tetano
Editor, Senior Moderator
FASEB J
. 2022 May;36 Suppl 1.
doi: 10.1096/fasebj.2022.36.S1.0R714.
Assessing the effects of COVID-19-induced hypoxia and ACE2 signaling on the human blood-brain barrier in-vitro
Iqra Pervaiz[SUP] 1 [/SUP], Sree Pooja Varahachalam[SUP] 1 [/SUP], Abraham J Al-Ahmad[SUP] 1 [/SUP]
Affiliations
Abstract
Introduction: Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV2). The portal of entry for the virus is peptidase ACE2, a major key player of the renin-angiotensin-aldosterone system (RAAS), resulting in severe lung injury. Although COVID-19 mainly manifests as an acute respiratory distress syndrome (ARDS), there is increasing evidence of neurological symptoms in patients infected by COVID-19. Yet, there is limited understanding of how COVID-19 impacts the central nervous system (CNS). We speculate that such neurological symptoms maybe a consequence of a dysfunction of the blood-brain barrier (BBB) with our central hypothesis that the neurological effects of SARS CoV-2 are driven by chronic hypoxic stress-impaired ACE2 at the BBB.
Methods: An in-vitro human induced pluripotent stem cells (hiPSCs) BBB model was used in the study. Such model was exposed to various levels of hypoxia (1,5 and 10%) for up to 24 hours. In addition, normoxic cells were treated with Angiotensin II (AngII) or Angiotensin 1-7 (degradation by product of AngII by ACE2). Changes in the barrier function was assessed using TEER, permeability to fluorescein and tight junction staining. Changes in ACE2 and MasR expression was assessed by immunofluorescence, whereas ACE2 shedding and HIF-1 alpha expression was assessed by ELISA.
Results: Mild (10%) hypoxia was sufficient to induce the loss of barrier function. Secretion of ACE2 under hypoxia followed a biphasic pattern, with highest levels at 5% and 10%. Ang II and Ang1-7 had little effect on the barrier function under normoxic condition. The hypoxic exposure induced shedding of the membrane bound ACE2 and molecular mechanism of hypoxic exposure in regulation of ACE2 occurs in a HIF1α- dependent manner.
Discussion: Our preliminary data suggest that our human model of the BBB responds to hypoxia and express critical components of RAAS. Both ACE2 and MasR negatively respond to mild hypoxia followed by a decreased barrier function with no changes in tight junction complex. Such loss was correlated with increased ACE2 shedding in HIF1α- dependent manner. We are currently investigating role of Ang 1-7 in rescuing barrier function under hypoxic stress.
. 2022 May;36 Suppl 1.
doi: 10.1096/fasebj.2022.36.S1.0R714.
Assessing the effects of COVID-19-induced hypoxia and ACE2 signaling on the human blood-brain barrier in-vitro
Iqra Pervaiz[SUP] 1 [/SUP], Sree Pooja Varahachalam[SUP] 1 [/SUP], Abraham J Al-Ahmad[SUP] 1 [/SUP]
Affiliations
- PMID: 35560704
- DOI: 10.1096/fasebj.2022.36.S1.0R714
Abstract
Introduction: Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV2). The portal of entry for the virus is peptidase ACE2, a major key player of the renin-angiotensin-aldosterone system (RAAS), resulting in severe lung injury. Although COVID-19 mainly manifests as an acute respiratory distress syndrome (ARDS), there is increasing evidence of neurological symptoms in patients infected by COVID-19. Yet, there is limited understanding of how COVID-19 impacts the central nervous system (CNS). We speculate that such neurological symptoms maybe a consequence of a dysfunction of the blood-brain barrier (BBB) with our central hypothesis that the neurological effects of SARS CoV-2 are driven by chronic hypoxic stress-impaired ACE2 at the BBB.
Methods: An in-vitro human induced pluripotent stem cells (hiPSCs) BBB model was used in the study. Such model was exposed to various levels of hypoxia (1,5 and 10%) for up to 24 hours. In addition, normoxic cells were treated with Angiotensin II (AngII) or Angiotensin 1-7 (degradation by product of AngII by ACE2). Changes in the barrier function was assessed using TEER, permeability to fluorescein and tight junction staining. Changes in ACE2 and MasR expression was assessed by immunofluorescence, whereas ACE2 shedding and HIF-1 alpha expression was assessed by ELISA.
Results: Mild (10%) hypoxia was sufficient to induce the loss of barrier function. Secretion of ACE2 under hypoxia followed a biphasic pattern, with highest levels at 5% and 10%. Ang II and Ang1-7 had little effect on the barrier function under normoxic condition. The hypoxic exposure induced shedding of the membrane bound ACE2 and molecular mechanism of hypoxic exposure in regulation of ACE2 occurs in a HIF1α- dependent manner.
Discussion: Our preliminary data suggest that our human model of the BBB responds to hypoxia and express critical components of RAAS. Both ACE2 and MasR negatively respond to mild hypoxia followed by a decreased barrier function with no changes in tight junction complex. Such loss was correlated with increased ACE2 shedding in HIF1α- dependent manner. We are currently investigating role of Ang 1-7 in rescuing barrier function under hypoxic stress.