Bart R. Thomson

Objectives To validate whether cerebrospinal fluid oxyhaemoglobin (CSF-Hb), measured from external ventricular or lumbar drains, is associated with secondary brain injury (SAH-SBI) after aneurysmal subarachnoid haemorrhage (aSAH), and to assess its value as a real-time monitoring biomarker. Design Preregistered multicentre prospective observational cohort study. Setting Eight neurosurgical tertiary centres in Switzerland, Germany, and Austria, between August 2021 and June 2024. Participants 366 patients with aSAH (mean age 58 years; 65% women). Of these, 260 provided cerebrospinal fluid (CSF) samples via external ventricular drain (EVD; 2,467 samples, median 10 days per patient) and 66 via lumbar drain (LD; 379 samples, median 6 days). Interventions Daily CSF samples were collected via EVD or LD from day 1 to day 14 after haemorrhage; no therapeutic interventions were tested. Main outcome measures CSF-Hb and its metabolites were analysed post hoc in a blinded manner. The primary outcome was SAH-SBI, defined as a composite of angiographic vasospasm (aVSP), delayed cerebral ischaemia (DCI), and delayed ischaemic neurological deficits (DIND), assessed daily over 14 days. Secondary outcomes included temporal CSF-Hb profiles and associations with aneurysm location, haematoma volume, intraventricular haemorrhage, chronic hydrocephalus, and 3-month functional outcome. Results CSF-Hb showed a delayed peak pattern: concentrations were low after aSAH, rose to a maximum on day 10 (EVD-derived CSF-Hb median 11.3 μM, IQR 2.64 to 25.90), and then declined. Larger haematoma volume (pConclusions In this preregistered multicentre validation study, EVD-derived CSF-Hb did not perform as a robust real-time monitoring biomarker for SAH-SBI, showing limited same-day discrimination and substantial between-centre heterogeneity. These findings argue against clinical implementation of CSF-Hb point-measurement as a single-parameter biomarker. In contrast, CSF methaemoglobin remained consistently associated with SAH-SBI, supporting the mechanistic relevance of haemolysis-related pathways. Future work using the HeMoVal biobank will apply multi-marker, pathway-level analyses to define haemolysis-related biomarker signatures and provide a platform for robust external validation of future candidates. Study registration ClinicalTrials.gov NCT04998370 ; date of registration 10 August 2021.

Secondary brain injury (SBI) occurs with a lag of several days post-bleeding in patients with aneurysmal subarachnoid hemorrhage (aSAH) and is a strong contributor to mortality and long-term morbidity. aSAH-SBI coincides with cell-free hemoglobin (Hb) release into the cerebrospinal fluid. This temporal association and convincing pathophysiological concepts suggest that CSF-Hb could be a targetable trigger of SBI. However, sparse experimental evidence for Hb’s neurotoxicity in vivo defines a significant research gap for clinical translation. We modeled the CSF-Hb exposure observed in aSAH patients in conscious sheep, which allowed us to assess neurological functions in a gyrencephalic species. Twelve animals were randomly assigned for 3-day bi-daily intracerebroventricular (ICV) injections of either Hb or Hb combined with the high-affinity Hb scavenger protein haptoglobin (Hb-Hp, CSL888). Repeated CSF sampling confirmed clinically relevant CSF-Hb concentrations. This prolonged CSF-Hb exposure over 3 days resulted in disturbed movement activity, reduced food intake, and impaired observational neuroscores. The Hb-induced neurotoxic effects were significantly attenuated when Hb was administered with equimolar haptoglobin. Preterminal magnetic resonance imaging (MRI) showed no CSF-Hb-specific structural brain alterations. In both groups, histology demonstrated an inflammatory response and revealed enhanced perivascular histiocytic infiltrates in the Hb-Hp group, indicative of adaptive mechanisms. Heme exposure in CSF and iron deposition in the brain were comparable, suggesting comparable clearance efficiency of Hb and Hb-haptoglobin complexes from the intracranial compartment. We identified a neurological phenotype of CSF-Hb toxicity in conscious sheep, which is rather due to neurovascular dysfunction than structural brain injury. Haptoglobin was effective at attenuating CSF-Hb-induced neurological deterioration, supporting its therapeutic potential.

Background Secondary brain injury (SBI) occurs with a lag of several days post-bleeding in patients with aneurysmal subarachnoid hemorrhage (aSAH) and is a strong contributor to mortality and long-term morbidity. aSAH-SBI coincides with cell-free hemoglobin (Hb) release into the cerebrospinal fluid. This temporal association and convincing pathophysiological concepts suggest that CSF-Hb could be a targetable trigger of SBI. However, sparse experimental evidence for Hb’s neurotoxicity in vivo defines a significant research gap for clinical translation. Methods We modeled the CSF-Hb exposure observed in aSAH patients in conscious sheep, which allowed us to assess neurological functions. Twelve animals were randomly assigned for three-day bi-daily intracerebroventricular (ICV) injections of either Hb or Hb combined with the high-affinity Hb scavenger protein haptoglobin (Hb-Hp, CSL888). Results Repeated CSF sampling confirmed clinically relevant CSF-Hb concentrations. This prolonged CSF-Hb exposure over three days resulted in disturbed movement activity, reduced food intake, and impaired observational neuroscores. The Hb-induced neurotoxic effects were significantly attenuated when Hb was administered with equimolar haptoglobin. Preterminal magnetic resonance imaging (MRI) showed no CSF-Hb-specific structural brain alterations. In both groups, histology demonstrated an inflammatory response and revealed enhanced perivascular histiocytic infiltrates in the Hb-Hp group, indicative of adaptive mechanisms. Heme exposure in CSF and iron deposition in the brain were comparable, suggesting comparable clearance efficiency of Hb and Hb-haptoglobin complexes from the intracranial compartment. Conclusion We identified a neurological phenotype of CSF-Hb toxicity in conscious sheep, which is rather due to neurovascular dysfunction than structural brain injury. Haptoglobin was effective at attenuating CSF-Hb-induced neurological deterioration, supporting its therapeutic potential.

In recent years, insect husbandry has seen an increased interest in order to supply in the production of raw materials, food, or as biological/environmental control. Unfortunately, large insect rearings are susceptible to pathogens, pests and parasitoids which can spread rapidly due to the confined nature of a rearing system. Thus, it is of interest to monitor the spread of such manifestations and the overall population size quickly and efficiently. Medical imaging techniques could be used for this purpose, as large volumes can be scanned non-invasively. Due to its 3D acquisition nature, computed tomography seems to be the most suitable for this task. This study presents an automated, computed tomography-based, counting method for bee rearings that performs comparable to identifying all Osmia cornuta cocoons manually. The proposed methodology achieves this in an average of 10 seconds per sample, compared to 90 minutes per sample for the manual count over a total of 12 samples collected around lake Zurich in 2020. Such an automated bee population evaluation tool is efficient and valuable in combating environmental influences on bee, and potentially other insect, rearings.

The functional neurological outcome of patients with intracerebral hemorrhage (ICH) strongly relates to the degree of secondary brain injury (ICH-SBI) evolving within days after the initial bleeding. Different mechanisms including the incitement of inflammatory pathways, dysfunction of the blood–brain barrier (BBB), activation of resident microglia, and an influx of blood-borne immune cells, have been hypothesized to contribute to ICH-SBI. Yet, the spatiotemporal interplay of specific inflammatory processes within different brain compartments has not been sufficiently characterized, limiting potential therapeutic interventions to prevent and treat ICH-SBI. Using a whole-blood injection model in mice, we systematically characterized the spatial and temporal dynamics of inflammatory processes after ICH using 7-Tesla magnetic resonance imaging (MRI), spatial RNA sequencing (spRNAseq), functional BBB assessment, and immunofluorescence average-intensity-mapping. We identified a pronounced early response of the choroid plexus (CP) peaking at 12 to 24h, that was characterized by inflammatory cytokine expression, epithelial and endothelial expression of leukocyte adhesion molecules, and the accumulation of leukocytes. In contrast, we observed a delayed secondary reaction pattern at the injection site (striatum) peaking at 96h, defined by gene expression corresponding to perilesional leukocyte infiltration and correlating to the delayed signal alteration seen on MRI. Pathway analysis revealed a dependence of the early inflammatory reaction in the CP on toll-like receptor 4 (TLR4) signaling via myeloid differentiation factor 88 (MyD88). TLR4 and MyD88 knockout mice corroborated this observation, lacking the early upregulation of adhesion molecules and leukocyte infiltration within the CP 24h after whole-blood injection. In conclusion, we report a biphasic brain reaction pattern after ICH with a MyD88-TLR4-dependent early inflammatory response of the CP, preceding inflammation, edema and leukocyte infiltration at the lesion site. Pharmacological targeting of the early CP-activation might harbor the potential to modulate the development of ICH-SBI. Graphical abstractAkeret, Buzzi et al. characterized the spatiotemporal dynamics after striatal whole blood injection in mice using magnetic resonance imaging (MRI), spatial RNA sequencing (spRNAseq), functional blood-brain barrier (BBB) assessment, and immunofluorescence average intensity mapping (IF). They report a biphasic brain reaction pattern with an early MyD88-TLR4-dependent inflammatory response of the CP, which preceded secondary inflammation and leukocyte infiltration at the perilesional site.

Purpose Cell-free hemoglobin in the cerebrospinal fluid (CSF-Hb) may be one of the main drivers of secondary brain injury after aneurysmal subarachnoid hemorrhage. Haptoglobin scavenging of CSF-Hb has been shown to mitigate cerebrovascular disruption. Using digital subtraction angiography (DSA) and blood oxygenation-level dependent cerebrovascular reactivity imaging (BOLD-CVR) the aim was to assess the acute toxic effect of CSF-Hb on cerebral blood flow and autoregulation, as well as to test the protective effects of haptoglobin. Methods DSA imaging was performed in eight anesthetized and ventilated sheep (mean weight: 80.4 kg) at baseline, 15, 30, 45 and 60 minutes after infusion of hemoglobin (Hb) or co-infusion with haptoglobin (Hb:Haptoglobin) into the left lateral ventricle. Additionally, 10 ventilated sheep (mean weight: 79.8 kg) underwent BOLD-CVR imaging to assess the cerebrovascular reserve capacity. Results DSA imaging did not show a difference in mean transit time or cerebral blood flow. Wholebrain BOLD-CVR compared to baseline decreased more in the Hb group after 15 minutes (Hb vs Hb:Haptoglobin: −0.03 ±0.01 vs −0.01 ±0.02) and remained diminished compared to Hb:Haptoglobin group after 30 minutes (Hb vs Hb:Haptoglobin: −0.03 ±0.01 vs 0.0 ±0.01), 45 minutes (Hb vs Hb:Haptoglobin: −0.03 ±0.01 vs 0.01 ±0.02) and 60 minutes (Hb vs Hb:Haptoglobin: −0.03 ±0.02 vs 0.01 ±0.01). Conclusion It is demonstrated that CSF-Hb toxicity leads to rapid cerebrovascular reactivity impairment, which is blunted by haptoglobin co-infusion. BOLD-CVR may therefore be further evaluated as a monitoring strategy for CSF-Hb toxicity after aSAH.

Purpose: Registration of pre‐ and intraoperative images is a crucial step of surgical liver navigation, where rigid registration of vessel centerlines is currently commonly used. When using 3D ultrasound (US), accuracy during navigation might be influenced by the size of the intraoperative US volume, yet the relationship between registration accuracy and US volume size is understudied. In this study, we specify an optimal 3D US volume size for registration using varying volumes of liver vasculature. While previous studies measured accuracy at registered fiducials, in this work, accuracy is determined at the target lesion which is clinically the most relevant structure.Methods: Three‐dimensional US volumes were acquired in 14 patients after laparotomy and liver mobilization. Manual segmentation of vasculature and centerline extraction was performed. Intraoperative and preoperative vasculature centerlines were registered with coherent point drift, using different sub‐volumes (sphere with radius r = 30, 40, …, 120 mm). Accuracy was measured by fiducial registration error (FRE) between vessel centerlines and target registration error (TRE) at the center of the target lesion.Results: The lowest FRE for vessel registration was reached with r = 50 mm (6.5 ± 2.5 mm), the highest with r = 120 mm (7.1 ± 2.1 mm). Clinical accuracy at the target lesion, resulted most accurate (TRE = 8.8 ± 5.0 mm) in sub‐volumes with a radius of 50 mm. Smaller US sub‐volumes resulted in lower average TREs when compared to larger US sub‐volumes (Pearson's correlation coefficient R = 0.91, p < 0.001).Conclusion: Our results indicate that there is a linear correlation between US volume size and registration accuracy at the tumor. Volumes with radii of 50 mm around the target lesion yield higher accuracy (p < 0.05) (Trial number IRBd18032, 11 September 2018).

Secondary brain injury after aneurysmal subarachnoid hemorrhage (SAH-SBI) is a significant contributor to poor outcomes in patients after rupture of an intracranial aneurysm. The lack of diagnostic biomarkers and novel drug targets represent an unmet need. Prior experimental evidence has suggested cell-free hemoglobin in the cerebrospinal fluid (CSF-Hb) as a pathophysiological driver of SAH-SBI. The aim of this study was to investigate the clinical and pathophysiological association between CSF-Hb and SAH-SBI. We prospectively enrolled 47 consecutive patients and collected daily CSF samples within 14 days after aneurysm rupture. There was very strong evidence for a positive association between CSF-Hb and SAH-SBI. The diagnostic accuracy of CSF-Hb for SAH-SBI markedly exceeded that of established methods (area under the curve: 0.89 [0.85-0.92]). Temporal LC-MS/MS CSF proteomics demonstrated that erythrolysis accompanied by an adaptive macrophage response are the two dominant biological processes occurring in the CSF space after aneurysm rupture. To further investigate the pathophysiology between CSF-Hb and SAH-SBI, we explored the vasoconstrictive and lipid peroxidation activities of Hb ex-vivo. These experiments revealed critical inflection points overlapping CSF-Hb concentration thresholds in patients with SAH-SBI. Selective Hb depletion and in-solution neutralization by the Hb-scavenger haptoglobin or the heme-scavenger hemopexin efficiently attenuated the vasoconstrictive and lipid peroxidation activities of CSF-Hb in patient CSF. Collectively, the clinical association between high CSF-Hb levels and SAH-SBI, the underlying pathophysiological rationale, and the favorable effects of haptoglobin and hemopexin in ex-vivo experiments position CSF-Hb as a highly attractive biomarker and potential drug target.