The pathophysiology of myocardial injury that results from cardiac ischemia and reperfusion (I/R) is incompletely understood. Experimental evidence from murine models indicates that innate immune mechanisms including complement activation via the classical and lectin pathways are crucial. Whether factor B (fB), a component of the alternative complement pathway required for amplification of complement cascade activation, participates in the pathophysiology of myocardial I/R injury has not been addressed. We induced regional myocardial I/R injury by transient coronary ligation in WT C57BL/6 mice, a manipulation that resulted in marked myocardial necrosis associated with activation of fB protein and myocardial deposition of C3 activation products. In contrast, in fB-/- mice, the same procedure resulted in significantly reduced myocardial necrosis (% ventricular tissue necrotic; fB-/- mice, 20 ± 4%; WT mice, 45 ± 3%; P < 0.05) and diminished deposition of C3 activation products in the myocardial tissue (fB-/- mice, 0 ± 0%; WT mice, 31 ± 6%; P<0.05). Reconstitution of fB-/- mice with WT serum followed by cardiac I/R restored the myocardial necrosis and activated C3 deposition in the myocardium. In translational human studies we measured levels of activated fB (Bb) in intracoronary blood samples obtained during cardio-pulmonary bypass surgery before and after aortic cross clamping (AXCL), during which global heart ischemia was induced. Intracoronary Bb increased immediately after AXCL, and the levels were directly correlated with peripheral blood levels of cardiac troponin I, an established biomarker of myocardial necrosis (Spearman coefficient = 0.465, P < 0.01). Taken together, our results support the conclusion that circulating fB is a crucial pathophysiological amplifier of I/R-induced, complement-dependent myocardial necrosis and identify fB as a potential therapeutic target for prevention of human myocardial I/R injury.
Preeclampsia is a leading cause of maternal and fetal morbidity and mortality. Bb, the active fragment of complement factor B (fB), has been reported to be a predictor of preeclampsia. However, conflicting results have been found by some investigators. We hypothesized that the disagreement in findings may be due to the racial/ethnic differences among various study groups, and that fB activation is significant in women of an ethnic minority with preeclampsia. We investigated the maternal and fetal levels of Bb (the activated fB fragment) in pregnant women of an ethnic minority with or without preeclampsia. We enrolled 291 pregnant women (96% of an ethnic minority, including 78% African–American). Thirteen percent of these were diagnosed with preeclampsia. Maternal venous blood was collected from all participants together with fetal umbilical cord blood samples from 154 deliveries in the 291 women. The results were analyzed using the Mann–Whitney U test and multivariate analyses. Maternal Bb levels were significantly higher in the preeclamptic group than in the nonpreeclamptic group. Levels of Bb in fetal cord blood were similar in both groups. Subgroup analyses of African–American patients’ results confirmed the study hypothesis that there would be a significant increase in Bb in the maternal blood of the preeclamptic group and no increase in Bb in the fetal cord blood of this group. These results suggest that a maternal immune response through complement fB might play a role in the development of preeclampsia, particularly in African–American patients.
Purpose: Harmonized animal models are an indispensable tool for the development of safe and effective medical countermeasures (MCMs) against radiation injury, and rhesus macaques (referred herein as NHPs) play a critical role in FDA approval of radiation medical countermeasures for acute and delayed radiation syndromes. Reliance on such models requires that they be well characterized, which consists, in part, of a reproducible dose to mortality response relationship (DRR). However, data describing the DRR for both male and female NHPs from the same study are scarce. Furthermore, the level of supportive care and the use of blood transfusions may shift the DRR, yet such information can be difficult to compare across publications. To address these knowledge gaps, the DRRs of two different NHP total body irradiation (TBI) models are compared in this paper, one which is reliant on the use of male animals provided blood transfusions, and the other which incorporates both sexes wherein animals are not provided transfusions. Materials and methods: Studies were conducted using NHPs (Macacca mulatta) receiving TBI, with survival reported over a 60 days. Two primary studies, incorporating both male and female animals not receiving blood transfusions as a provision of supportive care, were compared to two previously published studies, which incorporated only male animals provided blood transfusions as a part of the supportive care regimen. Criterion for euthanasia, and all other provisions of supportive care were comparable. Linear probit plots estimating the lethal dose (LD) and upper and lower limits of the 95% confidence interval (CI) for 10, 30, 50, 70 and 90% mortality, were compared between individual studies and the two models presented. Results: Comparison of probit estimates reveals two important findings. (1) Females have higher mortality than males at identical radiation doses, and (2) blood transfusions increased survival of male animals at lower doses but not at high doses of radiation exposure. Conclusions: The use of single sex animal models may lead to an incomplete understanding of potential sex differences in the dose to mortality response of the TBI model. Consistent use of both sexes and type of supportive care will improve the transferability and reliability of NHP-TBI models currently in use, assist in the selection of radiation doses for single dose lethality studies, and allow investigators to determine the effectiveness of a particular MCM.
The NOD/SCID/IL2Rγnull (NSG) mouse is a relevant model for toxicology and tumorigenicity studies evaluating human cell therapies. Data was compiled from toxicology study control NSG mice exposed to gamma irradiation (0 or 200 cGy) or busulfan. Retrospective data evaluation included mortality, clinical observations, body weights, hematology, and external and internal macroscopic observations. There was no mortality in any of the 129 toxicology control (irradiated and non-irradiated) mice up to the 20-week observation period. Mortalities occurred between Days 1 and 25 among animals given busulfan ≥25 mg/kg/day at 1 or 2 doses via intraperitoneal (i.p.) injection. There were 4/10, 6/10 and 4/10 deaths at 25, 30 and 35 mg/kg/day busulfan, respectively. Busulfan-treated mice presented with dose-dependent clinical signs including signs of anemia in some individuals. Hematology, including white blood cell (WBC) and neutrophil (NEUT) counts, from irradiated mice at Weeks 12 and 20 revealed comparable values to non-irradiated animals. In contrast, irradiated mice treated with a positive control (HL-60) were euthanized prior to Week 12. There were no irradiation-related differences in macroscopic observations with lymphoid atrophy identified comparably in irradiated and non-irradiated groups. These results suggest that irradiation was suitable for conditioning to enable cell engraftment in NSG mice in the context of regulatory toxicology and tumorigenicity studies. Busulfan administered at 20 mg/kg/day for 2 days, i.p. was also well-tolerated, and it could be considered for toxicology studies of genetically modified human cells.
The pathophysiology of myocardial injury resulting from cardiac ischemia and reperfusion (I/R) is incompletely understood. Experimental evidence from murine models indicates that innate immune mechanisms including complement activation via the classical and lectin pathways are crucial. Whether factor B (fB), a component of the alternative complement pathway required for amplification of complement cascade activation, participates in the pathophysiology of myocardial I/R injury has not been addressed. We induced regional myocardial I/R injury by transient coronary ligation in WT mice, a manipulation that resulted in marked myocardial necrosis associated with upregulated myocardial expression of fB gene, activation of fB protein, and myocardial deposition of C3 activation products. In contrast, in fB−/− mice, the same procedure resulted in significantly reduced myocardial necrosis and diminished deposition of C3 activation products in the myocardial tissue. To extend the analysis to humans we measured levels of activated fB (Bb) in intracoronary blood samples obtained during cardio-pulmonary bypass surgery before and after aortic cross clamping (AXCL), during which global heart ischemia was induced. Intracoronary Bb increased immediately after AXCL, the levels were directly related to the length of the AXCL and correlated with peripheral blood levels of cardiac troponin I, an established biomarker of myocardial necrosis. Taken together, our results support the conclusion that fB is a crucial pathophysiological amplifier of I/R-induced, complement-dependent myocardial necrosis and identify fB as a potential therapeutic target for prevention of human myocardial I/R injury.
Myocardial ischemia/reperfusion (I/R) elicits an acute inflammatory response involving complement factors. Previous animal studies showed that circulation complement C3 was deposited in the ischemic myocardium flooded with oxygenated blood upon reperfusion. Recently, we reported that myocardial necrosis was decreased in C3-/- mice after heart I/R. The current study used in the same heart model to test the effect of C3 on myocardial apoptosis. Our results showed that myocardial apoptosis was increased in C3-/- mice after heart I/R. Further, comparative proteomics analyses found that cytochrome c was present in the myocardial C3-complex following I/R. These results indicate that C3 can interact with cytochrome c in the cytosol of cardiomyocytes during myocardial I/R, which may sequester cytochrome c and thus reduce the number of cells undergoing apoptosis. In summary, our findings raise the possibility of a new mechanism affecting cell death relevant to pathologic conditions such as ischemia: a circulating innate immune factor, i.e. complement, can interact with intracellular factor(s), and influence the types of cell death that occur.
Introduction Cancer patients are at an increased risk of moderate-to-severe obstructive sleep apnea (OSA). The STOP-Bang score is a commonly used screening questionnaire to assess risk of OSA in the general population. We hypothesize that cancer-relevant features, like radiation therapy (RT), may be used to determine the risk of OSA in cancer patients. Machine learning (ML) with non-parametric regression is applied to increase the prediction accuracy of OSA risk. Methods Ten features namely STOP-Bang score, history of RT to the head/neck/thorax, cancer type, cancer stage, metastasis, hypertension, diabetes, asthma, COPD, and chronic kidney disease were extracted from a database of cancer patients with a sleep study. The ML technique, K-Nearest-Neighbor (KNN), with a range of k values (5 to 20), was chosen because, unlike Logistic Regression (LR), KNN is not presumptive of data distribution and mapping function, and supports non-linear relationships among features. A correlation heatmap was computed to identify features having high correlation with OSA. Principal Component Analysis (PCA) was performed on the correlated features and then KNN was applied on the components to predict the risk of OSA. Receiver Operating Characteristic (ROC) - Area Under Curve (AUC) and Precision-Recall curves were computed to compare and validate performance for different test sets and majority class scenarios. Results In our cohort of 174 cancer patients, the accuracy in determining OSA among cancer patients using STOP-Bang score was 82.3% (LR) and 90.69% (KNN) but reduced to 89.9% in KNN using all 10 features mentioned above. PCA + KNN application using STOP-Bang score and RT as features, increased prediction accuracy to 94.1%. We validated our ML approach using a separate cohort of 20 cancer patients; the accuracies in OSA prediction were 85.57% (LR), 91.1% (KNN), and 92.8% (PCA + KNN). Conclusion STOP-Bang score and history of RT can be useful to predict risk of OSA in cancer patients with the PCA + KNN approach. This ML technique can refine screening tools to improve prediction accuracy of OSA in cancer patients. Larger studies investigating additional features using ML may improve OSA screening accuracy in various populations Support (if any):
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