Oxygen tension monitoring with cutaneous electrodes in adults

Two commercially available kits for determining blood ammonia were compared, an enzymatic method requiring a sample volume of 100mul and a nonenzymatic method requiring 1 ml of sample. Calibration curves for both kits were linear to 270 mumol/l, however correlation studies on plasma samples revealed a lack of agreement between methods (r equal to 0.825, Y equal to 0.76X +51). Improved correlation occurs when protein interference is eliminated from the enzymatic procedure. Finally, the nonenzymatic method is modified to accommodate 100 mul of sample and compared to the original method.     

Oxygen tension of the skin of ischemic legs

Using a Kontron Roche Transcutaneous Oxygen Monitor, we measured oxygen tension on the skin of the legs at three sites in patients with peripheral vascular disease and group of controls. Significant decreases in oxygen tension occurred in the patient groups, which correlated well with ankle systolic pressure, with differences between those with claudication and those with rest pain. These results suggest that in limbs with claudication, significant skin hypoxia may exist during rest in spite of reportedly normal skin and muscle blood flow. The progressive decrease in skin oxygen tension down a limb with occlusive vascular disease may play a significant role in skin healing.     

Oxygen tension profiles in isolated hamster retractor muscle at different temperatures

Oxygen tension (P0(2)) profiles within unperfused hamster retractor muscles were obtained at 25, 30, and 37 degrees by using sharpened, recessed oxygen microelectrodes. The microelectrode was driven vertically into freshly excised muscle lying on a flat, impermeable boundary inside a diffusion chamber. Intramuscular P0(2) profiles were measured as a function of electrode depth in 10-mu m steps during both inward and outward penetrations when the upper surface of the muscles was exposed to humidified gases containing 10, 21, 50, and 100% 0(2). The ratio of the 0(2) consumption (M) to the 0(2) permeability (K, Krogh diffusion coefficient = D alpha, diffusion coefficient-solubility product) was estimated by curve-fitting the experimental steady-state distribution of 0(2) through muscles to the analytic solution of the diffusion equation assuming that M obeys zero-order kinetics and K is constant, uniform, and independent of P0(2). The ratios of M/K were independent of temperature and were found to be independent of surface P0(2) and muscle thickness. The average value of M/K was 3.9 +/- 0.45 (SE; n = 30) x 10(5) mm Hg/cm(2), which is consistent with that estimated from previous measurements of M and D using different non-steady-state techniques (Bentley et aL, 1993). These results are consistent with other in vitro 0(2) consumption measurements (Sullivan and Pittman, 1984) and do not provide evidence for nonclassical respiratory activity in resting mammalian skeletal muscle.     

Genes expressed after retinoic acid-mediated differentiation of embryoid bodies are likely to be expressed during embryo development

In order to test if retinoic acid-mediated differentiation of embryoid bodies can be used as an in vitro preselection method for ES cell lines generated by gene trap, we correlated gene expression after in vitro differentiation and in 11.5-day embryos. Fifty-two genes captured by gene trap and expressed in undifferentiated embryonic stem cells were analyzed. Most genes expressed after differentiation in vitro were also expressed during embryo development. In order to correlate the expression patterns in vitro and in vivo, the in vitro expression in the center and in the periphery of the embryoid body outgrowths was observed. This allowed us to distinguish, according to in vitro expression, not expressed genes from those expressed widely in 11.5-day embryos. Consequently, with this parameter we increased the probability to obtain the restricted expression patterns in vivo. This study demonstrates the potential of the differentiation procedure in combination with the gene trap to select in vitro for genes expressed during embryo development.     

活套高度闭环和张力闭环控制投入时机的选择

热连轧过程中利用位置环起套时经常会造成活套位置超调,从而造成带钢头部被拉窄拉薄。但是通过跟踪套量变化曲线,可以预先判断活套与带钢接触的时机,从而快速退出位置环,减小由于活套位置环起套超调造成的拉钢。针对活套起套过程,提出一种确定活套高度闭环和张力闭环投入时机的算法,经过现场实际应用,效果明显,使起套过程控制获得极大改善,确认这是一种行之有效的方法。文中还就该算法在起套过程中遇到的几个问题进行了探讨。     

Oxygen tension in the oviduct and uterus of rhesus monkeys, hamsters and rabbits

Oxygen tension was measured using flexible polarographic microelectrodes within the oviductal and uterine lumen in rhesus monkeys (n = 9), golden hamsters (n = 21) and rabbits (n = 6), during the reproductive cycle (monkey), during oestrus and pseudopregnancy (hamsters, rabbits) and during pregnancy (hamsters). In general, oxygen tensions in each species were much less than half of atmospheric O2, ranging from high values of about 60 mm Hg (8.7% O2) in the rabbit oviduct, rabbit and hamster uterus, to as low as 11 mm Hg (1.5% O2) in the monkey uterus. Oxygen tensions did not vary significantly between left and right sides of the reproductive tracts (all species), nor between pregnant and pseudopregnant states nor between oviduct and uterus (hamsters). Differences owing to reproductive stage were found in the monkey oviduct, hamster oviduct and uterus, and rabbit uterus. Oxygen tensions were consistently very low (11-14 mm Hg) in the monkey uterus throughout the menstrual cycle. In hamsters and rabbits, intrauterine O2 decreased significantly at about the normal time of blastocyst formation and implantation, to 37 mm Hg (5.3% O2) and 24 mm Hg (3.5% O2), respectively. This study indicates that embryos develop in vivo under low oxygen concentrations, especially during the peri-implantation period. The data have implications for investigations of embryo metabolism and for improving embryo development in vitro.     

The superantigen toxic shock syndrome toxin-1 induces CD40 ligand expression and modulates IgE isotype switching

Isotype switching to IgE requires two signals. The first signal is provided by the cytokines IL-4 or IL-13, and the second signal is delivered by the interaction between the B cell antigen CD40 and its ligand (CD40L) which is expressed on activated T cells. Since superantigens have been shown to activate T cells, we examined the effect of the superantigen toxic shock syndrome toxin-1 (TSST-1) on CD40L expression on T cells. TSST-1 induced expression of CD40L in both freshly isolated T cells and in T cell lines expanded by re-stimulation with TSST-1. CD40L was preferentially expressed in the V beta 2 subset of T cells expanded by TSST-1. We next examined the effect of TSST-1 on IgE synthesis by human peripheral blood mononuclear cells (PBMC). Addition of TSST-1 to PBMC inhibited IL-4-induced IgE synthesis in a dose-dependent manner. This inhibition was reversed partly by adding a neutralizing antibody to IFN-gamma. In contrast, TSST-1 induced high amounts of IgE synthesis in the presence of IL-4 at low T:B cell ratios (0.5:10 to 4:10), a condition which circumvents the inhibitory effect of IFN-gamma. TSST-1 induction of IgE synthesis was inhibited by a mAb to CD40L. These results indicate that superantigens induce CD40L expression in T cells and cause isotype switching in B cells which is mediated by CD40L-CD40 interaction.     

Oxygen tension alters the effects of cytokines on the megakaryocyte, erythrocyte, and granulocyte lineages

Whereas patients with multiple myeloma continue to relapse after autologous transplantation and are unlikely to be cured, the probability of progression is less after allogeneic transplantation and a proportion of patients may be cured. This is attributable to an immunologically mediated graft-versus-myeloma (GVM) effect which is akin to the well-known graft-versus-leukemia effect. The available clinical and experimental evidence strongly support the existence of GVM, but it is not known whether GVM is separable from graft-versus-host disease (GVHD) in practice. The best way to exploit GVM reactions is unclear, and the morbidity and mortality associated with GVHD undermine long-term survival. There is usually a time lag of a few weeks between immune intervention and disease response. There is a propensity for extramedullary disease recurrence in patients whose marrow disease is controlled with immunologic manipulation. Exploration of GVM outside conventional allogeneic transplantation or after autologous transplantation is necessary to increase the number of patients likely to benefit from this phenomenon and to make it safer. This article reviews the currently available literature on the subject.     

Myosin regulatory light chain modulates the Ca2+ dependence of the kinetics of tension development in skeletal muscle fibers

To determine the role of myosin regulatory light chain (RLC) in modulating contraction in skeletal muscle, we examined the rate of tension development in bundles of skinned skeletal muscle fibers as a function of the level of Ca(2+) activation after UV flash-induced release of Ca(2+) from the photosensitive Ca(2+) chelator DM-nitrophen. In control fiber bundles, the rate of tension development was highly dependent on the concentration of activator Ca(2+) after the flash. There was a greater than twofold increase in the rate of tension development when the post-flash [Ca(2+)] was increased from the lowest level tested (which produced a steady tension that was 42% of maximum tension) to the highest level (producing 97% of maximum tension). However, when 40-70% of endogenous myosin RLC was extracted from the fiber bundles, tension developed at the maximum rate, regardless of the post-flash concentration of Ca(2+). Thus, the Ca(2+) dependence of the rate of tension development was eliminated by partial extraction of myosin RLC, an effect that was partially reversed by recombination of RLC back into the fiber bundles. The elimination of the Ca(2+) dependence of the kinetics of tension development was specific to the extraction of RLC rather than an artifact of the co-extraction of both RLC and Troponin C, because the rate of tension development was still Ca(2+) dependent, even when nearly 50% of endogenous Troponin C was extracted from fiber bundles fully replete with RLC. Thus, myosin RLC appears to be a key component in modulating Ca(2+) sensitive cross-bridge transitions that limit the rate of force development after photorelease of Ca(2+) in skeletal muscle fibers.     

Development of mouse embryonic stem (ES) cells: IV. Differentiation to mature T and B lymphocytes after implantation of embryoid bodies into nude mice

Mouse embryonic stem (ES) cells in culture can differentiate into late stages of many lineage-committed precursor cells. Under appropriate organ-culture conditions, ES cells differentiate into lymphoidlike cells at a stage equivalent to lymphoid cells found in fetal liver. These hematopoietic precursors are located in cup-shaped structures found in some embryoid bodies; we called such embryoid bodies "ES fetuses." In this study, we have followed the maturation of hematopoietic cells after implantation of ES fetuses into nude mice for 3 weeks. ES-cell-derived lymphoid cells-pre-B cells, mature B cells, and mature T cells were found in all lymphoid organs. Interestingly, there was also an increase of T cells of host origin. Because native nude mouse lack thymus, these T cells might be educated by thymuslike epithelium generated from ES fetuses. Practical applications of this combined in vitro and in vivo system are discussed.     

The embryoid body as a novel in vitro assay system for antiangiogenic agents

Tumor progression necessitates the induction of blood vessels that converge upon the tumor and enhance the diffusibility of oxygen and nutrients. Approaches to treat cancer by antiangiogenic therapy are therefore straightforward, and there is a great need for suitable in vitro systems to test antiangiogenic agents. In the present study, embryoid bodies (EBs) differentiated from totipotent mouse embryonic stem (ES) cells and cultivated using the spinner flask technique are introduced as an in vitro system for antiangiogenesis research. ES cells effectively differentiated endothelial cells within the three-dimensional tissue of EBs. The total area of capillary-like structures, which were positive for CD31 (platelet endothelial cell adhesion molecule, PECAM-1), was assessed by confocal laser scanning microscopy and image analysis of a series of optical sections. Endothelial differentiation occurred between Day 4-5 and Day 8 of EB development. Within 7 days, 100% of EBs contained capillary-like structures. Suramin, tamoxifen, tetrahydrocortisol, and a combination of tetrahydrocortisol and heparin were tested for their antiangiogenic capacity in the EB system and were found to efficiently inhibit endothelial differentiation. Diffusion studies of a 10-kd 2',7'-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein (BCECF)-dextran and the fluorescent, amphiphilic agent doxorubicin in avascular and vascularized EBs revealed that the endothelial structures formed functional vessels that facilitated diffusion. The diffusion coefficient D for doxorubicin was 296 x 10(-9) cm2 s(-1) in vascularized 8-day-old EBs, ie, about 10-fold larger than in avascular 3-day-old EBs (18 x 10(-9) cm2 s(-1)) and EBs treated with suramin (14 x 10(-9) cm2 s(-1)), tamoxifen (13.5 x 10(-9) cm2 s(-1)), and tetrahydrocortisol/heparin (18.5 x 10(-9) cm2 s(-1)). Consequently, avascular EBs treated with antiangiogenic agents developed central necrosis, which was absent in vascularized EBs. Our findings indicate that EBs are a suitable in vitro model system to study the effects of antiangiogenic agents in a three-dimensional tissue context. Furthermore, EBs provide a unique model to investigate the diffusion of anticancer agents in a tissue in both the avascular and vascularized states.     

Genetic dissection of a mammalian replicator in the human beta-globin locus

The timing and localization of DNA replication initiation in mammalian cells are heritable traits, but it is not known whether initiation requires specific DNA sequences. A site-specific recombination strategy was used to show that DNA sequences previously identified as replication initiation sites could initiate replication when transferred to new chromosomal locations. An 8-kilobase DNA sequence encompassing the origin of DNA replication in the human beta-globin locus initiated replication in the simian genome. Specific deletions within the globin origin did not initiate replication in these chromosomal sites. These data suggest that initiation of DNA replication in mammalian cells requires specific sequence information and extend the replicon hypothesis to higher eukaryotes.     

Mutagenesis of retroviral vectors transducing human beta-globin gene and beta-globin locus control region derivatives results in stable transmission of an active transcriptional structure

Retrovirus-mediated gene transfer of the human beta-globin gene into hematopoietic stem cells is an attractive approach to the therapy of human beta-globin gene disorders. However, expression of the transduced beta-globin gene linked to its proximal cis-acting sequences (-0.8 to +0.3 kb from the cap site) is considerably below the level required for a significant therapeutic effect. The discovery of the beta-locus control region (beta-LCR), organized in four major DNase I hypersensitive sites far upstream of the human beta-like globin gene cluster, provided a potential means to achieve a high level of expression of a linked human beta-globin gene, but initial attempts to incorporate beta-LCR derivatives in retroviral vectors resulted in the production of low-titer viruses with multiple rearrangements of the transmitted proviral structures. We now describe how extensive mutagenesis of the transduced beta-globin gene, eliminating a 372 bp intronic segment and multiple reverse polyadenylation and splicing signals, increases viral titer significantly and restores stability of proviral transmission upon infection of cell lines and bone marrow-repopulating cells. These optimized vectors have enabled us to analyze the expression properties of various retrovirally transduced beta-LCR derivatives in dimethylsulfoxide-induced murine erythroleukemia cells and to achieve ratios of human beta-globin/murine beta maj-globin mRNA, on a per gene basis, as high as 80%.     

Accumulation of alpha- and beta-globin messenger RNAs in mouse erythroleukemia cells

The accumulation of alpha- and beta-globin mRNA sequences in murine erythroleukemia cells (MELC) treated with various inducers has been studied using specific alpha- and beta-globin complementary DNAs (cDNAs). In cells cultured with dimethylsulfoxide (Me2SO), hexamethylene bisacetamide (HMBA) or butyric acid, accumulation of alpha-globin mRNA is detectable after 16, 12 and 8 hr of culture, respectively. An increase in beta-globin mRNA sequences is not detected until 20-24 hr after culture. In cells exposed to hemin, both alpha- and beta-globin mRNAs are detectable by 6 hr of culture, and a constant ratio of alpha/beta-mRNA is maintained during induction. In maximally induced cells, the alpha/beta-globin mRNA ratios are approximately 1 in cells induced by Me2SO and HMBA, and 0.66 and 0.3-0.50 in cells induced by butyric acid and hemin, respectively. Thus different inducers of erythroid differentiation in MELC lead to different times of onset of the expression of alpha- and beta-like genes. In addition, the relative accumulation of alpha- and beta-globulin mRNAs in induced cells differs with various types of inducers.     

Laryngeal foreign bodies in children

Laryngeal foreign bodies (F.B.) in children are relative rare, especially in infants under one year of age, and the diagnosis and removal are difficult. The history, the clinical and radiologic findings can be misleading. Seventeen patients with laryngeal F.B. (10 of them under one year of age) are presented, and the diagnostic problems and treatment are discussed. Foreign bodies were either objects of a sharp and thin quality, e.g. an eggshell fragment, or large, e.g. a piece of meat, causing apnea and death.