scholarly journals Nonviral Gene Delivery to Mesenchymal Stem Cells Using Cationic Liposomes for Gene and Cell Therapy

2010 ◽  
Vol 2010 ◽  
pp. 1-12 ◽  
Author(s):  
C. Madeira ◽  
R. D. Mendes ◽  
S. C. Ribeiro ◽  
J. S. Boura ◽  
M. R. Aires-Barros ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Gene Delivery ◽  
Cationic Liposome ◽  
Nonviral Gene Delivery ◽  
Great Promise ◽  
Full Potential ◽  
Genetically Engineer ◽  
Or Gene ◽  
First Time

Mesenchymal stem cells (MSCs) hold a great promise for application in several therapies due to their unique biological characteristics. In order to harness their full potential in cell-or gene-based therapies it might be advantageous to enhance some of their features through gene delivery strategies. Accordingly, we are interested in developing an efficient and safe methodology to genetically engineer human bone marrow MSC (BM MSC), enhancing their therapeutic efficacy in Regenerative Medicine. The plasmid DNA delivery was optimized using a cationic liposome-based reagent. Transfection efficiencies ranged from ~2% to ~35%, resulting from using a Lipid/DNA ratio of 1.25 with a transgene expression of 7 days. Importantly, the number of plasmid copies in different cell passages was quantified for the first time and ~20,000 plasmid copies/cell were obtained independently of cell passage. As transfected MSC have shown high viabilities (>90%) and recoveries (>52%) while maintaining their multipotency, this might be an advantageous transfection strategy when the goal is to express a therapeutic gene in a safe and transient way.

scholarly journals Enhanced PRL-1 expression in placenta-derived mesenchymal stem cells accelerates hepatic function via mitochondrial dynamics in a cirrhotic rat model

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Jae Yeon Kim ◽  
Jong Ho Choi ◽  
Ji Hye Jun ◽  
Sohae Park ◽  
Jieun Jung ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Gene Delivery ◽  
Cell Therapy ◽  
Delivery Systems ◽  
Nonviral Gene Delivery ◽  
Early Gene ◽  
Next Generation ◽  
Atp Production ◽  
Mitochondrial Functions

Abstract Background Placenta-derived mesenchymal stem cells (PD-MSCs) have been highlighted as an alternative cell therapy agent that has become a next-generation stem cell treatment. Phosphatase of regenerating liver-1 (PRL-1), an immediate early gene, plays a critical role during liver regeneration. Here, we generated enhanced PRL-1 in PD-MSCs (PD-MSCsPRL-1, PRL-1+) using lentiviral and nonviral gene delivery systems and investigated mitochondrial functions by PD-MSCPRL-1 transplantation for hepatic functions in a rat bile duct ligation (BDL) model. Methods PD-MSCsPRL-1 were generated by lentiviral and nonviral AMAXA gene delivery systems and analyzed for their characteristics and mitochondrial metabolic functions. Liver cirrhosis was induced in Sprague-Dawley (SD) rats using common BDL for 10 days. PKH67+ naïve and PD-MSCsPRL-1 using a nonviral sysyem (2 × 106 cells/animal) were intravenously administered into cirrhotic rats. The animals were sacrificed at 1, 2, 3, and 5 weeks after transplantation and engraftment of stem cells, and histopathological analysis and hepatic mitochondrial functions were performed. Results PD-MSCsPRL-1 were successfully generated using lentiviral and nonviral AMAXA systems and maintained characteristics similar to those of naïve cells. Compared with naïve cells, PD-MSCsPRL-1 improved respirational metabolic states of mitochondria. In particular, mitochondria in PD-MSCsPRL-1 generated by the nonviral AMAXA system showed a significant increase in the respirational metabolic state, including ATP production and mitochondrial biogenesis (*p < 0.05). Furthermore, transplantation of PD-MSCsPRL-1 using a nonviral AMAXA system promoted engraftment into injured target liver tissues of a rat BDL cirrhotic model and enhanced the metabolism of mitochondria via increased mtDNA and ATP production, thereby improving therapeutic efficacy. Conclusions Our findings will further our understanding of the therapeutic mechanism of enhanced MSCs and provide useful data for the development of next-generation MSC-based cell therapy and therapeutic strategies for regenerative medicine in liver disease.

scholarly journals Nonviral Gene Delivery of Growth and Differentiation Factor 5 to Human Mesenchymal Stem Cells Injected into a 3D Bovine Intervertebral Disc Organ Culture System

2013 ◽  
Vol 2013 ◽  
pp. 1-9 ◽  
Author(s):  
Christian Bucher ◽  
Amiq Gazdhar ◽  
Lorin M. Benneker ◽  
Thomas Geiser ◽  
Benjamin Gantenbein-Ritter
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Gene Transfer ◽  
Gene Delivery ◽  
Intervertebral Disc ◽  
Organ Culture ◽  
Nonviral Gene Delivery ◽  
Human Mscs ◽  
Differentiation Factor ◽  
Gdf5 Gene

Intervertebral disc (IVD) cell therapy with unconditioned 2D expanded mesenchymal stem cells (MSC) is a promising concept yet challenging to realize. Differentiation of MSCs by nonviral gene delivery of growth and differentiation factor 5 (GDF5) by electroporation mediated gene transfer could be an excellent source for cell transplantation. Human MSCs were harvested from bone marrow aspirate and GDF5 gene transfer was achieved byin vitroelectroporation. Transfected cells were cultured as monolayers and as 3D cultures in 1.2% alginate bead culture. MSC expressed GDF5 efficiently for up to 21 days. The combination of GDF5 gene transfer and 3D culture in alginate showed an upregulation of aggrecan and SOX9, two markers for chondrogenesis, and KRT19 as a marker for discogenesis compared to untransfected cells. The cells encapsulated in alginate produced more proteoglycans expressed in GAG/DNA ratio. Furthermore, GDF5 transfected MCS injected into an IVD papain degeneration organ culture model showed a partial recovery of the GAG/DNA ratio after 7 days. In this study we demonstrate the potential of GDF5 transfected MSC as a promising approach for clinical translation for disc regeneration.

scholarly journals Enhanced PRL-1 expression in placenta-derived mesenchymal stem cells accelerates hepatic function via mitochondrial dynamics in a cirrhotic rat model

2020 ◽  
Author(s):  
Jae Yeon Kim ◽  
Jong Ho Choi ◽  
Ji Hye Jun ◽  
Sohae Park ◽  
Jieun Jung ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Gene Delivery ◽  
Cell Therapy ◽  
Delivery Systems ◽  
Nonviral Gene Delivery ◽  
Early Gene ◽  
Next Generation ◽  
Atp Production ◽  
Mitochondrial Functions

Abstract Background: Placenta-derived mesenchymal stem cells (PD-MSCs) have been highlighted as an alternative cell therapy agent that has become a next-generation stem cell treatment. Phosphatase of regenerating liver-1 (PRL-1), an immediate early gene, plays a critical role during liver regeneration. Here, we generated enhanced PRL-1 in PD-MSCs (PD-MSCsPRL-1, PRL-1+) using lentiviral and nonviral gene delivery systems and investigated mitochondrial functions by PD-MSCPRL-1 transplantation for hepatic functions in a rat bile duct ligation (BDL) model. Methods: PD-MSCsPRL-1 were generated by lentiviral and nonviral AMAXA gene delivery systems and analyzed for their characteristics and mitochondrial metabolic functions. Liver cirrhosis was induced in Sprague-Dawley (SD) rats using common BDL for 10 days. PKH67+ naïve and PD-MSCsPRL-1 were intravenously administered into cirrhotic rats using a nonviral system (2x106 cells/animal). The animals were sacrificed at 1, 2, 3, and 5 weeks after transplantation and engraftment of stem cells, and histopathological analysis and hepatic mitochondrial functions were performed. Results: PD-MSCsPRL-1 were successfully generated using lentiviral and nonviral AMAXA systems and maintained characteristics similar to those of naïve cells. Compared with naïve cells, PD-MSCsPRL-1 improved respirational metabolic states of mitochondria. In particular, mitochondria in PD-MSCsPRL-1 generated by the nonviral AMAXA system showed a significant increase in the respirational metabolic state, including ATP production and mitochondrial biogenesis (*p<0.05). Furthermore, transplantation of PD-MSCsPRL-1 using a nonviral AMAXA system promoted engraftment into injured target liver tissues of a rat BDL cirrhotic model and enhanced the metabolism of mitochondria via increased mtDNA and ATP production, thereby improving therapeutic efficacy. Conclusions: Our findings will further our understanding of the therapeutic mechanism of enhanced MSCs and provide useful data for the development of next-generation MSC-based cell therapy and therapeutic strategies for regenerative medicine in liver disease.

scholarly journals Characterization of Human Mesenchymal Stem Cells Isolated from the Testis

2018 ◽  
Vol 2018 ◽  
pp. 1-9 ◽  
Author(s):  
Letizia De Chiara ◽  
Elvira Smeralda Famulari ◽  
Sharmila Fagoonee ◽  
Saskia K. M. van Daalen ◽  
Stefano Buttiglieri ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Pluripotent Stem Cells ◽  
Embryonic Stem ◽  
Human Mesenchymal Stem Cells ◽  
Spermatogonial Stem Cells ◽  
Great Promise ◽  
Human Testis ◽  
First Time

Mesenchymal stem cells hold great promise for regenerative medicine as they can be easily isolated from different sources such as adipose tissue, bone marrow, and umbilical cord blood. Spontaneously arising pluripotent stem cells can be obtained in culture from murine spermatogonial stem cells (SSCs), while the pluripotency of the human counterpart remains a matter of debate. Recent gene expression profiling studies have demonstrated that embryonic stem cell- (ESC-) like cells obtained from the human testis are indeed closer to mesenchymal stem cells (MSCs) than to pluripotent stem cells. Here, we confirm that colonies derived from human testicular cultures, with our isolation protocol, are of mesenchymal origin and do not arise from spermatogonial stem cells (SSCs). The testis, thus, provides an important and accessible source of MSCs (tMSCs) that can be potentially used for nephrotoxicity testing in vitro. We further demonstrate, for the first time, that tMSCs are able to secrete microvesicles that could possibly be applied to the treatment of various chronic diseases, such as those affecting the kidney.

scholarly journals Enhanced PRL-1 expression in placenta-derived mesenchymal stem cells accelerates hepatic function via mitochondrial dynamics in a cirrhotic rat model

2020 ◽  
Author(s):  
Jae Yeon Kim ◽  
Jong Ho Choi ◽  
Ji Hye Jun ◽  
Sohae Park ◽  
Jieun Jung ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Gene Delivery ◽  
Cell Therapy ◽  
Delivery Systems ◽  
Nonviral Gene Delivery ◽  
Early Gene ◽  
Next Generation ◽  
Atp Production ◽  
Mitochondrial Functions

Abstract Background: Placenta-derived mesenchymal stem cells (PD-MSCs) have been highlighted as an alternative cell therapy agent that has become a next-generation stem cell treatment. Phosphatase of regenerating liver-1 (PRL-1), an immediate early gene, plays a critical role during liver regeneration. Here, we generated enhanced PRL-1 in PD-MSCs (PD-MSCsPRL-1, PRL-1+) using lentiviral and nonviral gene delivery systems and investigated mitochondrial functions by PD-MSCPRL-1 transplantation for hepatic functions in a rat bile duct ligation (BDL) model.Methods: PD-MSCsPRL-1 were generated by lentiviral and nonviral AMAXA gene delivery systems and analyzed for their characteristics and mitochondrial metabolic functions. Liver cirrhosis was induced in Sprague-Dawley (SD) rats using common BDL for 10 days. PKH67+ naïve and PD-MSCsPRL-1 were intravenously administered into cirrhotic rats using a nonviral system (2x106 cells/animal). The animals were sacrificed at 1, 2, 3, and 5 weeks after transplantation and engraftment of stem cells, and histopathological analysis and hepatic mitochondrial functions were performed.Results: PD-MSCsPRL-1 were successfully generated using lentiviral and nonviral AMAXA systems and maintained characteristics similar to those of naïve cells. Compared with naïve cells, PD-MSCsPRL-1 improved respirational metabolic states of mitochondria. In particular, mitochondria in PD-MSCsPRL-1 generated by the nonviral AMAXA system showed a significant increase in the respirational metabolic state, including ATP production and mitochondrial biogenesis (*p<0.05). Furthermore, transplantation of PD-MSCsPRL-1 using a nonviral AMAXA system promoted engraftment into injured target liver tissues of a rat BDL cirrhotic model and enhanced the metabolism of mitochondria via increased mtDNA and ATP production, thereby improving therapeutic efficacy.Conclusions: Our findings will further our understanding of the therapeutic mechanism of enhanced MSCs and provide useful data for the development of next-generation MSC-based cell therapy and therapeutic strategies for regenerative medicine in liver disease.

scholarly journals Tissue Harvesting Site Effect on the Canine Adipose Stromal Vascular Fraction Quantity and Quality

Animals ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 460
Author(s):  
Hanan Hendawy ◽  
Akiko Uemura ◽  
Danfu Ma ◽  
Ryosuke Namiki ◽  
Haney Samir ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Sampling Site ◽  
Stromal Vascular Fraction ◽  
Site Effect ◽  
Falciform Ligament ◽  
Great Promise ◽  
Trypan Blue Exclusion ◽  
Trypan Blue Exclusion Test ◽  
Clusters Of Differentiation

Mesenchymal stem cells (MSCs) constitute a great promise for regenerative therapy, but these cells are difficultly recovered in large amounts. A potent alternative is the stromal vascular fraction (SVF), non-cultured MSCs, separated from adipose tissue (AT). We aim to evaluate AT harvesting site effect on the SVF cells’ quantity and quality in dogs. Subcutaneous abdominal fat, falciform ligament and peri-ovarian fat were sampled. After SVF isolation, the trypan blue exclusion test and a hemocytometer were used to assess the cell viability and cellular yield. SVF cells were labeled for four surface antigenic markers, clusters of differentiation CD90, CD44, CD29, and CD45, and then examined by flow cytometry. Semi-quantitative RT-PCR was used to evaluate the gene expression of the former markers in addition to OCT-4 and CD34. SVF cells in the peri-ovarian AT recorded the highest viability% (99.63 ± 0.2%), as well as a significantly higher cellular yield (36.87 ± 19.6 × 106 viable cells/gm fat, p < 0.001) and a higher expression of adipose-derived mesenchymal stem cells AD-MSCs surface markers than that of other sites. SVF cells from the peri-ovarian site revealed a higher expression of MSC markers (CD90, CD44, and CD29) and OCT-4 compared to the other sites, with weak CD45 and CD34 expressions. The positive OCT-4 expression demonstrated the pluripotency of SVF cells isolated from different sites. To conclude, the harvesting site is a strong determinant of SVF cells’ quantity and quality, and the peri-ovarian site could be the best AT sampling site in dogs.

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