Effects of hemoadsorption with a novel adsorbent on sepsis: in vivo and in vitro study.

BACKGROUND/AIMS: Hemoadsorption may improve outcomes for sepsis by removing circulating cytokines. We tested a new sorbent used for hemoadsorption. METHODS: CTR sorbent beads were filled into columns of three sizes: CTR0.5 (0.5 ml), CTR1 (1.0 ml) and CTR2 (2.0 ml) and tested using IL-6 capture in vitro. Next, rats were subjected to cecal ligation and puncture and randomly assigned to hemoadsorption with CTR0.5, CTR1, CTR2 or sham treatment. Plasma biomarkers were measured. RESULTS: In vitro, IL-6 removal was accelerated with increasing bead mass. In vivo, TNF, IL-6, IL-10, high mobility group box1, and cystatin C were significantly lower 24 h after CTR2 treatment. Seven-day survival rate was 50, 64, 63, and 73% for the sham, CTR0.5, CTR1, CTR2, respectively. CONCLUSION: CTR appeared to have a favorable effect on kidney function despite no immediate effects on cytokine removal. However, CTR2 beads did result in a late decrease of cytokines.

Hollow fiber membrane modification with functional zwitterionic macromolecules for improved thromboresistance in artificial lungs.

Respiratory assist devices seek optimized performance in terms of gas transfer efficiency and thromboresistance to minimize device size and reduce complications associated with inadequate blood biocompatibility. The exchange of gas with blood occurs at the surface of the hollow fiber membranes (HFMs) used in these devices. In this study, three zwitterionic macromolecules were attached to HFM surfaces to putatively improve thromboresistance: (1) carboxyl-functionalized zwitterionic phosphorylcholine (PC) and (2) sulfobetaine (SB) macromolecules (mPC or mSB-COOH) prepared by a simple thiol-ene radical polymerization and (3) a low-molecular weight sulfobetaine (SB)-co-methacrylic acid (MA) block copolymer (SBMAb-COOH) prepared by reversible addition-fragmentation chain transfer (RAFT) polymerization. Each macromolecule type was covalently immobilized on an aminated commercial HFM (Celg-A) by a condensation reaction, and HFM surface composition changes were analyzed by X-ray photoelectron spectroscopy. Thrombotic deposition on the HFMs was investigated after contact with ovine blood in vitro. The removal of CO2 by the HFMs was also evaluated using a model respiratory assistance device. The HFMs conjugated with zwitterionic macromolecules (Celg-mPC, Celg-mSB, and Celg-SBMAb) showed expected increases in phosphorus or sulfur surface content. Celg-mPC and Celg-SBMAb experienced rates of platelet deposition significantly lower than those of unmodified (Celg-A, >95% reduction) and heparin-coated (>88% reduction) control HFMs. Smaller reductions were seen with Celg-mSB. The CO2 removal rate for Celg-SBMAb HFMs remained comparable to that of Celg-A. In contrast, the rate of removal of CO2 for heparin-coated HFMs was significantly reduced. The results demonstrate a promising approach to modifying HFMs using zwitterionic macromolecules for artificial lung devices with improved thromboresistance without degradation of gas transfer.

Modeling competitive cytokine adsorption dynamics within hemoadsorption beads used to treat sepsis.

Extracorporeal blood purification is a promising therapeutic modality for sepsis, a potentially fatal, dysfunctional immunologic state caused by infection. Removal of inflammatory mediators such as cytokines from the blood may help attenuate hyper-inflammatory signaling during sepsis and improve patient outcomes. We are developing a hemoadsorption device to remove cytokines from the circulating blood using biocompatible, porous sorbent beads. In this work, we investigated whether competitive adsorption of serum solutes affects cytokine removal dynamics within the hemoadsorption beads. Confocal laser scanning microscopy (CLSM) was used to quantify intraparticle adsorption profiles of fluorescently labeled IL-6 in horse serum, and results were compared to predictions of a two component competitive adsorption model. Supraphysiologic IL-6 concentrations were necessary to obtain adequate CLSM signal, therefore unknown model parameters were fit to CLSM data at high IL-6 concentrations, and the fitted model was used to simulate cytokine adsorption behavior at physiologically relevant levels which were below the microscopy detection threshold. CLSM intraparticle IL-6 adsorption profiles agreed with predictions of the competitive adsorption model, indicating displacement of cytokine by high affinity serum solutes. However, competitive adsorption effects were predicted using the model to be negligible at physiologic cytokine concentrations associated with hemoadsorption therapy.

Characterizing accelerated capture of deoligomerized TNF within hemoadsorption beads used to treat sepsis.

Sepsis is a systemic inflammatory response to infection, characterized by overexpression of cytokines in the circulating blood. Removal of cytokines and other inflammatory mediators from the blood may help attenuate systemic inflammation during sepsis and improve patient outcomes. In this work, we examined the dynamics of TNF capture within porous, polymeric sorbent beads used in a cytokine adsorption device. We sought to quantify how perturbation of TNF oligomeric structure accelerates TNF removal within the device. TNF was incubated with 10% DMSO for 24 h, which promoted complete monomerization of trimeric TNF, and accelerated TNF capture within the sorbent device compared with native TNF; removal halftime = 13.3 ± 1.5 min versus 112.8 ± 13.3 min, respectively. Intramolecular crosslinking stabilized the trimeric TNF structure and prevented DMSO monomerization. Results demonstrate that TNF is an unstable oligomeric molecule that can be dissociated into its smaller monomeric constituents to facilitate faster capture by hemoadsorption beads. Strategies to promote localized TNF deoligomerization at the sorbent surface may significantly accelerate TNF capture rates from the circulating blood using hemoadsorption as a treatment for sepsis. This concept could be extended to improve removal of other oligomeric molecules using size exclusion filtration materials for a variety of disease states.

IL-6 adsorption dynamics in hemoadsorption beads studied using confocal laser scanning microscopy.

Sepsis is characterized by a systemic inflammatory response caused by infection, and can result in organ failure and death. Removal of inflammatory mediators such as cytokines from the circulating blood is a promising treatment for severe sepsis. We are developing an extracorporeal hemoadsorption device to remove cytokines from the blood using biocompatible, polymer sorbent beads. In this study, we used confocal laser scanning microscopy (CLSM) to directly examine adsorption dynamics of a cytokine (IL-6) within hemoadsorption beads. Fluorescently labeled IL-6 was incubated with sorbent particles, and CLSM was used to quantify spatial adsorption profiles of IL-6 within the sorbent matrix. IL-6 adsorption was limited to the outer 15 microm of the sorbent particle over a relevant clinical time period, and intraparticle adsorption dynamics was modeled using classical adsorption/diffusion mechanisms. A single model parameter, alpha = q(max) K/D, was estimated by fitting CLSM intensity profiles to our mathematical model, where q(max) and K are Langmuir adsorption isotherm parameters, and D is the effective diffusion coefficient of IL-6 within the sorbent matrix. Given the large diameter of our sorbent beads (450 microm), less than 20% of available sorbent surface area participates in cytokine adsorption. Development of smaller beads may accelerate cytokine adsorption by maximizing available surface area per bead mass.