Selective Removal of Endotoxin from a DNA Solution by Cross-linked Cyclodextrin Beads

Sakata, Masayo; Yoshimura, Kazuki; Sakamoto, Itsumi; Todokoro, Masami; Kunitake, Masashi

doi:10.2116/analsci.27.213

Cited by 14 publications

(10 citation statements)

References 12 publications

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“…This conclusion was challenged by a recent report showing that βCD may act as a scavenger for endotoxin (Sakata et al . ). To examine the interplay between βCD, LPS and cells in more detail, we conducted two different sets of experiments.…”

Section: Resultsmentioning

confidence: 97%

“…D , schematic diagram of the interaction of the acyl chains of lipid A with the hydrophobic cavity of βCD (according to Sakata et al . ). * P < 0.05 versus βCD washout.…”

Section: Resultsmentioning

confidence: 97%

“…However, recently it has been reported that cyclodextrins remove endotoxin from DNA solutions (Sakata et al . ). It was proposed that the fatty acids of lipid A may be partially adsorbed into the hydrophobic cavity of βCD.…”

Section: Discussionmentioning

confidence: 97%

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Inhibition of cardiac pacemaker channel hHCN2 depends on intercalation of lipopolysaccharide into channel‐containing membrane microdomains

Klöckner

Rueckschloss

Großmann

et al. 2014

The Journal of Physiology

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Key pointsr The regulation of cardiac function is seriously impaired in severe inflammatory diseases. One characteristic of this dysfunction is a strong reduction in heart rate variability (HRV) so that the cardiac cycle is more regular. This phenomenon is strongly correlated with an unfavourable prognosis in patients with systemic inflammation.r Although the depression in HRV can be partially explained by the interplay between cardiac pacemaker channels and lipopolysaccharide (LPS) liberated from the outer walls of Gram-negative bacteria, the underlying mechanism is still elusive.r Using HEK293 cells stably expressing a cardiac pacemaker channel, we demonstrate that only intact LPS molecules can intercalate into target cell membranes and then directly interact with extracellular parts of pacemaker channels. Intracellular signalling cascades do not contribute to LPS-dependent channel modulation.r The present results help to elucidate how LPS interacts with pacemaker channels to attenuate the regularity of the cardiac cycle.Abstract Depressed heart rate variability in severe inflammatory diseases can be partially explained by the lipopolysaccharide (LPS)-dependent modulation of cardiac pacemaker channels. Recently, we showed that LPS inhibits pacemaker current in sinoatrial node cells and in HEK293 cells expressing cloned pacemaker channels, respectively. The present study was designed to verify whether this inhibition involves LPS-dependent intracellular signalling and to identify structures of LPS responsible for pacemaker current modulation. We examined the effect of LPS on the activity of human hyperpolarization-activated cyclic nucleotide-gated channel 2 (hHCN2) stably expressed in HEK293 cells. In whole-cell recordings, bath application of LPS decreased pacemaker current (I hHCN2 ) amplitude. The same protocol had no effect on channel activity in cell-attached patch recordings, in which channels are protected from the LPS-containing bath solution. This demonstrates that LPS must interact directly with or close to the channel protein. After cleavage of LPS into lipid A and the polysaccharide chain, neither of them alone impaired I hHCN2 , which suggests that modulation of channel activity critically depends on the integrity of the entire LPS molecule. We furthermore showed that β-cyclodextrin interfered with LPS-dependent channel modulation predominantly via scavenging of lipid A, thereby abrogating the capability of LPS to intercalate into target cell membranes. We conclude that LPS impairs I hHCN2 by a local mechanism that is restricted to the vicinity of the channels. Furthermore, intercalation of lipid A into target cell membranes is a prerequisite for the inhibition that is suggested to depend on the direct interaction of the LPS polysaccharide chain with cardiac pacemaker channels.

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Section: Resultsmentioning

confidence: 97%

“…D , schematic diagram of the interaction of the acyl chains of lipid A with the hydrophobic cavity of βCD (according to Sakata et al . ). * P < 0.05 versus βCD washout.…”

Section: Resultsmentioning

confidence: 97%

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Inhibition of cardiac pacemaker channel hHCN2 depends on intercalation of lipopolysaccharide into channel‐containing membrane microdomains

Klöckner

Rueckschloss

Großmann

et al. 2014

The Journal of Physiology

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show abstract

“…Furthermore, Sakata et al have also indicated that LPS can selectively be removed from solutions containing DNA. Copolymer particles composed of γ-cyclodextrinchloromethyloxirane (γ-CyD/CMO, Figure 18) were used to selectively remove LPS from a DNA solution [95]. It was determined that using a 1,6-hexamethylenediisocyanate (HMDI) crosslinker, rather than the CMO crosslinker, allowed for higher incorporation of CyD during polymer preparation [96].…”

Section: Lipopolysaccharidesmentioning

confidence: 99%

Polymer antidotes for toxin sequestration

Weisman

Chou

O’Brien

et al. 2015

Advanced Drug Delivery Reviews

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“…It was also reported that this method provided ease of subsequent plasmid DNA purification. Due to to proteinendotoxin interactions, endotoxin removal from protein solutions requires techniques that result in strong interactions with endotoxin, such as affinity chromatography, which has proven to be one of the most effective methods (Sakata et al, 2011;Anspach 2001;Li et al, 2011;Wei et al, 2007); however, this method is not highly reproducible and may be followed by a significant loss of the product being purified.…”

Section: Cortez and Co-workers (2004a; 2004b) Extracted Xylitol Dehydmentioning

confidence: 99%

Utilização de sistemas poliméricos de duas fases aquosas (SPDFA) compostos por polietileno glicol/ácido poliacrílico (PEG/APA) para extração de ácido clavulânico

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Selective Removal of Endotoxin from a DNA Solution by Cross-linked Cyclodextrin Beads

Cited by 14 publications

References 12 publications

Inhibition of cardiac pacemaker channel hHCN2 depends on intercalation of lipopolysaccharide into channel‐containing membrane microdomains

Inhibition of cardiac pacemaker channel hHCN2 depends on intercalation of lipopolysaccharide into channel‐containing membrane microdomains

Polymer antidotes for toxin sequestration

Utilização de sistemas poliméricos de duas fases aquosas (SPDFA) compostos por polietileno glicol/ácido poliacrílico (PEG/APA) para extração de ácido clavulânico

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