Pilar Alves-Martinez, Monica Garcia-Alloza

There are around 15 million preterm newborns (PT) every year (Atienza-Navarro et al., 2020). With these figures in mind, prematurity represents a major health problem worldwide and it is a leading cause of infant mortality, accounting for up to 35% of all deaths among newborns (Atienza-Navarro et al., 2020) and up to 18% of the deaths among children under 5 years of age. Although advances in neonatology have significantly reduced mortality rates associated with prematurity, related comorbidities and sequelae remain a crucial societal and economic burden (Alves-Martinez et al., 2022). PT are born at a critical time in brain development, thus they have an immature central nervous system characterized by low neuronal migration, poor myelination, impaired grey matter growth matter or a fragile vascular structure of the germinal matrix (Atienza-Navarro et al., 2020), among others. The germinal matrix is a highly vascularized subependymal region adjacent to the ventricle. This tissue is prone to bleeding, due to the high vascularisation of the region and the hemodynamic instability of the PT (Atienza-Navarro et al., 2020), causing a germinal matrixintraventricular hemorrhage (GM-IVH). GM-IVH is the most common intracerebral hemorrhage of the PT and affects up to 20–30% of these patients (Alves-Martinez et al., 2022). When GMIVH occurs, neuronal loss, white matter lesions and inflammation are observed. The extension of the lesions, as well as related problems and sequelae, are highly dependent on the severity of the bleeding (Atienza-Navarro et al., 2020) and may include short- and long-term, neurological, sensory, cognitive and motor disabilities or neuropsychiatric disorders (Atienza-Navarro et al., 2020).

Whereas the health and societal relevance of GMIVH seems out of question, there are no successful treatments to prevent or ameliorate GM-IVH (Alves-Martinez et al., 2022) and patients are in a tremendous need of new therapeutic approaches that may limit GM-IVH brain pathology and complications. In this sense, caffeine is regularly used to treat apnea in PT (Di Martino et al., 2020) and previous studies have shown its antiinflammatory and neuroprotective properties (Yang et al., 2022). Caffeine is a methylxanthine with a similar structure to adenosine (Kolahdouzan and Hamadeh, 2017); thus, caffeine acts as an antagonist of adenosine and can bind to its receptors (A1, A2, A3 and A4). Specifically, A2 receptors are highly expressed in the brain, where they are involved in the inflammatory cascade, leading to microglial activation and the generation of reactive oxygen species and inflammatory mediators ultimately contributing to neuronal death (Kolahdouzan and Hamadeh, 2017), which relates to these receptors with different neurodegenerative processes. Therefore, caffeine might provide an interesting approach to limiting brain neurodegeneration and inflammation in GM-IVH by blocking adenosine receptors. Nevertheless, the number of studies addressing the effects of caffeine in GM-IVH of the PT is still limited, hampering the understanding of the treatment. Thus, the recent study by Alves-Martinez et al. (2022) provides new insights into the neuroprotective role of caffeine. Authors show that caffeine treatment reduces neuronal loss and inflammation; it has a positive effect on proliferation and neurogenesis and improves cognitive function in a murine model of the disease.

Clinical trial with caffeine for PT complications:Several epidemiological studies have reported the beneficial effects of some drugs on GMIVH, including prophylactic vitamin K, angiogenic inhibitors, or endothelial growth factor R2 inhibitors, among others (Alves-Martinez et al., 2022). Nevertheless, further studies are needed before they can be used in the Neonatology clinical care unit. On the other hand, previous studies have also shown the neuroprotective and anti-inflammatory effects of caffeine; though, to our knowledge, studies focusing on the direct effect of caffeine on GM-IVH are limited. Clinical trials usually address other complications of prematurity (mostly apnea of prematurity), while the effects of caffeine on GM-IVH are largely secondary outcomes. Most studies converge on the safety of caffeine as both a short- and longterm treatment and a previous study with 2006 PT infants showed that patients using caffeine to treat apnea of prematurity had better survival rates and cognitive performances at 18 and 21 months of age (Schmidt et al., 2017). Another study on a cohort of 70 very low-weight PT has shown that caffeine treatment results in an improvement in white matter microstructural development (Doyle et al., 2010). Furthermore, caffeine treatment in PT may improve motor function by 5 years of age (Schmidt et al., 2017) and improve cognitive skills by 11 years of age (Murner-Lavanchy et al., 2018). Interestingly, early administration of caffeine may be neuroprotective and reduce GMIVH incidence. In this sense, a clinical trial with 986 infants ≤ 32 gestational weeks with respiratory distress syndrome (Borszewska-Kornacka et al., 2017) (magnetic resonance imaging and neurodevelopment in PT after administration of high doses of caffeine. ClinicalTrials. gov. Identifier: NCT00809055) showed that early (in the first 24 hours of life) administration of caffeine resulted in the reduced need of ventilatory support, mortality and incidence of prematurity-related complications, including GM-IVH. Whereas the benefits were more robust in the group with less severe GM-IVH, the results also revealed that late administration of caffeine (day 2+ of life) significantly limits the positive effects of the treatment, showing that caffeine may not only be neuroprotective but it may also have preventive effects, by reducing GM-IVH incidence when administered early.

Effect of caffeine in a murine model of GMIVH of the PT:Previous studies have shown the neuroprotective and anti-inflammatory effects of caffeine. These effects seem to be mediated by caffeine binding to adenosine receptors that are expressed in several cell types in central nervous system, including astrocytes, oligodendrocytes and microglia (Kolahdouzan and Hamadeh, 2017). Caffeine blockade of adenosine receptors reduces inflammation and apoptosis, by inhibiting adenosine-mediated activation of protein Gαs, and consequent activation of adenylyl cyclase, reducing the cAMP formation and protein kinase A activation (Kolahdouzan and Hamadeh, 2017). On the other hand, adenosine binding to adenosine 2A receptor also results in microglial activation by glutamate release, as glutamate reuptake is inhibited and extracellular signalregulated kinase is up-regulated (Kolahdouzan and Hamadeh, 2017). Therefore, caffeine also limits these deleterious effects on the central nervous system. Caffeine treatment ameliorates the inflammatory damage caused by hypoxiaischemia in neonatal rodent models by inhibiting NLRP3 (NOD-, LRR- and pyrin domain-containing protein 3) inflammasome activation, reducing the release of inflammatory factors or reducing microglial activation (Yang et al., 2022) and the number of amoeboid microglia (Di Martino et al., 2020). In addition, caffeine reduces the number of reactive astrocytes (Di Martino et al., 2020), the release of oxidative stress markers, the expression of inflammatory cytokines and the number of apoptotic cells (Endesfelder et al., 2017). Interestingly, other studies show an improvement in long-term cognitive function in neonatal rat models of hypoxia-ischemia (Potter et al., 2018).Animal models that resemble GM-IVH mostly rely on the intraventricular administration of autologous blood or collagenase (Atienza-Navarro et al., 2020). Bacterial collagenase is an enzyme with proteolytic action that degrades the extracellular matrix of brain blood vessels, thus producing cerebral hemorrhage. Mice with GM-IVH present reduced cortical size and ventriculomegaly, an indicator of brain atrophy and a relevant characteristic of GM-IVH, as well as neuronal loss (Alves-Martinez et al., 2022). Neuron population is also affected in both. Small vessel damage is accompanied by a significant inflammatory response (Segado-Arenas et al., 2018). The damage to the subventricular zone also results in reduced proliferation and neurogenesis, limiting the regenerative process after the insult (Segado-Arenas et al., 2018). Interestingly, peripheral markers of brain damage, such as gelsolin levels (Segado-Arenas et al., 2018), are significantly reduced, supporting the utility of the model. Moreover, cognitive impairment, affecting learning and memory capacities can be detected in young adults, supporting the chronic behavioral limitations (Figure 1A).

Given the suitability of the model, this approach provides a relevant tool to specifically assess the role of caffeine on GM-IVH of the PT. Therefore, a recent study has analyzed the possible direct effect of caffeine by administering 10 or 20 mg/Kg of caffeine for 3 consecutive days after inducing a GM-IVH in P7 mice. Caffeine treatment results in a significant improvement in short and longterm pathological and behavioral complications in this animal model (Alves-Martinez et al., 2022). Caffeine reduces cortical thinning and ventricle enlargement, altogether limiting brain atrophy. Similarly, caffeine ameliorated brain injury in a hypoxic-ischemic neonatal model in rodents, an effect proposed to be mediated by the blockade of the adenosine A1 receptor (Alexander et al., 2013). In addition, another study of prenatal hypoxic-ischemia in rodents described a reduction of tissue damage after caffeine treatment (Di Martino et al., 2020). Besides, a general improvement in neuronal wellbeing is observed and tau hyperphosphorylation and neuronal curvature are improved, while the neuronal population is preserved in the cortex after the treatment with caffeine. Previous studies have reported similar results in other murine models, supporting the implication of adenosine 2A receptors in tau phosphorylation and showing that genetic or pharmacological blockade of adenosine 2A receptors results in decreased tau hyperphosphorylation (Laurent et al., 2016). Likewise, the neuronal population is preserved by caffeine treatment and proliferation and neurogenesis in the subventricular zone are restored. In line with these observations, caffeine enhances neuron survival and inhibits the accumulation of hypoxia inducible factor-1α after hypoxic injury, both in primary cultures and in neonatal mouse pups (Li et al., 2019). Furthermore, other studies have shown amelioration in dementia-induced impaired neurogenesis in Alzheimer’s disease mouse models (Stazi et al., 2021). Caffeine treatment effectively limits vascular pathology by reducing the presence of hemorrhages in the subventricular zone and the cortex. Also, the inflammatory response is ameliorated and microglia load is reduced after caffeine treatment, in line with previous observations showing the capacity of caffeine to inhibit NLRP3 inflammasome activation, promote microglial anti-inflammatory phenotype and decrease proinflammatory cytokine release in a hypoxic-ischemic model in neonatal rats (Yang et al., 2022). More importantly, better performances are observed when episodic and working memories are assessed (Alves-Martinez et al., 2022). Whereas it is not possible to point towards a single cause, it is feasible that the pleiotropic effects of caffeine may account for observed improvements at the cognitive level (Figure 1B).

Figure 1｜Schematic effects of caffeine in a murine model of GM-IVH of the preterm newborn.

Ultimately, these results contribute to characterizing the brain pathology of GM-IVH of the PT and to further understanding the positive effects that caffeine treatment may have. Since the number of studies addressing the role of caffeine in GM-IVH of the PT is still limited, the fact that the treatment successfully limits neuronal damage, vascular alterations, inflammation and cognitive problems support to continue the studies to fully characterize the use of caffeine to reduce complications and sequelae associated with GM-IVH of the PT.

The present work was supported by the following funds: To PAM: predoctoral fellowship. Instituto de Investigacion Biomedica de la Provincia de Cadiz (INIBICA). To MGA: Agencia Andaluza del Conocimiento. Proyectos I + D + I—Programa Operativo FEDER Andalucia 2014–2020 fondos FEDER (SOL201800107189-TRA). Agencia Estatal de Investigacion. Ministerio de Educacion y Ciencia. Programa Estatal de Generacion de Conocimiento y Fortalecimiento Cientifico y Tecnologico del

Sistema de I + D + i y del Programa Estatal de I + D + i Orientada a los Retos de la Sociedad, del Plan Estatal de Investigacion Cientifica y Tecnica y de Innovacion 2017–2020 (PID2020-115499RB-I00/AEI/10.13039/501100011033).

*Correspondence to:Monica Garcia-Alloza, PhD, monica.garcia@uca.es.

https://orcid.org/0000-0003-1610-4114(Monica Garcia-Alloza)

Date of submission:September 24, 2022

Date of decision:November 19, 2022

Date of acceptance:December 8, 2022

Date of web publication:January 30, 2023

https://doi.org/10.4103/1673-5374.363830

How to cite this article:Alves-Martinez P, Garcia-Alloza M (2023) Effect of caffeine in the intraventricular hemorrhage of the preterm newborn. Neural Regen Res 18(9):1942-1943.

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