Influence of hypoxia on wavelength dependence of differential pathlength and near-infrared quantification

Continuous wave near-infrared spectroscopy (NIRS) measurements of cardiac correlated changes in attenuation in the adult human head were computed using a Fourier analysis technique that eliminates the positive error bias associated with the magnitude of the Fourier coefficient. These attenuation changes were used to determine wavelength dependence of differential pathlength, DP(lambda), at four stages during progressive hypoxia (21, 17, 13 and 9% FIO2) in normal volunteers. The effects of incorporating DP(lambda) into NIRS algorithms to compute relative concentration changes and absolute concentration of oxyhaemoglobin and deoxyhaemoglobin are discussed. Because variations in DP(lambda) are restricted to wavelengths below 780 nm, absolute concentration calculations are influenced by hypoxia-induced changes while relative concentrations are unaffected. However, even accounting for changes in DP(lambda) did not allow computation of physiologically reasonable absolute concentrations of the haemoglobin species.     

A simple technique for evaluation of methods of cell separation

A simple method is described for labelling cells with fluorescein and using them in artificial mixtures to assess cell separation procedures. The method facilitates the examination of the variables in a separation procedure. It is thus possible to tailor a separation procedure (for example panning with monoclonal antibody) to suit the specific requirements of the experiment.     

Cardiovascular effects of cerebroventricular ouabain perfusion in the adult dog.

Cerebroventricular perfusion with artificial cerebrospinal fluid containing 10(-5) M ouabain was performed in adult dogs in order to describe the time course of the cardiovascular effect of intraventricular ouabain and to evaluate treatments to eliminate the cardiovascular effect. The central effect of ouabain caused a 56% increase in blood pressure above control values and a 35% increase in heart rate with various cardiac arrhythmias. Both alpha- and beta-adrenergic blocking drugs given intravenously.altered the pressure and rate effects ou ouabain, whereas vagotomy attenuated the effect.     

CD4+ T cell-mediated rejection of major histocompatibility complex class I-disparate grafts: A role for alloantibody

Experimental studies of the T cell requirement for rejection of class I major histocompatibility complex (MHC)-disparate grafts have generated controversy over both the autonomy of CD8⁺ T cells and the mechanism whereby CD4⁺ T cells are able to independently mediate rejection. In this study of rejection of RT1A^a class I MHC-disparate rat cardiac and skin allografts by high-responder PVG RT1^u recipients, we show that elimination of CD8⁺ T cells [by anti-CD8 monoclonal antibody (mAb) administration in vivo] fails to prolong graft survival, whereas partial depletion of CD4⁺ T cells (by anti-CD4 mAb treatment) markedly delays rejection of class I-disparate heart grafts, and marginally prolongs survival of skin grafts. Anti-CD4-treated PVG-RT1^u athymic nude rats reconstituted with CD8⁺ T cells failed to reject class I-disparate skin grafts for several weeks and eventual rejection correlated with re-emergence of a small number of donor derived CD4⁺ T cells. Conversely, anti-CD8-treated nude rats reconstituted with CD4⁺ T cells alone rapidly rejected class I-disparate skin grafts. Passive transfer of anti-class I immune serum to anti-CD4-treated euthymic recipients promptly restored their ability to specifically reject a class I-disparate heart graft. Similarly, passive transfer of immune serum to PVG-RT1^u nude rats bearing skin allografts caused destruction of class I-disparate but not third-party grafts. These results demonstrate that CD4⁺ T cells are both necessary and sufficient to cause rejection of class I-disparate heart and skin grafts in this model and that CD4⁺ T cell-dependent alloantibody plays a decisive role in effecting rejection.     

Experimental infection of chimpanzees with antihemophilic (factor VIII) materials: recovery of virus-like particles associated with non-A, non-B hepatitis.

Non-A, non-B viral hepatitis was transmitted to four colony-born chimpanzees by infusion of three lots of antihemophilic factor (factor VIII) implicated in the transmission of non-A, non-B hepatitis to two human recipients. All four inoculated animals showed histopathological evidence of viral hepatitis, and all demonstrated significant ALT elevations between seven and one-half weeks after inoculation. Acute-phase plasma from one of the infected chimpanzees (no. 771) was shown to induce non-A, non-B hepatitis in two other chimpanzees approximately three weeks after their inoculation. In addition, an acute-phase open liver wedge biopsy obtained from animal no. 771 was processed and examined by immune electron microscopy (IEM) for virus-like particles with convalescent serum from a serologically confirmed case of non-A, non-B hepatitis. Twenty-five to 30 nm (mean = 27 nm) diameter virus-like particles that were either "full" or "empty" were identified in this liver preparation by IEM. Two additional chimpanzees inoculated with a cesium chloride gradient fraction of an isopycnically banded liver homogenate (animal no. 771) also developed elevated ALT activity two to two and one-half weeks later. Our findings have experimentally verified that commercially produced factor VIII materials can induce non-A, non-B hepatitis in champanzees and that the disease can be subpassaged in these animals by inoculation of either acute-phase plasma or liver. These results also provide evidence for the association of 27 nm-diameter virus-like particles with non-A, non-B viral hepatitis.     

Hypoplasia of the cerebellar vermis and corpus callosum in thrombocytopenia with absent radius syndrome on MRI studies

Thrombocytopenia with absent radius (TAR) syndrome is infrequently (7%) associated with mental retardation. In those cases, the mental deficiency is presumed to be a consequence of intracranial hemorrhage due to the thrombo-cytopenia. We report on 2 infants with TAR syndrome. One had developmental delay with evidence of cerebral dysgenesis by magnetic resonance imaging (MRI). Such findings have not been noted in the literature, but may not have been investigated in most cases. The other infant with TAR syndrome, who has had normal psychomotor development, has a normal brain on MRI scan. Detailed neuroimaging studies, preferably MRI, should be considered in the evaluation of patients with TAR syndrome, especially when there are documented signs of developmental delay, with or without a history of intracranial hemorrhage. © 1994 Wiley-Liss, Inc.     

Controlled alterations in the physical and biological properties of R17 bacteriophage induced by gunaidine hydrochloride

The effects of guanidine hydrochloride on the physical and biological properties of R17 phage were studied to ascertain how these effects relate to the structure of the phage particle. The dialysis of phage against 3.0 M guanidine at 4 C resulted in a loss of infectivity and subtle changes in the sedimentation properties of the resulting non-infectious particles. Upon increasing the temperature of the reaction from 4 C to 37 C, additional physical modifications were found to occur in a sequential manner, giving rise to three unique particle forms followed by the eventual breakdown of phage structure. These three modified phage forms were found to have S values of 78, 58, and 44, respectively, to contain all three macromolecules characteristic of R17 phage, and to have the buoyant density of normal phage. The RNA of these particles were sensitive to RNase, and negative stains readily penetrated their interior, suggesting that changes had occurred in the capsid. The 44S particles in particular were found to be highly unstable as judged by electron microscopic and analytical ultracentrifugational studies. Various stages in the disassembly of these particles were identified.Protein fragments directly released from 44S particles were found to be morphologically similar to 11S subunits produced during the breakdown of phage in higher concentrations of guanidine. The morphological identify of the subunits released from 44S particles and the 11S protein subunits was supported by the similarity of the two types of subunits in forming branched chains, in the arrangement of subunits within the chains, and by their cosedimentation in sucrose gradients.Further analysis of the 11S subunits showed the following characteristics: (1) three morphological configurations with the most common form having dimensions of 106 × 50 Å; (2) a sedimentation rate of 10.5–11.4S; (3) molecular weight of 240,000 daltons. On the basis of these properties and on the known properties of phage coat protein, a model is proposed to explain the structure of these subunits and their possible relalationship to the structure of the phage particle. Additional support for the proposed model of phage structure is provided by electron microscopic observations and the recently discovered functions of the A (maturation) protein.     

Detergent-resistant membrane microdomains facilitate Ib oligomer formation and biological activity of Clostridium perfringens iota-toxin

Clostridium perfringens iota-toxin consists of two separate proteins identified as a cell binding protein, iota b (Ib), which forms high-molecular-weight complexes on cells generating Na(+)/K(+)-permeable pores through which iota a (Ia), an ADP-ribosyltransferase, presumably enters the cytosol. Identity of the cell receptor and membrane domains involved in Ib binding, oligomer formation, and internalization is currently unknown. In this study, Vero (toxin-sensitive) and MRC-5 (toxin-resistant) cells were incubated with Ib, after which detergent-resistant membrane microdomains (DRMs) were extracted with cold Triton X-100. Western blotting revealed that Ib oligomers localized in DRMs extracted from Vero, but not MRC-5, cells while monomeric Ib was detected in the detergent-soluble fractions of both cell types. The Ib protoxin, previously shown to bind Vero cells but not form oligomers or induce cytotoxicity, was detected only in the soluble fractions. Vero cells pretreated with phosphatidylinositol-specific phospholipase C before addition of Ib indicated that glycosylphosphatidyl inositol-anchored proteins were minimally involved in Ib binding or oligomer formation. While pretreatment of Vero cells with filipin (which sequesters cholesterol) had no effect, methyl-beta-cyclodextrin (which extracts cholesterol) reduced Ib binding and oligomer formation and delayed iota-toxin cytotoxicity. These studies showed that iota-toxin exploits DRMs for oligomer formation to intoxicate cells.