A human haemopoietic cell line (K562) which exhibits various erythroid characteristics, has been utilised as a model system for studying erythropoietic differentiation. We have analysed the alterations in expression of cell membrane determinants which accompany the induction of Hb synthesis. We confirm that the K562 cell line exhibits a number of erythroid features: it expresses immunologically detectable membrane proteins, glycophorin A and spectrin and can ber induced, by addition of haemin or n-butyrate, to synthesise Hb. N and i-like blood group activities are demonstrable in uninduced K562, but band 3, ABH and other major alloantigens characteristic of mature erythrocytes are lacking. The acquisition of cell surface antigens typifying erythroid or other haemopoietic lineages has not been demonstrated following induction, although certain properties not associated with morphologically recognisable red cell precursors are lost. The relevance of these findings to the foetal nature and possible multipotentiality of the K562 cell line are discussed.
The second data release of the LOFAR Two-Metre Sky Survey (LoTSS) covers 27% of the northern sky, with a total area of ~5700 deg 1 . The high angular resolution of LOFAR with Dutch baselines (6 arcsec) allows us to carry out optical identifications of a large fraction of the detected radio sources without further radio followup; however, the process is made more challenging by the many extended radio sources found in LOFAR images as a result of its excellent sensitivity to extended structure. In this paper we present source associations and identifications for sources in the second data release based on optical and near-infrared data, using a combination of a likelihood-ratio cross-match method developed for our first data release, our citizen science project Radio Galaxy Zoo: LOFAR, and new approaches to algorithmic optical identification, together with extensive visual inspection by astronomers. We also present spectroscopic or photometric redshifts for a large fraction of the optical identifications. In total 4 116 934 radio sources lie in the area with good optical data, of which 85% have an optical or infrared identification and 58% have a good redshift estimate. We demonstrate the quality of the dataset by comparing it with earlier optically identified radio surveys. This is by far the largest ever optically identified radio catalogue, and will permit robust statistical studies of star-forming and radio-loud active galaxies.
The study of osteoclast integrins has been previously hampered by the lack of a source of large numbers of purified osteoclasts. Osteoclastoma, a human giant cell tumor of bone, supplied a rich source of osteoclasts within a tissue containing many diverse cell types. Osteoclastoma integrin immunostaining confirmed the presence of the integrin alpha v beta 3 complex and the alpha 2 and beta 1 integrin subunits on osteoclasts. However, weak integrin expression, for example with alpha v beta 5, was difficult to interpret. Purification with magnetic beads coated with vitronectin receptor monoclonal antibody (13C2) enabled osteoclast membranes to be isolated with high purity and yield (57%) from osteoclastoma tissue. Positively (osteoclast-enriched) selected membranes were biochemically assessed for integrin expression by immunoprecipitation and visualization by non-radioactive enhanced chemiluminescence. alpha 1, alpha 4, alpha 6, alpha 8, alpha M, alpha X, gpIIb, beta 4, beta 6, and beta 8 integrin chains were undetectable at a sensitivity of 1 ng. alpha 3, alpha 5, alpha L, beta 2, and alpha v beta 5 were found in the negatively selected osteoclastoma tissue but not in the positively purified osteoclast membranes. The presence of alpha v beta 1 and alpha 2 beta 1 dimers was demonstrated biochemically on the immunoisolated osteoclast membranes. Osteoclast alpha v beta 3 isolation by Arg-Gly-Asp (RGD) affinity chromatography for NH2-terminal amino acid sequencing confirmed that the osteoclast vitronectin receptor was identical to that previously characterized on other cell types. In situ hybridization using human alpha v riboprobes in osteoclasts from human and rodent bone further demonstrated the high level and specificity of expression of alpha v vitronectin receptor in osteoclasts.
The main function of collagen is mechanical, hence there is a fundamental scientific interest in experimentally investigating the mechanical and structural properties of collagen fibrils on the nanometre scale. Here, we present a novel atomic force microscopy (AFM) based scraping technique that can dissect the outer layer of a biological specimen. Applied to individual collagen fibrils, the technique was successfully used to expose the fibril core and reveal the presence of a D-banding-like structure. AFM nanoindentation measurements of fibril shell and core indicated no significant differences in mechanical properties such as stiffness (reduced modulus), hardness, adhesion and adhesion work. This suggests that collagen fibrils are mechanically homogeneous structures. The scraping technique can be applied to other biological specimens, as demonstrated on the example of bacteria.
Parathyroid hormone-related protein (PTHrP) is not required for osteoclastogenesis during embryonic development; however, after birth it has been shown to regulate osteoclast formation during tooth eruption. Our study explores the hypothesis that PTHrP also may regulate osteoclast differentiation in the regenerating skeletal tissues of deer antlers, bones capable of complete regeneration. Osteoclast-like multinucleated cells (MNCs) formed spontaneously in micromass cultures derived from antler cartilage and these cells had the phenotypic characteristics of osteoclasts. PTHrP and receptor activator of NF-kappaB ligand (RANKL) stimulated antler osteoclast formation although the effect of RANKL was less marked than that of PTHrP. The addition of osteoprotegerin (OPG) only partially decreased (by approximately 65%) the number of osteoclasts in PTHrP-treated cultures. To determine whether PTHrP also potentially could have direct effects on antler osteoclasts, we studied, by confocal microscopy, the expression of the type I PTH/PTHrP receptor (PTH1R) in MNCs cultured on glass and found the receptor protein to have a nuclear localization. In situ hybridization showed that antler MNCs also expressed PTH1R and PTHrP messenger RNAs (mRNAs). PTHrP was immunolocalized in MNCs cultured on glass but was undetectable in cells resorbing a dentine substrate. In tissue sections of antler cartilage, PTHrP and PTH1R were expressed in vitronectin receptor-positive (VNR+) osteoclast-like cells localized in the perivascular stroma. Thus, these data show that PTHrP plays a role in the regulation of osteoclast differentiation in regenerating skeletal tissues and that PTHrP can have effects on osteoclastogenesis that are independent of RANKL synthesis. Ours is the first study to describe the expression of the type I PTH/PTHrP receptor in mammalian osteoclasts at a protein and mRNA level, which indicates that PTHrP also may have a direct effect on osteoclasts. This also is the first study to show a nuclear localization of the PTHIR in cells of the osteoclast lineage, although the functional significance of this observation has yet to be established.
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