Human umbilical cord mesenchymal stem cell transplantation restores damaged ovaries.

Ovarian injury because of chemotherapy can decrease the levels of sexual hormones and potentia generandi of patients, thereby greatly reducing quality of life. The goal of this study was to investigate which transplantation method for human umbilical cord mesenchymal stem cells (HUMSCs) can recover ovarian function that has been damaged by chemotherapy. A rat model of ovarian injury was established using an intraperitoneal injection of cyclophosphamide. Membrane-labelled HUMSCs were subsequently injected directly into ovary tissue or tail vein. The distribution of fluorescently labelled HUMSCs, estrous cycle, sexual hormone levels, and potentia generandi of treated and control rats were then examined. HUMSCs injected into the ovary only distributed to the ovary and uterus, while HUMSCs injected via tail vein were detected in the ovary, uterus, kidney, liver and lung. The estrous cycle, levels of sex hormones and potentia generandi of the treated rats were also recovered to a certain degree. Moreover, in some transplanted rats, fertility was restored and their offspring developed normally. While ovary injection could recover ovarian function faster, both methods produced similar results in the later stages of observation. Therefore, our results suggest that transplantation of HUMSCs by tail vein injection represents a minimally invasive and effective treatment method for ovarian injury.

Comparison of cell proliferation, apoptosis, cellular morphology and ultrastructure between human umbilical cord and placenta-derived mesenchymal stem cells.

Research in mesenchymal stem cells (MSCs) is mainly focused on applications for treatments of brain and spinal cord injury as well as mechanisms underlying effects of MSCs. However, due to numerous limitations, there is little information on selection of appropriate sources of MSCs for transplantation in clinical applications. Therefore, in this study we compared various properties of human umbilical cord-derived MSCs (HUCMSCs) with human placenta-derived MSCs (HPDMSCs), including cell proliferation, apoptosis, cellular morphology, ultrastructure, and their ability to secrete various growth factors (i.e. vascular endothelial growth factor, insulin-like growth factors-1, and hepatocyte growth factor), which will allow us to select appropriate MSC sources for cellular therapy. Cell culture, flow cytometry, transmission electron microscope (TEM) and atomic force microscope (AFM) were used for assessment of HUCMSCs and HPDMSCs. Results showed that the two types of cells appeared slightly different when they were observed under AFM. HUCMSCs appeared more fibroblast-like, whereas HPDMSCs appeared as large flat cells. HUCMSCs had higher proliferative rate and lower rate of apoptosis than HPDMSCs (p<0.05). However, HPDMSCs secreted more of the three growth factors than HUCMSCs (p<0.05). Results of TEM revealed that the two types of MSCs underwent active metabolism and had low degree of differentiation, especially HUCMSCs. Results of AFM showed that HUCMSCs had stronger ability of mass transport and cell migration than HPDMSCs. However, HPDMSCs displayed stronger adhesive properties than HUCMSCs. Our findings indicate that different sources of MSCs have different properties, and that care should be taken when choosing the appropriate sources of MSCs for stem cell transplantation.

In vitro release and in vitro-in vivo correlation for silybin meglumine incorporated into hollow-type mesoporous silica nanoparticles.

BACKGROUND: The purpose of this study was to develop a sustained drug-release model for water-soluble drugs using silica nanoparticles. METHODS: Hollow-type mesoporous silica nanoparticles (HMSNs) were prepared using Na(2)CO(3) solution as the dissolution medium for the first time. The water-soluble compound, silybin meglumine, was used as the model drug. The Wagner-Nelson method was used to calculate the in vivo absorption fraction. RESULTS: The results of transmission electron microscopy and nitrogen adsorption revealed that the empty HMSNs had uniformly distributed particles of size 50-100 nm, a spherical appearance, a large specific surface area (385.89 ± 1.12 m(2)/g), and ultralow mean pore size (2.74 nm). The highly porous structure allowed a large drug-loading rate (58.91% ± 0.39%). In 0.08 M Na(2)CO(3) solution, silybin meglumine-loaded HMSNs could achieve highly efficacious and long-term sustained release for 72 hours in vitro. The results of in vitro-in vivo correlation revealed that HMSNs in 0.08 M Na(2)CO(3) solution had a correlation coefficient R(2) value of 0.9931, while those of artificial gastric juice and artificial intestinal juice were only 0.9287 and 0.7689, respectively. CONCLUSION: The findings of in vitro-in vivo correlation indicate that HMSNs together with Na(2)CO(3) solution could achieve an excellent linear relationship between in vitro dissolution and in vivo absorption for 72 hours, leading to a promising model for sustained release of water-soluble drugs.

Oral bioavailability of silymarin formulated as a novel 3-day delivery system based on porous silica nanoparticles.

The purpose of this study was to develop porous silica nanoparticles (PSNs) as a carrier to improve oral bioavailability of poorly water-soluble drugs, using silymarin as a model. PSNs were synthesized by reverse microemulsion and ultrasonic corrosion methods. A 3-day release formulation consisting of a silymarin solid dispersion, a hydrophilic gel matrix and silymarin-loaded PSNs was prepared. In vitro release studies indicated that both the silymarin-loaded PSNs and the 3-day release formulation showed a typical sustained-release pattern over a long period, about 72 h. The in vivo studies revealed that the 3-day release formulation gave a significantly higher plasma concentration and larger area under the concentration-time curves than commercial tablets when orally administered to beagle dogs. This implies that the prepared 3-day release formulation significantly enhanced the oral bioavailability of silymarin, suggesting that PSNs can be used as promising drug carriers for oral sustained release systems. Thus providing a technically feasible approach for improving the oral bioavailability and long-term efficacy of poorly soluble drugs.

Incorporating pTGF-ß1/calcium phosphate nanoparticles with fibronectin into 3-dimensional collagen/chitosan scaffolds: efficient, sustained gene delivery to stem cells for chondrogenic differentiation.

The objective of this study was to prepare a 3-dimensional nanoparticle gene delivery system (3D-NGDS) based on collagen/chitosan scaffolds, in which plasmid transforming growth factor beta 1 (TGF-ß1)/calcium phosphate nanoparticles mixed with fibronectin (FN) were used to transfect mesenchymal stem cells (MSCs). Scanning electron microscopy was used to characterise the microstructure of 3-dimensional collagen/chitosan scaffolds. An analysis performed to quantify the TGF-b1 concentrations in MSC cultures revealed that the MSCs transfected with the 3D-NGDS showed remarkably high levels of TGF-b1 over long periods, retaining a concentration of TGF-b1 of approximately 10 ng/mL within two weeks, with the highest level (12.6 ng/mL) being observed on the 6th day. An immunohistochemistry analysis for collagen type II revealed that much higher production of collagen II from the 9th to 15th day was observed in the 3D-NGDS-transfected MSCs than that in MSCs transfected by the Lipofectamine 2000 method. The glycosaminoglycan content of the 3D-NGDS was comparable to those treated with TGF-ß1 as well as TGF-ß1 plus dexamethasone, and was significantly higher than those treated with free plasmid and Lipofectamine 2000. A remarkable type I collagen expression inhibition of the 3D-NGDS at day 21 was observed via ELISA. These results suggested that transfection with the 3D-NGDS could successfully induce MSC chondrogenic differentiation in vitro without dexamethasone. In summary, the 3D-NGDS could be developed into a promising alternative method to transfer exogenous nucleic acid to MSCs in clinical trials.

Efficient gene delivery to mesenchymal stem cells by an ethylenediamine-modified polysaccharide from mulberry leaves.

This study investigates the use of a natural polysaccharide isolated from mulberry leaves as a nonviral gene vector. Ethylenediamine is chemically grafted to the backbone of a polysaccharide from mulberry leaves (MPS) to acquire nucleic acid binding affinity. A particle-size observation indicates that the cationic mulberry leaf polysaccharide (CMPS) can efficiently combine with plasmid transforming growth factor ß1 (TGF-ß1) to form nanoscaled particles. In addition, the electrophoresis assay indicates a retarded plasmid migration when the CMPS/pTGF-ß1 weight ratio is increased to 30:1. The in vitro cell transfection experiment is performed based on bone marrow mesenchymal stem cells (MSCs) derived from rat femurs and tibias, and the findings reveal that the complex with a CMPS/pTGF-ß1 weight ratio of 50:1 exhibits the highest cell transfection effect, which is significantly higher than that of branched poly(ethyleneimine) (PEI) (25 kDa; p = 0.001, Student's t-test) and slightly higher than Lipofectamine 2000. Moreover, the cytotoxicity assay also demonstrates that all of these tested complexes and the plasmid TGF-ß1 are nontoxic to mesenchymal stem cells (MSCs). The results of the living cell imaging confirm that more of the CMPS/plasmid TGF-ß1 nanoparticles can be taken up and at a faster rate by the MSCs than by the positive control Lipofectamine 2000; these data are consistent with the transfection efficiency data. Together, these results suggest that the CMPS/pTGF-ß1 nanoparticle can potentially be developed into a promising alternative for the transfer of therapeutic genes into cells.

Encapsulation of plasmid DNA in calcium phosphate nanoparticles: stem cell uptake and gene transfer efficiency.

BACKGROUND: The purpose of this study was to develop calcium phosphate nanocomposite particles encapsulating plasmid DNA (CP-pDNA) nanoparticles as a nonviral vector for gene delivery. METHODS: CP-pDNA nanoparticles employing plasmid transforming growth factor beta 1 (TGF-ß1) were prepared and characterized. The transfection efficiency and cell viability of the CP-pDNA nanoparticles were evaluated in mesenchymal stem cells, which were identified by immunofluorescence staining. Cytotoxicity of plasmid TGF-ß1 and calcium phosphate to mesenchymal stem cells were evaluated by MTT assay. RESULTS: The integrity of TGF-ß1 encapsulated in the CP-pDNA nanoparticles was maintained. The well dispersed CP-pDNA nanoparticles exhibited an ultralow particle size (20-50 nm) and significantly lower cytotoxicity than Lipofectamine™ 2000. Immunofluorescence staining revealed that the cultured cells in this study were probably mesenchymal stem cells. The cellular uptake and transfection efficiency of the CP-pDNA nanoparticles into the mesenchymal stem cells were higher than that of needle-like calcium phosphate nanoparticles and a standard calcium phosphate transfection kit. Furthermore, live cell imaging and confocal laser microscopy vividly showed the transportation process of the CP-pDNA nanoparticles in mesenchymal stem cells. The results of a cytotoxicity assay found that both plasmid TGF-ß1 and calcium phosphate were not toxic to mesenchymal stem cells. CONCLUSION: CP-pDNA nanoparticles can be developed into an effective alternative as a nonviral gene delivery system that is highly efficient and has low cytotoxicity.