Fatemeh Amiri

In vitro augmentation of mesenchymal stem cells viability in stressful microenvironments: In vitro augmentation of mesenchymal stem cells viability

Mesenchymal stem cells (MSCs) are under intensive investigation for use in cell-based therapies because their differentiation abilities, immunomodulatory effects, and homing properties offer potential for significantly augmenting regenerative capacity of many tissues. Nevertheless, major impediments to their therapeutic application, such as low proliferation and survival rates remain as obstacles to broad clinical use of MSCs. Another major challenge to evolution of MSC-based therapies is functional degradation of these cells as a result of their exposure to oxidative stressors during isolation. Indeed, oxidative stress-mediated MSC depletion occurs due to inflammatory processes associated with chemotherapy, radiotherapy, and expression of pro-apoptotic factors, and the microenvironment of damaged tissue in patients receiving MSC therapy is typically therapeutic not favorable to their survival. For this reason, any strategies that enhance the viability and proliferative capacity of MSCs associated with their therapeutic use are of great value. Here, recent strategies used by various researchers to improve MSC allograft function are reviewed, with particular focus on in vitro conditioning of MSCs in preparation for clinical application. Preconditioning, genetic manipulation, and optimization of MSC culture conditions are some examples of the methodologies described in the present article, along with novel strategies such as treatment of MSCs with secretome and MSC-derived microvesicles. This topic material is likely to find value as a guide for both research and clinical use of MSC allografts and for improvement of the value that use of these cells brings to health care.

Lipocalin 2 decreases senescence of bone marrow-derived mesenchymal stem cells under sub-lethal doses of oxidative stress

The regenerative potential of mesenchymal stem cells (MSCs) is impaired by cellular senescence, a multi factorial process that has various functions. However, pathways and molecules involved in senescence have not been fully identified. Lipocalin 2 (Lcn2) has been the subject of intensive research, due to its contribution to many physiological and pathophysiological conditions. The implication of Lcn2 has been reported in many conditions where senescence also occurs. In the present study, we evaluated the role of Lcn2 in the occurrence of senescence in human bone marrow-derived mesenchymal stem cells (hB-MSCs) under oxidative conditions. When hB-MSCs were genetically engineered to over-express Lcn2 (MSC-Lcn2) and exposed to H2O2, the proliferation rate of the cells increased. However, the number of colonies and the number of cells that made up each colony in both MSC-V and MSC-Lcn2 cells decreased compared to those cultivated under normal conditions. Our results revealed that over-expression of recombinant Lcn2 in hB-MSCs decreases senescence induced by H2O2 treatment. Senescent cells were observed in aged hB-MSCs; however, no alteration in the expression level of Lcn2 was detected compared to earlier passages. Finally, a higher amount of Lcn2 protein was detected in the plasma of the elderly than in young people. Our findings suggest that Lcn2 might restore the health and regeneration potential of MSCs by decreasing senescence.

Down-regulation of the autophagy gene, ATG7, protects bone marrow-derived mesenchymal stem cells from stressful conditions.

Background Mesenchymal stem cells (MSCs) are valuable for cell-based therapy. However, their application is limited owing to their low survival rate when exposed to stressful conditions. Autophagy, the process by which cells recycle the cytoplasm and dispose of defective organelles, is activated by stress stimuli to adapt, tolerate adverse conditions, or trigger the apoptotic machinery. This study aimed to determine whether regulation of autophagy would affect the survival of MSCs under stress conditions. Methods Autophagy was induced in bone marrow-derived MSCs (BM-MSCs) by rapamycin, and was inhibited via shRNA-mediated knockdown of the autophagy specific gene, ATG7. ATG7 expression in BM-MSCs was evaluated by reverse transcription polymerase chain reaction (RT-PCR), western blot, and quantitative PCR (qPCR). Cells were then exposed to harsh microenvironments, and a water-soluble tetrazolium salt (WST)-1 assay was performed to determine the cytotoxic effects of the stressful conditions on cells. Results Of 4 specific ATG7-inhibitor clones analyzed, only shRNA clone 3 decreased ATG7 expression. Under normal conditions, the induction of autophagy slightly increased the viability of MSCs while autophagy inhibition decreased their viability. However, under stressful conditions such as hypoxia, serum deprivation, and oxidative stress, the induction of autophagy resulted in cell death, while its inhibition potentiated MSCs to withstand the stress conditions. The viability of autophagy-suppressed MSCs was significantly higher than that of relevant controls (P<0.05, P<0.01 and P<0.001). Conclusion Autophagy modulation in MSCs can be proposed as a new strategy to improve their survival rate in stressful microenvironments.

Autophagy-Modulated Human Bone Marrow-Derived Mesenchymal Stem Cells Accelerate Liver Restoration in Mouse Models of Acute Liver Failure

Background: Mesenchymal stem cells (MSCs) have been recently received increasing attention for cell-based therapy, especially in regenerative medicine. However, the low survival rate of these cells restricts their therapeutic applications. It is hypothesized that autophagy might play an important role in cellular homeostasis and survival. This study aims to investigate the regenerative potentials of autophagy-modulated MSCs for the treatment of acute liver failure (ALF) in mice. Methods: ALF was induced in mice by intraperitoneal injection of 1.5 ml/kg carbon tetrachloride. Mice were intravenously infused with MSCs, which were suppressed in their autophagy pathway. Blood and liver samples were collected at different intervals (24, 48 and 72 h) after the transplantation of MSCs. Both the liver enzymes and tissue necrosis levels were evaluated using biochemical and histopathological assessments. The survival rate of the transplanted mice was also recorded during one week. Results: Biochemical and pathological results indicated that 1.5 ml/kg carbon tetrachloride induces ALF in mice. A significant reduction of liver enzymes and necrosis score were observed in autophagy-modulated MSC-transplanted mice compared to sham (with no cell therapy) after 24 h. After 72 h, liver enzymes reached their normal levels in mice transplanted with autophagy-suppressed MSCs. Interestingly, normal histology without necrosis was also observed. Conclusion: Autophagy suppression in MSCs ameliorates their liver regeneration potentials due to paracrine effects and might be suggested as a new strategy for the improvement of cell therapy in ALF.

Targeting delivery of lipocalin 2-engineered mesenchymal stem cells to colon cancer in order to inhibit liver metastasis in nude mice.

One of the major obstacles in cancer therapy is the lack of anticancer agent specificity to tumor tissues. The strategy of cell-based therapy is a promising therapeutic option for cancer treatment. The specific tumor-oriented migration of mesenchymal stem cells (MSCs) makes them a useful vehicle to deliver anticancer agents. In this study, we genetically manipulated bone marrow-derived mesenchymal stem cells with their lipocalin 2 (Lcn2) in order to inhibit liver metastasis of colon cancer in nude mice. Lcn2 was successfully overexpressed in transfected MSCs. The PCR results of SRY gene confirmed the presence of MSCs in cancer liver tissue. This study showed that Lcn2-engineered MSCs (MSC-Lcn2) not only inhibited liver metastasis of colon cancer but also downregulated the expression of vascular endothelial growth factor (VEGF) in the liver. Overall, MSCs by innate tropism toward cancer cells can deliver the therapeutic agent, Lcn2, and inhibit cancer metastasis. Hence, it could be a new modality for efficient targeted delivery of anticancer agent to liver metastasis.

Positive selection of Wharton's jelly-derived CD105+ cells by MACS technique and their subsequent cultivation under suspension culture condition: A simple, versatile culturing method to enhance the multipotentiality of mesenchymal stem cells

Abstract Objective Whartons jelly (WJ), an appropriate source of mesenchymal stem cells (MSCs), has been shown to have a wide array of therapeutic applications. However, the WJ-derived MSCs are very heterogeneous and have limited expression of pluripotency markers. Hence, improvement of their culture condition would promote the efficiency of WJ-MSCs. This study aims to employ a simple method of cultivation to obtain WJ-MSCs which express more pluripotency markers. Methods CD105+ cells were separated by magnetic-associated (activated) cell sorting from umbilical cord mucous tissue. CD105+ cells were added to Methocult medium diluted in α-minimum essential medium (α-MEM) and seeded in poly(2-hydroxyethyl methacrylate) (poly-HEMA)-coated plates for suspension culture preparation. Differentiation capacity of isolated cells was evaluated in the presence of differentiation-inducing media. The expression of pluripotency markers such as Oct3/4, Nanog, and Sox2 was also analyzed by RT-PCR and western blot techniques. Moreover, immunocytochemistry was performed to detect alpha-smooth muscle actin (antigene) (α-SMA) protein. Results WJ-MSCs grew homogeneously and formed colonies when cultured under suspension culture conditions (Non-adhesive WJ-MSCs). They maintained their growth ability in both adherent and suspension cultures for several passages. Non-adhesive WJ-MSCs expressed Oct3/4, Nanog, and Sox2 both at transcriptional and translational levels in comparison to those cultured in conventional adherent cultures. They also expressed α-SMA protein. Discussion In this study, we isolated WJ-MSCs using a slightly modified culture condition. Our simple non-genetic method resulted in a homogeneous population of WJ-MSCs, which highly expressed pluripotency markers. Conclusion In the future, more multipotent WJ-MSCs can be harnessed as a non-embryonic source of MSCs in MSC-based cell therapy.

Thermodynamics of the Spin-1/2 Two-Leg Ladder Compound (C_5H_{12}N)_2CuBr_4

The thermodynamic behavior of the spin

Lipocalin 2 enhances mesenchymal stem cell-based cell therapy in acute kidney injury rat model

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