Adult human subcutaneous adipose tissue contains cells with intriguing multilineage developmental plasticity, much like marrow-derived mesenchymal stem cells. Putative stem or progenitor cells from fat have been given many different names in the literature, reflecting an early and evolving consensus regarding their phenotypic characterization. The study reported here used microarrays to evaluate over 170 genes relating to angiogenesis and extracellular matrix in undifferentiated, early-passage human adipose-derived adherent stromal (hADAS) cells isolated from three separate donors. The hADAS populations unanimously transcribed 66% of the screened genes, and 83% were transcribed by at least two of the three populations. The most highly transcribed genes relate to functional groupings such as cell adhesion, matrix proteins, growth factors and receptors, and proteases. The transcriptome of hADAS cells demonstrated by this work reveals many similarities to published profiles of bone marrow mesenchymal stem cells (MSCs).In addition, flow analysis of over 24 hADAS cell surface proteins (n = 7 donors) both confirms and expands on the existing literature and reveals strong intergroup correlation, despite an inconsistent nomenclature and the lack of standardized protocols for cell isolation and culture. Finally, based on flow analysis and reverse transcription polymerase chain reaction studies, our results suggest that hADAS cells do not express several proteins that are implicated as markers of "stemness" in other stem cell populations, including telomerase, CD133, and the membrane transporter ABCG2. Stem Cells 2005;23:412-423

Laminin promotes epithelial cell adhesion in part through a site of nine amino acids CDPGYIGSR on the B1 chain. Using smaller synthetic peptides from this sequence as well as various peptides with amino acid substitutions, we find that the minimum sequence necessary for efficient cell adhesion as well as receptor binding is YIGSR. The deletion of tyrosine or the substitution of arginine in the peptides resulted in a significant loss of activity. The presence of an amide group on the terminal arginine of either peptide increases activity significantly. YIGSR is active in promoting the adhesion of a variety of epithelial cells; however, it is inactive with chondrocytes, fibroblasts, and osteoblasts.

Cranial sutures play a critical role in calvarial morphogenesis, serving as growth centers during skull development. Both biomechanical tensile forces originating in the cranial base and biochemical factors present in dura mater have been postulated as determinants of suture morphogenesis and patency. A rat transplant model free of the putative biomechanical influence of the dura and cranial base was used to investigate the role of the dura mater in both the initial morphogenesis and maintenance of sutures during skull growth. Day 19 fetal presumptive (F19) and day 1 neonatal differentiated (Nl) coronal sutures, including associated frontal and parietal bones, were transplanted with or without underlying dura mater to the center of adult parietal bones. After 1,2, and 3 weeks, transplanted tissues were examined histologically and histomorphometrically to determine whether sutures formed and whether they were obliterated by ossification in the absence of dura mater. Both F19 and N1 sutures remained patent for 2 weeks either in the presence or the absence of transplant dura mater. However, at 3 weeks, in the absence of transplant dura mater, sutures were obliterated by bone, while in the presence of dura mater sutures resisted ossification, demonstrating an essential requirement for interactions with the transplant dura mater in maintaining functional sutures. Both F19 and N1 transplants showed comparable bone growth (cross-sectional surface area), regardless of the presence of transplant dura mater. These experiments suggest that tissue interactions of a biochemical nature, rather than biomechani-, cal forces generated through the cranial base, are required to maintain the suture as a non-ossified growth center. Furthermore, while the presence of dura mater was essential for maintenance of suture patency, fetal dura mater was not required for initial suture formation. 0 1993 Wiley-Liss, Inc.

Peripheral nerve transection occurs commonly in traumatic injury, causing deficits distal to the injury site. Conduits for repair currently on the market are hollow tubes; however, they often fail due to slow regeneration over long gaps. To facilitate increased regeneration speed and functional recovery, the ideal conduit should provide biochemically relevant signals and physical guidance cues, thus playing an active role in regeneration. To that end, laminin and laminin–polycaprolactone (PCL) blend nanofibers were fabricated to mimic peripheral nerve basement membrane. In vitro assays established 10% (wt) laminin content is sufficient to retain neurite-promoting effects of laminin. In addition, modified collector plate design to introduce an insulating gap enabled the fabrication of aligned nanofibers. The effects of laminin content and fiber orientation were evaluated in rat tibial nerve defect model. The lumens of conduits were filled with nanofiber meshes of varying laminin content and alignment to assess changes in motor and sensory recovery. Retrograde nerve conduction speed at 6 weeks was significantly faster in animals receiving aligned nanofiber conduits than in those receiving random nanofiber conduits. Animals receiving nanofiber-filled conduits showed some conduction in both anterograde and retrograde directions, whereas in animals receiving hollow conduits, no impulse conduction was detected. Aligned PCL nanofibers significantly improved motor function; aligned laminin blend nanofibers yielded the best sensory function recovery. In both cases, nanofiber-filled conduits resulted in better functional recovery than hollow conduits. These studies provide a firm foundation for the use of natural–synthetic blend electrospun nanofibers to enhance existing hollow nerve guidance conduits.

The precision and repeatability offered by computer-aided design and computer-numerically controlled techniques in biofabrication processes is quickly becoming an industry standard. However, many hurdles still exist before these techniques can be used in research laboratories for cellular and molecular biology applications. Extrusion-based bioprinting systems have been characterized by high development costs, injector clogging, difficulty achieving small cell number deposits, decreased cell viability, and altered cell function post-printing. To circumvent the high-price barrier to entry of conventional bioprinters, we designed and 3D printed components for the adaptation of an inexpensive 'off-the-shelf' commercially available 3D printer. We also demonstrate via goal based computer simulations that the needle geometries of conventional commercially standardized, 'luer-lock' syringe-needle systems cause many of the issues plaguing conventional bioprinters. To address these performance limitations we optimized flow within several microneedle geometries, which revealed a short tapered injector design with minimal cylindrical needle length was ideal to minimize cell strain and accretion. We then experimentally quantified these geometries using pulled glass microcapillary pipettes and our modified, low-cost 3D printer. This systems performance validated our models exhibiting: reduced clogging, single cell print resolution, and maintenance of cell viability without the use of a sacrificial vehicle. Using this system we show the successful printing of human induced pluripotent stem cells (hiPSCs) into Geltrex and note their retention of a pluripotent state 7 d post printing. We also show embryoid body differentiation of hiPSC by injection into differentiation conducive environments, wherein we observed continuous growth, emergence of various evaginations, and post-printing gene expression indicative of the presence of all three germ layers. These data demonstrate an accessible open-source 3D bioprinter capable of serving the needs of any laboratory interested in 3D cellular interactions and tissue engineering.