Angela D. Burnette

Pretransplant Donor-Specific and Non-Specific Immune Parameters Associated With Early Acute Rejection

Background. New immunosuppression protocols have resulted in decreased rates of biopsy-proven acute rejection; however, it is unclear whether recipients without biopsy-proven acute rejection are still at risk for immune complication and chronic allograft dysfunction. The aim of our studies was to determine whether pretransplant immune parameters were associated with posttransplant early acute rejection, unstable creatinine courses, and poor graft outcome. Methods. Immune parameters, including human leukocyte antigen (HLA) mismatch, HLA-specific antibodies, global CD4+ cellular response as measured by intracellular adenosine triphosphate (iATP) synthesis, and IFN-&ggr; precursor frequencies to donor or third-party cells as measured by ELISPOT were determined for a total of 126 kidney recipients treated with a protocol, including rapid discontinuation of prednisone. Results. The donor specific pretransplant parameters of HLA class I mismatches (P=0.04) and total HLA mismatches (P=0.04) with the donor as well as the pretransplant HLA-donor specific antibodies (P=0.002) were associated with biopsy-proven acute rejection. Higher pretransplant iATP levels, a donor nonspecific parameter, were found associated with biopsy proven acute rejection (P=0.04). Pretransplant iATP levels were significantly greater for recipients with early unstable creatinine levels (P=0.01). Recipients with a pretransplant iATP value greater than 375 ng/ml were 3.67 times more likely to experience acute rejection (P=0.03). Conclusions. Pretransplant assessment of donor specific and nonspecific immune parameters may identify recipients who can benefit from closer clinical and immunological surveillance to allow for tailored immunsuppression and selective intervention aimed at optimizing both short and long-term graft outcome.

RNA Aptamer Therapy for Vaso-Occlusion in Sickle Cell Disease

Patients with sickle cell disease (SCD) often suffer painful vaso-occlusive episodes caused in part by the adhesion of sickle erythrocytes (SS-RBC) to the vascular endothelium. To investigate inhibition of SS-RBC adhesion as a possible treatment for vaso-occlusion, 2 adhesion molecules, α(v)β(3) and P-selectin, were targeted by high-affinity RNA aptamers. An in vitro flow chamber assay was used to test the antiadhesion activity of α(v)β(3) aptamer clone 17.16. Human SS-RBC were passed across a confluent monolayer of thrombin-stimulated human umbilical vein endothelial cells (HUVEC) at a constant rate. α(v)β(3) aptamer reduced SS-RBC adhesion to activated endothelial cells to the level seen with untreated HUVEC. An aptamer reactive with complement component 8 was used as a negative control and exerted no inhibition, confirming the specificity of α(v)β(3) aptamer (P=0.04). At 2 dyn/cm(2) shear stress, 30 nM α(v)β(3) aptamer showed maximal effect in decreasing SS-RBC adhesion to HUVEC. The antiadhesive activity of the P-selectin aptamer clone PF377 was also tested using HUVEC pretreated with IL-13 to upregulate expression of P-selectin as seen in activated endothelial cells. At 1 dyn/cm(2) shear stress, 60 nM of P-selectin aptamer had antiadhesion activity similar to heparin, a known inhibitor of SS-RBC adhesion to P-selectin. A negative control did not prevent adhesion (P=0.05). These data show the potential utility of aptamers to block endothelial adhesion molecules to prevent or treat vaso-occlusion in SCD.

Scavenging nucleic acid debris to combat autoimmunity and infectious disease

Significance Pathological inflammation resulting from autoimmune, inflammatory, and infectious diseases remains the major cause of morbidity and mortality around the world. We explored the utility of nucleic acid-binding polymers to limit pathological activation of the innate immune response by scavenging nucleic acid-containing debris. We evaluated such polymers following local and systemic delivery for treatment of cutaneous and systemic lupus erythematosus in mice and observed dramatic therapeutic benefit. Remarkably, such treatment does not limit the ability of lupus-prone mice to combat influenza infection. Thus, by selectively targeting activators of the pathogenic immune response rather than the effectors of the innate immune system, nucleic acid scavengers are able to limit pathological inflammation without compromising one’s ability to fight a viral infection. Nucleic acid-containing debris released from dead and dying cells can be recognized as damage-associated molecular patterns (DAMPs) or pattern-associated molecular patterns (PAMPs) by the innate immune system. Inappropriate activation of the innate immune response can engender pathological inflammation and autoimmune disease. To combat such diseases, major efforts have been made to therapeutically target the pattern recognition receptors (PRRs) such as the Toll-like receptors (TLRs) that recognize such DAMPs and PAMPs, or the downstream effector molecules they engender, to limit inflammation. Unfortunately, such strategies can limit the ability of the immune system to combat infection. Previously, we demonstrated that nucleic acid-binding polymers can act as molecular scavengers and limit the ability of artificial nucleic acid ligands to activate PRRs. Herein, we demonstrate that nucleic acid scavengers (NASs) can limit pathological inflammation and nucleic acid-associated autoimmunity in lupus-prone mice. Moreover, we observe that such NASs do not limit an animal’s ability to combat viral infection, but rather their administration improves survival when animals are challenged with lethal doses of influenza. These results indicate that molecules that scavenge extracellular nucleic acid debris represent potentially safer agents to control pathological inflammation associated with a wide range of autoimmune and infectious diseases.