Joseph W. Franses

Mechanisms of Tissue Uptake and Retention in Zotarolimus-Coated Balloon Therapy

Background— Drug-coated balloons are increasingly used for peripheral vascular disease, and, yet, mechanisms of tissue uptake and retention remain poorly characterized. Most systems to date have used paclitaxel, touting its propensity to associate with various excipients that can optimize its transfer and retention. We examined zotarolimus pharmacokinetics. Methods and Results— Animal studies, bench-top experiments, and computational modeling were integrated to quantify arterial distribution after zotarolimus-coated balloon use. Drug diffusivity and binding parameters for use in computational modeling were estimated from the kinetics of zotarolimus uptake into excised porcine femoral artery specimens immersed in radiolabeled drug solutions. Like paclitaxel, zotarolimus exhibited high partitioning into the arterial wall. Exposure of intimal tissue to drug revealed differential distribution patterns, with zotarolimus concentration decreasing with transmural depth as opposed to the multiple peaks displayed by paclitaxel. Drug release kinetics was measured by inflating zotarolimus-coated balloons in whole blood. In vivo drug uptake in swine arteries increased with inflation time but not with balloon size. Simulations coupling transmural diffusion and reversible binding to tissue proteins predicted arterial distribution that correlated with in vivo uptake. Diffusion governed drug distribution soon after balloon expansion, but binding determined drug retention. Conclusions— A large bolus of zotarolimus releases during balloon inflation, some of which pervades the tissue, and a fraction of the remaining drug adheres to the tissue–lumen interface. As a result, the duration of delivery modulates tissue uptake where diffusion and reversible binding to tissue proteins determine drug transport and retention, respectively.

Matrix-embedded endothelial cells are protected from the uremic milieu

BACKGROUND Endothelial cells (ECs) embedded in 3D matrices [matrix-embedded endothelial cells (MEECs)] of denatured collagen implanted around vascular access anastomoses preserve luminal patency. MEEC implant efficacy depends on embedded EC health. As the uremic milieu inhibits proliferation and induces apoptosis of ECs, we examined whether uremia might impact MEECs. METHODS ECs grown on 2D gelatin-coated polystyrene tissue culture plates (gTCPS) or in MEEC were treated with sera pooled from 20 healthy control or uremic patients with end-stage renal disease. EC viability was examined using 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyl tetrazolium bromide assay, cell counting and Trypan blue exclusion. Media conditioned (CM) with 2 and 3D-supported ECs were examined for its potential to inhibit vascular smooth muscle cell (vSMC) proliferation using (3)[H] thymidine incorporation and cyclin D1 expression. ECs grown on gTCPS were treated with uremic serum filtered through matrices to examine if matrices retain uremic toxins or whether EC effects were cell mediated. RESULTS Uremic serum significantly reduced viability and number of live, and increased dead ECs when grown on gTCPS, but not in MEECs. EC survival correlated with vSMC inhibition. While CM from ECs grown in gTCPS with uremic serum inhibited vSMC proliferation no better than uremic serum alone (22 versus 27%), MEEC CM inhibited vSMC proliferation by 47% (P = 0.0004). Cyclin D1 expression tracked with indices of vSMC proliferation. There was no significant difference in EC viability between EC treated with matrix-filtered or unfiltered uremic serum. CONCLUSION The viability, number and efficacy of MEECs were preserved in uremic serum compared to those of ECs on gTCPS. MEECs are protected from uremic toxicity, not from retention of uremic toxins by matrices, but likely from intrinsic changes in EC sensitivity to uremia. MEECs implanted at vascular access sites should inhibit neointimal hyperplasia in uremia. This study underscores the robustness of matrix embedding as a cell protectant, especially in hostile environments like uremia.

Detection and Analysis of Circulating Epithelial Cells in Liquid Biopsies From Patients With Liver Disease

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Matrix‐Embedded Endothelial Cells Attain a Progenitor‐Like Phenotype

Culture of endothelial cells (ECs) embedded in 3D scaffolds of denatured collagen has shown tremendous therapeutic potential in clinical trials of tissue repair. It is postulated that these matrix‐embedded ECs (MEECs) attain a differential phenotype similar to early progenitor forms, which cannot be attained in 2D culture. MEECs are compared to 2D‐ECs and endothelial progenitor cells (EPCs) by secretome, phenotype, and genetic fingerprint, and are found to be altered from 2D‐ECs on all levels, adopting an EPC‐like phenotype. This manifests in elevation of CD34 expression—a progenitor cell marker—and protein secretion and gene expression profiles that are similar to EPCs. Even more striking is that EPCs in 2D lose their phenotype, evident by the loss of CD34 expression, but are able to regain expression over time when embedded in the same 3D matrices, suggesting that future in vitro EPC work should use ME‐EPCs to recapitulate in vivo phenotype. These findings elucidate the relationship between EPCs and the substratum‐dependent regulation imparted by ECs which is critical to understand in order to optimize MEEC therapy and propel it into the clinic.