ABSTRACT
Background. Data regarding cell-mediated immunological differences in children across COVID-19 clinical spectrum are limited. We prospectively investigated Spike-protein specific cellular immunity in children with symptomatic COVID-19 or MIS-C, by single cell cytokine expression profiling. Methods. PBMCs fromchildrenwithMIS-C, symptomaticCOVID-19 (onemonth after hospitalization) and healthy controls were isolated prospectively. Cell suspensions were divided into two quantitatively equal samples (a negative control-unstimulated and a positive-stimulated).Cells stimulationwas performed usingSARS-CoV-2 Spike antigenic peptidesmix (Peptivator SARS-CoV-2 Prot-S). Cells of each sample were stained with fluorochrome monoclonal antibodies against 8 surface markers (CD3, CD4, CD8, CD14, CD19, CD137, CD197, CD45RA) and 6 intracellular markers (IL-4, IL-2, IL-17, IFN-gamma, TNF-alpha, Granzyme B). Viability was assessed by 7AAD. Stained cell preparations were analysed using 13 colour Flow Cytometry (DX Flex, Beckman Coulter). Flow cytometric analysis was performed using Kaluza 2.1 Software. Results. Sixteen children (4 MIS-C, 8 post-COVID-19 and 4 healthy controls) were included in the study. The mean age (+/-SD) of the participants was 11.22 years (+/-3.48). Children with MIS-C had significantly higher mean (+/-SD) values of CD8+IFN-gamma/million CD3+ [226.68 (134.92) vs 45.43 (57.28);P: 0.033] and median (IQR) of CD8+IFN-gamma/ml [156.38 (184.13) vs 26.34 (82.36);P: 0.033] compared to symptomatic COVID-19 children. Compared to healthy controls, MIS-C patients had significantly higher median (IQR) values of CD4+IL-2/million CD3+ [923.12 (3777.95) vs 97.59 (71.71);P: 0.042] and CD4+IL-2/ml median (IQR) [626.33 (2744.98) vs 169.3 (173.91);P: 0.042]. No other significant differences were observed regarding other immunological markers in 3 study groups. Conclusion. IFN-gamma production by CD8+ T cells is highly expressed in children with MIS-C compared to hospitalized children one month after acute COVID-19 and could consist possible immunological biomarker. Further studies are needed in order to elucidate the pathophysiological basis of these findings.
ABSTRACT
Background: Pulmonary embolism (PE) is reported in around2.6-8.9% of COVID-19 hospitalized adult patients, but it is very rare in children and adolescents with symptomatic infection from SARS-CoV-2. Aims: This is the case of a15-year-old male patient with COVID-19 complicated by deep vein thrombosis (DVT) who developed PE. Methods: The patient presented with fever and a four-day history of pain and swelling of the right lower limb, without any history of injury. It is notable that15 days before admission, he experienced diarrhea, vomiting and low-grade fever for 48 hours. The adolescent boy had short stature and was rather overweighted for his age and gender (body mass index:25.2 kg/m2), while over the last two years he was receiving anastrozole (aromatase inhibitor), for height increase. Results: On admission, due to pain, limited mobility of the right limb and edema of the ipsilateral knee, a triplex ultrasound was performed that revealed DVT extended from the right iliac to the popliteal vein. RT-PCR for SARS-CoV-2 was positive, while the rest of the laboratory results showed a prolongation in prothrombin time (16.3 seconds, normal values:10-14 sec), elevated d-dimers (10 mg/dl, n.v.: < 0.5 mg/dl), high levels of factor VIII (214 IU/dl, n.v: 50-150 IU/dl), low levels of antithrombin (36 IU/ml, n.v.: 80-120 IU/ml) and increased ferritin (346 μg/L, n.v.:10-150 μg/L). Initial management consisted of antibiotic therapy plus anticoagulation with subcutaneous low molecular weight heparin (LMWH) i.e. tinzaparin in therapeutic dose. Soon after admission the patient developed severe hypotension with low diastolic blood pressure, refractive to IV normal saline boluses while his oxygen saturation dropped to 94% few hours later. A CT pulmonary angiogram (CTPA) was performed revealing a big thrombus with longitudinal diameter of3 cm, in the left pulmonary artery, establishing the diagnosis of PE on the ground of DVT. After PE diagnosis, the patient was transferred to the intensive care unit (ICU) for 48 hours, and subsequently at the special SARS-CoV-2 ward for 4 weeks. He completed a ten-day course of intravenous dexamethasone and anticoagulation treatment was switched to oral warfarin, after completion of three weeks of LWMH. Apart from COVID-19 whose hypercoagulable physis is already well-established the patient had additional risk factors predisposing to thromboembolic episodes. More precisely, he was overweight and under aromatase inhibitor therapy (which can be procoagulant by increasing testosterone levels). Moreover, the preexisting symptoms from the gastrointestinal system, could have predisposed him to DVT in view of dehydration and increased viscosity. On top of it, it is noteworthy that the patient had excessive screen time for his tele-education the last three weeks prior to the onset of VTE, meaning there were long periods of immobilization. Additionally, during the course of COVID-19 infection he had low levels of antithrombin, a serious prothrombotic condition attributed to the disease. Summary/Conclusion: This case underlines the fact that pediatric patients with COVID-19, are predisposed to the development of thromboembolic events, especially in the presence of preexisting prothrombotic risk factors. Larger studies in pediatric population with SARS-CoV-2 infection are needed, for the establishment of recommendations regarding risk evaluation, hemostatic monitoring and application of anticoagulation in outpatient, as well as hospitalized patients.