Edward L. Snyder

Tissue-engineered vascular grafts transform into mature blood vessels via an inflammation-mediated process of vascular remodeling

Biodegradable scaffolds seeded with bone marrow mononuclear cells (BMCs) are the earliest tissue-engineered vascular grafts (TEVGs) to be used clinically. These TEVGs transform into living blood vessels in vivo, with an endothelial cell (EC) lining invested by smooth muscle cells (SMCs); however, the process by which this occurs is unclear. To test if the seeded BMCs differentiate into the mature vascular cells of the neovessel, we implanted an immunodeficient mouse recipient with human BMC (hBMC)-seeded scaffolds. As in humans, TEVGs implanted in a mouse host as venous interposition grafts gradually transformed into living blood vessels over a 6-month time course. Seeded hBMCs, however, were no longer detectable within a few days of implantation. Instead, scaffolds were initially repopulated by mouse monocytes and subsequently repopulated by mouse SMCs and ECs. Seeded BMCs secreted significant amounts of monocyte chemoattractant protein-1 and increased early monocyte recruitment. These findings suggest TEVGs transform into functional neovessels via an inflammatory process of vascular remodeling.

Cytokine generation in stored platelet concentrates.

BACKGROUND: Cytokines, because of the nature of their immunoinflammatory actions, are potential mediators of the symptom complex of nonhemolytic transfusion reactions. One possible source of cytokines in the transfusion setting is the stored blood component itself.

Reduction of febrile but not allergic reactions to RBCs and platelets after conversion to universal prestorage leukoreduction.

BACKGROUND:  Between January 1995 and November 1998, at Yale‐New Haven Hospital, 25 percent of RBCs transfused were processed through prestorage or bedside leukoreduction filters, chosen on a per patient basis (selective leukoreduction [SLR]). Between January 1995 and July 1999, 30 percent of platelet concentrates (PCs) were infused through bedside leukoreduction filters. In an attempt to decrease febrile nonhemolytic transfusion reactions (FNHTR), a change was made from SLR to universal prestorage leukoreduction (UPL) for RBCs between November 1998 and December 1999 and for random donor PCs between July 1999 and January 2000. FNHTR and allergic transfusion reactions (ATR) reported from January 1995 through December 2002 were reviewed.

Apoptotic activity in stored human platelets

BACKGROUND: Platelets possess some of the machinery required for apoptotic cell death. However, disruption of mitochondria function, implicated in several models of cell death, has not been extensively studied in platelets. Mitochondrial viability and several other measures of apoptotic death in stored and experimentally stressed platelets were evaluated.

Occurrence of the Release Reaction during Preparation and Storage of Platelet Concentrates

To determine the degree of damage occurring during preparation and storage of platelet concentrates, the percent release of B‐thromboglobulin (BTG) and percent leakage of the cytosolic protein lactic dehydrogenase was determined sequentially from phlebotomy to the end of storage for 72 h at 20–24°C. The effect of storage temperature, pH, and radiation was also evaluated. The results showed that during preparation of platelet concentrate a large degree of release was found after resuspension of the platelet button formed after the high‐speed centrifugation. During storage the percent BTG release increased from 18.1 to 40.2% (p<0.05). The percent release seen during storage at 4°C (72 h) was 19.2%, while that seen for platelets subjected to temperature cycling at 4–37 °C was 24.9%. Both of these values were significantly less (p<0.05) than that seen for concentrates stored at room temperature. A negative correlation between pH and BTG release was found (r=‐0.64). Irradiation to 10,000 rad did not induce the release reaction or lactic dehydrogenase leakage. We conclude that the degree of in vitro platelet release is dependent on the preparative manipulations, and gentler protocols for preparation and storage of platelets should be investigated.

Induction of human tumor-loaded dendritic cells

A preferred anti‐cancer vaccine would be tumor‐specific, simple to rapidly construct and safe to administer. It would permit immunization against a spectrum of the tumors distinctive antigens, without requiring their prior identification. Toward these goals, we describe a modification of standard extracorporeal photopheresis (ECP) which initiates, within a single day, both monocyte‐to‐dendritic cell (DC) differentiation and malignant cell apoptosis. The transition of monocytes to immature DCs was identified by the expression of cytoplasmic CD83 and membrane CD36 in the absence of membrane CD14 staining, as well as induction of membrane CD83 expression. Differentiating DCs were avidly phagocytic and engulfed apoptotic malignant T cells. Differentiating DCs were capable of stimulating significant proliferation of normal alloreactive lymphocyte responders, indicting increased expression of membrane MHC class II molecules. This approach provides a clinically practical means of developing tumor‐loaded cells that have initiated the transition to DCs without the requirement of exogenous cytokines, excessive cellular manipulation or isolation. Construction of DC vaccines using this methodology can be generalized to other diseases and may offer a novel approach for improved cancer immunotherapy.

Optimizing Autologous Stem Cell Mobilization Strategies to Improve Patient Outcomes: Consensus Guidelines and Recommendations

Autologous hematopoietic stem cell transplantation (aHSCT) is a well-established treatment for malignancies such as multiple myeloma (MM) and lymphomas. Various changes in the field over the past decade, including the frequent use of tandem aHSCT in MM, the advent of novel therapies for the treatment of MM and lymphoma, and the addition of new stem cell mobilization techniques, have led to the need to reassess current stem cell mobilization strategies. Mobilization failures with traditional strategies are common and result in delays in treatment and increased cost and resource utilization. Recently, plerixafor-containing strategies have been shown to significantly reduce mobilization failure rates, but the ideal method to maximize stem cell yields and minimize costs associated with collection has not yet been determined. A panel of experts convened to discuss the currently available data on autologous hematopoietic stem cell mobilization and transplantation and to devise guidelines to optimize mobilization strategies. Herein is a summary of their discussion and consensus.

Cytokine generation in stored, white cell‐reduced, and bacterially contaminated units of red cells

BACKGROUND: Proinflammatory cytokines were measured in the supernatant portion of stored, bacterially contaminated, and/or white cell (WBC)‐ reduced units of red cells (RBCs). Previous studies from this laboratory and others have shown that cytokines are generated in platelet concentrates during storage. This earlier work has been expanded to the study of stored RBCs.

Clinical safety of platelets photochemically treated with amotosalen HCl and ultraviolet A light for pathogen inactivation: the SPRINT trial

BACKGROUND:  A photochemical treatment (PCT) method utilizing a novel psoralen, amotosalen HCl, with ultraviolet A illumination has been developed to inactivate viruses, bacteria, protozoa, and white blood cells in platelet (PLT) concentrates. A randomized, controlled, double‐blind, Phase III trial (SPRINT) evaluated hemostatic efficacy and safety of PCT apheresis PLTs compared to untreated conventional (control) apheresis PLTs in 645 thrombocytopenic oncology patients requiring PLT transfusion support. Hemostatic equivalency was demonstrated. The proportion of patients with Grade 2 bleeding was not inferior for PCT PLTs.

Activation in stored platelet concentrates: correlation between membrane expression of P‐selectin, glycoprotein IIb/IIIa, and beta‐ thromboglobulin release

By using two distinct measurements of alpha‐degranulation (surface P‐ selectin [alpha‐granule membrane protein‐140] expression and beta‐ thromboglobulin [beta‐TG] release) and quantitation of glycoprotein (GP) IIb/IIIa surface density, stored platelet concentrates were evaluated to determine a) which method of measuring platelet alpha‐ granule release was more sensitive in detecting early platelet activation; b) whether Day 1 levels of activation predicted the extent of activation or cell lysis on Day 5 of storage; and c) whether changes in surface GPIIb/IIIa density were primarily dependent on platelet activation. By using samples from paired and unpaired units stored for 1, 3, and 5 days, four observations could be made. 1) A flow cytometric assay for the percentage of P‐selectin‐positive platelets was more sensitive for early detection of platelet activation than was measurement of beta‐TG release. This finding was most likely due to enhanced sensitivity in detecting platelets that had undergone partial alpha‐granule release. 2) Total P‐selectin expression correlated with beta‐TG release, which indicated that the extent of alpha‐granule membrane fusion with the external platelet membrane was proportional to the amount of alpha‐granule contents released into the supernatant. 3) All of the activation measurements on Day 1 predicted the activation values, but did not predict the degree of cell lysis (measured by lactate dehydrogenase discharge), on Day 5 of storage. 4) Surface GPIIb/IIIa density was increased on the subset of P‐selectin‐positive platelets as compared with the P‐selectin‐negative subset at all times during storage, but, within each subset, GPIIb/IIIa surface density did not significantly increase over the time of storage.