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1.
A. M. Liberati E. Ballatori M. Fizzotti M. Schippa L. Cini S. Cinieri M. G. Proietti R. Di Marzio M. Senatore F. Grignani 《Cancer immunology, immunotherapy : CII》1988,26(1):87-93
Summary A total of 19 Hodgkin's disease (HD) patients (12 male, 7 female) aged 26–67 years, who had been in complete unmaintained remission for 6 months or more when the study was initiated, were randomly given 50 mg thymostimulin (TS) i.m. daily (G1) or every other day (G2) for 35 days. A third group (G3) was not treated. Then TS, at the same dose was administered twice a week for the following 22 weeks in patients both initially receiving loading or intermittent TS treatment. When compared with age-and sex-matched controls, as a group, the patients' circulating OKT
3
+
, OKT
4
+
, OKT
11
+
and E-AETR+ cells were depressed (P<0.001 for both proportions and absolute numbers), whereas their OKT
8
+
cell population was not. Following 5 weeks of daily TS administration, the proportions and numbers of all T cell fractions significantly increased in G1 patients (P<0.03 for all the comparisons tested), while following intermittent TS treatment (G2) only the proportions of OKT
3
+
and OKT
11
+
cells (P<0.03), but not of other T cell fractions, significantly increased. In addition, no significant changes in the absolute numbers of T cell fractions were observed in this group of patients. Furthermore, no spontaneous variations in the T cell pool size occurred in untreated patients. TS maintenance therapy did not produce any further improvement in the size of overall T cells and T cell subsets but sustained percentage and absolute numbers of these cells during administration and the absolute number of T cells even after discontinuation of therapy. The TS-induced improvement in the T cell pool was not associated with any change in the size of circulating non-T lymphocytes and monocytes. In vitro phytohemagglutinin-induced interleukin-2 (IL-2) and gamma-interferon (IFN-) synthesis was assessed in 11 patients (3 G1, 4 G2, and 4 G3). Although it was not statistically significant, a rise in IL-2 and IFN- production was observed in TS-treated patients, but not in untreated controls. TS failed to exert any effect on the serum circulating levels of neopterin, type I and II IFN, beta-2 microglobulin (B2-M) and immunoglobulins (Ig). TS can thus improve defective T cell frequences and numbers and may modulate IL-2 and IFN- production. 相似文献
2.
The formation of metarhodopsin II in various bovine rhodopsin preparations (rod outer segment (ROS) suspensions and rhodopsin-detergent solutions) was measured by means of flash spectrophotometry. The half-lifetime and formation of metarhodopsin II in ROS did not depend on the calcium concentration in the range of less than 10–9 M (using EGTA or EDTA) to 15×10–3 M calcium at pH values of 5.0, 7.1, and 9.0 (Table 1).The regeneration of rhodopsin from opsin by adding 11-cis retinal to ROS-suspensions and rhodopsin digitonin solutions was measured spectrophotometrically. It was not substantially different in either saline, one containing less than 10–7 M calcium (by adding EGTA), the other containing 10–3 M calcium (Table 2).Abbreviations A
absorption
- A
absorption change
- CTAB
N-Cetyl-N,N,N-trimethylammoniumbromide
- E700
extinction at =700 nm
- EDTA
ethylenediamine-NNNN-tetraacetic acid
- EGTA
2,2-ethylenedioxybis [ethyliminodi (acetic acid)]
- MI
metarhodopsin I
- MII
metarhodopsin II
- Rh
rhodopsin
- ROS
rod outer segment
This work is based upon a Ph. D. dissertation (Nöll, 1974) and was presented in part at the Jahrestagung der Deutschen Gesellschaft für Biophysik, Freiburg, Germany, October 1974 相似文献
3.
L. Bergmann P. S. Mitrou K. C. Weber W. Kelker 《Cancer immunology, immunotherapy : CII》1984,17(2):112-116
Summary In 42 patients with untreated or treated multiple myeloma (MM) or benign monoclonal gammopathy (BMG) the lymphocytes and T lymphocyte subsets were determined by monoclonal antibodies and other surface markers.In untreated MM, the T cells (1077/l vs 1439/l, P<0.01) and especially the OKT4+ lymphocytes (700/l vs 950/l, P<0.05) were significantly reduced compared with a control group. The OKT8+ cells were slightly but not significantly decreased.In previously treated MM, the loss of T cells was more pronounced than in the untreated group and was primarily caused by a further reduction of OKT4+ cells. Patients with BMG revealed decreased OKT8+ lymphocytes (304/l vs 502/l, P<0.001), whereas the OKT4+ cells were within the normal range. Therefore, the OKT4/OKT8 ratio was significantly elevated compared with that in untreated MM patients and normal controls (3.31 vs 2.06 vs 2.13; P<0.005).To sum up, in MM the results revealed a reduction of T cells, mainly of OKT4+ cells, which is intensified by chemotherapy and persists even after a long therapy-free interval. The different findings of T cell subsets in BMG and MM may be a helpful criterion to differentiate between BMG and MM. 相似文献
4.
S. J. D. Karlish 《Journal of bioenergetics and biomembranes》1980,12(3-4):111-136
Conformational changes have been studied in (Na,K) ATPase labeled at or near the ATP binding region with fluorescein following incubation with fluorescein isothiocyanate (FITC). One or two fluorescein groups are bound per ATPase molecule. (Na,K) ATPase activity, phosphorylation from ATP, and nucleotide binding are abolished in labeled enzyme, but phosphorylation from inorganic phosphate or K-phosphatase activity are only partially inactivated. The fluorescein groups are incorporated only into the 96 KD catalytic chain of the (Na,K) ATPase, and presence of ATP during the incubation with FITC protects against the incorporation and inhibition of enzymic activity. Upon trypsin treatment of labeled membranes the fluorescein appears first in a 58 KD fragment and eventually is released into the medium. The fluorescein-labeled (Na,K) ATPase shows a large quenching of fluorescence (15–20%) on conversion of the E1 or E1 · Na conformation in cation-free or Na+-rich media to the E2 · (K) form in K+ (or congeners Tl+, Rb+, Cs+, NH
4
+
) rich media. Cation titrations suggest that K+ and Na+ ions compete at a single binding site and stabilize E1 · Na or E2 · (K) respectively;K
K0.23 mM,K
Na1.2 mM. The rate of the conformational transition E2 · (K) E1 · Na is slow,k=0.3 sec–1, but contrary to previous experience [7, 8] ATP does not stimulate this rate. The rate of the transitions E1 + K+ E2 · (K) rises sharply with K+ concentration and shows saturation behavior, from which ak
max286 sec–1 andK
k74 mM are deduced. The data support and extend the previous suggestion that K+ ions bound initially at a low-affinity (probably cytoplasm oriented) site in state E1 are trapped in the occluded form E2 · (K) by the conformational change poised far (K
c1000) in the direction of E2 · (K). It is proposed in addition that at least two binding sites for K+ exist at the cytoplasmic surface of isolated (Na,K) ATPase in state E1 but a large difference in affinities precludes detection in fluorescence titrations of more than one site. A variety of ligands in addition to K+ produce fluorescence-quenched or E2 forms of the labeled (Na,K) ATPase. These include Mg2+ plus inorganic phosphate, without or with K+ ions (E2P or E2P · K) or with ouabain (E2-ouabain or E2P · ouabain). Na+ ions antagonize these effects. The collected data support the notion that there may be many subspecies of the E1 and E2 forms (either phosphorylated or nonphosphorylated) with different numbers of Na+ and/or K+ ions bound or occluded, each subspecies having a characteristic ability to catalyze reactions and/or transport cations. The relationship between the conformational changes in fluorescein-labeled enzyme and the subunit structure of the (Na,K) ATPase is discussed with particular reference to half of the site models for ATP hydrolysis. 相似文献
5.
The present report demonstrates that NK effector cells present in the T-cell population and the null cell population of human peripheral blood mononuclear cells (PBMC) possess distinct phenotypes as defined by monoclonal antibodies. All the E? null cell-type effectors are OKM1+ OKT3?. Conversely the E+ T-cell-type effectors are OKM1? OKT3?. These results were obtained by fractionation of PBMC by means of anti-F(ab′)2 immunoabsorbent combined with E-rosette sedimentation. Each cell subset obtained was then treated with monoclonal antibody OKT3 or OKM1, and C. The NK activity of each cell population was quantitatively titrated against 51Cr-labeled JM target cells. 相似文献
6.
Expression of Ly-6A/E alloantigens in thymocyte and T-lymphocyte subsets: variability related to the Ly-6
a and Ly-6
b haplotypes 总被引:2,自引:0,他引:2
Elaine K. Codias Carolyn Cray Ruben D. Baler Robert B. Levy Thomas R. Malek 《Immunogenetics》1989,29(2):98-107
We have studied the cellular basis for differential expression of the Ly-6A/E alloantigen on T cells obtained from mice of the Ly-6
a (10–20% Ly-6A/E
+) and Ly-6
b (50–60% Ly-6A/E
+) haplotypes. During T-cell ontogeny only a small fraction (< 12 %) of thymocytes expressed Ly-6A/E. By 4 weeks of age adult levels of Ly-6A/E bearing lymphocytes were seen in peripheral lymphoid tissue. Immunohistochemical studies of the thymus revealed that Ly-6A/E+ cells were located predominantly in the medulla with small clusters of Ly-6A/E+ cells throughout the cortex. Consistent with this result, phenotypic studies showed that in the adult thymus the majority of Ly-6A/E expression was on mature CD4+ CD8– and CD4– CD8+ cortisone-resistant and precursor CD4– CD8– thymocytes. However, a much higher percentage of CD4+ CD8– and CD4– CD8– thymocytes as well as CD4+ CD8– peripheral T cells expressed Ly-6A/E from Ly-6
b mice. Furthermore, although gamma interferon induced increased Ly-6A/E expression in certain thymocyte and T-cell subsets, this induction functioned preferentially for cells obtained from Ly-6
b mice. Studies using F1 hybrid mice (Ly-6
a × Ly-6
b) indicated that the basal level of Ly-6A/E expression on these subsets appeared to be under codominant genetic control, whereas gamma interferon-induced regulation of Ly-6A/E expression appeared to be under dominant genetic control. Collectively, these results suggest that the expression of Ly-6A/E on a particular T-cell subset is established in the thymus and is a stable characteristic of each haplotype. In addition, the low levels of Ly-6A/E expression for the Ly-6
a haplotype appear to be partially due to the inability of the majority of resting CD4+ T cells to express Ly-6A/E and to the relatively poor induction of this protein by gamma interferon. 相似文献
7.
Cross-flow filtration (CFF) has been investigated as a method of separating filamentously growing fungal cells and purifying the polysaccharide produced. The effects of transmembrane pressure, module geometry (e.g. channel height or tube diameter), tangential feed velocity and cell as well as polysaccharide concentration are discussed. Apart from these experiments, influences by the recirculation pump used are shown.List of Symbols
b
f
fouling index
-
b
factor refering to the behaviour of the sublayer
-
C kg · m–3
concentration
-
C
g
kg · m–3
solute concentration at the membrane
-
C
b
kg · m–3
solute concentration in the bulk phase
-
D s-1
shear rate
-
k m · s–1
mass-transfer coefficient
-
K mPa · sn
consistency index
-
n
flow behaviour index
-
P
w m3 · s–1 · m–2
rate of permeation
-
P
w1 m3 · s–1 · m–2
rate of permeation at 1 minute
-
P
w
m3 · s–1 · m–2
rate of permeation at the beginning
-
p
Pa pressure
-
Q m2
largest cross-section of a particle
-
q m2
smallest cross-section of a particle
-
Re
Reynolds number
-
R
f
–1
fouling resistance
-
R
m
m–1
membrane resistance
-
t s
time
-
w m · s–1
tangential feed velocity
Greek Symbols
friction factor
-
pTM Pa
transmembrane pressure
-
mPa · s
shear viscosity
-
sp
specific viscosity (rel. increase of viscosity sp=rel-1)
- [] m3· kg–1
intrinsic viscosity
-
w
m2 · s–1
kinematic viscosity
-
kg · m–3
density
Indices
b
bulk
-
cell
cells
-
f
fouling
-
g
gelling
-
PS
polysaccharide
-
rel
relative
-
sp
specific
-
w
water 相似文献
8.
Arabella Mazzocchi Andrea Anichini Chiara Castelli Marialuisa Sensi Francesca Poli Carlo Russo Giorgio Parmiani 《Cancer immunology, immunotherapy : CII》1990,32(1):13-21
Summary The specificity analysis of a CD3+, WT31+, CD8+ cytotoxic T lymphocyte (CTL) clone (CTL 49), isolated from peripheral blood lymphocytes of a melanoma patient (no. 665) after mixed lymphocyte culture with an HLA-A2+ allogeneic lymphoblastoid cell line (VSKB-LCL), revealed that CTL 49 could lyse, in addition to HLA-A2+ lines, autologous HLA-A2– melanoma (Me665/2) and K562 targets. Killing of VSKB-LCL, but not of Me665/2, could be inhibited by anti-CD3 and by anti-HLA-A2 antibodies or by modulation of the CD3 complex. Cold-target competition studies showed that K562, but not VSKB-LCL, could compete with Me665/2 for lysis by CTL 49. However, unlike K562, Me665/2 could be lysed by CTL 49 in a Ca2+-independent fashion in 4 h and 18 h assays. CTL 49 expressed mRNA specific for tumor necrosis factor (TNF) and, to a lesser extent, for lymphotoxin (TNF). Exposure of the clone to anti-CD3 antibodies induced the expression of interferon(IFN)--specific and the up-regulation of TNF- and TNF-specific mRNA. Antibodies to TNF, TNF and IFN reduced the lysis of Me665/2, but not of K562, by CTL 49 in 18-h cytotoxic assays. Antibodies to TNF and to IFN almost completely inhibited the lysis seen on Me665/2 (but not on K562), in 96-h assays, by supernatants isolated from VSKB-LCL- or anti-CD3-stimulated CTL 49 cells. Taken together, these data indicate that major-histocompatibility-complex-independent lysis of autologous tumor cells and of natural killer reference targets by the same alloreactive T cell clone are activities related at the level of target recognition but distinct at the level of the lytic hit. Thus, efficient lysis of autologous tumor cells results from a complex mechanism based upon direct effector-target interaction as well as on cytokine-mediated cytolytic effects. 相似文献
9.
Protein turnover was examined, using tritiated water, in various 2-cm regions of 7-11-d-old, first leaves of barley (Hordeum vulgare). Differences were found between the regions in their protein turnover and their responses to stress. The rate constant for degradation for total protein was the same throughout the leaf and the average half-life (t1/2) of protein=approx. 220 h. Only in the older regions did a 24-h pulse of3H2O preferentially label protein with a t1/2 (90 h) considerably shorter than the t1/2 for total protein. Soluble protein was degraded faster than insoluble protein and contained an appreciable short-lived protein component observable by short-pulse labelling. The rate of protein synthesis was greatest in the cells of the youngest region and declined as each region aged. The mean rate of protein synthesis over the 4-d period was 4 and 7 nmol h-1 of amino-N with respect to the regions 1–3 and 7–9 cm from the leaf tip. Seedlings, stressed by adding polyethylene glycol (2.0 MPa) to the roots, showed a marked loss of protein from the older leaf regions with only small losses in the younger regions. Amino acids accumulated in the younger region continuously whereas in the older region little accumulation occurred until day 3 of stress when proline levels increased. Protein synthesis was decreased by between 30% and 50% in all leaf regions. In the region 1–3 cm from the leaf tip, the rate of protein degradation of total protein was enhanced and equalled the rate of degradation of 24-h-pulse-labelled protein which was not itself significantly affected by stress (t1/2=approx. 90 h). In the region 3–5 cm, the degradation of both 4-d and 24-h-labelled protein was enhanced by stress to rates similar to those found in the region 1–3 cm. This was largely through increases in the degradation of the insoluble protein, but the degradation of soluble protein was also raised. Protein degradation in the region 7–9 cm was not affected by stress.Abbreviations t1/2
average half-life
- PEG
polyethylene glycol 相似文献
10.
Bent Vestergaard-Bogind Per Stampe Palle Christophersen 《The Journal of membrane biology》1987,95(2):121-130
Summary The conductance of the Ca2+-activated K+ channel (g
K(Ca)) of the human red cell membrane was studied as a function of membrane potential (V
m
) and extracellular K+ concentration ([K+]ex). ATP-depleted cells, with fixed values of cellular K+ (145mm) and pH (7.1), and preloaded with 27 m ionized Ca were transferred, with open K+ channels, to buffer-free salt solutions with given K+ concentrations. Outward-current conductances were calculated from initial net effluxes of K+, correspondingV
m
, monitored by CCCP-mediated electrochemical equilibration of protons between a buffer-free extracellular and the heavily buffered cellular phases, and Nernst equilibrium potentials of K ions (E
K) determined at the peak of hyperpolarization. Zero-current conductances were calculated from unidirectional effluxes of42K at (V
m
–E
K)0, using a single-file flux ratio exponent of 2.7. Within a [K+]ex range of 5.5 to 60mm and at (V
m
–E
K) 20 mV a basic conductance, which was independent of [K+]ex, was found. It had a small voltage dependence, varying linearly from 45 to 70 S/cm2 between 0 and –100 mV. As (V
m
–E
K) decreased from 20 towards zero mVg
K(Ca) increased hyperbolically from the basic value towards a zero-current value of 165 S/cm2. The zero-current conductance was not significantly dependent on [K+]ex (30 to 156mm) corresponding toV
m
(–50 mV to 0). A further increase ing
K(Ca) symmetrically aroundE
K is suggested as (V
m
–E
K) becomes positive. Increasing the extracellular K+ concentration from zero and up to 3mm resulted in an increase ing
K(Ca) from 50 to 70 S/cm2. Since the driving force (V
m
–E
K) was larger than 20 mV within this range of [K+]ex this was probably a specific K+ activation ofg
K(Ca). In conclusion: The Ca2+-activated K+ channel of the human red cell membrane is an inward rectifier showing the characteristic voltage dependence of this type of channel. 相似文献
11.
A M Silverstein 《Cellular immunology》1982,67(2):396-409
The induction of differentiation in human malignant T-lymphoblastic cell lines MOLT-3 and Jurkat by the tumor promoter, 12-O-tetradecanoylphorbol 13-acetate (TPA) was examined using the monoclonal antibodies OKT3, OKT4, OKT6, and OKT8 which are known to react with human T-cell differentiation antigens. It was found that in the presence of nanomolar concentrations of TPA the proportion of OKT3+ (mature T-cell marker) cells increased while the proportion of OKT4+, OKT6+, and OKT8+ (relatively immature T-cell markers) cells decreased. These changes in the distribution of the OKT antigens in MOLT-3 cells were found to be more prominent with MOLT-3 cells than when the Jurkat cells were used. In studies using a double labeling approach it was found that although the OKT3+ and E-rosette-positive (E+) cells appeared to belong to the same subpopulations of MOLT-3 cells, the OKT3 antigen was probably not related to the receptor for sheep erythrocytes because adsorption of the OKT3 antibody did not block E-rosette formation. Studies using the DNA synthesis inhibitor, arabinosylcytidine (ara-C) also indicate that DNA synthesis was not required for the induction of more mature T-cell antigens in the malignant T-cell lines by TPA. These studies, taken together with our earlier reports, support the conclusion that namomolar concentrations of TPA can induce differentiation in these malignant T-cell lines. Furthermore we have shown that the T-cell hybridoma antibodies are useful markers to detect differentiation changes in human T cells. 相似文献
12.
Summary In the NaK-ATPase proteoliposomes (PLs), the NaK-pump activity, Na+ uptake, and ATP hydrolysis were apparently enhanced by carbonyl cyanidem-chlorophenylhydrazone (CCCP) and other ionophores without ion gradients. These ionophore effects were not cation specific. Without ionophores, the PL's ATPase activity fell to its steady-state value within 3 sec at 15°C. This decrease in activity disappeared in the presence of CCCP. Since CCCP is believed to enhance proton mobility across the lipid bilayer and dissipate membrane potential (V
m
), we postulated that aV
m
build-up partially inhibits the PLs by changing the conformation of the NaK-pump, and that CCCP eliminated this partial inhibition. Since this activation required extracellular K+ and high ATP concentration in the PLs, CCCP must affect the conversion between the phosphorylated forms of NaK-ATPase (EP); this step has been suggested by Goldschlegger et al. (1987) to be the voltage-sensitive step (J. Physiol. (London)
387:331–355). Although cytoplasmic K+ accelerated the change of ADP-and K+-sensitive EP (E*P) to K+-sensitive ADP-insensitive EP (E2P), CCCP did not compete with cytoplasmic K+ when cytoplasmic Na+ was saturated. When the PLs were phosphorylated with 20 m ATP and 20 m palmitoyl CoA instead of with high concentration of ATP, CCCP increased the E*P content and decreased the ADP-sensitive K+-insensitive EP (E1P). The results described above suggest that CCCP affects the E1P to E*P change in the E1PE*PE2P conversion and that this reaction step is inhibited byV
m
. 相似文献
13.
The present study was conducted to characterize the possible interaction of Al3+ and Fe2+ with synthetic melanin in the potentiation of lipid peroxidation in liposomes and rat caudate-putamen homogenates. Al3+ stimulated melanin-initiated lipid peroxidation as measured by the production of 2-thiobarbituric acid-reactive substances (TBARS) and conjugated dienes. The effect of Al3+ was dependent on melanin (10–100 g/ml) and Al3+ (2.5–250 M) concentrations and no synergism between Fe2+ and Al3+ was observed. The prooxidant effect of Al3+ was partially inhibited by superoxide dismutase indicating the involvement of O
2
-
. Ga3+ and Be2+ which can increase NADH oxidation in the presence of O
2
-
, also were shown to stimulate melanin-initiated TBARS production. Based on the effect of Al3+ and other non redox metals, we suggest that Al3+ does not act through either the induction of melanin free radicals, or the induction of changes in membrane physical properties. Results show that Al3+ enhances melanin-initiated lipid peroxidation in part through an interaction with O
2
-
generated from the autoxidation of melanin. We speculate that Al3+ contributes to neuromelanin-mediated oxidative damage in dopaminergic neurons and subsequent neuronal degeneration and death in Parkinson's disease. 相似文献
14.
Liana Preciado-Patt Liora Cahalon Rami Hershkovitz Ofer Lider Mordechai Pras Mati Fridkin 《Letters in Peptide Science》1998,5(5-6):349-355
Serum amyloid A (SAA), an acute-phase reactant, exists naturally as a minor protein in the sera of healthy individuals. However, its levels in sera are increased markedly during various transient and chronic inflammatory diseases, often concomitantly with accumulation at inflicted sites. SAA is synthesized mainly in the liver following the synergistic action of cytokines, mainly tumor necrosis factor- (TNF-) and interleukin-1 and -6 (IL-1 and IL-6). It was already shown by us that upon interaction with SAA or amyloid A (AA), the extracellular matrix (ECM) and laminin induced the adhesion of resting human CD4+ T-cells in an apparently 1-integrin-mediated manner. Herein we have shown that the SAA–ECM complex modulates the regulation of cytokine synthesis by human T-lymphocytes. The SAA–ECM complex dramatically enhanced the release of TNF- by human T-cells in a dose-dependent manner, reaching its maximal effect in the presence of 100 M recombinant SAA. The SAA domain, responsible for the enhanced release of TNF- by human T-lymphocytes, is apparently the amyloid A protein (AA, i.e. SAA2–82). Specifically, TNF- enhanced secretion is mediated through intimate interactions of SAA/AA, with laminin. Thus, the ECM serving as a temporary anchorage site for SAA and AA seems to be involved in regulating TNF- secretion and the recruitment and accumulation of immunocytes in extravascular, inflammatory compartments. 相似文献
15.
Objectives
Several recent lines of evidence indicate that endothelial microparticles are a new biomarker that can be used to monitor endothelial dysfunction in coronary artery disease (CAD). However, data concerning the detection of small microparticles (diameter <0.5 µm) are lacking. The aim of this study was to detect small-size endothelial microparticles (SEMPs) in CAD patients to monitor endothelial dysfunction.Methods
In total, 19 CAD patients and 14 healthy subjects were recruited. The absolute numbers and percentages of CD31+/CD42b− SEMPs and CD62E+ SEMPs were determined by flow cytometry. Clinical parameters were also recorded.Results
The mean percentage of CD62E+ SEMPs was higher in the CAD patient group than in the healthy subject group. The area under the receiver operating characteristic curve of the percentage of CD62E+ SEMPs was 0.795, and the cut-off value was 1.35. There was no correlation between the percentage of CD62E+ SEMPs and various clinical parameters.Conclusion
The percentage of CD62E+ SEMPs is a potential biomarker for monitoring endothelial function in CAD. 相似文献16.
An anion channel that only allows outward current flow (anion influx) has been identified in protoplasts derived from wheat (Triticum aestivum L., Triticum turgidum L.) roots. The anion outward rectifier (anion OR) measured by patch-clamp of whole cells activated very quickly, usually reaching a steady-state level in less than 100 ms and was easily distinguished from the cation outward rectifier (cation OR) which activated more slowly during membrane depolarisation. The anion OR is permeable to NO
3
–
and Cl–, moderately permeable to I–, and relatively impermeable to H2PO4/– and ClO4/–. An anomalous mole-fraction effect between ClO4/
– and Cl– was observed on the outward current, indicating that the channel is a multi-ion pore. The anion OR is gated by both voltage and external anion concentration such that it activates near to the equilibrium potential for the permeant anion. It activated at more negative membrane potentials when NO
3
–
was substituted for Cl– in the external medium, indicating that the channel may function to allow NO
3
–
influx under luxuriant external NO
3
–
concentrations. For most experiments, K+ and Cl– were the main cation and anion in solution, and under these conditions it appeared likely that the anion OR functioned in membrane-potential regulation by facilitating a Cl– influx at membrane potentials more positive than the chloride reversal potential (ECl). If ECl was more negative than the K+ reversal potential (EK) then the anion OR dominated but both the anion and cation ORs occurred together when the membrane potential difference (Vm) was positive of both ECl and EK. The cation OR was inhibited by increasing external Cl– concentrations, but the anion OR was not affected by external K+ or Na+ concentration. The anion-transport inhibitors, zinc and phenylglyoxal were ineffective in blocking the anion OR. 4,4-Di-isothiocyanostilbene-2, 2-disulfonic acid (DIDS) irreversibly blocked about 34% of the current when applied extracellularly at a concentration of 25 M, and about 69% at a concentration of 200 M. However, DIDS (200 M) also occasionally acted as an irreversible blocker of the cation OR. Perchlorate blocked irreversibly 75% of the current at an external concentration of 10 mM and did not block the cation OR. Whole-cell currents also indicated that the anion OR was insensitive to external pH (pH=5–7) and calcium concentration ([Ca2+]=0.1–10 mM). Increasing intracellular calcium concentration significantly increased the occurrence of the fast outward current in whole cells (P < 0.005, X2 test). With approximately 10 nM calcium inside the cell the anion outward current was observed in 64% (n = 45) of cells and with 50 nM calcium inside the cell the anion current was observed in 88% (n = 69) of cells. Single-anion OR channels observed in outside-out patches had a conductance in 300 mM KCl (external) of about 4 pS. When voltage pulses were applied to outside-out patches the average currents were similar to those observed in whole cells. The significance of the anion OR as a likely route for Cl uptake in high salinities is discussed.Abbreviations Bath
solution bathing the extracellular face of the membrane
- DIDS
(4,4-diisothiocyanostilbene-2,2-disulfonic acid)
- Ex
reversal potential for ion x
- OR
outward rectifier
-
Pip
solution inside the pipette
- TEACl
(tetraethyl-ammonium chloride)
- Vm
membrane potential difference
We thank the Australian Research Council for financial support, G.P. Findlay and A. Garrill for helpful discussions, and K. Morris and D. Mackenzie for expert technical assistance. M.S. was supported by an Australian Postgraduate Research Award. 相似文献
17.
Ian Henry Lambert Else Kay Hoffmann Finn Jørgensen 《The Journal of membrane biology》1989,111(2):113-131
Summary The fluorescence intensity of the dye 1,1-dipropyloxadicarbocyanine (DiOC3-(5)) has been measured in suspensions of Ehrlich ascites tumor cells in an attempt to monitor their membrane potential (V
m
) under different ionic conditions, after treatment with cation ionophores and after hypotonic cell swelling. Calibration is performed with gramicidin in Na+-free K+/choline+ media, i.e., standard medium in which NaCl is replaced by KCl and cholineCl and where the sum of potassium and choline is kept constant at 155mm. Calibration by the valinomycin null point procedure described by Lariset al. (Laris, P.C., Pershadsingh, A., Johnstone, R.M., 1976,Biochim. Biophys. Acta
436:475–488) is shown to be valid only in the presence of the Cl–-channel blocker indacrinone (MK196). Distribution of the lipophilic anion SCN– as an indirect estimation of the membrane potential is found not to be applicable for the fast changes inV
m
reported in this paper. Incubation with DiOC3-(5) for 5 min is demenstrated to reduce the Cl– permeability by 26±5% and the NO
3
–
permeability by 15±2%, while no significant effect of the probe could be demonstrated on the K+ permeability. Values forV
m
, corrected for the inhibitory effect of the dye on the anion conductance, are estimated at –61±1 mV in isotonic standard NaCl medium, –78±3 mV in isotonic Na+-free choline medium and –46±1 mV in isotonic NaNO3 medium. The cell membrane is depolarized by addition of the K+ channel inhibitor quinine and it is hyperpolarized when the cells are suspended in Na+-free choline medium, indicating thatV
m
is generated partly by potassium and partly by sodium diffusion. Ehrlich cells have previously been shown to be more permeable to nitrate than to chloride. Substituting NO
3
–
for all cellular and extracellular Cl– leads to a depolarization of the membrane, demonstrating thatV
m
is also generated by the anions and that anions are above equilibrium. Taking the previously demonstrated single-file behavior of the K+ channels into consideration, the membrane conductances in Ehrlich cells are estimated at 10.4 S/cm2 for K+, 3.0 S/cm2 for Na+, 0.6 S/cm2 for Cl– and 8.7 S/cm2 for NO
3
–
. Addition of the Ca2+-ionophore A23187 results in net loss of KCl and a hyperpolarization of the membrane, indicating that the K+ permeability exceeds the Cl– permeability also after the addition of A23187. The K+ and Cl– conductances in A23187-treated Ehrlich cells are estimated at 134 and 30 S/cm2, respectively. The membrane potential is depolarized in hypotonically swollen cells, confirming that the increase in the Cl– permeability following hypotonic exposure exceeds the concommitant increase in the K+ permeability. In control experiments where the membrane potentialV
m
=E
K
=E
Cl
=E
Na
, it is demonstrated that cell volume changes has no significant effect on the fluorescence signal, apparently because of a large intracellular buffering capacity. The increase in the Cl– conductances is 68-fold when cells are transferred to a medium with half the osmolarity of the standard medium, as estimated from the net Cl– efflux and the change inV
m
. The concommitant increase in the K+ conductance, as estimated from the net K+ efflux, is only twofold. 相似文献
18.
Summary Mercury concentration in intraoral air and urine of seven females with dental amalgam was measured before and after intake of one hard-boiled egg. A considerable decrease in mercury concentration in intraoral air was found. Twenty women with about equal dental amalgam status, with or without subjective symptoms related to dental amalgam, were also studied. Mercury concentrations in intraoral air and urine were measured. For all the 27 women the basal intraoral air concentration of mercury ranged over 0.6–10.4 g/m3 (median value 4.3 g/m3). This corresponds to a release of 0.02–0.38 ng/s (median value 0.16 ng/s). In urine, the mercury concentration varied from < 0.8–6.9 g/g creatinine (median value 1.9 g/g creatinine). Data from both parameters were significantly correlated to the total number of teeth areas with dental amalgam. Protein values in urine indicated no renal damage. Maximum concentrations of mercury vapour in intraoral air for the 27 women who had chewed chewing gum for 5 min varied between 2–60 g Hg/m3 (median value 19 g Hg/m3). This corresponds to 0.07–2.20 ng Hg/s and a median value of 0.70 ng Hg/s. 相似文献
19.
In an effort to generate an A.CA mouse expressing Ed, the Ead gene has been introduced into A. CA mice which lack the major histocompatibility complex (MHC) class II E molecule. Flow cytometric analysis shows cell surface expression of the E chain on lymphocytes and macrophages in the transgenic mice. Analysis of T-cell receptor (Tcr) genes deleted in some E-expressing mouse strains demonstrates that T cells expressingTcrb-VS are partially deleted in these transgenic mice while those expressingTcrb-V8 andTcrb-11 are not. In addition, the expressed Ed chain can promote Mycoplasma arthriditis mitogen (MAM)-induced T-cell proliferation. The expression of the Ea chain, presumably as an Aß
fEd heterodimer, can alter the peripheral T-cell repertoire and T-cell reactivity to a microbial superantigen. 相似文献
20.
Young Ah Lee Hang-Rae Kim Jeong Seon Lee Hae Woon Jung Hwa Young Kim Gyung Min Lee Jieun Lee Ji Hyun Sim Sae Jin Oh Doo Hyun Chung Choong Ho Shin Sei Won Yang 《PloS one》2015,10(12)