated Intervertebral Discs of Patients with Modic Type 1 Changes Be Treated with Mesenchymal Stem Cells?

Low back pain (LBP) has been among the leading causes of disability for the past 30 years. This highlights the need for improvement in LBP management. Many clinical trials focus on developing treatments against degenerative disc disease (DDD). The multifactorial etiology of DDD and associated risk factors lead to a heterogeneous patient population. It comes as no surprise that the outcomes of clinical trials on intradiscal mesenchymal stem cell (MSC) injections for patients with DDD are inconsistent. Intradiscal MSC injections have demonstrated substantial pain relief and significant disability-related improvements, yet they have failed to regenerate the intervertebral disc (IVD). Increasing evidence suggests that the positive outcomes in clinical trials might be attributed to the immunomodulatory potential of MSCs rather than to their regenerative properties. Therefore, patient stratification for inflammatory DDD phenotypes may (i) better serve the mechanisms of action of MSCs and (ii) increase the treatment effect. Modic type 1 changes—pathologic inflammatory, fibrotic changes in the vertebral bone marrow—are frequently observed adjacent to degenerated IVDs in chronic LBP patients and represent a clinically distinct subpopulation of patients with DDD. This review discusses whether degenerated IVDs of patients with Modic type 1 changes should be treated with an intradiscal MSC injection.     

Can Gram-Negative Bacteria Develop Resistance to Antimicrobial Blue Light Treatment?

Antimicrobial blue light (aBL) treatment is considered low risk for the development of bacterial resistance and tolerance due to its multitarget mode of action. The aim of the current study was to demonstrate whether tolerance development occurs in Gram-negative bacteria. We evaluated the potential of tolerance/resistance development in Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa and demonstrated that representative Gram-negative bacteria may develop tolerance to aBL. The observed adaption was a stable feature. Assays involving E. coli K-12 tolC-, tolA-, umuD-, and recA-deficient mutants revealed some possible mechanisms for aBL tolerance development.