Side-effects of high pressure irrigation

This study was undertaken to identify side-effects of high pressure irrigation. Standardized surgical wounds made in Yorkshire pigs were subjected to high pressure syringe and pulsatile irrigation. As a result of these treatments, fluids were disseminated into the adjacent tissue of the wound, predominantly in a lateral direction. Bacteria did not accompany this disseminated fluid and apparently were filtered out by the surface tissues. This treatment results in a tissue injury which impairs its defenses, making the wound more susceptible to infection. However, the remarkable cleansing capacity of high pressure irrigation appears to outweigh this side-effect, since heavily contaminated wounds subjected to this treatment heal per primum without infection.     

A heterologous system for detecting eukaryotic enzymes which synthesize pseudouridine in transfer ribonucleic acids

tRNA pseudouridylation activities have been detected in embryonic mouse cell fractions and in extracts from HeLa, mouse L-cell and baby hamster kidney (BHK) cell lines. These activities were identified by the use of heterologous reaction systems, with tRNA from hisT strains of Salmonella typhimurium as substrate. hisT mutants are defective for an enzyme that forms psi residues in the anticodon region of many tRNAs and accumulate undermodified species of tRNA. The pseudouridylation activity from BHK cells has been examined in detail and quantitated by a modified tritium release assay (Cortese, R., Kammen, H.O., Spengler, S.J., and Ames, B.N. (1974) J. Biol. Chem. 249, 1103-1108). Maximal rates of tritium release required a suitable cationic environment (optimally, a combination of Mg2+ and NH4+) and a thiol reductant. The activity was totally inhibited in the presence of thiol-reactive reagents, such as 5,5'-dithiobis(2-nitrobenzoic acid) and p-chloromercuribenzoate. A major portion of this 3H release activity was associated with psi modification reactions. This conclusion stems from the following observations: (a) BHK extracts preferentially catalyzed a release of 3H from hisT [5-3H]tRNA, rather than from similarly labeled wild type tRNA; (b) this activity was specific for protons attached to C5 of the pyrimidine rings; no release of 3H was obtained with hisT or wild type [6-3H]tRNA as substrate; (c) the reaction products of hisT tRNA with BHK enzyme were examined by reverse phase column chromatography of tRNAPhe isoacceptors on RPC-5 columns. The enzyme modified both of the principal isoacceptors of hisT tRNAPhe to an equal extent, yielding products indistinguishable from wild type tRNAPhe. Significant levels of 3H release were obtained by the action of enzyme on wild type [5-3H]tRNA, even after gel filtration of the enzyme. This suggests that the enzyme may be able to hypermodify certain species of wild type S. typhimurium tRNA. The activities for wild type tRNA and hisT tRNA appeared to be associated with the same enzyme.     

Increased stem cell somatic mutation in the non-neoplastic colorectal mucosa of patients with familial adenomatous polyposis

Colorectal tumorigenesis in familial adenomatous polyposis (FAP) results from somatic mutation of either the normal APC allele or another growth control gene in epithelial cells bearing a germline APC defect. The rate at which tumors develop is therefore dependent on the somatic mutation frequency; it is not known whether this is normal or elevated in FAP. We aimed to quantify stem cell somatic mutation in FAP, comparing it with hereditary nonpolyposis colorectal cancer (HNPCC) and Crohn's disease (CD). Stem cell somatic mutation frequency was studied in 47 FAP patients, 5 HNPCC patients, and 13 CD patients, all younger than 49 years, by quantifying crypt-restricted loss of O-acetyltransferase activity in sections of morphologically normal colonic mucosa from individuals heterozygous for this monogenically inherited polymorphism. Median stem cell somatic mutation frequency was significantly higher in FAP than HNPCC (4.2 x 10(-4) v 1.4 x 10(-4), Mann-Whitney U, P < .02). The level in CD (4.0 x 10(-4)) was similar to FAP. Mutated crypts occurred in groups more frequently in FAP (22%) than HNPCC (12%) or CD (10%), suggesting an increase in stem cell division associated with crypt fission in FAP. We conclude that stem cell somatic mutation frequency is raised in non-neoplastic colorectal mucosa in FAP. This is probably related to increased stem cell proliferation and contributes to the high rate of tumor formation in this condition.     

GnRH-stimulated glycosylation (proximal and distal) of luteinizing hormone by cultured rat pituitary cells

In the present work, the effects of GnRH on the translation (by [14C]leucine incorporation; [14C]Leu-LH) and the glycosylation of LH by rat pituitary cells in primary culture were established. The use of specific markers as radioactive precursors made it possible to discriminate the action of the neurohormone on proximal glycosylation (by[3H]mannose incorporation; [3H]Man-LH) as well as distal glycosylation (by [3H]galactose incorporation; [3H]Gal-LH) in the course of synthesis and release of LH. Pituitary cells from ovariectomized adult rats were incubated for different periods between 0 and 5 h in medium containing [14C]Leu plus [3H]Man or [14C]Leu plus [3H]Gal with or without 10 nM GnRH. GnRH increased synthesis and release of newly synthesized LH. The magnitude of the stimulatory effect on the kinetics of [14C]Leu (slope = 63.58; 158% of control) and [3H]Man (slope = 75.15; 161%) incorporation to LH was similar. The action of the neurohormone appears to be exerted on translation, the increased [3H]Man incorporation being a secondary phenomenon arising from the greater amount of available polypeptide chains as acceptors of the polymannose core. However, a direct effect of GnRH on proximal glycosylation cannot be excluded. GnRH also stimulated the kinetics of release of [14C]Leu-LH (slope = 6.14; 236% of control) and [3H]Man-LH (slope = 8.06; 191%). Comparatively, the effect of GnRH on [3H]Gal-LH was detected earlier than that on LH labeled with the other precursors; increases in rates of production (slope = 71.57; 278% of control) and release (slope = 32.08; 494%) were higher than those in [14C]Leu- and [3H]Man-LH kinetics, indicating that GnRH acts specifically on this distal step of LH glycosylation. GnRH enhanced the relative terminal glycosylation ([3H]Gal/[14C]Leu ratio) of total and release LH without modifying the relative proximal glycosylation ([3H]Man/[14C]Leu ration) of the hormone. We conclude that GnRH can induce not only changes in the quantity (greater number of molecules) but also in the quality (molecules more glycosylated) of the secreted LH by acting directly at translation and distal glycosylation level.     

Spectrum of aerobic endurance running performance in eleven inbred strains of rats

Mice homozygous for a disruption in the Mdr2 gene (Mdr2 (-/-) mice) lack the Mdr2 P-glycoprotein (P-gp) in the canalicular membrane of the hepatocyte and are unable to excrete phosphatidylcholine into the bile. These mice develop a nonsuppurative cholestatic liver disease, presumably caused by the high concentrations of free cytotoxic bile acids in bile. We generated transgenic mice that express the human homolog of Mdr2, MDR3, specifically in the liver by the use of an albumin promoter. In these mice the MDR3 P-gp is exclusively located in the canalicular membrane of hepatocytes and phospholipid excretion into bile is restored. Mice that contain the same amount of MDR3 P-gp as that of Mdr2 P-gp in wild-type mice, also excrete the same amount of phospholipids. No histopathological abnormalities were observed in the livers of these mice. In mice that express MDR3 at a higher or lower level, the phospholipid excretion correlated with the amount of MDR3 P-gp. We conclude that the human MDR3 P-gp is functionally homologous to the murine Mdr2 P-gp and that it can fully replace this P-gp in Mdr2 (-/-) mice, restoring the excretion of phospholipids into the bile. The phospholipid excretion is limited by the amount of MDR3 or Mdr2 P-gp. The excretion of cholesterol is not tightly coupled to the excretion of phospholipids in these mice, because a very low phospholipid excretion level is sufficient to give almost wild-type cholesterol excretion into the bile.     

Interferon alpha inhibits a Src-mediated pathway necessary for Shigella-induced cytoskeletal rearrangements in epithelial cells

Shigella flexneri, the causative agent of bacillary dysentery, has the ability to enter nonphagocytic cells. The interferon (IFN) family of cytokines was found to inhibit Shigella invasion of cultured epithelial cells. We show here that IFN-alpha inhibits a Src-dependent signaling cascade triggered by Shigella that leads to the reorganization of the host cell cytoskeleton. Immunofluorescence studies showed that IFN-alpha inhibits Shigella-induced actin polymerization required for bacterial entry into cells. Phosphorylation of cortactin, a Src-substrate specifically tyrosyl-phosphorylated during Shigella entry, was inhibited by IFN-alpha. Overexpression of a dominant interfering form of pp60c-src led to inhibition of Shigella-induced cytoskeletal rearrangements and decreased cortactin phosphorylation indicating a role for Src in Shigella entry. Also, Shigella uptake in cells that expressed constitutively active Src was unaffected by IFN-alpha treatment. We conclude that Src kinase activity is necessary for Shigella invasion of epithelial cells and that IFN-alpha inhibits this Src-dependent signaling pathway.     

Mammography grid performance

PURPOSE: To measure directly the grid performance of mammography units for the range of breast thicknesses and x-ray tube potentials encountered in clinical practice. MATERIALS AND METHODS: Contrast improvement factors and Bucky factors were determined for four mammographic units as a function of x-ray tube potential (25, 30, and 35 kVp), phantom thickness (2, 4, and 8 cm) and, on one unit, three target-filter combinations. Three units used a linear grid; one, a cellular grid. Two methods were used for nongrid measurements. RESULTS: For all units tested, contrast improvement factor increased with increased phantom thickness and with increased kilovolt peak level for the 8-cm-thick phantom and changed little with kilovolt peak level for 2- and 4-cm-thick phantoms. At 25 and 30 kVp, contrast improvement factor performance with the linear grids was comparable; with the cellular grid, it was 5%-10% higher. In all cases, the Bucky factor increased with increased phantom thickness and decreased with increased tube potential. CONCLUSION: Differences in grid performance exist. At 25 and 30 kVp, the cellular grid exhibited superior contrast improvement factor performance, whereas one of the linear grids exhibited superior Bucky factor performance. Measured contrast improvement and Bucky factors are dependent on nongrid technique. Cassette tunnels introduce scatter and should not be used with nongrid or magnification techniques.     

The effects of low-frequency ultrasound on Staphylococcus epidermidis

Low frequency ultrasound (LFUS) significantly enhances skin permeability to a variety of drugs; however, its bacterial effects have not been well studied. Staphylococcus epidermidis organisms were grown and standardized to 10(5) cfu/ml 24 h prior to investigation and suspended in normal saline. LFUS was applied with two probes immersed in the bacterial suspensions over a range of suspension volumes, intensities, and exposure times. The suspension temperature was measured, and a sample was removed, streaked onto blood agar plates, and incubated at 37 degrees C for 24 h. Quantitative bacterial counts were then obtained. LFUS resulted in significant reductions in bacterial counts that correlated with fluid temperature. Probe size and ultrasound intensity appeared to affect bacterial counts, but were also correlated with temperature. Bacterial growth was minimal with temperatures exceeding 45 degrees C. While LFUS can reduce bacterial counts, these conditions have the potential to cause burns in humans.