RESUMEN
In vitro studies on adherent cells require a process of passage to dissociate the cells from the culture substrate using enzymes or other chemical agents to maintain cellular activity. However, these proteolytic enzymes have a negative influence on the viability and phenotype of cells. The mesenchymal stem cell (MSC)-like cell line, C3H10T1/2, adhered, migrated, and proliferated to the same extent on newly designed microporous titanium (Ti) membrane and conventional culture dish, and spontaneous transfer to another substrate without enzymatic or chemical dissociation was achieved. The present study pierced a 10 µm-thick pure Ti sheet with 25 µm square holes at 75 µm intervals to create a dense porous structure with biomimetic topography. The pathway of machined holes allowed the cells to access both sides of the membrane frequently. In a culture with Ti membranes stacked above- and below-seeded cells, cell migration between the neighboring membranes was confirmed using the through-holes of the membrane and contact between the membranes as migration routes. Furthermore, the cells on each membrane migrated onto the conventional culture vessel. Therefore, a cell culture system with enzyme-free passaging was developed.
RESUMEN
The surface topography of Titanium (Ti) combined toughness and biocompatibility affects the attachment and migration of cells. Limited information of morphological characteristics, formed by precise machining in micron order, is currently available on the Ti that could promote osteoconduction. In the present study, a pure Ti membrane was pierced with precise 25 µm square holes at 75 µm intervals and appear burrs at the edge of aperture. We defined the surface without burrs as the "Head side" and that with burrs as the "Tail side". The effects of the machining microtopography on the proliferation and differentiation of the preosteoblasts (MC3T3-E1 cells) were investigated. The cells were more likely to migrate to, and accumulate in, the aperture of holes on the head side, but grew uniformly regardless of holes on the tail side. The topography on the both surfaces increased osteopontin gene expression levels. Osteocalcin expression levels were higher on the head side than one on the blank scaffold and tail side (p < 0.05). The osteocalcin protein expression levels were higher on the tail side than on the head side after 21 days of cultivation, and were comparable to the proportion of the calcified area (p < 0.05). These results demonstrate the capacity of a novel microporous Ti membrane fabricated using a precise mechanical punching process to promote cell proliferation and activity.
RESUMEN
A compound refractive lens made of nickel and designed for focusing high-energy synchrotron X-rays is presented. The lens consists of 600 parabolic grooves and focuses X-rays in one plane only (planar lens). The lenses made and investigated by us earlier exhibited low transmission and irregularities in the focused beam profile. Since then, improvements in lens manufacturing technology have been made. The present lens gives an almost Gaussian profile and produces four times higher intensity at its maximum compared with the intensity of primary X-ray beams of 174â keV.
RESUMEN
We assessed whether the adenovirus-mediated gene transfer of triple human complement regulating proteins (hCRPs) to the porcine aortic endothelium (PAE), could possibly exert a synergistic effect to inhibit human complement activation. Adenovirus vectors, encoding E.Coli beta-galactosidase (AxCALacZ), human membrane cofactor protein (MCP) (AxCAMCP), decay-accelerating factor (DAF) (AxCADAF), and CD59 (AxCACD59) were produced by the COS-TPC method. AxCALacZ was transfected to porcine aortic endothelium cells (PAECs) under various multiplicities of infection (MOI) to determine the efficiency of adenovirus-mediated gene transfer by 5-bromo-4-chloro-3-indolyl beta- D-galactopyranoside (X-gal) staining. The mRNA expressions of transfected CRPs were examined by reverse transcriptase-polymerase chain reaction (RT-PCR). Cellular damage to the PAEC was assessed by an MTT assay. PAEC was most efficiently transfected with the LacZ gene at 10(3) MOI/60-min incubation time (89.1%). In all samples transfected with the CRP gene, the corresponding mRNAs were detected in the RT-PCR. In the MTT assay, PAECs co-cultured with 20% human serum, showed the highest cellular viability after gene transfer of triple CRPs (117.7%), when compared with those of marker LacZ, single or double CRPs. The adenovirus-mediated multiple gene transfer of CRPs may thus be an efficient method for suppressing complement activation in the porcine-to-human model of hyperacute rejection.
