Abstract
Background
We aimed to characterize a preclinical model of intraventricular hemorrhage-induced brain damage (IVH-BD) in extremely low birth weight newborns (ELBWN), to identify potential therapeutic targets based on its pathophysiology.
Methods
IVH was induced in 1-day-old (P1) Wistar rats by left periventricular injection of clostridium collagenase (PVCC). At P6, P14, and P45 IVH-BD (area of damage, motor and cognitive deficits, Lactate/N-acetylaspartate ratio), white matter injury (WMI: ipsilateral hemisphere and corpus callosum atrophy, oligodendroglial population and myelin basic protein signal reduction), blood–brain barrier (BBB) dysfunction (occludin and Mfsd2a expression, Gadolinium leakage) and inflammation (TNFα, TLR4, NFkB, and MMP9 expression; immune cell infiltration), excitotoxicity (Glutamate/N-acetylaspartate), and oxidative stress (protein nitrosylation) were assessed. Sham animals were similarly studied.
Results
IVH-BD leads to long-term WMI, resulting in motor and cognitive impairment, thus reproducing IVH-BD features in ELBWN. BBB dysfunction with increased permeability was observed at P6 and P14, coincident with an increased inflammatory response with TLR4 overexpression, increased TNFα production, and increased immune cell infiltration, as well as increased excitotoxicity and oxidative stress.
Conclusions
This model reproduced some key hallmarks of IVH-BD in ELBWN. Inflammation associated with BBB dysfunction appears as relevant therapeutic target to prevent IVH-BD-induced WMI.
Impact
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Paraventricular injection of clostridium collagenase (PVCC) to 1-day-old Wistar rats uniquely reproduced the neuroimaging, histologic and functional characteristics of intraventricular hemorrhage-induced brain damage (IVH-BD) in extremely low birth weight newborns (ELBWN).
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PVCC-induced IVH triggered a prolonged inflammatory response associated with blood–brain barrier increased permeability, which in turn facilitates the infiltration of inflammatory cells.
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Thus, PVCC led to white matter injury (WMI) resulting in long-term motor and cognitive impairment.
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This model offers a valuable tool to obtain further insight into the mechanisms of IVH-BD in ELBWN and proposes some key therapeutic targets.
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Data availability
The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.
References
Ballabh, P. Pathogenesis and prevention of intraventricular hemorrhage. Clin. Perinatol. 41, 47–67 (2014).
Brouwer, M. J. et al. Effects of posthemorrhagic ventricular dilatation in the preterm infant on brain volumes and white matter diffusion variables at term-equivalent age. J. Pediatr. 168, 41–49 (2016).
Bolisetty, S. et al. Intraventricular hemorrhage and neurodevelopmental outcomes in extreme preterm infants. Pediatrics 133, 55–62 (2014).
Garton, T., Hua, Y., Xiang, J., Xi, G. & Keep, R. F. Challenges for intraventricular hemorrhage research and emerging therapeutic targets. Expert Opin. Ther. Targets 21, 1111–1122 (2017).
Zhao, X. et al. Distinct patterns of intracerebral hemorrhage-induced alterations in NF-κB subunit, iNOS, and COX-2 expression. J. Neurochem. 101, 652–663 (2007).
Romantsik, O., Bruschettini, M. & Ley, D. Intraventricular hemorrhage and white matter injury in preclinical and clinical studies. Neoreviews 20, 636–652 (2019).
Linsell, L., Malouf, R., Morris, J., Kurinczuk, J. J. & Marlow, N. Prognostic factors for cerebral palsy and motor impairment in children born very preterm or very low birthweight: a systematic review. Dev. Med. Child Neurol. 58, 554–569 (2016).
Fowlie, P. W., Davis, P. G. & McGuire, W. Prophylactic intravenous indomethacin for preventing mortality and morbidity in preterm infants. Cochrane Database Syst. Rev. CD000174 (2010).
Balasubramaniam, J. & Del Bigio, M. R. Animal models of germinal matrix hemorrhage. J. Child Neurol. 21, 365–371 (2006).
Alles, Y. C. J. et al. A novel preclinical rodent model of collagenase-induced germinal matrix/intraventricular hemorrhage. Brain Res. 1356, 130–138 (2010).
Lekic, T. et al. Rodent neonatal germinal matrix hemorrhage mimics the human brain injury, neurological consequences, and post-hemorrhagic hydrocephalus. Exp. Neurol. 236, 69–78 (2012).
Manaenko, A., Chen, H., Zhang, J. H. & Tang, J. Comparison of different preclinical models of intracerebral hemorrhage. Acta Neurochir. Suppl. 111, 9–14 (2011).
Segado-Arenas, A. et al. Cognitive impairment and brain and peripheral alterations in a murine model of intraventricular hemorrhage in the preterm newborn. Mol. Neurobiol. 55, 4896–4910 (2018).
Rolland, W. B. et al. Fingolimod confers neuroprotection through activation of Rac1 after experimental germinal matrix hemorrhage in rat pups. J. Neurochem. 140, 776–786 (2017).
Andersson, E. A., Rocha‐ferreira, E., Hagberg, H., Mallard, C. & Ek, C. J. Function and biomarkers of the blood-brain barrier in a neonatal germinal matrix haemorrhage model. Cells 10, 1677 (2021).
Semple, B. D., Blomgren, K., Gimlin, K., Ferriero, D. M. & Noble-Haeusslein, L. J. Brain development in rodents and humans: Identifying benchmarks of maturation and vulnerability to injury across species. Prog. Neurobiol. 106–107, 1–16 (2013).
Martins, C. A. et al. Neuroprotective effect of ACTH on collagenase-induced peri-intraventricular hemorrhage in newborn male rats. Sci. Rep. 10, 17734 (2020).
Paxinos, G. & Watson, C. The Rat Brain in Stereotaxic Coordinates 3rd edn (Academic Press, 1997).
Ceprián, M. et al. Cannabidiol reduces brain damage and improves functional recovery in a neonatal rat model of arterial ischemic stroke. Neuropharmacology 116, 151–159 (2017).
Ceprián, M. et al. Cannabidiol administration prevents hypoxia-ischemia-induced hypomyelination in newborn rats. Front. Pharmacol. 10, 1131 (2019).
Pazos, M. R. et al. Mechanisms of cannabidiol neuroprotection in hypoxic-ischemic newborn pigs: role of 5HT(1A) and CB2 receptors. Neuropharmacology 71, 282–291 (2013).
Barata, L. et al. Neuroprotection by cannabidiol and hypothermia in a piglet model of newborn hypoxic-ischemic brain damage. Neuropharmacology 146, 1–11 (2019).
Truettner, J. S., Bramlett, H. M. & Dietrich, W. D. Posttraumatic therapeutic hypothermia alters microglial and macrophage polarization toward a beneficial phenotype. J. Cereb. Blood Flow Metab. 37, 2952–2962 (2017).
Mecha, M., Carrillo-Salinas, F. J., Feliú, A., Mestre, L. & Guaza, C. Microglia activation states and cannabinoid system: therapeutic implications. Pharmacol. Ther. 166, 40–55 (2016).
Dean, J. M. et al. Strain-specific differences in perinatal rodent oligodendrocyte lineage progression and its correlation with human. Dev. Neurosci. 33, 251–260 (2011).
Volpe, J. J. The encephalopathy of prematurity-brain injury and impaired brain development inextricably intertwined. Semin. Pediatr. Neurol. 16, 167–178 (2009).
Newville, J., Jantzie, L. & Cunningham, L. Embracing oligodendrocyte diversity in the context of perinatal injury. Neural Regen. Res. 12, 1575 (2017).
Dorner, R. A., Burton, V. J., Allen, M. C., Robinson, S. & Soares, B. P. Preterm neuroimaging and neurodevelopmental outcome: a focus on intraventricular hemorrhage, post-hemorrhagic hydrocephalus, and associated brain injury. J. Perinatol. 38, 1431–1443 (2018).
Thompson, D. K. et al. Corpus callosum alterations in very preterm infants: perinatal correlates and 2year neurodevelopmental outcomes. Neuroimage 59, 3571–3581 (2012).
Segovia, K. N. et al. Arrested oligodendrocyte lineage maturation in chronic perinatal white matter injury. Ann. Neurol. 63, 520–530 (2008).
Kriegstein, A. & Alvarez-Buylla, A. The glial nature of embryonic and adult neural stem cells. Annu. Rev. Neurosci. 32, 149–184 (2009).
Choi, E.-K. et al. Animal models of periventricular leukomalacia. Lab. Anim. Res. 27, 77–84 (2011).
Yang, Y.-R. et al. Mfsd2a (major facilitator superfamily domain containing 2a) attenuates intracerebral hemorrhage-induced blood-brain barrier disruption by inhibiting vesicular transcytosis. J. Am. Heart Assoc. 6, e005811 (2017).
Saunders, N. R., Dziegielewska, K. M., Møllgård, K. & Habgood, M. D. Physiology and molecular biology of barrier mechanisms in the fetal and neonatal brain. J. Physiol. 596, 5723–5756 (2018).
Zimmer, J., Kristensen, B. W., Jakobsen, B. & Noraberg, J. Excitatory amino acid neurotoxicity and modulation of glutamate receptor expression in organotypic brain slice cultures. Amino Acids 19, 7–21 (2000).
Lin, S. et al. Heme activates TLR4-mediated inflammatory injury via MyD88/TRIF signaling pathway in intracerebral hemorrhage. J. Neuroinflammation 9, 548 (2012).
Bhalala, U. S., Koehler, R. C. & Kannan, S. Neuroinflammation and neuroimmune dysregulation after acute hypoxic-ischemic injury of developing brain. Front. Pediatr. 2, 1–12 (2015).
Zhao, H., Garton, T., Keep, R. F., Hua, Y. & Xi, G. Microglia/macrophage polarization after experimental intracerebral hemorrhage. Transl. Stroke Res. 6, 407–409 (2015).
Rempe, R. G. Matrix metalloproteinases in the brain and blood–brain barrier: versatile breakers and makers. J. Cereb. Blood Flow Metab. 36, 1481–1507 (2016).
Svedin, P., Hagberg, H., Sävman, K., Zhu, C. & Mallard, C. Matrix metalloproteinase-9 gene knock-out protects the immature brain after cerebral hypoxia-ischemia. J. Neurosci. 27, 1511–1518 (2007).
Qin, W. et al. Melatonin protects blood-brain barrier integrity and permeability by inhibiting matrix metalloproteinase-9 via the NOTCH3/NF-kB pathway. Aging 11, 11391–11415 (2019).
Xue, M. & Del Bigio, M. R. Comparison of brain cell death and inflammatory reaction in three models of intracerebral hemorrhage in adult rats. J. Stroke Cerebrovasc. Dis. 12, 152–159 (2003).
Acknowledgements
We thank the Flow Cytometry Unit of Hospital Clinico San Carlos-IdISSC for excellent technical assistance and Jason Willis-Lee MITI for medical writing assistance.
Funding
This work was supported by the PI19/00927 project, integrated into the Plan National de I + D + I, AES 2017-2020; funded by the ISCIII and co-funded by the European Regional Development Fund (ERDF) “A way to make Europe.”
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A.D.P.: conceptualization, methodology, investigation, data curation, writing—original draft. M.V., C.V., D.C.: investigation, data curation. E.F.-V.: conceptualization, investigation, resources. A.G.-R.: conceptualization, methodology, investigation, writing—original draft. J.M.-O.: conceptualization, methodology, formal analysis, resources, writing—original draft, writing—review and editing.
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Del Pozo, A., Villa, M., Vargas, C. et al. Intraventricular hemorrhage induces inflammatory brain damage with blood–brain barrier dysfunction in immature rats. Pediatr Res 93, 78–88 (2023). https://doi.org/10.1038/s41390-022-02062-3
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DOI: https://doi.org/10.1038/s41390-022-02062-3
