ENDOTHELIAL INFLAMMATION AND DYSFUNCTION INDUCED BY SARS-COV-2 SPIKE PROTEIN 1 INVOLVE ADAM17 AND INTERFERON ACTIVATION PATHWAYS

Objective: COVID19 is associated with vascular inflammation. IFN-alpha (IFNa) and IFN-lambda3 (IFNl3) are potent cytokines produced in viral infections. Their effects involve interferon-stimulated genes (ISGs) and may influence expression of angiotensin-converting enzyme 2 (ACE2), the receptor for S-protein (S1P) of SARS-CoV-2. We hypothesized that S1P-induced immune/inflammatory responses in endothelial cells (EC) are mediated via IFN-activated pathways Design and methods: Human ECs were stimulated with S1P (1 mg/mL), IFNa (100ng/mL) or IFNl3 (100IU/mL). Because ACE2, ADAM17 and TMPRSS2 are important for SARS-CoV-2 infection, we used inhibitors of ADAM17 (marimastat, 3.8 nM), ACE2 (MLN4760, 440pM), and TMPRSS2 (camostat, 50 mM). Gene and protein expression was investigated by real-time PCR and immunoblotting, respectively. Vascular function was assessed in mesenteric arteries from wild-type (WT) normotensive and hypertensive (LinA3) mice and in ISG15-deficient (ISG15KO) mice. Results: S1P increased expression of IFNa (3-fold), IFNl3 (4-fold) and ISGs (2-fold) in EC (p < 0.05). EC responses to IFNa (ISG15: 16-fold) were greater than to IFNl3 (ISG15: 1.7-fold) (p < 0.05). S1P increased gene expression of IL-6 (1.3-fold), TNFa (6.2-fold) and IL-1b (3.3-fold), effects that were amplified by IFNs. Only the ADAM17 inhibitor marimastat inhibited S1P effects. IFNa and IFNl3 increase protein expression of ADAM17 (27%) and TMPRSS2 (38%). No changes were observed on ACE2 expression. This was associated with increased phosphorylation of Stat1 (134%), Stat2 (102%), ERK1/2 (42%). EC production of IL-6 was increased by IFNa (1,230pg/mL) and IFNl3 (1,124pg/mL) vs control (591pg/mL). Nitric oxide generation and eNOS phosphorylation (Ser1177) were reduced by IFNa (40%) and IFNl3 (40%). Vascular functional responses demonstrated that endothelium-dependent vasorelaxation (% Emax) in vessels from WT-mice stimulated with IFNa (67%) and IFNl3 (71%) were reduced vs control (82%) (p < 0.05). Responses were not altered in vessels from ISG15KO mice. Increased contraction was observed only in vessels from hypertensive mice treated with IFNa (9.1 ± 0.5mN vs control: 7.3 ± 0.3mN) (p < 0.05). Conclusions: In ECs, S1P, IFNa and IFNl3 increased ISG15 and IL-6 by mechanisms dependent on ADAM17. IFNs amplifies endothelial cell inflammatory responses and induced vascular dysfunction through ISG15-dependent mechanisms, with augmented effects in hypertension. Our findings demonstrate that S1P induces immune/inflammatory responses that may be important in endotheliitis associated with COVID-19. This may be especially important in the presence of cardiovascular risk factors, including hypertension.

ACE2 IS INVOLVED IN SARS-COV-2 INDUCED ENDOTHELIAL CELL INFLAMMATION INDEPENDENT OF VIRAL REPLICATION

Objective: COVID-19 association with cardiovascular disease is thought to be due to endothelial cell inflammation. ACE2 interactions with SARS-CoV-2 spike protein S1 subunit is important to viral infection. Here we questioned whether SARS-CoV-2 induces vascular inflammation via ACE2 and whether this is related to viral infection. Design and Methods: Human microvascular endothelial cells (EC) were exposed to recombinant S1p (rS1p) 0.66 ug/mL for 10 min, 5 h and 24 h. Gene expression was assessed by RT-PCR and levels of IL6 and MCP1, as well as ACE2 activity, were assessed by ELISA. Expression of ICAM1 and PAI1 was assessed by immunoblotting. ACE2 activity was blocked by MLN4760 (ACE2 inhibitor) and siRNA. Viral infection was assessed by exposing Vero E6 (kidney epithelial cells;pos ctl) and EC to 105 pfu of SARS-CoV-2 where virus titre was measured by plaque assay. Results: rS1p increased IL6 mRNA (14.2 ± 2.1 vs. C:0.61 ± 0.03 2-ddCT) and levels (1221.2 ± 18.3 vs. C:22.77 ± 3.2 pg/mL);MCP1 mRNA (5.55 ± 0.62 vs. C:0.65 ± 0.04 2-ddCT) and levels (1110 ± 13.33 vs. C:876.9 ± 33.4 pg/mL);ICAM1 (17.7 ± 3.1 vs. C:3.9 ± 0.4 AU) and PAI1 (5.6 ± 0.7 vs. C: 2.9 ± 0.2), p < 0.05. MLN4760, but not rS1p, decreased ACE2 activity (367.4 ± 18 vs. C: 1011 ± 268 RFU, p < 0.05) and blocked rS1p effects on ICAM1 and PAI1. ACE2 siRNA blocked rS1p-induced IL6 release, ICAM1, and PAI1 responses as well as rS1p-induced NFkB activation. EC were not susceptible to SARS-CoV-2 infection, while the virus replicated well in Vero E6. Conclusion: rS1p induces an inflammatory response through ACE2 in endothelial cells;an effect that was independent of viral infection.

Venous Thromboembolism in Patients Hospitalized for Sars-Cov-2: A Systematic Review

Introduction: The Coronavirus Disease 2019 (COVID-19), caused by SARS-CoV-2 virus, according to stipulations, claimed about 18,2 million lives until December 31, 2021. The COVID-19 generates a hypercoagulability state. Therefore, the main guidelines recommend thromboprophylaxis in hospitalized COVID-19 patients. However, doubts remain about the use of Low-Molecular-Weight Heparin (LMWH). Aim(s): To analyze the incidence of venous thromboembolism in hospitalized patients with COVID-19 in the regime of thromboprophylaxis with LMWH. Method(s): This systematic review was conducted following the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) rules. EMBASE and PubMed were the selected databases. The following health descriptors were used: "venous thromboembolism" "COVID-19", "coronavirus disease 2019", "anticoagulant"and "anticoagulants agent". Two authors independently performed a systematic literature search in databases. A search encompassed that was published between 01/01/2019 until 31/12/2021. The studies eligible for inclusion had a population over 18 years old, with confirmed diagnosis of COVID-19, hospitalized, and all population did use exclusively LMWH Articles were excluded if they did not provide data about Prophylactic Anticoagulation Regimen (PAR) and if they are case reports. Two authors (E.O.C and J.R.N.P) realized the data collection process, in an independent form. Result(s): Our search found 202 articles, of which 80 were duplicates. A total of 122 titles were excluded after reading the title and . Therefore, 17 articles were selected for reading the full-text. In this step, 5 titles were excluded, because they don't give information about the PAR and 3 because they don't use LMWH exclusively. So, we selected 9 titles. A total of 1614 participants, from 8 countries, submitted to prophylaxis with LMWH. Being that 160 participants (9.91%) had a venous thromboembolic event. 6 articles gave information about bleeding that occurred with 43 participants (3%). Discussion(s): According to our article, the presence of venous thrombotic events was found in 18% to 37% of hospitalized patients who had not used prophylactic anticoagulation. The greater susceptibility to thrombotic events in patients infected with SARS-CoV-2 remains under study, however some hypotheses have already been pointed out. Among them, virus entry into cells through angiotensin S receptors (ACE2), widely expressed in lung cells, myocardium and other endothelial cells. Injury to these cells triggers an inflammatory process with the release of more cytokines and consequent pro-coagulant stimulus. Added to this factor is the ability of the virus to bind to ACE2 receptors to cause abnormal activation of the renin-angiotensin axis, leading to platelet aggregation and increased thromboembolic risk. The group of patients positive for VTE and infected by SARS-CoV-2 had a decrease in lymphocyte count and an increase in serum levels of D-Dimer. Conclusion(s): The proportion of thrombotics events in hospitalized patients diagnosed with COVID-19 is 18% to 37%, without prophylactic anticoagulation. Our research demonstrated that less than half of the population submitted to prophylaxis anticoagulation evolved to venous thromboembolic events. Copyright © 2022

Atypical Hemolytic Uremic Syndrome due to Factor H Antibodies Following COVID-19

Introduction: Atypical hemolytic uremic syndrome (aHUS) is a form of thrombotic microangiopathy (TMA) characterized by dysregulated complement activation. Antibodies to factor H (anti-FH), a regulator of the alternative complement pathway, are a recognized cause of aHUS particularly in children. We present the case of an elderly patient who developed aHUS following COVID-19. Case Description: A 74-year-old male presented with weakness, petechial rash involving extremities and diarrhea for 2 weeks. Prior history included hepatitis C infection status-post treatment 2 years ago with associated cirrhosis. Three weeks ago, the patient had been diagnosed with COVID-19. His symptoms of sore throat, cough and fever had by now resolved. Initial investigations showed leukocytosis and AKI with an active urinary sediment and nephrotic range proteinuria (Fig 1). Hemoglobin and platelets were normal and a blood smear was negative for hemolysis. Imaging revealed small bowel enteritis suggestive of an infectious or vasculitic process. Infectious workup returned negative. Autoimmune serologies revealed a borderline positive ANA, low C3 and low-normal C4. Renal biopsy revealed diffuse endothelial injury with swollen endothelial cells, focal mesangiolysis and glomerular basement membrane duplication. Hence, pulse dose steroids were started and complement function panel sent. Soon after steroid initiation, the patient's renal function, leukocytosis and rash improved. Ultimately, complement testing returned positive for anti-FH. At follow-up, renal function had returned to baseline with continued steroid taper. Discussion(s): COVID-19 is associated with TMA likely due to endothelial toxicity or complement pathway dysregulation. Our patient had no prior history of renal or hematologic disease. Given the chronology of events, it is likely that COVID-19 triggered formation of anti-FH, in turn leading to development of aHUS in our patient.

Identification of a red cell hemostatic mechanism causing COVID-19 microvascular obstruction

Background: Severe COVID-19 infection is associated with a wide spectrum of clinical manifestations, leading to systemic thromboinflammation and multiorgan dysfunction. The primary cause of multiorgan damage is widespread endothelial injury, leading to microangiopathy and organ ischemia. The molecular mechanisms by which ischemic endothelial cells causes microvascular obstruction remains ill defined. Aim(s): Identification of distinct microvascular occlusion mechanisms in COVID-19. Method(s): The microvasculature of multiple organs from patients dying from COVID-19, myocardial infarction or stroke were analyzed by H&E, immunohistochemistry, SEM and CLEM. Animal models of gut ischemia and stroke were also examined. Intravital confocal microscopy examined endothelial injury and microvascular obstruction mechanisms mediated by platelets, red cells and fibrin. Result(s): We demonstrate the existence of a distinct microvascular hemostatic mechanism mediated by hemolyzed red blood cells (RBC), independent of platelets and fibrin. Extensive RBC hemolysis was apparent in the microvasculature of COVID-19 patients and in humans with major organ ischemia, leading to widespread microvascular obstruction. This RBC hemostatic mechanism was triggered by organ ischemia and associated with localized accumulation of hemolyzed RBCs at sites of endothelial necroptosis. RBC hemolysis was impaired in animals lacking the necroptosis mediator, MLKL or the C9 component of complement, indicating the involvement of cell intrinsic and extrinsic membrane lytic processes. Intravital microscopy revealed that the RBC hemostatic mechanism was triggered by the fragmentation of lyzed RBCs and the deposition of RBC membranes on the surface of dying endothelial cells, forming an endovascular sealant that prevents interstitial bleeding. Conclusion(s): Our studies demonstrate the existence of a previously unidentified microvascular hemostatic mechanism mediated by hemolyzed RBCs. Dysregulation of this hemostatic mechanism is linked to microvascular obstruction and bleeding in COVID-19 and ischemic diseases.

Recovery after COVID-19 and persisting endothelial cell activation and hypercoagulability-The prospective observational ROADMAP-post COVID-19 study

Background: Hypercoagulable state and endothelial cell activation are common alterations in patients with COVID-19. Nevertheless, the hypothesis of persistent hypercoagulability and endothelial cell activation following recovery from COVID-19 remains an unresolved issue. Aim(s): To investigate the persistence of endothelial cell activation and hypercoagulability after recovery from COVID-19. Patients/ Methods. COVID-19 survivors (n = 208) and 30 healthy individuals were enrolled in this study. Method(s): The following biomarkers were measured: Procoagulant phospholipid-dependent clotting time (PPL-ct), D-Dimer, fibrin monomers (FM), free Tissue factor pathway inhibitor (free-TFP) I, heparinase, and soluble thrombomodulin (sTM). Antibodies against SARS-CoV- 2 (IgG and IgA) were also measured. Result(s): The median interval between symptom onset and screening for SARS-CoV- 2 antibodies was 62 days (IQR = 22 days). Survivors showed significantly higher levels of D-Dimers, FM, TFPI, and heparanase as compared to that of the control group. Survivors had significantly shorter PPL-ct. Elevated D-dimer was associated with older age. Elevated FM was associated with female gender. Elevated heparanase was independently associated with male gender. Decreased Procoag-PPL clotting time was associated with female gender. One out of four of COVID-19 survivors showed increase at least one biomarker of endothelial cell activation or hypercoagulability. Conclusion(s): Two months after onset of COVID-19, a significant activation of endothelial cells and in vivo thrombin generation persists in at least one out of four survivors of COVID-19. The clinical relevance of these biomarkers in the diagnosis and follow-up of patients with long COVID-19 merits to be evaluated in a prospective clinical study.

Expression of extracellular vesicles in plasma and cerebrospinal fluid is associated with the procoagulant state in patients with neurological complications of COVID-19

Background: Extracellular vesicles (EVs) have been described to be associated with hemostatic disturbances in different clinical settings. (Table Presented) Aims: In this study we have investigated EVs in plasma from patients with COVID-19 in relation to the activation of coagulation. Moreover, we assessed the presence of EVs in the cerebrospinal fluid (CSF) of patients suffering neurological symptoms during COVID-19. Method(s): Eighteen patients with COVID-19 and neurological symptom admitted to the Uppsala University Hospital, Sweden were included. Median age of the patients was 64 (39-85) years, 39% were women. Twenty-one aged matched healthy individuals were included as controls. Informed consent was obtained. EVs derived from platelets (CD61+), neutrophils (MPO+) and endothelial cells (CD51/61+);together with EVs-expressing phosphatidylserine (PS+), tissue factor (CD142+), complement components C5b-9 (TCC+), C3a and C4d were determined by flow cytometry. Overall hemostasis potential (OHP), including overall coagulation potential (OCP) and overall fibrinolytic potential (OFP) were measured and scanning electron microscopy of fibrin clots was performed. Result(s): Significantly higher OCP (p < 0.01) and OHP (p < 0.001) and lower OFP (p < 0.05) were observed in Covid-19 patients (p < 0.05), compared to controls. Denser fibrin structure was found in COVID-19 patients (Figure 1). Increased concentrations of PS+, MPO+, CD61+ and TCC+ EVs were found in plasma from Covid-19 patients compared to healthy controls, and the concentrations of PS+, CD61+ and TCC+ EVs were positively correlated with OCP and OHP in Covid-19 patients. The presence of CD61+, CD51/61+, MPO+ EVs and EVs exposing PS and TCC was identified in the CSF obtained from 17 patients (Figure 2). Conclusion(s): Procoagulant state together with elevated levels of circulating EVs of different cell origin was found in patients with Covid-19. The unique finding of this study is the presence of EVs in CSF of Covid-19 patients with neurologic manifestations. EVs may represent potentially novel biomarkers of blood-brain barrier damage during SARS-COV- 2 infection.

Biomarkers in COVID-19: Association with disease severity and outcomes

Background: In coronavirus disease 2019 (COVID-19) the need for intervention increases with disease severity and a risk prediction model that incorporates biomarkers would be beneficial for identifying patients for treatment escalation. Aim(s): To investigate biomarkers changes associated with disease severity and outcomes (mortality, thrombosis). Method(s): COVID-19 patients were sampled between April 15 and May 31 2020. Disease severity was assessed by World Health Organization (WHO) ordinal scale. 132 systemic biomarkers were investigated by routine and multiplex assays and statistical analysis performed to characterise the biomarker profile of COVID-19 patients associated with disease severity, duration, survival and thrombosis. Result(s): The study enrolled 150 COVID-19 positive adults and 16 healthy volunteers. The average age was 64 years, 59% were male, 85% had co-morbidities, 33% had a thrombotic event, and 13% died. A cross comparative analysis of biomarkers identified 13 biomarkers common to severity, mortality and thrombosis with significant correlation;including endothelial dysfunction (VWF, tPA, TFPI), hypercatabolism (low albumin, Hb, FXIII) and inflammatory response (IL-8, Osteopontin). Similarly, 14 biomarkers associated with severity and mortality included pro-inflammatory cytokines and their receptors (sTNFRII, STNFRI, sIL2a, IL6, MIP1a), neutrophils (elevated WBC, Neutrophils, TIMP1) and tissue remodelling (SCGF, EG3A). Nine biomarkers common across severity and thrombosis were angiogenesis (VEGF, LYVE1, Follistatin), acute phase response (SAP, AGP) and clot formation (Fibrinogen and PAPs). Conclusion(s): The biomarker profile associated with poorer outcomes indicates an inflammatory response, endothelial cell disruption, hypercoagulability and hypercatabolism. This study has identified several biomarkers that may be useful indicators of disease severity and progression. Further work is needed to determine how these may be used to direct clinical management. (Figure Presented).

A role for Von Willebrand Factor-ADAMTS13 axis imbalance and abnormal angiogenesis in long COVID syndrome

Background: Acute COVID-19 is associated with marked endotheliopathy, VWF-ADAMTS13 axis imbalance and abnormal pulmonary angiogenesis. Persistent endotheliopathy and elevated VWF levels have also been reported in convalescent COVID-19 patients. Aim(s): We investigated the hypothesis that altered pulmonary microvascular architecture may persist in COVID-19 convalescence, resulting in ongoing endothelial cell (EC) activation and VWF-ADAMTS13 axis imbalance, possibly contributing to Long COVID pathogenesis. Method(s): 50 patients (median age 50 years, 60% male, median 68 days post acute COVID-19) were reviewed. Six-minute- walk tests (6MWT) were performed (median 6MWT distance 430m) and plasma samples collected. Plasma VWF:Ag and ADAMTS13 levels were measured by ELISA, and angiogenesis markers assessed by membrane-based antibody array. Result(s): Plasma VWF:Ag levels were significantly elevated in convalescent COVID-19 patients compared to controls (1.1 vs. 0.84 IU/ml;p = 0.004), with 30% (15/50) having VWF:Ag levels above the upper limit of normal. In contrast, plasma ADAMTS13 was significantly reduced in convalescent COVID-19 (median 467 ng/ml vs. 636 ng/ ml p < 0.001). ADAMTS13 levels were significantly lower in those who required hospitalization for acute COVID-19 compared with those managed as outpatients (median 454 ng/ml vs. 513 ng/ml, p = 0.04). Overall, the VWF/ADAMTS13 ratio was significantly elevated in convalescent COVID-19 compared with controls (2.1 vs. 1.1 p = 0.0002) and interestingly was elevated in patients with reduced 6MWT distance (distance >=430 m or <430 m: 1.8 vs. 2.4, p = 0.02). In total, 15 angiogenesis markers were elevated in convalescent COVID-19 compared to controls. An additional 17 angiogenesis (Figure Presented) markers were unique to convalescent COVID-19 and were not found in control plasma (Table 1). Conclusion(s): Collectively, these novel findings demonstrate that endotheliopathy is sustained for months following acute COVID-19 in some patients. As a result, plasma VWF levels are significantly increased;ADAMTS13 levels reduced, and there is ongoing dysregulation of angiogenesis. Further studies will be required to define whether these alterations play a role in Long COVID pathogenesis.

Inflammatory convalescent plasma to treat COVID-19: Impact of amotosalen/UVA pathogen reduction technology

Background: Blood product in therapeutic transfusion are now commonly acknowledged to present biologically active constituents during processes of preparation. In the midst of worldwide COVI-19 pandemic, preliminary evidence, suggest that convalescent plasma may lessen the severity of COVID-19, particularly concerning patients with profound B-cell lymphopenia and prolonged COVID-19 symptoms. Aim(s): This study examined the influence of photochemical pathogen reduction treatment (PRT) using amotosalen-HCl and UVA light vs untreated control convalescent plasma (n = 72 -paired samples) -cFFP. Method(s): This study investigated the soluble inflammatory factors: SCD40L, IFN-alpha, IFN-beta, IFN-gamma, IL-1 beta, IL-6, IL-8, IL-10, IL-18, TNF-alpha and ex-vivo inflammatory bioactivity on endothelial cells. Result(s): We observed that IL-8 concentrations were significantly decreased in cFFP w PRT, whereas IL-18 concentration was increased. We observed after activation with cFFP w PRT and w/o PRT no significant modulation of IL-6 released by endothelial cells. CD54 and CD31 expression in the presence of cFFP (w or w/o PRT) is close to negative controls, even if CD54 and CD31 were significant decreased in presence of cFFP w vs w/o PRT. Conclusion(s): It appears valuable to carry on investigations, of IL-18 and IL-8, on both the physiopathology of PRT convalescent plasma treated and post marketing clinical trials. Further research, including a careful clinical evaluation of CCP-treated patients, will be required to further define the clinical relevance of these findings.

Inflammatory markers and auto-Abs to type I IFNs in COVID-19 Convalescent Plasma

Background: COVID-19 convalescent plasma (CCP) contains neutralizing anti-SARS- CoV- 2 antibodies that may be useful as COVID-19 passive immunotherapy in patients at risk of developing severe disease. Aim(s): Such plasma from convalescent patients may also have additional immune-modulatory properties when transfused to COVID-19 patients. Method(s): CCP (n = 766) were compared to control non-convalescent plasma (n = 166) for soluble inflammatory markers, ex-vivo inflammatory bioactivity on endothelial cells, neutralizing auto-Ab to type I IFNs, and reported adverse events in the recipients. Result(s): CCP exhibited significantly higher IL-6 and TNF-alpha (0.531+/-0.04 vs 0.271+/-0.04;p = 0.0061 and 0.900+/-0.07 vs 0.283+/-0.07 pg/ml;p < 0.0001), respectively) and lower IL-10 (0.731+/-0.07 vs 1.22+/-0.19 pg/ ml, p = 0.0034) levels than control plasma. Other inflammatory markers as well as ex-vivo bioactivity did not differ significantly between CCP and control plasma. Neutralizing auto-Abs against type I IFNs were detected in 14/766 (1.8 %) CCP. They were not associated with reported adverse events when transfused (n = 14). Inflammatory markers and bioactivity in CCP with or without auto-Ab, or in CCP associated or not with adverse events in transfused patients, did not differ significantly. Overall, CCP exhibited moderately increased inflammatory markers compared to control plasma with no discernable differences in ex-vivo bioactivity. Auto-Ab to type I IFNs, detected in a small fraction of CCP, were not associated with reported adverse events or differences in inflammatory markers. Conclusion(s): Further defining the clinical relevance of these findings will require further studies including careful clinical evaluation of patients treated with CCP.

Endothelial cell activation, Weibel-Palade body secretion and enhanced angiogenesis in severe COVID-19

Background: Severe COVID-19 is associated with marked endothelial cell (EC) activation that plays a key role in immunothrombosis and pulmonary microvascular occlusion. However, the biological mechanisms through which SARS-CoV-2 causes EC activation and damage remain poorly defined. Aim(s): We investigated EC activation in patients with acute COVID-19, and in particular focused on how proteins stored within Weibel-Palade bodies (WPBs) may impact key aspects of disease pathogenesis. Method(s): 39 patients with confirmed COVID-19 were recruited. Weibel-Palade body biomarkers [von Willebrand factor (VWF), angiopoietin-2 (Ang-2) and osteoprotegerin (OPG)] and soluble thrombomodulin (sTM) levels were determined. In addition, EC activation and angiogenesis were assessed in the presence or absence of COVID-19 plasma incubation. Result(s): Markedly elevated plasma VWF:Ag, Ang-2, OPG and sTM levels were observed in acute COVID-19 patients. The increased levels of both sTM and WPB components (VWF, OPG and Ang-2) correlated with COVID-19 severity. Incubation of COVID-19 plasma with ECs triggered enhanced VWF secretion and increased Ang-2 expression (Figure 1). In keeping with the autopsy reports of intussusceptive angiogenesis, treatment with COVID-19 plasma also caused significantly increased EC angiogenesis (Figure 1). Conclusion(s): We propose that as COVID-19 develops, progressive loss of TM and increased sTM, as well as increased Ang-2 expression result in loss of EC quiescence, WPB exocytosis, and a local pro-angiogenic state.

Chorangioma of placenta: Single center analysis

Background: The most common benign neoplasms of the placenta are non-trophoblastic tumours. They include chorioangioma, teratoma, leiomyoma and hepatocellular adenoma. Chorioangioma is the most common subtype. The incidence of chorangioma is 0.5-1.0%. Small chorangiomas are clinically insignificant. Giant chorioangioma is rare tumours, measuring more than 4 cm in diameter and seen in association with elderly primi, twin pregnancy, hypertension, diabetics, and female fetus. Giant chorioangioma is associated with complications that can affect the mother, fetus, or neonate. Aim(s): This is a retrospective study of chorangioma cases seen at Institute of Pathology, Medical faculty, Skopje during a 10-year period from 2012 to 2021. Method(s): Macroscopic findings, gestational weeks at delivery, maternal age and outcome of pregnancy were evaluated. Histological and immunochistochemical analyses of the placental chorangiomas were preformed. Results & Conclusion(s): In period of 10 years 6 causes of placental chorangioma were clinically identified and histopathologically confirmed. Four cases were giant chorangiomas measuring from 8 to 13 cm. One of the cases was identified as chorangioma of the umbilical cord. Three of the cases shows potential maternal risk factors like primary infertility, extreme obesity and post COVID status. The median gestational age of delivery was 33 + 6 weeks and 39.6 weeks, respectively. The maternal age range was between 25 to 34 years. Four of the pregnancies had favourable outcome with no complications during and after birth. One was with foetal distress and one with premature delivery. Microscopic examination of the mass showed numerous proliferative thin walled capillaries lined by flattened endothelium and separated by fibrous stroma. This was further confirmed by IHC for CD34, which showed strong reactivity of endothelial cells.

CORONA VIRUS, ITS EFFECTS AND SOLUTION-A REVIEW

The year 2019 is a outbreak year during which the whole globe has suffered from Covid19 pandemic which has been spotted initially in China and later spread to the whole world;as a result of this viral disease, the whole world had shut down affecting billions of people but till today the Covid battle is on and people are suffering not only from this disease but also in terms of economy, starving being jobless etc. This paper briefs about Corona virus, its types, and structure;the replication and spreading of this virus, Covid19 detection methods, research on vaccination developed across the world to curb this virus;virus impact on various sectors, precautions to be taken to stay away from this virus and Ayurvedic remedy for it. The waves of corona had taken many lives on the globe & have its effect on life style of people. To curb this virus, prevention vaccination has to be found and we people must change in a way so that we could avoid future consequences for the upcoming generation. Keywords.

Pathophysiology and Management of Hypercoagulation in Infectious Diseases

Numerous systemic infections may have hypercoagulation as one of the complications, which may range from asymptomatic presentation of elevation of biochemical markers of coagulation such as that of fibrin and thrombin generation, to a much severe, symptomatic, life-threatening, disseminated intravascular coagulation (DIC), which results in the formation of thrombi in the microvasculature of various organs. This phenomenon contributes to increase in morbidity and mortality in various infectious diseases. The current review discusses various mechanisms of hypercoagulation during infections such as tissue factor activation, endothelial cell activation, inhibition of physiological anticoagulant pathways, and fibrinolysis inhibition. The review also discusses pathophysiological changes in the coagulation system and its management in the recent pandemic of COVID-19. The article also discusses role of various parenteral and oral anticoagulants in the management of infectious diseases. The review provides clinical data on various anticoagulants used during hospitalization and extended prophylaxis for the management of venous thromboembolism in various infections. Methodology Because this is a review of published literature and no humans or animals were involved, ethical committee approval was not required and patient consent was not required.

CLOTTING AND COVID-19: IT IS NOT ALWAYS VTE

SESSION TITLE: COVID-19 Case Report Posters 2 SESSION TYPE: Case Report Posters PRESENTED ON: 10/19/2022 12:45 pm - 01:45 pm INTRODUCTION: Coronavirus disease 2019 (COVID-19) is an enveloped positive stranded RNA virus that affects multiple organ systems in the body. COVID-19 venous and thromboembolic events are well documented;however, few reports of arterial thrombosis exist. Arterial embolism is reported to occur in one to five percent of patients. We present a case of a patient who experienced arterial thromboembolism. CASE PRESENTATION: A 38-year-old woman with a history of diabetes, hypertension, and recent COVID-19 pneumonia three weeks prior presented to the hospital for lower extremity weakness, paresthesias, and pain in her bilateral lower extremities. Upon examination, she was found to have bilateral cold feet, lack of sensation to toes or plantar aspect of feet, nondopplerable pedal or dorsalis pedis pulses bilaterally, dopplerable femoral pulses bilaterally. A CT angiogram of the abdomen with bilateral runoff revealed distal abdominal aortic and bilateral iliac artery thrombus, thrombus in bilateral runoff arteries. She was evaluated by vascular and started on a heparin drip. She underwent bilateral iliofemoral thromboembolectomy and bilateral iliac stents. Surgery recommended allowing demarcation in the outpatient setting, however, due to intractable pain vascular surgery determined that bilateral below the knee amputations were necessary. She underwent testing for possible hypercoagulable state and was found to have an elevated cardiolipin antibody and lupus anticoagulant (LA) screen positive which are reported in 50% of critically ill COVID-19 patients, though the clinical or pathological value of these results are unclear at this time. DISCUSSION: Several mechanisms for hypercoagulability in COVID-19 infection have been postulated. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) binds to host angiotensin converting enzyme 2 (ACE2) proteins. ACE2 receptors can be found throughout multiple organs and specifically on endothelial cells. ACE2 maintains the endothelial integrity of vessels. Coagulation testing in COVID-19 patients reveal increased prothrombin, activated partial thromboplastin time (aPTT), platelet counts, fibrinogen levels. Increased inflammation and cytokine release lead to a hypercoagulable state. Studies have shown that cardiolipin antibodies and LA positivity are higher in patients with COVID-19 which predispose patients to venous and arterial thrombosis. CONCLUSIONS: Venous thrombosis is often considered in patients with COVID-19 and clotting complications, however, due to the growing number of case reports regarding arterial thrombosis – arterial complications must be considered in the differential. Further research regarding the mechanism of arterial thrombosis are required to better understand the pathogenesis and develop targeted therapies to prevent occurrence of arterial thrombosis. Reference #1: Cheruiyot I, Kipkorir V, Ngure B, Misiani M, Munguti J, Ogeng'o J. Arterial Thrombosis in Coronavirus Disease 2019 Patients: A Rapid Systematic Review. Ann Vasc Surg. 2021;70:273-281. doi:10.1016/j.avsg.2020.08.087 Reference #2: Current and novel biomarkers of thrombotic risk in COVID-19: a Consensus Statement from the International COVID-19 Thrombosis Biomarkers Colloquium ;Nature Reviews Cardiology Reference #3: Taha, M., & Samavati, L. (2021). Antiphospholipid antibodies in COVID-19: a meta-analysis and systematic review. RMD open, 7(2), e001580. https://doi.org/10.1136/rmdopen-2021-001580 DISCLOSURES: No relevant relationships by Gretchen Grosch No relevant relationships by Stephanie Link No relevant relationships by Soophia Naydenov No relevant relationships by Tanner Wallen

A RARE CASE OF COVID-19-INDUCED AORTITIS

SESSION TITLE: Post-COVID-19 Infection Complications SESSION TYPE: Case Report Posters PRESENTED ON: 10/17/2022 12:15 pm - 01:15 pm INTRODUCTION: Aortitis is a type of vasculitis that refers to inflammation of the aortic wall. Most common causes are rheumatologic disorders and bacterial infection. Here, we report a viral cause of aortitis induced by COVID-19. CASE PRESENTATION: A 73 year old female with history of coronary artery disease, chronic kidney disease, COPD, hypertension, pulmonary embolism on Eliquis and abdominal aortic aneurysm (AAA) status post repair presented with acute hypoxemia secondary to Covid-19 pneumonia. Of note, patient was vaccinated against COVID-19. CT abdomen at admission demonstrated a known infrarenal AAA with increased degree aortic wall thickening, concerning for aortitis. Aortitis was initially thought to be due to endovascular infection from possible bacteremia rather than surgical site infection as the patient had the AAA repair almost a year prior. Given that bacterial aortitis could result in death, blood cultures were obtained and she was started on Vancomycin and Rocephin. Rapid Plasma Reagin was ordered to rule out syphilis. She had titer of 1-2 which was thought to be false positive as fluorescent treponemal antibody absorption test was negative. After blood cultures and inflammatory markers were negative, antibiotics were discontinued. Aortitis was attributed to COVID. Patient was treated with DEXA-ARDS protocol. Repeat CT abdomen after 8 days no longer showed gross evidence of aortitis. Patient was discharged home with home healthcare. DISCUSSION: Aortitis, a rare complication of COVID-19, has been reported. A proposed mechanism of this pathogenesis involves acute endotheliitis, where endothelial cells infected by virions become infiltrated by neutrophils and mononuclear cells, leading to apoptosis and lymphocytic endotheliitis [1]. Later, these arteries move through the stages of an accelerated karyolysis, accumulation of apoptotic bodies, caspase granules, and fibrinoid substances, leading to leukocytoclastic vasculitis [1]. This inflammatory reaction is followed by deposition of polyclonal antigen-antibody immune complexes, which is a type III hypersensitivity acute vasculitis [2]. Our patient's history of AAA repair predisposed her to increased endothelial dysfunction. After other bacterial infectious causes and post-surgical complications were ruled out, patient was treated with steroids. Most cases of COVID induced aortitis have been treated with prednisone that required treatment for around 1 month [3]. Here, we present a patient treated with DEXA-ARDS with resolution of aortitis. CONCLUSIONS: Due to the novelty, the understanding of exact pathogenesis and long term effects of COVID-19 induced aortitis is limited. However, our case does serve to support prior case reports of COVID-19 aortitis that showed clinical and radiologic response to steroids. Further research is warranted to diagnose and treat aortitis in order to avoid life threating complications. Reference #1: Varga, Zsuzsanna, et al. "Endothelial cell infection and endotheliitis in COVID-19.” The Lancet 395.10234 (2020): 1417-1418. Reference #2: Roncati, Luca, et al. "Type 3 hypersensitivity in COVID-19 vasculitis.” (2020): 108487-108489. Reference #3: Dhakal, Pravash et al. "Aortitis in COVID-19.” IDCases vol. 24 (2021): e01063. doi:10.1016/j.idcr.2021.e01063 DISCLOSURES: No relevant relationships by Jessica Lee No relevant relationships by Thong Ngo No relevant relationships by Marrian Sedrak No relevant relationships by Hena Yagnik

COVID-19-ASSOCIATED PULMONARY MUCORMYCOSIS PRESENTING AS SPONTANEOUS PNEUMOTHORAX: A RARE PRESENTATION

SESSION TITLE: COVID-19 Case Report Posters 1 SESSION TYPE: Case Report Posters PRESENTED ON: 10/17/2022 12:15 pm - 01:15 pm INTRODUCTION: Mucormycosis is an angio-invasive fungal infection with substantial morbidity and mortality. While diabetes and immune suppression remain well-known risk factors for mucormycosis, COVID-19 is now emerging as its independent predictor. CASE PRESENTATION: A 43-year-old male, with a history of hyperlipidemia and alcoholism, presented to the hospital with complaints of progressive dyspnea on exertion, productive cough, intermittent fever, anorexia, and chest pain over the course of 2 weeks. About 5 weeks prior to the current presentation, he was tested positive for COVID-19 by a polymerase chain reaction (PCR) based test and remained in quarantine at home. He was not vaccinated against COVID-19. He had no known immunosuppressive disease. On initial examination, he was ill-appearing and had a temperature of 101 F, blood pressure 138/83 mmHg, respiratory rate 22/minute, pulse 102/minute, and saturation of 91% on 2 L nasal cannula oxygen. A computerized tomography (CT) scan of the chest revealed small bilateral pneumothorax (2 cm and 5mm) along with extensive ground-glass opacifications in all lobes. In the next 24 hours, the right-sided pneumothorax progressed to tension pneumothorax requiring pigtail pleural drainage catheter placement. The drained pleural fluid had more than 100,000/uL total nucleated cells (91% neutrophils, 2% lymphocytes, and 1% eosinophils) and ultimately cultures grew Rhizopus spp. He was started on intravenous liposomal amphotericin-B infusion (5 mg/kg daily). On hospital discharge, he was switched to oral posaconazole (started with loading 300 mg delayed-release tablet twice a day, followed by 300 mg dosing of delayed-release posaconazole tablets daily) to complete the long term treatment course. DISCUSSION: Most of the reported cases of mucormycosis in COVID-19 were in patients with either diabetes or receiving steroids. This is a rare presentation of COVID-19–associated pulmonary mucormycosis (CAPM) as spontaneous pneumothorax, in the absence of known immunosuppression history. COVID-19 results in a considerable increase in cytokines, particularly interleukin-6 (IL-6), which increase free iron by increasing ferritin levels due to increased synthesis and decreased iron transport. Also, concomitant acidosis increases free iron by reducing the ability of transferrin to chelate iron and this available iron becomes a considerable resource for mucormycosis. [1] Also, Mucorales adheres to and invades endothelial cells by specific recognition of the host receptor glucose-regulator protein 78 (GRP-78). Acidosis associated with severe COVID-19 triggers GRP-78 and fungal ligand spore coating homolog (CotH) protein expression on endothelial cells, both contributing to angioinvasion, hematogenous dissemination, and tissue necrosis. [2] CONCLUSIONS: Mucormycosis can present as spontaneous pneumothorax after recent COVID-19 and clinicians should be aware of rare clinical presentation. Reference #1: Singh AK, Singh R, Joshi SR, et al. Mucormycosis in COVID-19: A systematic review of cases reported worldwide and in India. Diabetes Metab Syndr Clin Res Rev 2021;15:102146. doi:10.1016/j.dsx.2021.05.019 Reference #2: Baldin C, Ibrahim AS. Molecular mechanisms of mucormycosis—The bitter and the sweet. PLOS Pathog 2017;13:e1006408. doi:10.1371/journal.ppat.1006408 DISCLOSURES: No relevant relationships by Faran Ahmad No relevant relationships by AYESHA BATOOL No relevant relationships by Zachary DePew No relevant relationships by Neil Mendoza

The aftermath of the COVID-19 pandemic on access to corneal transplant procedures in Morocco

Purpose : The COVID-19 pandemic led to disrupted corneal supply to countries tributary to foreign eye bank donations. The authors carried out the first retrospective, observational study to assess the impact of the pandemic on corneal tissue supply and quality in Morocco, by comparing the surgical patterns during the pre-pandemic time and the first year of the pandemic at the only tertiary eye center performing keratoplasty in Morocco. Methods : A retrospective analysis comparing the years 2019 and 2020 ( January 1st to December 31st) was performed based on electronic health records database from the department of Ophthalmology at Cheikh Zaid International Hospital.Quantitative analysis evaluated the numbers of keratoplasties performed, the number procedures for therapeutic or tectonic purposes, the numbers of delivered corneal tissue, numbers of patients on the waiting list. Quality assessment of the grafts was based on the mean endothelial cell density count provided by the supplier, prior to corneal surgery. The comparison was based on a Chi2 test with p<0.05 considered statistically significant. Statistical study was performed using Jamovi (The Jamovi project2021,Version 1.6[Computer Software]) Results : Out of a total of 345 patients registered on the corneal transplant list on 2019, 252 patients underwent keratoplasty (73%) with 4,4 % for tectonic or therapeutic purposes, whereas out of 507 patients on the list on 2020, 160 received keratoplasty (31,55%) with 13,7% in a context of emergency. The waiting list grew from 293 patients in 2019 to 453 in 2020 marking an increase of 21,4 %. A statistically significant decrease in the number of corneal tissue received at our center was noted, with a mean of 4,8 (+/-4,23) per week in 2019 and 2,9 (+/-3,02) per week in 2020 (p=0,02).All corneal tissue was exclusively provided by three US eye banks. Mean endothelial cell density count prior to surgery was not statistically different between 2019 ( 2699 c/mm2 +-255) and 2020 ( 2668 c/mm2266),(p= 0,22). Conclusions : These results reflect the imbalance between corneal demand and supply creating a disruption of the corneal tissue chain in Morocco. Given the exceptional activity of the local Eye bank of Morocco due to numerous challenges, and the scarcity of centers that perform keratoplasty, Morocco witnesses a gap between corneal blindness and access to keratoplasty which was further enlarged during the pandemic.