Study design. Our trial was designed to evaluate the impact of plasmapheresis therapy in the outcome of severe COVID-19 and cytokine release syndrome at one medical center in Mexico City. The study was designed and performed according to ethical guidelines of the 1975 Declaration of Helsinki, and approved by the Local Committees of Research, Ethics in Research and Biosafety of the Centro Médico Nacional ‘20 de Noviembre’ ISSSTE, Mexico City (Protocol ID No. 09-136.2021). All participants provided written informed consent.
Study population. Hospitalized patients between April-August 2020. Patients were aged 16 to 65 years old, diagnosed with SARS-CoV-2 infection, as confirmed by typical tomographic findings according to Radiological Society of North America and/or qRT-PCR confirmation. Clinical presentation of Acute Respiratory Distress Syndrome and increased levels of interleukine-6 > 40 pg/mL, ferritin > 500 ng/mL, C reactive protein (CRP) > 60 mg/L, erythrosedimentation rate > 40 m/sec; and/or lymphopenia < 1.0 x10/L. Patients were excluded if they had SOFA score > 11 points, active bleeding, platelet count < 50,000 cels and/or hypofibrinogenemia < 80 mg/dL. Written informed consent was obtained from all the patients or from a legal representative if they were unable to provide consent.
Standard therapy. It was based on the use of chloroquine, azithromycin, dexamethasone, supplementary oxygen; as well as tocilizumab and intravenous immunoglobulin. This therapy was common for the control and PLEX group, while PLEX group received tocilizumab and immunoglobulin before PLEX therapy.
Plasmapheresis. A double lumen central venous catheter was placed, either at jugular or femoral vein approaches. The plasmapheresis therapy was performed with a membrane-based system, using PrismaFlex CRRT and a TPE 1000–2000 filter according to body surface. Exchange plasma volumes of 1.5 times the estimated circulating plasma volume, according to Kapplan's formula (Plasma volume (lts) = 0.065 x Weight (kg) x (1 - Hematocrit [%])). The blood flow rate was set at range 75–150 ml/min. Replacement solution consisted of 3% albumin, at a flow rate started at 100 mL/hr, and increased up to maximum of 1500 mL/hr; while 2 fresh frozen plasma were transfused at the end of each session. Anticoagulation was performed at doses of 30–40 IU/Kg/hr of unfractionated heparin. A second plasmapheresis session was systematically performed 48 hours after the first session. Blood samples were obtained from catheter blood before and after every session, for cytokine determination.
Cytokine determination. Blood samples were centrifuged and 500mcL of serum are used for cytokine determination. Briefly, serum was combined with surface-bound capture anti-IL6 polyclonal antibody and alkaline phosphatase system developer as tracer (IL6 Bead Pack, Diagnostic Products Corporation, LA Cal. USA), in a immulite 2000 automatized immunoassay (Siemens), working within a range of 3-870 pg/mL. This system works under certification ISO 13485:2003.
Lung damage determination. One day before starting treatment and ten days after the last PLEX, the volume of lung involvement was calculated through tomographic volumetric assessment. Non-contrast enhanced chest CT imaging was performed using 2 CT scanners (Siemens SOMATOM drive and Siemens SOMATOM emotion scanners, Siemens Healthineers, Germany). Imaging reconstructions were performed with a 1-mm thickness slices without interstice gap. Lung segmentation was performed using the Alma Medical workstation version 5.0. For determination of non-affected lung parenchyma volume (NLV), automated segmentation tool was selected, and a reference attenuation range between − 1000 and − 600 HU was designated. Vascular structures, airways, and pathologic opacities were excluded. For determination of Lung Opacities Segmentation-Lung Opacities Volume (LOV): Initial automated segmentation was attempted using thresholding-based methods. A reference attenuation range between − 500 HU and 20 HU was selected. Then, the semi-automated option was selected to perform a region-based segmentation to adjust lesion boundaries. Volumes from each side were added to calculate total NLV and LOV. Total lung volume was calculated adding NLV + LOV.
Outcomes. The primary outcome was all-cause mortality within 60 days after inclusion. Secondary outcomes were the free mechanical-ventilation days, changes in SOFA score, decrease of pro-inflammatory markers at day 7, hospital length-of-stay, and decrease of lung’s volume involvement.
Statistical analysis. Data distribution was assessed by Kolmogorov-Smirnoff test. Then, qualitative and quantitative data were resumed as n(%) and mean ± SD, respectively. Inferential analyses were performed by either chi square, one-way independent T-test, or U-Mann-Whitney. Kaplan-Meyer curves were constructed. Risk estimation was evaluated through OR and CI95%. Statistical significance was considered at p < 0.05.