Abstract
Introduction: Recent advances in preventive care, such as hydoxyurea and prophylactic antibiotics, have reduced the mortality of children with sickle cell disease (SCD) s in developed countries. Yet, the chronic hemolytic anemia and recurrent vaso-occlusive crises result in systemic inflammation and coagulopathy. Markers of coagulation activation correlate with painful crises, acute chest syndrome, stroke, venous thromboembolism, pulmonary hypertension, left ventricular diastolic heart disease, and sickle nephropathy. These complications result in end-organ failure that causes increased morbidity and mortality in adult SCD patients. We have shown that tissue factor (TF), the primary initiator of extrinsic coagulation, contributes to inflammation and coagulation in mouse models of SCD. (1,2). It has also been demonstrated that long-term reduction in thrombin protects from cardiopulmonary dysfunction and reduces mortality of sickle cell mice (3). Recent work from our laboratory demonstrates that high molecular weight kininogen (HK) promotes thrombin generation and inflammation in sickle mice. HK is proteolytically cleaved into bradykinin and cleaved HK fragments (HKf) by kallikrein and other proteases. HKf induces TF expression and activity on monocytes dependent on Mac-1 (CD11b/CD18). We found that Mac-1 inhibition attenuates thrombin generation and inflammation in sickle cell mice.
Hypothesis: Long-term HK deficiency in sickle cell mice will attenuate TF-mediated coagulation and inflammation, and protect against end-organ damage and mortality.
Methods and Results: To evaluate the effect of long-term HK deficiency on outcomes in sickle cell disease, we used bone marrow from Townes sickle (SS) and wild type (AA) mice to generate chimeras in lethally irradiated HK+/+ (WT) and HK-/- (KO) mice to create AA/WT, AA/KO, SS/WT, and SS/KO mice. Efficient reconstitution of bone marrow was confirmed by hemoglobin electrophoresis. Eight months after chimeras were generated, endpoints were assessed. SS/WT mice had early mortality (median survival 209 days, 6/23 mice survived to 250 days); HK deficiency significantly prolonged survival in SS mice (median survival 240 days, 24/29 mice survived to 250 days; p<0.01). Plasma levels of interleukin-6 were significantly higher in SS/WT mice compared to AA/WT controls (21 ± 3.7 ng/mL vs 6.6 ± 2.2 ng/mL in AA/WT, p<0.001); HK deficiency attenuated this increase (5.1 ± 1.2 ng/mL, p<0.001). The neutrophil-lymphocyte ratio was also elevated in SS/WT mice (0.47 ± 0.004 vs 0.3 ± 0.05 in AA/WT, p<0.05), yet not in the SS/KO group (0.25 ± 0.07, p<0.05), indicating that HK deficiency protects against inflammation in SS mice. Analysis of urine for renal injury markers revealed that SS/WT mice had elevated urine albumin/creatinine ratios (652 ± 34 mg albumin/g creatinine vs. 276 ± 54, p<0.01), which was significantly decreased in SS/KO mice (321 ± 49, p<0.01). Moreover, SS/WT mice had significantly reduced urine osmolality compared to AA/WT controls (2116 mOsm/kg vs 1208 mOsm/kg, p<0.05), which was reversed in SS/KO mice (1842 mOsm/kg, p<0.05). This suggests that HK deficiency protects against kidney injury and preserves urinary concentrating ability. We also observed increases in the relative left ventricle (LV+S/BW; 4.9 ± 0.2 vs 3.3 ± 0.1, p<0.05) and right ventricle (RV/BW; 1.4 ± 0.4 vs 0.99 ± 0.09, p<0.05) size in SS/WT mice compared to AA/WT controls, which was prevented in SS/KO mice (LV+S/BW: 4.0 ± 0.2, p<0.05 and RV/BW: 1.1 ± 0.06, p<0.05).
Conclusions: These data indicate that HK deficiency attenuates chronic inflammation, kidney failure, and heart hypertrophy, and improves survival of sickle cell mice.
Disclosures
No relevant conflicts of interest to declare.
High Molecular Weight Kininogen: A Review of the Structural Literature
