Immuno-Thrombotic Complications of COVID-19: Implications for Timing of Surgery and Anticoagulation.

Early in the coronavirus disease 2019 (COVID-19) pandemic, global governing bodies prioritized transmissibility-based precautions and hospital capacity as the foundation for delay of elective procedures. As elective surgical volumes increased, convalescent COVID-19 patients faced increased postoperative morbidity and mortality and clinicians had limited evidence for stratifying individual risk in this population. Clear evidence now demonstrates that those recovering from COVID-19 have increased postoperative morbidity and mortality. These data-in conjunction with the recent American Society of Anesthesiologists guidelines-offer the evidence necessary to expand the early pandemic guidelines and guide the surgeon's preoperative risk assessment. Here, we argue elective surgeries should still be delayed on a personalized basis to maximize postoperative outcomes. We outline a framework for stratifying the individual COVID-19 patient's fitness for surgery based on the symptoms and severity of acute or convalescent COVID-19 illness, coagulopathy assessment, and acuity of the surgical procedure. Although the most common manifestation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is COVID-19 pneumonitis, every system in the body is potentially afflicted by an endotheliitis. This endothelial derangement most often manifests as a hypercoagulable state on admission with associated occult and symptomatic venous and arterial thromboembolisms. The delicate balance between hyper and hypocoagulable states is defined by the local immune-thrombotic crosstalk that results commonly in a hemostatic derangement known as fibrinolytic shutdown. In tandem, the hemostatic derangements that occur during acute COVID-19 infection affect not only the timing of surgical procedures, but also the incidence of postoperative hemostatic complications related to COVID-19-associated coagulopathy (CAC). Traditional methods of thromboprophylaxis and treatment of thromboses after surgery require a tailored approach guided by an understanding of the pathophysiologic underpinnings of the COVID-19 patient. Likewise, a prolonged period of risk for developing hemostatic complications following hospitalization due to COVID-19 has resulted in guidelines from differing societies that recommend varying periods of delay following SARS-CoV-2 infection. In conclusion, we propose the perioperative, personalized assessment of COVID-19 patients' CAC using viscoelastic hemostatic assays and fluorescent microclot analysis.

Fibrin Strands Will Grow from Soluble Fibrin and Hang Up in an In Vitro Microcirculatory Viscoelastic Model: Is This a Major Cause of COVID-19 Associated Coagulopathy?

Viscoelastic testing (VET) by both TEG and ROTEM has demonstrated hypercoagulability early in corona virus disease 2019 (COVID-19) associated coagulopathy (CAC). Additional VET studies demonstrated fibrinolytic shutdown late in a majority of severely ill COVID-19 patients with an associated elevation of d-dimer. Elevated d-dimer confirms that coagulation, followed by fibrinolysis, has occurred. These findings imply that, during CAC, three enzymes-thrombin, Factor XIIIa and plasmin-must have acted in sequence. However, limitations in standard VET analyses preclude exploration of the earliest phases of clot induction, as well as clot formation and clot dissolution in flowing blood. Herein, we describe a novel method illuminating aspects of this unexplored area. In addition, we created an in vitro blood flow model in which the interactions of thrombin, Factor XIII and plasmin with fibrinogen can be studied, allowing the determination of soluble fibrin (SF), the highly unstable form of fibrin that precedes the appearance of a visible clot. This model allows the determination of the SF level at which fibrin microclots begin to form.

A macrophage attack culminating in microthromboses characterizes COVID 19 pneumonia.

INTRODUCTION: A neutrophilic infiltrate characterizes bacterial pneumonia. Macrophage infiltration is similarly characteristic of the viral pneumonia caused by SARS-CoV-2. These infiltrating macrophages, while phagocytic and capable of engulfing virus laden alveolar cells, are also rich in tissue factor-a thromboplastin. This prothrombotic aspect likely explains how a respiratory virus whose malign effects should be confined to the oropharynx, bronchi and lungs, can cause a panoply of extra-pulmonary organ disorders. Elevated ferritin levels in ICU Covid 19 patients, and elevated acute phase proteins suggest immune overreaction. Elevated d-dimers implicate clotting as well. This evidence links hyperactive innate immunity (macrophage lung infiltrates) with the elevated levels of oligomeric fibrin present in the bloodstream of these patients. METHODS: An in-house assay measuring oligomeric (soluble) fibrin (also referred to as soluble fibrin monomer complexes or SFMC) in whole blood, previously developed for monitoring incipient disseminated intravascular coagulation (DIC) during liver transplantation, was made available to COVID ICU attendings. Since SFMC constitutes the input to intravascular fibrin clots and d-dimer reflects fibrin clot dissolution, it was thought that the two tests, run in tandem along with assays of immune activation, might clarify the frequency and possibly the cause of DIC in patients with severe COVID-19 pneumonia. RESULTS: Classical DIC with intravascular clotting and thrombocytopenia was documented only rarely. However, early in the pandemic shortly after the assay was made available, it identified three patients undergoing acute defibrination. In each patient virtually all of the body's fibrinogen was transformed into SFMC over 3-4 days and deposited somewhere in the vasculature without any gross clots being detected. CONCLUSIONS: Three COVID-19 patients with evidence of a hyperactive immune response (elevated ferritin and acute phase proteins) defibrinated while blood levels of SFMC were being monitored. SFMC levels that were five times higher than normal appeared in the circulation during the defibrination process. SFMC at these levels may precipitate as showers of microclots, damaging heart, kidney, brain, and so forth.

Extreme Plasma Dilution Decreases Heparin and Protamine Cardiopulmonary Bypass Requirements: A Case Report on a Jehovah's Witness Patient.

The primary focus of cardiopulmonary bypass management in Jehovah's Witness patients is the need to conserve blood. A consequence of these strategies inevitably results in hemodilution that is frequently extreme enough to dilute clotting factors and potentially impair coagulation. The purpose of this case report is to demonstrate that a hemodiluted patient requires less heparin to sustain anticoagulation and less protamine to reverse heparin at cardiopulmonary bypass termination. Patient harm may ensue unless the effects of extreme hemodilution are recognized.

Crotalus atrox venom preconditioning increases plasma fibrinogen and reduces perioperative hemorrhage in a rat model of surgical brain injury.

Perioperative bleeding is a potentially devastating complication in neurosurgical patients, and plasma fibrinogen concentration has been identified as a potential modifiable risk factor for perioperative bleeding. The aim of this study was to evaluate preconditioning with Crotalus atrox venom (Cv-PC) as potential preventive therapy for reducing perioperative hemorrhage in the rodent model of surgical brain injury (SBI). C. atrox venom contains snake venom metalloproteinases that cleave fibrinogen into fibrin split products without inducing clotting. Separately, fibrinogen split products induce fibrinogen production, thereby elevating plasma fibrinogen levels. Thus, the hypothesis was that preconditioning with C. atrox venom will produce fibrinogen spilt products, thereby upregulating fibrinogen levels, ultimately improving perioperative hemostasis during SBI. We observed that Cv-PC SBI animals had significantly reduced intraoperative hemorrhage and postoperative hematoma volumes compared to those of vehicle preconditioned SBI animals. Cv-PC animals were also found to have higher levels of plasma fibrinogen at the time of surgery, with unchanged prothrombin time. Cv-PC studies with fractions of C. atrox venom suggest that snake venom metalloproteinases are largely responsible for the improved hemostasis by Cv-PC. Our findings indicate that Cv-PC increases plasma fibrinogen levels and may provide a promising therapy for reducing perioperative hemorrhage in elective surgeries.

Association of plasma dilution with cardiopulmonary bypass-associated bleeding and morbidity.

OBJECTIVE: To investigate the relationship of cardiopulmonary bypass-associated plasma dilution with blood product transfusion and postoperative morbidity. DESIGN: Retrospective chart review. SETTING: Single academic medical center. PARTICIPANTS: Five hundred forty adults undergoing cardiac surgery between January 4, 2005 and September 19, 2007. INTERVENTIONS: Records were analyzed for demographics, blood volumes (BVs), and fluid balance. Plasma protein concentrations (% of baseline) at the end of bypass were calculated. The lowest and highest quartiles of plasma protein concentration were correlated with blood product administration and postoperative complications. MEASUREMENTS AND MAIN RESULTS: At the end of bypass, calculated plasma protein concentrations ranged from a low of 10% to a high of 111% of baseline. Concentrations below 45% of baseline were associated with increased blood product administration, longer ventilator support, and longer intensive care unit stay. CONCLUSIONS: Patient morbidity and likelihood of transfusion were associated with calculated plasma protein concentrations below 45% of baseline. Bleeding and administered fluids decrease both hematocrit and plasma proteins. Infusion of washed, salvaged blood or red blood cells raises hematocrit, but further dilutes clotting factors. If this dilution is excessive, coagulopathy may ensue. Patients with the smallest BVs are at greatest risk, but dilution can negatively impact patients with large BVs as well if the fluid used for cardiopulmonary bypass prime and anesthesia management represents a significant fraction of total BV.

Inhibiting platelet-stimulated blood coagulation by inhibition of mitochondrial respiration.

Platelets are important mediators of blood coagulation that lack nuclei, but contain mitochondria. Although the presence of mitochondria in platelets has long been recognized, platelet mitochondrial function remains largely unaddressed. On the basis of a small amount of literature that suggests platelet mitochondria are functional, we hypothesized that the inhibition of platelet mitochondria disrupts platelet function and platelet-activated blood coagulation. To test this hypothesis, members of the tetrazole, thiazole, and 1,2,3-triazole families of small molecule heterocycles were screened for the ability to inhibit isolated mitochondrial respiration and coagulation of whole blood. The families of heterocycles screened were chosen on the basis of the ability of the heterocycle family to inhibit a biomimetic model of cytochrome c oxidase (CcO). The strength of mitochondrial inhibition correlates with each compound's ability to deter platelet stimulation and platelet-activated blood clotting. These results suggest that for this class of molecules, inhibition of blood coagulation may be occurring through a mechanism involving mitochondrial inhibition.

An algorithm for preventing bypass-associated dilutional (BAD) coagulopathy: Theory and example.

OBJECTIVES: Crystalloid administered during cardiopulmonary bypass may significantly dilute clotting factor concentrations, particularly in low blood volume patients. Should the administered fluid (pump prime plus IVs) drop the clotting factor concentrations below approximately 38%, almost all patients will bleed, heparin levels will be overestimated and excessive neutralizing protamine will be administered. This combination can render blood virtually unclottable. This paper describes algorithms that quantify dilution risk and the maximum fluid that can be safely administered. A confirmatory calculation to prevent excessive protamine administration is also described.

Statistical clues to postoperative blood loss: moving averages applied to medical data.

With the advent of computerized databases, medical data has become easy to accumulate; however, effective use of this data continues to pose significant problems. In other circumstances, smoothing algorithms have been used to uncover non-obvious correlations, trends and relationships in noisy data. We have applied four such algorithms to a large dataset of postoperative blood replacement in cardiopulmonary bypass patients. When applied to this dataset, one of the algorithms proved surprisingly effective. It confirmed several previously observed correlations, and also provided an additional series of counterintuitive and apparently unrelated associations. These associations have been explored in an accompanying paper.

Postoperative bypass bleeding: a bypass-associated dilutional (BAD) coagulopathy?

A number of associations with post-bypass bleeding have been described in the accompanying paper. Herein we hypothesize that dilution is an underlying cause through a malign series of bypass-associated events. Heparinized blood behaves anomalously when diluted. Clotting times first shorten somewhat, then--as the dilution of whole blood approaches 50%--rapidly lengthen to unclottability. During cardiopulmonary bypass, low blood volume patients are at a significant risk of clotting factor dilution which will always be more severe than the level of whole blood dilution. If severe enough, this dilution may lower plasma clotting factors to a critical level and may result in excess protamine administration, secondary to overestimation of heparin. The presence of un-neutralized protamine combined with critically lowered clotting factors leads to marked coagulopathy.

Reinfusion of aspirated pericardial blood during CPB. Part I. Hypothesis: laparotomy sponges are a significant part of the CPB circuit?

Blood accumulating in the pericardial sac is routinely reinfused during cardiopulmonary bypass (CPB) surgery. Such reinfusion has been associated with an increased incidence of serious complications such as coagulopathy, systemic inflammation, and neurologic sequelae. We hypothesize that some of these complications occur because the reinfused blood has been exposed to and activated by laparotomy sponges used to elevate the heart during vein graft emplacement. Such laparotomy sponges expose accumulating pericardial blood to a large, raw, cotton surface with an area approximately five times that of the CPB circuit (excluding the biocompatible oxygenator membrane). Because the reinfused blood has been exposed to this surface, the sponge becomes, in essence, a significant-though inapparent-part of the CPB circuit. Steps should be taken to either eliminate the sponge or to reduce the area of this foreign surface and make it more biocompatible.

Reinfusion of aspirated pericardial blood during CPB. Part II. Laparotomy sponges are hazardous parts of the CPB circuit?

Usually, cotton laparotomy sponges are discarded when they become blood soaked. During bypass surgery, however, they are often wrung out into the pericardial sac and the contents of the sac are aspirated into the cardiopulmonary bypass (CPB) circuit. After cardiopulmonary bypass, many patients give evidence of mental confusion, excessive bleeding, and systemic inflammatory response syndrome (SIRS). We believe that a possible cause is reinfusion of blood that has been activated by contact with laparotomy sponges and contains fibrin microemboli and thrombin. Thrombin production, soluble fibrin (SF) development, and fibrinogen disappearance were measured, over time, in model systems composed of increasing amounts (0-10-20 microL) of tissue factor (TF) in 4 ml of anticoagulated blood distributed through the interstices of a 49-cm(2) laparotomy sponge. Clotting occurred in all of the sponges. Clotting was accelerated by the presence of TF. Without TF, clotting occurred in an average of 28 min (range 17-39). With 10 and 20 microl TF, clotting occurred in 20 (range 11-27) and 13 (range 10-16) min, respectively. Thrombin at a level of approximately 16 NIHU/ml whole blood was present in several donors for 10-20 min after clotting was complete. If 120 ml of blood was to be wrung from a full-sized laparotomy sponge at this point in time, it could contain as much as 2000 units of thrombin.

Avoiding regression analysis pitfalls when calibrating white blood cell differential counters.

Method comparison and calibration are important for clinical hematologists and for manufacturers of new methods and instruments. Automation of the white blood cell (WBC) differential count poses unique problems in this regard because the reference method is a microscopic manual cell count of 200 WBCs. A highly variable reference method can obscure the true relation between the reference and the test methods and reduce sensitivity to miscalibration. A computer model based on published cell distributions and on the statistical properties of proportions was created to enable testing of alternative approaches to calibration. Ordinary linear regression (OLR) and standard Deming regression (SDR) were applied to a comparison of the results obtained with a virtual cell counter (counting 2,000 WBCs) and that obtained with a virtual 200-cell reference method for monocytes (mean population percentage of 7.1%) and lymphocytes (mean population percentage of 29%). Both regression approaches can misrepresent the known mathematical relation between the methods. This problem almost always occurs when OLR is used and the proportion of the cell of interest is low, as in the case of monocytes (7.1% of the total count). OLR comparisons, however, introduce significant error even for cell counts of 29%. SDR performed much better for both cell types.

The effect of three commercial coaching courses on Step One USMLE performance.

BACKGROUND: A substantial industry exists to provide formal review courses for Step 1 of the United States Medical Licensing Examination (USMLE). There are limited data on the usefulness of these courses. AIM: To determine whether or not student participation in a commercial coaching course improves performance on Step 1 of the USMLE. METHODS: Scores achieved by 468 students on the National Board of Medical Examiners (NBME) Comprehensive Basic Science Examination (CBSE) were used to predict a score on Step 1 of the USMLE. The NBME is the organisation that prepares and administers the USMLE. Predicted USMLE scores were then regressed against the actual scores achieved by the students on Step 1. The students were divided into 2 groups: those who took a 3-4-week live commercial coaching course and those who studied on their own. RESULTS: The regression lines for the 369 students who studied on their own and the 99 students who took a commercial coaching course were statistically indistinguishable. The analysis was powerful enough to have picked up a difference of 1% on average (P = 0.05) or 2 questions out of the 350 constituting Step 1 of the USMLE. Neither the students who performed above average nor those who performed below average on the CBSE improved their performance on Step 1 as a result of the coaching courses. CONCLUSIONS: Students who take a live, 3-4-week commercial coaching course to improve performance on Step 1 of the USMLE do not achieve higher scores than students who study on their own. Students should strongly consider whether or not a substantial investment in time and money for a commercial coaching course is justified in the light of such meagre returns.

Rapid-SF: a rapid whole-blood screen for soluble fibrin monomer.

Thrombosis accounts for a high proportion of disability and death in the West. Although soluble fibrin (SF) assays have been shown to be good predictors of thrombosis, current quantitative assays are too complex or lengthy to provide timely results, while simpler methods are qualitative and lack sensitivity. We here describe a rapid, new, protamine-based whole-blood screening method for SF which is quantitative and suitable for point-of-care use. Citrated whole blood is mixed with reagent under controlled conditions and the time until development of an SF precipitate is measured. Negative samples remain precipitate-free for 300 seconds. Strongly positive samples develop precipitate in as little as 10 seconds. SF times are mathematically converted to arbitrary SF units (SFU). This Rapid-SF test provides a simple and reliable means of detecting the presence of SF, and is well-suited for whole-blood rapid screening in the emergency department, operating room or clinic.

Control of analyzer slope and intercept in the measurement of packed red cell volume (PCV): part II.

A majority of packed cell volume (PCV) assays performed in the United States are performed by multichannel hematology analyzers rather than by centrifugation of blood samples in glass capillary tubes. PCV results from both analytical approaches were compared to the recently described ICSH reference method to determine the effect of assay slope on clinical utility. Though not controllable by the end user, the slope of the PCV assay versus the reference was satisfactory when performed in centrifuged glass capillaries. In the small sample of multichannel analyzers surveyed, only two of six machines functioned as well in this respect as did the glass capillaries. The worst of the unsatisfactory multichannel analyzers would have identified correctly only about half of the anemic and the polycythemic patients in the 12,623 males surveyed in the Third National Health and Nutrition Examination Survey (NHANES III). The performance on the females in this U.S. Department of Health and Human Services study would have been similarly poor. It is important that manufacturers of multichannel analyzers attend to the slopes of their PCV assays if the results are to be diagnostically useful.

Control of analyzer slope and intercept in the measurement of packed red cell volume (PCV): part I.

Five multichannel hematology analyzers (four different models) were evaluated to determine the ability of analyzers to correctly measure packed cell volume (PCV) values across the clinically relevant range. A sixth analyzer (fifth model) was subsequently included using archival data. Twenty-two different blood samples with PCVs ranging from 0.2 to 0.6 L/L were run in duplicate on each analyzer. Duplicate reference PCV determinations were also done using the recently described ICSH-recommended reference method. Simultaneously, a simpler "surrogate reference" procedure was performed in duplicate. Analyzer values were compared to ICSH reference and to surrogate reference PCV values using Deming regression analysis. Compared to either reference, all analyzers showed a slope of less than 1.0, thereby overestimating the PCV at 0.2 L/L and underestimating the PCV at 0.6 L/L to some extent. This outcome was surprising. Deming regression slopes would be expected to average at 1.0. Instruments whose slopes are significantly less than 1.0 will show decreased sensitivity to both anemic and polycythemic patients. We know of no inherent reason why multichannel analyzers should behave in this fashion.