Assessment of necroptosis in the retina in a repeated primary ocular blast injury mouse model.

Primary blast injury (caused by the initial rapid increase in pressure following an explosive blast) to the retina and optic nerve (ON) causes progressive visual loss and neurodegeneration. Military personnel are exposed to multiple low-overpressure blast waves, which may be in quick succession, such as during breacher training or in combat. We investigated the necroptotic cell death pathway in the retina in a mouse repeated primary ocular blast injury (rPBI) model using immunohistochemistry. We further evaluated whether intravitreal injections of a potent necroptosis inhibitor, Necrostatin-1s (Nec-1s), protects the retina and ON axons by retinal ganglion cells (RGC) counts, ON axonal counting and optical coherence tomography (OCT) analysis of vitreous haze. Receptor interacting protein kinase (RIPK) 3, increased in the inner plexiform layer 2 days post injury (dpi) and persisted until 14 dpi, whilst RIPK1 protein expression did not change after injury. The number of degenerating ON axons was increased at 28 dpi but there was no evidence of a reduction in the number of intact ON axons or RNA-binding protein with multiple splicing (RBPMS)+ RGC in the retina by 28 dpi in animals not receiving any intravitreal injections. But, when intravitreal injections (vehicle or Nec-1s) were given there was a significant reduction in RBPMS+ RGC numbers, suggesting that rPBI with intraocular injections is damaging to RGC. There were fewer RGC lost after Nec-1s than vehicle injection, but there was no effect of Nec-1s or vehicle treatment on the number of degenerating axons. OCT analysis demonstrated no effect of rPBI on vitreous haze, but intravitreal injection combined with rPBI increased vitreous haze (P = 0.004). Whilst necroptosis may be an active cell death signalling pathway after rPBI, its inhibition did not prevent cell death, and intravitreal injections in combination with rPBI increased vitreous inflammation and reduced RBPMS+ RGC numbers, implying intravitreal injection is not an ideal method for drug delivery after rPBI.

Optical coherence tomography angiography analysis methods: a systematic review and meta-analysis.

Optical coherence tomography angiography (OCTA) is widely used for non-invasive retinal vascular imaging, but the OCTA methods used to assess retinal perfusion vary. We evaluated the different methods used to assess retinal perfusion between OCTA studies. MEDLINE and Embase were searched from 2014 to August 2021. We included prospective studies including ≥ 50 participants using OCTA to assess retinal perfusion in either global retinal or systemic disorders. Risk of bias was assessed using the National Institute of Health quality assessment tool for observational cohort and cross-sectional studies. Heterogeneity of data was assessed by Q statistics, Chi-square test, and I2 index. Of the 5974 studies identified, 191 studies were included in this evaluation. The selected studies employed seven OCTA devices, six macula volume dimensions, four macula subregions, nine perfusion analyses, and five vessel layer definitions, totalling 197 distinct methods of assessing macula perfusion and over 7000 possible combinations. Meta-analysis was performed on 88 studies reporting vessel density and foveal avascular zone area, showing lower retinal perfusion in patients with diabetes mellitus than in healthy controls, but with high heterogeneity. Heterogeneity was lowest and reported vascular effects strongest in superficial capillary plexus assessments. Systematic review of OCTA studies revealed massive heterogeneity in the methods employed to assess retinal perfusion, supporting calls for standardisation of methodology.

Modifications in Macular Perfusion and Neuronal Loss After Acute Traumatic Brain Injury.

Purpose: Traumatic brain injury (TBI) causes structural damage and functional impairment in the visual system, often with retinal ganglion cell (RGC) degeneration occurring without visual symptoms. RGC degeneration is associated with reduced retinal blood-flow, however, it is not known whether reductions in perfusion precede or are secondary to neurodegeneration. Methods: We conducted a prospective observational single-center case series. Patients were included if they were admitted to the hospital after acute TBI and underwent ophthalmic clinical examination, including optical coherence tomography (OCT) and OCT angiography (OCTA) acutely and at follow-up. Ganglion cell layer thickness (GCL) thickness, vascular density in the superficial vascular plexus (SVP), and intermediate capillary plexus (ICP) were quantified. Results: Twenty-one patients aged 20 to 65 years (mean = 38 years) including 16 men and 5 women were examined less than 14 days after moderate to severe TBI, and again after 2 to 6 months. Macular structure and perfusion were normal at baseline in all patients. Visual function was abnormal at baseline in three patients and subsequent neurodegeneration and loss of perfusion corresponded to baseline visual function abnormalities. Nine patients (43%) had reduced macular GCL thickness at follow up. Perfusion in the SVP strongly associated with local GCL thickness. The strongest association of the SVP metrics was the sum of vessel density (P < 0.0001). Conclusions: In cases of reduced visual function after TBI, macular perfusion remained normal until reductions in GCL thickness occurred, indicating that perfusion changes were secondary to local GCL loss.

Reliability of Optical Coherence Tomography Angiography Retinal Blood Flow Analyses.

Purpose: Investigate the association between the optical coherence tomography angiography (OCTA) metrics derived from different analysis programs to understand the comparability of studies using these different approaches. Methods: Secondary analysis of a prospective observational study (March 2018-September 2021). Forty-four right eyes and 42 left eyes from 44 patients were included. Patients were either undergoing upper gastrointestinal surgery with a critical care stay planned or were already in the critical care unit with sepsis. OCTA scans were obtained in an ophthalmology department or critical care setting. Fourteen OCTA metrics were compared within and between the programs, and agreement was measured by Pearson's R coefficient and intraclass correlation coefficient. Results: Correlation was highest between all Heidelberg metrics and Fractalyse (all >0.84), and lowest between Matlab skeletonized or foveal avascular zone metrics and all other measures (e.g., skeletal fractal dimension and vessel density at -0.02). Agreement between eyes was moderate to excellent in all metrics (0.60-0.90). Conclusions: The significant variability between metrics and programs used for OCTA analysis demonstrates that they are not interchangeable and supports a recommendation for perfusion density metrics to be reported as standard. Translational Relevance: Agreement between different OCTA analyses is variable and not interchangeable. The high agreement between non-skeletonized vessel density metrics suggests that these should be routinely reported.

Optical coherence tomography angiography as a surrogate marker for end-organ resuscitation in sepsis: A review.

Sepsis is a severe illness which results in alterations in the end organ microvascular haemodynamics and is associated with a high risk of mortality. There is currently no real-time method of monitoring microcirculatory perfusion during sepsis. Retinal microcirculation is closely linked to cerebral perfusion and may reflect systemic vascular alterations. Retinal perfusion can be assessed using the non-invasive imaging technique of optical coherence tomography angiography (OCTA). This narrative review aims to discuss the utility of using retinal imaging and OCTA in systemic illness and sepsis. OCTA can be used as a functional, non-invasive and real-time biomarker along with other haemodynamic parameters for assessing and managing patients with sepsis.

Effects of intravitreal injection of siRNA against caspase-2 on retinal and optic nerve degeneration in air blast induced ocular trauma.

Ocular repeated air blast injuries occur from low overpressure blast wave exposure, which are often repeated and in quick succession. We have shown that caspase-2 caused the death of retinal ganglion cells (RGC) after blunt ocular trauma. Here, we investigated if caspase-2 also mediates RGC apoptosis in a mouse model of air blast induced indirect traumatic optic neuropathy (b-ITON). C57BL/6 mice were exposed to repeated blasts of overpressure air (3 × 2 × 15 psi) and intravitreal injections of siRNA against caspase-2 (siCASP2) or against a control enhanced green fluorescent protein (siEGFP) at either 5 h after the first 2 × 15 psi ("post-blast") or 48 h before the first blast exposure ("pre-blast") and repeated every 7 days. RGC counts were unaffected by the b-ITON or intravitreal injections, despite increased degenerating ON axons, even in siCASP2 "post-blast" injection groups. Degenerating ON axons remained at sham levels after b-ITON and intravitreal siCASP2 "pre-blast" injections, but with less degenerating axons in siCASP2 compared to siEGFP-treated eyes. Intravitreal injections "post-blast" caused greater vitreous inflammation, potentiated by siCASP2, with less in "pre-blast" injected eyes, which was abrogated by siCASP2. We conclude that intravitreal injection timing after ocular trauma induced variable retinal and ON pathology, undermining our candidate neuroprotective therapy, siCASP2.

Stability of OCT and OCTA in the Intensive Therapy Unit Setting.

To assess the stability of retinal structure and blood flow measures over time and in different clinical settings using portable optical coherence tomography angiography (OCTA) as a potential biomarker of central perfusion in critical illness, 18 oesophagectomy patients completed retinal structure and blood flow measurements by portable OCT and OCTA in the eye clinic and intensive therapy unit (ITU) across three timepoints: (1) pre-operation in a clinic setting; (2) 24-48 h post-operation during ITU admission; and (3) seven days post-operation, if the patient was still admitted. Blood flow and macular structural measures were stable between the examination settings, with no consistent variation between pre- and post-operation scans, while retinal nerve fibre layer thickness increased in the post-operative scans (+2.31 µm, p = 0.001). Foveal avascular zone (FAZ) measurements were the most stable, with an intraclass correlation coefficient of up to 0.92 for right eye FAZ area. Blood flow and structural measures were lower in left eyes than right eyes. Retinal blood flow assessed in patients before and during an ITU stay using portable OCTA showed no systematic differences between the clinical settings. The stability of retinal blood flow measures suggests the potential for portable OCTA to provide clinically useful measures in ITU patients.

Rapid assessment of ocular drug delivery in a novel ex vivo corneal model.

Drug delivery by topical application has higher patient acceptance and lower morbidity than intraocular injection, but many ophthalmic treatments are unable to enter the eye or reach the posterior segment after topical application. The first stage towards posterior segment delivery after topical application is ocular surface penetration and existing models are in vivo or use large quantities of tissue. We therefore developed a novel ex vivo model using discs of porcine and human cornea and sclera (5 mm diameter) to assess penetration of a candidate neuroprotective siRNA. siRNA against caspase 2 or control solutions of known penetrance were applied to the corneal epithelial surface and trans-corneal penetration and corneal adsorbance measured at fixed time points. To demonstrate that leakage did not occur, we applied dextran blue, which should not penetrate the intact cornea and did not do so in our model. Fluorescein penetration (0.09%) was less than rhodamine B (6.98%) at 60 min. siCASP2 penetration was 0.01% by 60 min. When the applied siCASP2 was washed off after 2 min, (representing lacrimal drainage) 0.071% penetrated porcine cornea by 60 min and 0.0002% penetrated human cornea and 0.001% penetrated human sclera. Our ex vivo model rapidly and cost-effectively assesses transcorneal penetration of candidate topical therapies, allowing rates of trans-corneal penetration for potential therapies such as siRNA to be evaluated with small quantities of human or animal tissue.