Study design
This was a pre-planned secondary analysis of a Phase-II interventional single-arm pilot study conducted in the Pediatric Intensive Care Unit (PICU) at the Bambino Gesù Children’s Hospital in Rome, Italy. The study protocol underwent review and approval by the local Ethics Committee in July 2019 (Ethics Committee Reference: N376). Informed written consent was obtained from the next of kin or legal guardian for each patient enrolled in the study (9).
The patient recruitment and data collection period spanned from July 2019 to October 2021. Eligibility screening was conducted for all patients admitted to the PICU who presented with either microbiologically confirmed or suspected septic shock. The inclusion criteria for participation in the study were as follows: 1. Body weight ≥ 10 Kgs, 2. A diagnosis of septic shock as defined by the International Pediatric Consensus Conference criteria (10), and 3. The requirement for CRRT; CRRT was indicated either due to acute kidney injury, as defined by the Kidney Disease Improving Global Outcome (KDIGO) criteria (11), or the presence of fluid overload ≥ 10%. The study was registered in the clinical trials database under the identifier NCT05658588.
All study procedures were conducted in accordance with the ethical standards and guidelines set forth by the responsible committee for human experimentation, including institutional and national regulations. The study adhered to the principles outlined in the Helsinki Declaration, ensuring the protection of the rights, safety, and well-being of all participating patients.
Data collection and definition
Data collection for this study was managed through an electronic case report form (eCRF). At the time of patient admission (i.e. enrolment), a comprehensive set of information was documented, encompassing demographics, comorbidities, source of admission, primary diagnoses, PELOD-2 and VIS (12) (13), as well as the Pediatric Index of Mortality 3 (PIM-3) score (14). Continuous monitoring of invasive arterial pressure was initiated at baseline, which marked the onset of therapy, and persisted for a duration of 96 hours. Lactate levels, VIS, and PELOD-2 scores were assessed at multiple time points, specifically at baseline, 24, 48, 72 and 96 hours following the commencement of therapy.Sublingual microcirculation assessments were conducted daily for five consecutive days starting from the initiation of blood purification. A handheld vital microscope (HVM) utilizing incident dark field microscopy imaging technology (Braedius Medical, Huizen, The Netherlands) was employed to capture five video recordings at each designated time-point. Subsequently, the three videos with the highest quality were selected for subsequent analysis, in accordance with a recent consensus (15)(16). The videos underwent a meticulous offline analysis process, utilizing dedicated software (Analysis Manager V2, Braedius Medical, Huizen, The Netherlands) (17). Additionally, a qualitative assessment was conducted by two independent operators (16)(18). Intraobserver and interobserver variabilities are around 5% in our department (18).
Several critical microcirculatory parameters were computed, including total small vessel density (TVD), proportion of perfused vessels (PPV), and perfused microvascular density (PVD).
For the semi-quantitative evaluation of microcirculatory flow, the methodology described by Boerma et al. (19) was adopted. Each recorded image was divided into four equal quadrants, and flow within each quadrant was assigned a numerical score: no flow (score 0), intermittent flow (score 1), sluggish flow (score 2), and continuous flow (score 3). The microvascular flow index (MFI) was then determined based on the predominant flow type in each quadrant and subsequently averaged across all quadrants. Furthermore, the heterogeneity index (HI) of MFI was also computed (20). Patients who exhibited an improvement in microcirculation were categorized as "responders"; this classification was based on the attainment of specific criteria within 96 hours from the commencement of blood purification treatment, including achieving an MFI ≥ 2.5 and/or PPV exceeding 90% (2)(3)(19).
CytoSorb therapy
A hemodialysis catheter was carefully inserted into a central vein, with the choice of internal jugular or femoral placement contingent upon the patient's size and suitability; CRRT was carried out employing a standard hemofilter constructed from either polyarylethersulphone or ANST69. In addition, a CytoSorb (CytoSorbents Inc, New Jersey, USA) adsorber was integrated into the CRRT circuit, configured to operate in continuous veno-venous hemodiafiltration (CVVHDF) mode. The effluent dosage was maintained at 2000 ml/h*1.73 m2, ensuring effective blood purification.
Within the CRRT circuit, the CytoSorb cartridge was strategically positioned in series with the hemofilter, with routine replacement scheduled for every 24 hours. This arrangement was sustained for a maximum duration of 96 hours. Prior to each CRRT session, the circuit underwent a thorough saline solution flush and was primed with either albumin, blood, or saline, a decision made in accordance with the clinical judgment of the attending physicians.
To manage anticoagulation, unfractioned heparin (UFH) was administered through a continuous heparin sodium infusion, typically adjusted to maintain a post-filter activated clotting time (ACT) within the range of 160 to 180 seconds. In cases where contraindications to UFH were identified, regional citrate anticoagulation was judiciously employed as an alternative approach.
For patients requiring extracorporeal membrane oxygenation (ECMO) support, the placement of CRRT access and return was strategically positioned post-oxygenator and pre-pump to ensure compatibility and safety within the ECMO circuit.
Statistical analysis
For descriptive data, continuous variables are expressed as mean ± standard deviation (SD) or median and interquartile range (IQR), according to their distribution; categorical variables are expressed as count (n) or percentage (%). To evaluate changes between two timepoints the Mann-Whitney U test was performed. Spearman correlation was used to assess the association between microcirculation and hemodynamic parameters, both in all cohort and in the responders’ population. Differences were considered statistically significant at p < 0.05. All the tests were performed using NCSS 2021 Statistical Software (2021). NCSS, LLC. Kaysville, Utah, USA.