Francesco D'Angelo

Stem cell-biomaterial interactions for regenerative medicine

The synergism of stem cell biology and biomaterial technology promises to have a profound impact on stem-cell-based clinical applications for tissue regeneration. Biomaterials development is rapidly advancing to display properties that, in a precise and physiological fashion, could drive stem-cell fate both in vitro and in vivo. Thus, the design of novel materials is trying to recapitulate the molecular events involved in the production, clearance and interaction of molecules within tissue in pathologic conditions and regeneration of tissue/organs. In this review we will report on the challenges behind translating stem cell biology and biomaterial innovations into novel clinical therapeutic applications for tissue and organ replacements (graphical abstract).

Biocompatible Poly(L-lactide)/MWCNT Nanocomposites: Morphological Characterization, Electrical Properties, and Stem Cell Interaction

The promising perspectives of PLLA-based nanostructured biomaterials and their relevance in tissue engineering are reported. Nanocomposites based on PLLA and MWCNTs are developed with an MWCNT content ranging from 0 to 3 wt%. The electrical properties show a percolation threshold within a range of 0.21-0.33 wt% MWCNTs, and the conductivity increases by six orders of magnitude. The surface structure shows changes with the carbon nanotube concentration. The functional role of MWCNTs incorporation in terms of interactions with adult stem cells suggests that PLLA/MWCNT nanocomposites are suitable substrates for primary stem cell culture.

Poly (L-lactic acid)/calcium-deficient nanohydroxyapatite electrospun mats for bone marrow stem cell cultures

Electrospinning of bioresorbable polymers is a promising and valuable scaffolding technique. To improve its potential applications, the addition of specific fillers has been considered. This paper reports the fabrication of electrospun poly(L-lactic acid)/Ca-deficient-hydroxyapatite (PLLA/dHAp) mats, the content of nanosized d-HAp ranged between 1 and 8 wt%. All samples consisted of micrometric and submicrometric fibers, comprising 2D voids of 8 and 13 µm for PLLA and PLLA/d-HAp mats, respectively. The surface of the electrospun fibers was characterized by an uniform distribution of nanopores. Hybrid mats loaded with 1 wt% d-HAp showed the most homogeneous microstructure, differently from the mats loaded with 4 and 8 wt% d-HAp due to the presence of microagglomerates. The viscoelastic properties of PLLA/d-HAp hybrids were characterized by a decreasing trend of the storage modulus with increases in the nanofiller content. The microstructure, viscoelastic behavior, and cytocompatibility were investigated using murine bone marrow mesenchymal stem cells. On the basis of the biological data, the electrospun PLLA and PLLA/d-HAp mats can be regarded as potential scaffolds for bone marrow mesenchymal stem cells culture.

Efficient siRNA Delivery by the Cationic Liposome DOTAP in Human Hematopoietic Stem Cells Differentiating into Dendritic Cells

RNA interference technology is an ideal strategy to elucidate the mechanisms associated with human CD34+ hematopoietic stem cell differentiation into dendritic cells. Simple manipulations in vitro can unequivocally yield alloreactive or tolerogenic populations, suggesting key implications of biochemical players that might emerge as therapeutic targets for cancer or graft-versus-host disease. To knockdown proteins typically involved in the biology of dendritic cells, we employed an siRNA delivery system based on the cationic liposome DOTAP as the carrier. Freshly-isolated CD34+ cells were transfected with siRNA for cathepsin S with negligible cytotoxicity and transfection rates (>60%) comparable to the efficiency shown by lentiviral vectors. Further, cathepsin S knockdown was performed during both cell commitment and through the entire 14-day differentiation process with repeated transfection rounds that had no effect per se on cell development. Tested in parallel, other commercially-available chemical reagents failed to meet acceptable standards. In addition to safe and practical handling, a direct advantage of DOTAP over viral-mediated techniques is that transient silencing effects can be dynamically appraised through the recovery of targeted proteins. Thus, our findings identify DOTAP as an excellent reagent for gene silencing in resting and differentiating CD34+ cells, suggesting a potential for applications in related preclinical models.

Mechanotransduction: Tuning Stem Cells Fate

It is a general concern that the success of regenerative medicine-based applications is based on the ability to recapitulate the molecular events that allow stem cells to repair the damaged tissue/organ. To this end biomaterials are designed to display properties that, in a precise and physiological-like fashion, could drive stem cell fate both in vitro and in vivo. The rationale is that stem cells are highly sensitive to forces and that they may convert mechanical stimuli into a chemical response. In this review, we describe novelties on stem cells and biomaterials interactions with more focus on the implication of the mechanical stimulation named mechanotransduction.

Development of a new tool for 3d modeling for regenerative medicine

The effectiveness of therapeutic treatment based on regenerative medicine for degenerative diseases (i.e., neurodegenerative or cardiac diseases) requires tools allowing the visualization and analysis of the three-dimensional (3D) distribution of target drugs within the tissue. Here, we present a new computational procedure able to overcome the limitations of visual analysis emerging by the examination of a molecular signal within images of serial tissue/organ sections by using the conventional techniques. Together with the 3D anatomical reconstitution of the tissue/organ, our framework allows the detection of signals of different origins (e.g., marked generic molecules, colorimetric, or fluorimetric substrates for enzymes; microRNA; recombinant protein). Remarkably, the application does not require the employment of specific tracking reagents for the imaging analysis. We report two different representative applications: the first shows the reconstruction of a 3D model of mouse brain with the analysis of the distribution of the β-Galactosidase, the second shows the reconstruction of a 3D mouse heart with the measurement of the cardiac volume.

Knock‐down of HEXA and HEXB genes correlate with the absence of the immunostimulatory function of HSC‐derived dendritic cells

In an attempt to investigate whether the genetic defect in the HEXA and HEXB genes (which causes the absence of the lysosomal β‐N‐acetyl‐hexosaminidase), are related to the wide inflammation in GM2 gangliosidoses (Tay‐Sachs and Sandhoff disease), we have chosen the dendritic cells (DCs) as a study model. Using the RNA interference approach, we generated an in vitro model of HEXs knock‐down immunogenic DCs (i‐DCs) from CD34+‐haemopoietic stem cells (CD34+‐HSCs), thus mimicking the Tay‐Sachs (HEXA−/−) and Sandhoff (HEXB−/−) cells.

Nanocomposites Based on PLLA and Multi Walled Carbon Nanotubes Support the Myogenic Differentiation of Murine Myoblast Cell Line

We explored the effect of poly(L-lactic acid) (PLLA) containing various percentages (0.1, 0.5, 1, and 3 wt.%) of multi walled carbon nanotubes (MWCNTs) on the myogenic differentiation of C2C12 murine myoblast progenitor cells. We showed that all PLLA/MWCNTs nanocomposite materials support the myotubes formation more efficiently than neat PLLA as indicated by the high expression of the most significant myogenic markers: MyoD, Myosin Heavy Chain, dimension of myofibres, and fusion myogenic index. Interestingly, we note that both MyoD and myogenic fusion index levels were in the order 0.1 MWCNTs = 0.5 MWCNTs > 1 MWCNTs > 3 MWCNTs > neat PLLA, suggesting that the amount of MWCNTs influenced the cell differentiation.