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11.
Laura S. Rhoads Anne M. Danks IM John Anne Warner Robert L. Isaacson John Baust Robert G. Van Buskirk 《In vitro cellular & developmental biology. Animal》1993,29(3):208-214
Summary The possible role of extracellular calcium ([Ca+2]e) in cryopreservation-induced cytotoxicity was tested using Madin-Darby canine kidney (MDCK) cells and a fluorescent multiple
endpoint assay. MDCK cells maintained in 2 mM [Ca+2]e and treated with the calcium ionophore, ionomycin, increased their intracellular calcium ([Ca+2]i) as revealed by the calcium indicator dye, Fluo3 and the bottom-reading spectrofluorometer, CytoFluor 2300. The addition
of 10 mM [ethylene bis (oxyethylenenitrilo)]-tetraacetic acid (EGTA) to the extracellular medium before treatment with ionomycin blocked
this ionomycin-dependent increase in [Ca+2]i. A number of site and activity-specific fluorescent probes were surveyed to determine which indicator dye might best reveal
the ionomycin-induced cytotoxic events during this increase in [Ca+2]i. Although most dyes changed their emission profiles in response to calcium, neutral red was found to best reflect the loss
of [Ca+2]i homeostasis. The NR50 for a 15-min exposure to ionomycin in the presence of 2 mM [Ca+2]e was approximately 2μM ionomycin, but ionomycin had little apparent effect on neutral red retention when 10 mM EGTA was added to the extracellular medium. Thus it was clear that an increase in [Ca+2]i could be cytotoxic to MDCK cells and that neutral red could monitor this cytotoxic episode. To test if [Ca+2]e was similarly cytotoxic during cryopreservation, MDCK cells were subjected to cryopreservation in the presence of dimethylsulfoxide
(DMSO). In contrast to previous studies, plasma membrane integrity, not lysosomal function, seemed to best correlate with
cell survival subsequent to cryopreservation. In addition, decreasing [Ca+2]e had no discernable effect on the retention of plasma membrane indicator dyes, neutral red, or cell survival. It is concluded
that a) plasma membrane indicator dyes, not neutral red, might be better indicators of cytotoxicity occurring during cryopreservation;
b) DMSO might be toxic to lysosomes during cryopreservation of cultured cells; and c) although [Ca+2]e can contribute to cytotoxicity, the presence of [Ca+2]e might not influence cryopreservation-induced cytotoxicity. 相似文献
12.
We have identified a novel N -acetylgalactosaminyltransferase activity in
lactating bovine mammary gland membranes. Acceptor specificity studies and
analysis of products obtained in vitro by 400 MHz1H-NMR spectroscopy
revealed that the enzyme catalyses the transfer of N - acetylgalactosamine
(GalNAc) from UDP-GalNAc to acceptor substrates carrying a terminal,
beta-linked N -acetylglucosamine (GlcNAc) residue and establishes a
beta1-->4-linkage forming a GalNAcbeta1-->4GlcNAc ( N, N
'-diacetyllactosediamine, lacdiNAc) unit. Therefore, the enzyme can be
identified as a UDP-GalNAc:GlcNAcbeta-R beta1-->4-N-
acetylgalactosaminyltransferase (beta4-GalNAcT). This enzyme resembles
invertebrate beta4-GalNAcT as well as mammalian beta4-
galactosyltransferase (beta4-GalT) in acceptor specificity. It can,
however, be clearly distinguished from the pituitary hormone-specific
beta4-GalNAcT by its incapability of acting with an elevated activity on a
glycoprotein substrate carrying a hormone-specific peptide motif.
Furthermore, the GalNAcT activity appeared not to be due to a promiscuous
action of a beta4-GalT as could be demonstrated by comparing the
beta4-GalNAcT and beta4-GalT activities of the mammary gland, bovine
colostrum, and purified beta4-GalT, by competition studies with UDP-GalNAc
and UDP-Gal, and by use of an anti-beta4-GalT polyclonal inhibiting
antibody. Interestingly, under conditions where mammalian beta4-GalT forms
with alpha-lactalbumin (alpha-LA) the lactose synthase complex, the mammary
gland beta4-GalNAcT was similarly induced by alpha-LA to act on Glc with an
increased efficiency yielding the lactose analog GalNAcbeta1-->4Glc.
This enzyme thus forms the second example of a mammalian
glycosyltransferase the specificity of which can be modified by this milk
protein. It is proposed that the mammary gland beta4-GalNAcT functions in
the synthesis of lacdiNAc- based, complex-type glycans frequently occurring
on bovine milk glycoproteins. The action of this enzyme is to be considered
when aiming at the production of properly glycosylated protein
biopharmaceuticals in the milk of transgenic dairy animals.
相似文献
13.
Electron microscopy of ultrathin sections of leaves of symptomless Himalaya Giant blackberry and of the virus indicator species, Rubus macraei, showing severe leaf curl symptoms following graft inoculation with scions from this blackberry, detected highly flexuous virus‐like particles with an unusual ‘beaded’ structure. Such particles were restricted to a few vascular cells and were distinct from P‐protein common in some such cells. This virus, provisionally named Hawaiian rubus leaf curl virus (HRLCV), symptomlessly infected a wide range of Rubus species and cultivars. Badnavirus‐like bacilliform particles were observed in some cells of a single R. macraei plant showing leaf curl symptoms following graft inoculation with the causal agent of this disease symptom from Himalaya Giant blackberry after passage through red raspberry, but not in any other material. PCR with primer sets for the badnaviruses Rubus yellow net virus and Gooseberry veinbanding associated virus, showed that no Rubus sources studied contained these viruses. However, using a sequence‐specific primer set designed from the sequence of the product generated with a badnavirus degenerate primer set, a specific product was amplified from healthy plants of all of 16 raspberry cultivars and two Rubus species, but not from 16 blackberry cultivars (including cv. Himalaya Giant). All of these sources were free from viruses known to occur in Rubus. Sequence analysis of this product showed no homology with any known badnavirus, or with any other published sequences. It seems most likely therefore that a region of the raspberry genome has been amplified using the degenerate badnavirus primer set and that it is absent from the blackberry genome. 相似文献
14.
Sabine JM de Brouwer Floris W Kraaimaat Fred CGJ Sweep Marjonne CW Creemers Timothy RDJ Radstake Antoinette IM van Laarhoven Piet LCM van Riel Andrea WM Evers 《Arthritis research & therapy》2010,12(3):R89
Introduction
Stressful events are thought to contribute to the aetiology, maintenance and exacerbation of rheumatic diseases. Given the growing interest in acute stress responses and disease, this review investigates the impact of real-life experimental psychosocial, cognitive, exercise and sensory stressors on autonomic, neuroendocrine and immune function in patients with inflammatory rheumatic diseases. 相似文献15.
Complete nucleotide sequences of bovine alpha S2- and beta-casein cDNAs: comparisons with related sequences in other species 总被引:2,自引:0,他引:2
Stewart AF; Bonsing J; Beattie CW; Shah F; Willis IM; Mackinlay AG 《Molecular biology and evolution》1987,4(3):231-241
The nucleotide sequences corresponding to bovine alpha S2- and beta- casein
mRNAs have been determined by cDNA analysis. Both sequences appear to be
complete at their 5' ends. The nucleotide sequence of alpha S2-casein, when
compared with the corresponding cavine A sequence, helps to define the
boundaries of a large amino acid repeat (approximately 80 residues) whereas
comparisons with the nucleotide sequences of rat gamma- and mouse
epsilon-casein mRNAs also reveal extensive sequence similarities. An
alignment of these four sequences shows that the divergence of their
translated regions has been characterized by the duplication and deletion
of discrete segments of sequence that probably correspond to exons. A high
degree of nucleotide substitution is also found when the four sequences are
compared, except for well-conserved leader-peptide and phosphorylation-site
sequences and, to a lesser extent, the 5'-untranslated regions. Similar
comparison of the bovine and rat beta-caseins shows that their divergence
has involved a high rate of nucleotide substitution but that no major
insertions or deletions of sequence have occurred. The several splice sites
that have veen defined in the rat beta-casein gene are likely to have been
conserved in the bovine. The contrasting evolutionary histories of the
alpha- and beta-casein coding sequences correlate with the distinctive
functions of these proteins in the casein micelle system in milk.
相似文献
16.
ABEL TRUJILLO-OCAMPO HYUN-WOO CHO AMANDA C. HERRMANN WILFREDO RUIZ-VAZQUEZ ANDREW B. THORNTON HONG HE DAN LI MARIAM A. QAZILBASH QING MA STEVEN A. PORCELLI ELIZABETH J. SHPALL JEFFREY MOLLDREM JIN S. IM 《Cytotherapy》2018,20(8):1089-1101
Background aims
CD1d-restricted invariant natural killer (iNK) T cells are rare regulatory T cells that may contribute to the immune-regulation in allogeneic stem cell transplantation (ASCT). Here, we sought to develop an effective strategy to expand human iNK T cells for use in cell therapy to prevent graft-versus-host disease (GVHD) in ASCT.Methods
Human iNK T cells were first enriched from peripheral blood mononuclear cells (PBMCs) using magnetic-activated cell sorting separation, then co-cultured with dendritic cells in the presence of agonist glycolipids, alpha-galactosylceramide, for 2 weeks.Results
The single antigenic stimulation reliably expanded iNK T cells to an average of 2.8?×?107 per 5?×?108 PBMCs in an average purity of 98.8% in 2 weeks (N?=?24). The expanded iNK T cells contained a significantly higher level of CD4+ and central memory phenotype (CD45RA?CD62L+) compared with freshly isolated iNK T cells, and maintained their ability to produce both Th-1 (interferon [IFN]γ and tumor necrosis factor [TNF]α) and Th-2 type cytokines (interleukin [IL]-4, IL-5 and IL-13) upon antigenic stimulation or stimulation with Phorbol 12-myristate 13-acetate/ionomycin. Interestingly, expanded iNK T cells were highly autoreactive and produced a Th-2 polarized cytokine production profile after being co-cultured with dendritic cells alone without exogenous agonist glycolipid antigen. Lastly, expanded iNK T cells suppressed conventional T-cell proliferation and ameliorated xenograft GVHD (hazard ratio, 0.1266; P < 0.0001).Conclusion
We have demonstrated a feasible approach for obtaining ex vivo expanded, highly enriched human iNK T cells for use in adoptive cell therapy to prevent GVHD in ASCT. 相似文献17.
Robin IM Dunbar 《BMC biology》2007,5(1):21-3
The claim that differences in brain size across primate species has mainly been driven by the demands of sociality (the "social
brain" hypothesis) is now widely accepted. Some of the evidence to support this comes from the fact that species that live
in large social groups have larger brains, and in particular larger neocortices. Lindenfors and colleagues (BMC Biology 5:20) add significantly to our appreciation of this process by showing that there are striking differences between the two
sexes in the social mechanisms and brain units involved. Female sociality (which is more affiliative) is related most closely
to neocortex volume, but male sociality (which is more competitive and combative) is more closely related to subcortical units
(notably those associated with emotional responses). Thus different brain units have responded to different selection pressures. 相似文献
18.
Effect of self-association on the structural organization of partially folded proteins: inactivated actin 总被引:1,自引:0,他引:1 下载免费PDF全文
IM Kuznetsova AG Biktashev SY Khaitlina KS Vassilenko KK Turoverov VN Uversky 《Biophysical journal》1999,77(5):2788-2800
The propensity to associate or aggregate is one of the characteristic properties of many nonnative proteins. The aggregation of proteins is responsible for a number of human diseases and is a significant problem in biotechnology. Despite this, little is currently known about the effect of self-association on the structural properties and conformational stability of partially folded protein molecules. G-actin is shown to form equilibrium unfolding intermediate in the vicinity of 1.5 M guanidinium chloride (GdmCl). Refolding from the GdmCl unfolded state is terminated at the stage of formation of the same intermediate state. An analogous form, known as inactivated actin, can be obtained by heat treatment, or at moderate urea concentration, or by the release of Ca(2+). In all cases actin forms specific associates comprising partially folded protein molecules. The structural properties and conformational stability of inactivated actin were studied over a wide range of protein concentrations, and it was established that the process of self-association is rather specific. We have also shown that inactivated actin, being denatured, is characterized by a relatively rigid microenvironment of aromatic residues and exhibits a considerable limitation in the internal mobility of tryptophans. This means that specific self-association can play an important structure-forming role for the partially folded protein molecules. 相似文献
19.
RAC-3 is a NF-kappa B coactivator 总被引:8,自引:0,他引:8
20.
Rachel IM van Haaften Blanche Schroen Ben JA Janssen Arie van Erk Jacques JM Debets Hubert JM Smeets Jos FM Smits Arthur van den Wijngaard Yigal M Pinto Chris TA Evelo 《BMC bioinformatics》2006,7(1):200-15