Setting
This prospective, observational study was performed at a tertiary teaching university hospital (Oulu University Hospital) with a mixed medical-surgical ICU. All patients admitted to the ICU for sepsis or septic shock between 1 January 2012 and 31 December 2014 were screened for participation in this study, which included retinal angiography, measurements of retinal arterial filling time (RAFT), intraocular pressure (IOP), screening for delirium, and measuring blood proinflammatory and cerebral markers as well as hemodynamic parameters. Delirium was assessed using the Confusion Assessment Method for the ICU [12]. Level of consciousness was assessed with the Richmond Agitation-Sedation Scale (scores range from −5 to 4, with lower scores indicating less arousal, higher scores indicating more agitation, and 0 indicating an alert and calm state) [13]. The Glasgow Coma Scale was used to determine the level of consciousness of unsedated patients during imaging.
The inclusion criteria for patients were sepsis and septic shock according to the Surviving Sepsis Guidelines [14]. Patients with acute brain disease, psychiatric disorders, chronic alcoholism, or other types of encephalopathy, as well as those with various ophthalmological conditions, such as age-related macular degeneration, diabetic retinopathy, cataract, angle-closure glaucoma, or eye injuries, were excluded. The study was approved by the local ethics committee of the Northern Ostrobothnia Hospital District. After written informed consent was obtained from the patients or their relatives, patients were included in the study.
Patient management during the ICU stay
Patients with sepsis and septic shock were managed according to the Surviving Sepsis Campaign guidelines [14] and were administered antibiotic treatment according to our ICU antibiotic stewardship protocol. Macrohemodynamics were monitored by measuring arterial blood pressure and pulmonary arterial pressure. An ICU data management system (Centricity Critical Care*(8.1) SP7 (8.17.034); GE Healthcare, Barrington, IL, USA) was used to collect data concerning daily laboratory results, hemodynamic parameters, need for vasoactive or sedative agents, length of stay (LOS), presence of delirium, and severity-of-illness scoring (Acute Physiology and Chronic Health Evaluation II [APACHE II], Simplified Acute Physiology Score [SOFA]) after inclusion in the study. Surviving patients were met at the follow-up clinic 3–6 months after hospital discharge.
Interventions
Retinal angiography
Standardized retinal fluorescein angiographs and digital images were obtained for both eyes. Mydriasis was achieved 30 minutes after instilling topical tropicamide and phenylephrine hydrochloride eye drops. A Heidelberg Retina Angiograph 2 (HRA 2) camera was used in exceptional vertical alignment (HRA 2-00153; Heidelberg Engineering, Heidelberg, Germany). After 4 minutes, postangiography images were taken. The phase identified by laminar flow in the veins was determined as the endpoint of the arterial filling stage and start of the venous phase. The arterial filling time was measured from the patient’s right eye. The preliminary images were taken on the first possible day of the ICU stay or if the inclusion criteria were fulfilled during the ICU stay. The patients were reexamined 2–5 days later. The third angiography was performed 3–6 months after the hospital discharge. These postdischarge images were used as within-patient controls to detect pathological findings in fundus images during the ICU stay. The images were analyzed by an ophthalmologist blinded to the patients’ identity and any clinical data to interpret pathological findings.
Arterial filling time
RAFT was determined on the basis of digital fluorescein angiography using the HRA 2. In the analyses, we used the RAFT measured from the eye with worse microvascular retinal changes. The initial phase of the fluorescein angiography was determined, and the early filling stages were documented. The arterial phase was started when the central retinal artery began to fill. The median arterial filling time of the study population was 8.3 seconds. This median value was used as a cutoff point to divide the patient population into two groups: short retinal arterial filling time (SRAFT) less than 8.3 seconds and prolonged retinal arterial filling time (PRAFT) greater than 8.3 seconds.
Intraocular pressure measurement
IOP was measured in both eyes using an Icare PRO tonometer (1201775 TA 03 Icare® PRO; Icare Finland Oy, Vantaa, Finland). IOP measurements were performed at the time of each angiography and after 3–6 months at the post-ICU clinic. In patients with retinal abnormalities, the IOP in the eye with the most intense findings was included in the analyses, whereas the first measurement from the right eye was used for those patients without retinal abnormalities.
Laboratory data
Separated plasma was stored at −70 °C. The inflammatory cytokines tumor necrosis factor-α (TNF-α) and interleukin 6 (IL-6) as well as markers of brain dysfunction (neuron-specific enolase [NSE] and calcium-binding protein B [S100B]) were analyzed. Cytokine concentrations were determined using the MILLIPLEX® MAP Human Cytokine/Chemokine Magnetic Bead Panel (HCYTOMAG-60 K; EMD Millipore, Billerica, MA, USA). The lower detection limits were 0.7 pg/ml for TNF-α and 0.9 pg/ml for IL-6. The intra-assay coefficients of variation (CVs) for TNF-α and IL-6 were 2.6% and 2.0%, respectively, and the corresponding inter-assay CVs were 13% and 18.3%, respectively. S100B and NSE were measured using an immunochemiluminometric method (Elecsys 2010 analyzer; Roche Diagnostics GmbH, Mannheim, Germany).
Statistical analysis
The statistical analysis was performed using IBM SPSS Statistics version 22 software (IBM, Armonk, NY, USA). Proportional data are expressed as rate (count) and percent, and continuous variables are expressed as median and 25th and 75th percentiles. Proportional data were tested using Pearson’s chi-square test unless otherwise stated. Continuous variables were tested using a nonparametric Mann-Whitney U test and independent samples median test. Two-tailed P values less than 0.05 were considered statistically significant.
Outcome measures
The primary outcome measure was median RAFT. The secondary outcome endpoints were the IOP measurements and their relationship to RAFT and different pathological findings in the retina during sepsis. Also, demographic data, comorbidities, severity scores, and outcome data were compared between the SRAFT and PRAFT patient groups.