BACKGROUND: Human mesenchymal stem cells (hMSCs) are pluripotent cells that can differentiate to various mesenchymal cell types. Recently, a method to isolate hMSCs from bone marrow and expand them in culture was described. Here we report on the use of hMSCs as a platform for gene therapy aimed at bone lesions. METHODS: Bone marrow derived hMSCs were expanded in culture and infected with recombinant adenoviral vector encoding the osteogenic factor, human BMP-2. The osteogenic potential of genetically engineered hMSCs was assessed in vitro and in vivo. RESULTS: Genetically engineered hMSCs displayed enhanced proliferation and osteogenic differentiation in culture. In vivo, transplanted genetically engineered hMSCs were able to engraft and form bone and cartilage in ectopic sites, and regenerate bone defects (non-union fractures) in mice radius bone. Importantly, the same results were obtained with hMSCs isolated from a patient suffering from osteoporosis. CONCLUSIONS: hMSCs represent a novel platform for skeletal gene therapy and the present results suggest that they can be genetically engineered to express desired therapeutic proteins inducing specific differentiation pathways. Moreover, hMSCs obtained from osteoporotic patients can restore their osteogenic activity following human BMP-2 gene transduction, an important finding in the future planning of gene therapy treatment for osteoporosis.

Adenoviridae↗

PubMedDate not supplied

Combined transductional and transcriptional targeting of melanoma cells by artificial virus-like particles.

BACKGROUND: Artificial virus-like particles (AVPs) represent a novel type of liposomal vector resembling retroviral envelopes. AVPs are serum-resistant and non-toxic and can be endowed with a peptide ligand as a targeting device. The vitronectin receptor, alphavbeta3-integrin, is commonly upregulated on malignant melanoma cells. In the present study we investigated whether AVPs carrying cyclic peptides with an RGD integrin binding motif (RGD-AVPs) are suitable for the specific and efficient transduction of human melanoma cells. METHODS: Plasmid DNA was complexed with low molecular weight non-linear polyethyleneimine and packaged into anionic liposomes. Transduction efficiencies were determined after transient transfection of different cell lines in serum-free medium using green fluorescent protein or luciferase reporter genes. RESULTS: We demonstrated that RGD-AVPs transduced human melanoma cells with high efficiencies of > 60%. Efficient transduction was clearly dependent on the presence of the cyclic RGD ligand and was selective for melanoma cells. The specificity of the vector system could be further enhanced by using the melanocyte-specific tyrosinase promoter to drive transgene expression. CONCLUSION: Our findings suggest that the AVP technology is a useful approach for generating highly efficient and specific non-viral vectors for melanoma targeting, in particular in a setting of combined transductional and transcriptional targeting.

Amino Acid Sequence↗

PubMedDate not supplied

A critical discussion of intraclass correlation coefficients.

In general, intraclass correlation coefficients (ICC's) are designed to assess consistency or conformity between two or more quantitative measurements. They are claimed to handle a wide range of problems, including questions of reliability, reproducibility and validity. It is shown that care must be taken in choosing a suitable ICC with respect to the underlying sampling theory. For this purpose a decision tree is developed. It may be used to choose a coefficient which is appropriate for a specific study setting. We demonstrate that different ICC's may result in quite different values for the same data set, even under the same sampling theory. Other general limitations of ICC's are also addressed. Potential alternatives are presented and discussed, and some recommendations are given for the use of an appropriate method.

Analysis of Variance↗