Comparison the effects of hypoxia-mimicking agents on migration-related signaling pathways in mesenchymal stem cells.

Adipose-derived mesenchymal stem cells (Ad-MSCs) have been designated as the promising agents for clinical applications for easy accessibility, multi-linage differentiation and immunomodulation capacity. Despite this, optimal cell delivery conditions have remained as a clinical challenge and improvement of stem cell homing to the target organs is being considered as a major strategy in cell therapy systemic injection. It has been shown that homing of mesenchymal stem cells are increased when treated with physical or chemical hypoxia-mimicking factors, however, efficiency of different agents remained to be determined. In this study, hypoxia-mimicking agents, including valproic acid (VPA), cobalt chloride (CoCl2) and deferoxamine (DFX) were examined to determine whether they are able to activate signaling molecules involved in migration of Ad-MSCs in vitro. We report that Ad-MSCs treated by DFX resulted in a significantly enhanced mRNA expression of MAPK4 (associated with MAPK signaling pathway), INPP4B (associated with Inositol polyphosphate pathway), VEGF-A and VEGF-C (associated with cytokine-cytokine receptor pathways), IL-8 and its receptor, CXCR2 (associated with IL-8 signaling pathway). While the cells treated with VPA did not show such effects and CoCl2 only upregulated VEGF-A and VEGF-C gene expression. Furthermore, results of wound-healing assays showed migration capacity of Ad-MSCs treated with DFX significantly increased 8 and 24 h of the treatment. This study provides credible evidence around DFX, which might be an effective drug for pharmacological preconditioning of Ad-MSCs to boost their homing capacity and regeneration of damaged tissues though, activation of the migration-related signaling pathways.

Overexpression of MicroRNA-148b-3p stimulates osteogenesis of human bone marrow-derived mesenchymal stem cells: the role of MicroRNA-148b-3p in osteogenesis.

BACKGROUND: Mesenchymal stem cells (MSCs) are attractive choices in regenerative medicine and can be genetically modified to obtain better results in therapeutics. Bone development and metabolism are controlled by various factors including microRNAs (miRs) interference, which are small non-coding endogenous RNAs. METHODS: In the current study, the effects of forced miR-148b expression was evaluated on osteogenic activity. Human bone marrow-derived mesenchymal stem cells (BM-MSCs) were transduced with bicistronic lentiviral vector encoding hsa-miR-148b-3p or -5p and the enhanced green fluorescent protein. Fourteen days post-transduction, immunostaining as well as Western blotting were used to analyze osteogenesis. RESULTS: Overexpression of miR-148b-3p increased the osteogenic differentiation of human BM-MSCs as demonstrated by anenhancement of mineralized nodular formation and an increase in the levels of osteoblastic differentiation biomarkers, alkaline phosphatase and collagen type I. CONCLUSIONS: Since lentivirally overexpressed miR-148b-3p increased osteogenic differentiation capability of BM-MSCs, this miR could be applied as a therapeutic modulator to optimize bone function.

Phytosomal-curcumin antagonizes cell growth and migration, induced by thrombin through AMP-Kinase in breast cancer.

Here we explored the antitumor-activity of novel-formulated-form of curcumin (phytosomal-encapsulated-curcumin) or in combination with 5-FU in breast cancer. The antiproliferative activity was assessed in 2D and 3-dimensional cell-culture-model. The migratory-behaviors of the cells were determined by migration assay. The expression levels of CyclinD1,GSK3a/b, P-AMPK, MMP9, and E-cadherin were studied by qRT-PCR and/or Western blotting. The anti-inflammatory of nano-curcumin was assessed, while antioxidant activity was evaluated by malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), and total thiols (T-SH). To understand dynamic behavior of genes, we reconstructed a Boolean network, while the robustness of this model was evaluated by Hamming distance. phytosomal-curcumin suppressed cell-growth followed by tumor-shrinkage in 3D model through perturbation of AMP-activated protein kinase. Curcumin reduced the invasiveness of MCF-7 through perturbation of E-cadherin. Moreover, phytosomal-curcumin inhibited the tumor growth in xerograph model. Histological staining of tumor tissues revealed vascular disruption and RBC extravasation, necrosis, tumor stroma, and inflammation. Co-treatment of curcumin and 5-FU reduced the lipid-peroxidation and increased MDA/SOD level. Of note, curcumin reduced cyclinD-expression in breast cancer cell treated with thrombin, and activates AMPK in a time-dependent manner. Also suppression of AMPK abrogated inhibitory effect of phytosomal-curcumin on thrombin-induced cyclin D1 over-expression, suggesting that AMPK is essential for anti-proliferative effect of this agent in breast cancer. Our finding demonstrated that phytosomal-curcumin antagonizes cell growth and migration, induced by thrombin through AMP-Kinase in breast cancer, supporting further-investigations on the therapeutic potential of this novel anticancer agent in treatment of breast cancer.

Suppression of dsRNA response genes and innate immunity following Oct4, Stella, and Nanos2 overexpression in mouse embryonic fibroblasts.

The self-renewal capacity of germline derived stem cells (GSCs) makes them an ideal source for research and use in clinics. Despite the presence of active gene network similarities between embryonic stem cells (ESCs) and GSCs, there are unanswered questions regarding the roles of evolutionary conserved genes in GSCs. To determine the reprogramming potential of germ cell- specific genes, we designed a polycistronic gene cassette expressing Stella, Oct4 and Nanos2 in a lentiviral-based vector. Deep transcriptome analysis showed the activation of a set of pluripotency and germ-cell-specific markers and the downregulation of innate immune system. The global shut down of antiviral genes included MHC class I, interferon response genes and dsRNA 2'-5'-oligoadenylate synthetase are critical pathways that has been affected . Individual expression of each factor highlighted suppressive effect of Nanos2 on genes such as Isg15 and Oasl2. Collectively, to our knowledge this is the first report showing that Nanos2 could be considered as an immunosuppressive factor. Furthermore, our results demonstrate suppression of endogenous retrotransposons that harbor immune response but further analysis require to uncover the correlation between transposon suppression and immune response in germ cell development.

Using paracrine effects of Ad-MSCs on keratinocyte cultivation and fabrication of epidermal sheets for improving clinical applications.

Recent advances in wound healing have made cell therapy a potential approach for the treatment of various types of skin defects such as trauma, burns, scars and diabetic leg ulcers. Cultured keratinocytes have been applied to burn patients since 1981. Patients with acute and chronic wounds can be treated with autologous/allograft cultured keratinocytes. There are various methods for cultivation of epidermal keratinocytes used in cell therapy. One of the important properties of an efficient cell therapy is the preservation of epidermal stem cells. Mesenchymal Stem Cells (MSCs) are major regulatory cells involved in the acceleration of wound healing via induction of cell proliferation, angiogenesis and stimulating the release of paracrine signaling molecules. Considering the beneficial effects of MSCs on wound healing, the main aim of the present study is investigating paracrine effects of Adipose-derived Mesenchymal Stem Cell (Ad-MSCs) on cultivation of keratinocytes with focusing on preservation of stem cells and their differentiation process. We further introduced a new approach for culturing isolated keratinocytes in vitro in order to generate epidermal keratinocyte sheets without using a feeder layer. To do so, Ad-MSC conditioned medium was applied as an alternative to commercial media for keratinocyte cultivation. In this study, the expression of several stem/progenitor cell (P63, K19 and K14) and differentition (K10, IVL and FLG) markers was examined using real time PCR on days 7, 14 and 21 of culture in keratinocytes in Ad-MSC conditioned medium. P63 and α6 integrin expression was also evaluated via flow cytometry. The results were compared with control group including keratinocytes cultured in EpiLife medium and our data indicated that this Ad-MSC conditioned medium is a good alternative for keratinocyte cultivation and producing epidermal sheets for therapeutic and clinical purposes. The reasons are the expression of stem cell and differentiation markers and overcoming the requirement for feeder layer which leads to a xenograft-free transplantation. Besides, this approach has low cost and is easier to perform. However, more in vitro and in vivo experiments as well as safety evaluation required before clinical applications.

Long bone mesenchymal stem cells (Lb-MSCs): clinically reliable cells for osteo-diseases.

Mesenchymal stem cells (MSCs) have been designated as the most reliable cells in clinics to treat osteo-diseases because of their versatile nature. MSCs, isolated from long bone (Lb-MSCs) are rarely reported and named as RIA-MSCs because of the reamer-irrigator-aspirator (RIA) device. The potential of these cells in the treatment of non-union bone fractures made them the ideal candidates to be studied for clinical practices. In this work, effect of cryopreservation on the proliferation and differentiation capabilities of long bone MSCs (Lb-MSCs) has been studied. For this purpose, Lb-MSCs were isolated via RIA device and characterized using flow cytometry and differentiation assays. Cells were cryopreserved for 3, 6 and 12 months and thereafter were characterized using differentiation assays and genetic markers specific for osteogenic, chondrogenic, and adipogenic potential quantitatively by qRT-PCR. Lb-MSCs were found expressing MSC characteristic markers defining their identity. The population doubling time (PDT) was about 2.5 ± 0.5 days and colonies appeared after 7-10 days. Differentiation potential and gene expression of 3, 6 and 12 months cryopreserved Lb-MSCs were unaltered. The results show that cryopreservation did not have an effect on the differentiation potential of human Lb-MSCs. Therefore, our work offers Lb-MSCs as clinically cells for treating osteo-diseases.

Pluripotency induction in HEK293T cells by concurrent expression of STELLA, OCT4 and NANOS2.

Germline stem cells (GSCs) are attractive biological models because of their strict control on pluripotency gene expression, and their potential for huge epigenetic changes in a short period of time. Few data exists on the cooperative impact of GSC-specific genes on differentiated cells. In this study, we over-expressed 3 GSC-specific markers, STELLA, OCT4 and NANOS2, collectively designated as (SON), using the novel polycistronic lentiviral gene construct FUM-FD, in HEK293T cells and evaluated promoter activity of the Stra8 GSC marker gene We could show that HEK293T cells expressed pluripotency and GSC markers following ectopic expression of the SON genes. We also found induction of pluripotency markers after serum starvation in non-transduced HEK293T cells. Expression profiling of SON-expressing and serum-starved cells at mRNA and protein level showed the potential of SON factors and serum starvation in the induction of ESRRB, NANOG, OCT4 and REX1 expression. Additionally, the data indicated that the mouse Stra8 promoter could only be activated in a subpopulation of HEK293T cells, regardless of SON gene expression. We conclude that heterogeneous population of the HEK293T cells might be easily shifted towards expression of the pluripotency markers by ectopic expression of the SON factors or by growth in serum depleted media.

Injectable hydrogel delivery plus preconditioning of mesenchymal stem cells: exploitation of SDF-1/CXCR4 axis toward enhancing the efficacy of stem cells' homing.

Clinical applications of mesenchymal stem cells (MSCs) rely on their capacity to home and engraft in the appropriate target injury tissues for the long term. However, their homing efficiency has been observed to be very poor because of the lack or modifications of homing factors SDF-1α and CXCR4 receptors. Hence, this study was designed to investigate the homing and retention of pretreated human adipose tissue-derived MSCs (hASCs) from three different delivery routes in response to SDF-1α, released from chitosan-based injectable hydrogels. After stimulation of ASCs with a hypoxia mimicking agent, the expression level and functionality of CXCR4 were analyzed by flowcytometric analysis (FACS), transwell migration assay and qPCR. Then, the homing/retention of pretreated DiI-labeled hASCs were compared through three different in vivo delivery routes, 2 weeks after transplantation in Wistar rats. The cells were tracked histologically by fluorescent microscope and by PCR for human-specific CXCR4 gene. Results showed CXCR4 has dynamic expression pattern and pretreatment of hASCs significantly up-regulates CXCR4, leading to an increase in migration capacity toward 100 ng/mL SDF-1α in vitro and homing into the subcutaneously implanted hydrogel releasing SDF-1α in vivo. Furthermore, it seems that SDF-1α is particularly important in the retention of ASCs, in addition to its chemoattraction role. In summary, the delivery route in which the ASCs were mixed with the hydrogel rather than systemic delivery and local injection and preconditioning undertaken to increase CXCR4 expression concomitant with SDF-1α delivery by the injectable hydrogel, allowed for further homing/retention of ASCs. This might be a promising way to get better therapeutic outcomes in stem cell therapy.

Strategies to improve homing of mesenchymal stem cells for greater efficacy in stem cell therapy.

Stem/progenitor cell-based therapeutic approach in clinical practice has been an elusive dream in medical sciences, and improvement of stem cell homing is one of major challenges in cell therapy programs. Stem/progenitor cells have a homing response to injured tissues/organs, mediated by interactions of chemokine receptors expressed on the cells and chemokines secreted by the injured tissue. For improvement of directed homing of the cells, many techniques have been developed either to engineer stem/progenitor cells with higher amount of chemokine receptors (stem cell-based strategies) or to modulate the target tissues to release higher level of the corresponding chemokines (target tissue-based strategies). This review discusses both of these strategies involved in the improvement of stem cell homing focusing on mesenchymal stem cells as most frequent studied model in cellular therapies.

Extracellular vesicles and their cells of origin: Open issues in autoimmune diseases.

The conventional therapeutic approaches to treat autoimmune diseases through suppressing the immune system, such as steroidal and non-steroidal anti-inflammatory drugs, are not adequately practical. Moreover, these regimens are associated with considerable complications. Designing tolerogenic therapeutic strategies based on stem cells, immune cells, and their extracellular vesicles (EVs) seems to open a promising path to managing autoimmune diseases' vast burden. Mesenchymal stem/stromal cells (MSCs), dendritic cells, and regulatory T cells (Tregs) are the main cell types applied to restore a tolerogenic immune status; MSCs play a more beneficial role due to their amenable properties and extensive cross-talks with different immune cells. With existing concerns about the employment of cells, new cell-free therapeutic paradigms, such as EV-based therapies, are gaining attention in this field. Additionally, EVs' unique properties have made them to be known as smart immunomodulators and are considered as a potential substitute for cell therapy. This review provides an overview of the advantages and disadvantages of cell-based and EV-based methods for treating autoimmune diseases. The study also presents an outlook on the future of EVs to be implemented in clinics for autoimmune patients.

Introns and Their Therapeutic Applications in Biomedical Researches.

Context: Although for a long time, it was thought that intervening sequences (introns) were junk DNA without any function, their critical roles and the underlying molecular mechanisms in genome regulation have only recently come to light. Introns not only carry information for splicing, but they also play many supportive roles in gene regulation at different levels. They are supposed to function as useful tools in various biological processes, particularly in the diagnosis and treatment of diseases. Introns can contribute to numerous biological processes, including gene silencing, gene imprinting, transcription, mRNA metabolism, mRNA nuclear export, mRNA localization, mRNA surveillance, RNA editing, NMD, translation, protein stability, ribosome biogenesis, cell growth, embryonic development, apoptosis, molecular evolution, genome expansion, and proteome diversity through various mechanisms. Evidence Acquisition: In order to fulfill the objectives of this study, the following databases were searched: Medline, Scopus, Web of Science, EBSCO, Open Access Journals, and Google Scholar. Only articles published in English were included. Results & Conclusions: The intervening sequences of eukaryotic genes have critical functions in genome regulation, as well as in molecular evolution. Here, we summarize recent advances in our understanding of how introns influence genome regulation, as well as their effects on molecular evolution. Moreover, therapeutic strategies based on intron sequences are discussed. According to the obtained results, a thorough understanding of intron functional mechanisms could lead to new opportunities in disease diagnosis and therapies, as well as in biotechnology applications.

The Effects of Lycopene on Modulating Oxidative Stress and Liver Enzymes Levels in Metabolic Syndrome Patients: A Randomised Clinical Trial.

OBJECTIVE: The pathogenesis of metabolic syndrome (MetS) complications involves the excessive production of
reactive oxygen species, inflammation, and endothelial dysfunction. Due to Lycopene, a highly unstable structure and
its significant effects on modulating the metabolic system, there is a strong need for a formula that can increase its
stability. The aim of this study was to develop an approach for encapsulating Lycopene and investigate its effects on
inflammatory markers, oxidative stress, and liver enzymes in patients with MetS.
Materials and Methods: This study is a simple randomized, double-blind, objective-based clinical trial that involved
eighty subjects with MetS, who were equally and randomly assigned to two groups: one group received 20 mg of
Lycopene per day for 8 weeks, and the Placebo group followed the same protocol as the Lycopene group but received
a placebo instead of Lycopene. They were called Lycopene and placebo, respectively. During follow-up visits after 4
and 8 weeks, 20 ml of blood was collected for evaluation of liver enzymes and some inflammatory related markers.
Results: Prior to the assignment of volunteers to their respective groups, there were no notable differences in C-reactive
protein (CRP), serum liver enzymes, systolic and diastolic blood pressure, or pro-oxidant-antioxidant balance (PAB)
between the Lycopene and placebo groups. However, our subsequent analysis revealed a significant reduction in the
serum levels of CRP (P=0.001) and PAB (P=0.004) in the group that received Lycopene. Our encapsulated Lycopene
treatment was not associated with a significant difference in serum levels of alanine aminotransferase (ALT), aspartate
transferase (AST), or alkaline phosphatase (ALP) between our two groups.
Conclusion: This study investigated the impact of Lycopene on individuals with MetS, revealing a noteworthy
modulation effect on PAB and inflammation linked to MetS. However, no significant differences was demonstrated in
serum levels of ALT, AST and ALP between the studied group (registration number: IRCT20130507013263N3).

Targeting transforming growth factor beta (TGF-ß) using Pirfenidone, a potential repurposing therapeutic strategy in colorectal cancer.

The modulating factors within the tumor microenvironment, for example, transforming growth factor beta (TGF-ß), may limit the response to chemo and immunotherapy protocols in colorectal cancer (CRC). In the current study, the therapeutic potential of targeting the TGF-ß pathway using Pirfenidone (PFD), a TGF-ß inhibitor, either alone or in combination with five fluorouracil (5-FU) has been explored in preclinical models of CRC. The anti-proliferative and migratory effects of PFD were assessed by MTT and wound-healing assays respectively. Xenograft models were used to study the anti-tumor activity, histopathological, and side effects analysis. Targeting of TGF-ß resulted in suppression of cell proliferation and migration, associated with modulation of survivin and MMP9/E-cadherin. Moreover, the PFD inhibited TGF-ß induced tumor progression, fibrosis, and inflammatory response through perturbation of collagen and E-cadherin. Targeting the TGF-ß pathway using PFD may increase the anti-tumor effects of 5-FU and reduce tumor development, providing a new therapeutic approach to CRC treatment.

New windows to enhance direct reprogramming of somatic cells towards induced pluripotent stem cells.

Induced pluripotent stem cells are generated by direct reprogramming of somatic cells with the introduction of defined transcription factors or other means. Clinical applications of induced pluripotent stem cells are the latest of stem cell therapy approaches due to overcoming problems associated with insufficient cells from conventional sources and immune rejections. In practice, this is restricted by 4 major barriers including the use of genetic manipulations for delivering the reprogramming factors, low efficiency of this process, slow kinetics of the direct reprogramming, and potential for tumor development. Here, we review the latest achievements in improving reprogramming efficiency by alternative strategies. These alternatives mainly involve the replacement of genetic reprogramming factors with small molecules or other factors.

Mesenchymal Stem/Stromal Cells Overexpressing CXCR4R334X Revealed Enhanced Migration: A Lesson Learned from the Pathogenesis of WHIM Syndrome.

C-X-C chemokine receptor type 4 (CXCR4), initially recognized as a co-receptor for HIV, contributes to several disorders, including the WHIM (Warts, Hypogammaglobulinemia, Infections, and Myelokathexis) syndrome. CXCR4 binds to its ligand SDF-1 to make an axis involved in the homing property of stem cells. This study aimed to employ WHIM syndrome pathogenesis as an inspirational approach to reinforce cell therapies. Wild type and WHIM-type variants of the CXCR4 gene were chemically synthesized and cloned in the pCDH-513B-1 lentiviral vector. Molecular cloning of the synthetic genes was confirmed by DNA sequencing, and expression of both types of CXCR4 at the protein level was confirmed by western blotting in HEK293T cells. Human adipose-derived mesenchymal stem cells (Ad-MSCs) were isolated, characterized, and subjected to lentiviral transduction with Wild type and WHIM-type variants of CXCR4. The presence of copGFP-positive MSCs confirmed the high efficiency of transduction. The migration ability of both groups of transduced cells was then assessed by transwell migration assay in the presence or absence of a CXCR4-blocking agent. Our qRT-PCR results showed overexpression of CXCR4 at mRNA level in both groups of transduced MSCs, and expression of WHIM-type CXCR4 was significantly higher than Wild type CXCR4 (P<0.05). Our results indicated that the migration of genetically modified MSCs expressing WHIM-type CXCR4 had significantly enhanced towards SDF1 in comparison with Wild type CXCR4 (P<0.05), while it was reduced after treatment with CXCR4 antagonist. These data suggest that overexpression of WHIM-type CXCR4 could lead to enhanced and sustained expression of CXCR4 on human MSCs, which would increase their homing capability; hence it might be an appropriate strategy to improve the efficiency of cell-based therapies.

Potential inhibitory effect of lycopene on prostate cancer.

Studying prostate cancer is important due to its high annual incidences and mortality rates in the world. Although prostate cancer mortality rates are reduced using new therapy, complicated routes and side effects of these current drugs require a daily available treatment for prevention. Lycopene is a natural, prominent, and effective product which has a high value in diet. The anti-cancer effect, non-toxicity, safety and preventive or therapeutic roles of lycopene have been investigated in several studies. In the current review, we have collected information about the anti-cancer, anti-progressive and apoptotic effects of lycopene on prostate cancer. This article is a summary of the most important original and review articles on lycopene and its anticancer effects that are systematically categorized and presents information about the molecular structure, different sources, biological functions, and its in-vivo and in-vitro effects of lycopene on variety of cancerous and normal cells. The clinical studies provide a clear image for continuous use of this adjunctive dietary for different type of cancers, especially prostate cancer in men. In addition, this article discusses the various molecular pathways activated by lycopene that eventually prevent or suppress cancer. Lycopene has been found to effectively suppress the progression and proliferation, arrest in-cell cycle, and induce apoptosis of prostate cancer cells in both in-vivo and in-vitro conditions. Additionally, lycopene showed that it could modulate the signaling pathways and their protein for the treatment or prevention of prostate cancer.

MTA Enhances the Potential of Adipose-Derived Mesenchymal Stem Cells for Dentin-Pulp Complex Regeneration.

The aim of the current study was to investigate the effects of mineral trioxide aggregate (MTA) on the proliferation and differentiation of human adipose-derived mesenchymal stem cells (Ad-MSCs) as a surrogate cell source in futuristic stem-cell-based endodontic therapies. Human Ad-MSCs and mesenchymal stem cells derived from bone marrow (BM-MSCs) were isolated from liposuction waste adipose tissue and femur, respectively, and the effects of MTA-conditioned media on their viability, mineralization potential, and osteo/odontogenic differentiation capacity were subsequently evaluated. Alkaline phosphatase (ALP) activity, quantitative alizarin red S staining, and quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analyses were performed to investigate and compare the osteo/odontogenic induction potential of MTA on the Ad/BM-MSCs. The results of cytotoxicity assay revealed that at different concentrations, MTA-conditioned medium was not only biocompatible toward both cell types, but also capable of promoting cell proliferation. ALP activity assay showed that 0.2 mg/mL was the optimal concentration of MTA-conditioned medium for osteo/odontogenic induction in Ad/BM-MSCs. The expression of osteo/odontogenic gene markers was increased in Ad/BM-MSCs treated with 0.2 mg/mL MTA-conditioned media. Our results indicated that MTA can efficiently enhance the osteo/odontogenic potential of Ad-MSCs, and thus they can be considered as a better cell source for dentin-pulp complex regeneration. However, further investigations are required to test these potentials in animal models.

Application of mesenchymal stem cells to enhance non-union bone fracture healing.

ECM components include a number of osteoinductive and osteoconductive factors, which are involved in bone fracture healing. In this study, a combination of adipose derived mesenchymal stem cells (Ad-MSCs), cancellous bone graft (CBG), and chitosan hydrogel (CHI) was applied to the non-union bone fracture and healing effects were evaluated for the first time. After creation of animal models with non-union fracture in rats, they were randomly classified into seven groups. Radiography at 0, 2, 4, and 8 weeks after surgery, indicated the positive effects of Ad-MSCs + CBG + CHI and Ad-MSCs + CBG in treatment of bone fractures as early as 2 weeks after the surgery. These data were confirmed with both biomechanical and histological studies. Gene expression analyses of Vegf and Bmp2 showed a positive effect of Ad-MSCs on vascularization and osteogenic differentiation in all groups receiving Ad-MSCs, as shown by real-time PCR. Immunofluorescence analysis and RT-PCR results indicated existence of human Ad-MSCs in the fractured region 8 weeks post-surgery. In conclusion, we suggest that application of Ad-MSCs, CBG, and CHI, could be a suitable combination for osteoinduction and osteoconduction to improve non-union bone fracture healing. Further investigations are required to determine the exact mechanisms involved in this process before moving to clinical studies. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 107A: 301-311, 2019.

Augmented migration of mesenchymal stem cells correlates with the subsidiary CXCR4 variant.

Use of mesenchymal stem cells (MSCs) has been introduced as a promising tool, for structural and functional recovery of damaged tissues/organs. Studies have indicated that interactions between chemokine receptors and their ligands have a critical role in homing of MSCs to the site of injury. Although CXCR4 variants have been characterized, the exact role of each transcript in homing has remained unclear. In this study, cells were pretreated with various hypoxia-mimicking compounds (valproic acid, cobalt-chloride, and deferoxamine mesylate). Results indicated that both variants of CXCR4 were overexpressed after 24 hours of treatments and their expression could cooperatively induce and promote the cell migration. Moreover, deferoxamine mesylate was more effective in overexpression of variant A (lo), which resulted in higher level of CXCR4 protein and the highest rate of migration of the cells. In conclusion, our findings may have important potential implications in clinical applications, reinforcing the concept that manipulating the expression of specific CXCR4 variants may increase migration of MSCs.

MicroRNA-499a-5p Promotes Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells to Cardiomyocytes.

Since the adult mammalian heart has limited regenerative capacity, cardiac trauma, disease, and aging cause permanent loss of contractile tissue. This has fueled the development of stem cell-based strategies to provide the damaged heart with new cardiomyocytes. Bone marrow-derived mesenchymal stem cells (BM-MSCs) are capable of self-renewal and differentiation into cardiomyocytes, albeit inefficiently. MicroRNAs (miRNAs, miRs) are non-coding RNAs that have the potential to control stem cell fate decisions and are employed in cardiac regeneration and repair. In this study, we tested the hypothesis that overexpression of miR-499a induces cardiomyogenic differentiation in BM-MSCs. Human BM-MSCs (hBM-MSCs) were transduced with lentiviral vectors encoding miR-499a-3p or miR-499a-5p and analyzed by immunostaining and western blotting methods 14 days post-transduction. MiR-499a-5p-transduced cells adopted a polygonal/rod-shaped (myocyte-like) phenotype and showed an increase in the expression of the cardiomyocyte markers α-actinin and cTnI, as cardiogenic differentiation markers. These results indicate that miR-499a-5p overexpression promotes the cardiomyogenic differentiation of hBM-MSCs and may thereby increase their therapeutic efficiency in cardiac regeneration.