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Aims: To ascertain if human pericytes produce SPARC (acronym for Secreted Protein Acidic and Cysteine Rich), a matricellular protein implicated in the regulation of cell proliferation, migration, and cell-matrix interactions; clarify if SPARC expression in cardiac pericytes is modulated by hypoxia; and determine the functional consequences of SPARC silencing. Results: Starting from the recognition that the conditioned media (CM) of human pericytes promote proliferation and migration of cardiac stromal cells, we screened candidate mediators by mass-spectrometry analysis. Of the 14 high-confidence proteins (<1% FDR) identified in the bioactive fractions of the pericyte CM, SPARC emerged as the top-scored matricellular protein. SPARC expression was validated using ELISA and found to be upregulated by hypoxia/starvation in pericytes that express platelet-derived growth factor receptor α (PDGFRα). This subfraction is acknowledged to play a key role in extracellular matrix remodeling. Studies in patients with acute myocardial infarction showed that peripheral blood SPARC correlates with the levels of creatine kinase Mb, a marker of cardiac damage. Immunohistochemistry analyses of infarcted hearts revealed that SPARC is expressed in vascular and interstitial cells. Silencing of SPARC reduced the pericyte ability to secrete collagen1a1, without inhibiting the effects of CM on cardiac and endothelial cells. These data indicate that SPARC is enriched in the bioactive fraction of the pericyte CM, is induced by hypoxia and ischemia, and is essential for pericyte ability to produce collagen. Innovation: This study newly indicates that pericytes are a source of the matricellular protein SPARC. Conclusion: Modulation of SPARC production by pericytes may have potential implications for postinfarct healing.

The life-long story of the heart starts concomitantly with primary differentiation events occurring in multipotent progenitors located in the so-called heart tube. This initially tubular structure starts a looping process, which leads to formation of the final four-chambered heart with a primary contribution of geometric and position-associated cell sensing. While this establishes the correct patterning of the final cardiac structure, it also provides feedbacks to fundamental cellular machineries controlling proliferation and differentiation, thus ensuring a coordinated restriction of cell growth and a myocyte terminal differentiation. Novel evidences provided by embryological and cell engineering studies have clarified the relevance of mechanics-supported position sensing for the correct recognition of cell fate inside developing embryos and multicellular aggregates. One of the main components of this pathway, the Hippo-dependent signal transduction machinery, is responsible for cell mechanics intracellular transduction with important consequences for gene transcription and cell growth control. Being the Hippo pathway also directly connected to stress responses and altered metabolism, it is tempting to speculate that permanent alterations of mechanosensing may account for modifying self-renewal control in tissue homeostasis. In the present contribution, we translate these concepts to the aging process and the failing of the human heart, two pathophysiologic conditions that are strongly affected by stress responses and altered metabolism.

Periosteum contains mesenchymal stem cells (Pe-MSCs) that contribute to normal bone growth, healing, and turnover; understanding Pe-MSC capabilities may shed light over the treatment of bone defects using tissue engineering. Bone tissue regeneration needs in vitro bone precursors or stem cell coculture onto specific scaffolds but, despite extensive research in the field, very little is known about the matrix structure of the tissue-engineered tissues and the scaffold's effects on cell differentiation. To this purpose we have selected a clonal population (murine Pe-MSCs) that was seeded and differentiated onto an acellular bone scaffold. Cell differentiation was assessed after 3 months and 1 year by molecular, histological, biochemical, and biophysical analyses and results were compared with the same osteoinduced clonal cells cultured as cellular aggregates. Our data show that Pe-MSCs cultured onto acellular bone scaffold develop a complex three-dimensional matrix and an osteoblastic phenotype but do not produce hydroxyapatite (HA); moreover, they seem able to reabsorb the colonized bone scaffold. On the contrary, cells cultured as three-dimensional aggregates differentiate and produce osteoblastic markers and HA nanocrystals.

Grafts of tissue-engineered bone represent a promising alternative in the treatment of large and small bone defects. Current approaches are often badly tolerated by patients because of invasiveness, ethical problems, culture, and possibility of infection. Autologous grafts have been indicated as a solution to such problems. Because of tissue availability, many have proposed the use of cultured cells derived from bone marrow expanded in culture and induced to differentiate in bone tissue. Data reported in the literature show that it is possible to produce tissue substitutes in vitro indeed, but results are not always concordant regarding the in vitro produced bone quality. In the present work, we investigated bone formation in aggregates of human bone marrow-derived mesenchymal stem cells induced to differentiate in bone. After osteoinduction we characterized the mineral matrix produced using Fourier transform infrared spectroscopy, scanning electron microscopy, transmission electron microscopy, and X-ray powder diffraction. Cells were obtained from bone marrow, subjected to immunodepletion for CD3, CD11b, CD14, CD16, CD19, CD56, CD66b, and glycophorin A using RosetteSep and cultured in a new formulation of medium for four passages and then were allowed to form spontaneous aggregates. At the end of proliferation before aggregation, cells were analyzed by fluorescent activated cell sorting (FACS) for markers routinely used to characterize expanded mesenchymal stem cells and were found to be remarkably homogeneous for CD29 (99% ± 1%), CD73 (99% ± 1%), CD90 (95% ± 4%), CD105 (96% ± 4%), and CD133 (0% ± 1%) expression. Our results show that not only aggregated cells express the major markers of osteogenic differentiation, such as osteocalcin, osteonectin, osteopontin, and bone sialoprotein, but also the inorganic matrix is made of an apatite structurally and morphologically similar to native bone even without a scaffold.