A determinant of polyomavirus virulence enhances virus growth in cells of renal origin.

We have identified a strain of polyomavirus, Py(L), which is unusual in causing acute morbidity and early death after inoculation of newborn mice. We determined that these animals died of kidney failure associated with extensive, virus-mediated destruction of renal tissue. Interestingly, the Py(L) strain infects baby mouse kidney cell cultures more efficiently than do other strains.     

Degradation of extracellular matrix by the trophoblastic cells of first-trimester human placentas

First-trimester human placental villi were cultured on 3H-leucine-labeled extracellular matrices isolated from the PF HR9 and PYS-2 cell lines. Both cell lines produced an extracellular matrix that contained basement membrane-specific macromolecules, including type IV collagen, laminin and proteoglycan. Both matrices promoted outgrowth of cells from the villi which, according to morphological criteria, were identified as cytotrophoblastic cells. As the cells migrated from the attachment site, they caused a marked focal dissolution of the matrix which was accompanied by a concomitant release of 3H-labeled material into the media. Approximately half of this material chromatographed near the inclusion volume of Sephadex G-50, indicating that the labeled matrix components had been degraded. This phenomenon was dependent on the age of the placenta. Second-trimester placental villi also adhered to the matrix, but no areas of dissolution were formed and no significant amounts of radioactivity were released into the medium. These results suggest that culture of first-trimester human placental villi on extracellular matrices may be useful for the study of some of the early embryonic events leading to human implantation, during which the trophoblastic cells erode the uterine epithelium.     

Two-dimensional electrophoretic analysis of human erythrocyte cylindrin

In the absence of a peptidylproline substrate, the oxidative decarboxylation of 2-oxoglutarate by prolyl 4-hydroxylase (prolyl-glycyl-peptide,2-oxoglutarate:oxygen oxidoreductase (4-hydroxylating), EC 1.14.11.2) is stoicheiometrically coupled to the oxidation of ascorbate. The Km and Kd for O2 in this partial reaction are 1.5 mM, this value being one order of magnitude higher than the Km and Kd for O2 in the complete reaction in the presence of (Pro-Pro-Gly)5, indicating that in this case O2 can become enzyme-bound predominantly after the interaction of the peptide substrate with the enzyme. The Km values for 2-oxoglutarate in the partial and the complete reactions are the same. In the absence of both a peptide substrate and ascorbate 2 mol CO2 per mol enzyme are produced in the first 1-1.5 min, during which the enzyme becomes inactivated and, as shown earlier (De Jong , L., Albracht , S.P.J. and Kemp, A. (1982) Biochim. Biophys. Acta 704, 326-332) enzyme-bound Fe2+ becomes oxidized to Fe3+. The results are consistent with a mechanism in which a Fe2+O complex is the O-transferring intermediate involved in peptidylproline hydroxylation.     

Natural killer activity of kurloff cells: A direct demonstration on purified kurloff cell suspensions

In order to study their natural killer effect, guinea pig splenic Kurloff cells were fractionated by Percoll discontinuous density gradient centrifugation. Kurloff cells were collected and tested for cytotoxicity in a 24-hr chromium-release test. Comparison of different splenic cellular samples (of males or estrogenized females) with increasing percentage of Kurloff cells, revealed a highly significant positive correlation (r = 0.93, α < 0.01) with the cellular cytotoxicity developed against the K 562 target cells.     

A comparison of chromium-51 and iron-59 for estimating erythrocyte survival in the cat

Erythrocyte survival studies were conducted on eight, normal, healthy, 1-year-old male specific-pathogen-free cats using both chromium-51 and iron-59 simultaneously. The chromium-51 procedure gave a half-life value of 11.1 +/- 0.9 days. This was considerably lower than would be expected on the basis of the experimentally determined iron-59 erythrocyte survival time of 51.2 +/- 14.9 days. The results of this study indicated that there was considerable loss of the chromium-51 label in the cat other than that from senescence alone. An analysis of the chromium-51 disappearance curve indicated that there were two exponential disappearance rates for the chromium-51 label and, in the absence of cell death, approximately 67% of the label was lost with a rate constant of 0.02 per day and 33% was lost with a rate constant of 0.1 per day. An equation is presented which models the loss of chromium-51 label which could be used to calculate erythrocyte survival from a chromium-51 disappearance curve. Blood volume measurements, hemograms, bone marrow differential results, and iron kinetic values also were determined and the results presented. While a reasonable approximation of the erythrocyte life span could be made by correcting the chromium-51 values for losses other than senescence, the iron-59 procedure would be the preferred method in cats.     

Genetic analysis and molecular cloning of the Escherichia coli ruv gene

Summary The genetic organisation of the ruv gene, a component of the SOS system for DNA repair and recombination in Escherichia coli, was investigated. New point mutations as well as insertions and deletions were generated using transposon Tn10 inserted in eda as a linked marker for site specific mutagenesis, or directly as a mutagen. The mutations were ordered with respect to one another and previously isolated ruv alleles by means of transductional crosses. The direction of chromosome mobilization from ruv:: Mud(Ap^R lac)strains carrying F42lac ⁺ established that ruv is transcribed in a counterclockwise direction. Recombinant phages able to restore UV resistance to ruv mutants were identified, and the ruv ⁺ region was subcloned into a low copy number plasmid. The ruv ⁺ plasmid was able to correct the extreme radiation sensitivity and recombination deficiency of ruv recBC sbcB strains.     

Fetal bovine bone cells synthesize bone-specific matrix proteins

We isolated cells from both calvaria and the outer cortices of long bones from 3- to 5-mo bovine fetuses. The cells were identified as functional osteoblasts by indirect immunofluorescence using antibodies against three bone-specific, noncollagenous matrix proteins (osteonectin, the bone proteoglycan, and the bone sialoprotein) and against type 1 collagen. In separate experiments, confluent cultures of the cells were radiolabeled and shown to synthesize and secrete osteonectin, the bone proteoglycan and the bone sialoprotein by immunoprecipitation and fluorography of SDS polyacrylamide gels. Analysis of the radiolabeled collagens synthesized by the cultures showed that they produced predominantly (approximately 94%) type I collagen, with small amounts of types III and V collagens. In agreement with previous investigators who have employed the rodent bone cell system, we confirmed in bovine bone cells that (a) there was a typical cyclic AMP response to parathyroid hormone, (b) freshly isolated cells possessed high levels of alkaline phosphatase, which diminished during culture but returned to normal levels in mineralizing cultures, and (c) cells grown in the presence of ascorbic acid and beta-glycerophosphate rapidly produced and mineralized an extracellular matrix containing largely type I collagen. These results show that antibodies directed against bone-specific, noncollagenous proteins can be used to clearly identify bone cells in vitro.     

The scrotal myocutaneous flap

The scrotum is a thermoregulatory, well-vascularized structure formed by skin and nonstriated muscle with unique elastic properties. This makes it an ideal source of tissue coverage for problem wounds in its vicinity. Two patients in which scrotal musculocutaneous flaps were used are reported: one, a paraplegic, with a recurrent ischioperineal decubitus ulcer, and another with an ulcer of the penis with exposed Dacron graft previously placed to treat Peyronie's disease. After reviewing the anatomy of the scrotum and the existent literature, we studied scrotal vascularity in a fresh specimen by transillumination. Based on our experience, we conclude that this flap is easy to perform, reliable, and very useful for wounds around the perineal region.