Unraveling the Effects of Nicotine-Derived Nitrosamine Ketone (NNK) on Healthy and Neurodegenerative Brain via Intravital Imaging and Multiscale Ex Vivo Imaging Approaches.

Cigarette Smoke (CS) continues to be a leading cause for decline in quality of life and deaths globally. While the link to poor health and early death has been accepted for decades, it is increasingly recognized smoking may contribute to neurological disease. Epidemiologically, CS has been associated with neuroinflammation and neurological disorders, including Alzheimer’s disease, stroke, and multiple sclerosis. Nicotine-derived nitrosamine ketone (NNK) is a nicotine metabolite produced within the tobacco plant from combustion and metabolic breakdown of nicotine. Extensive cancer literature has established NNK as an important component of CS resulting in DNA mutation and ability to bind and activate nicotinic acetylcholine receptors. However, little is known about specific in vivo effects of NNK on the blood brain barrier (BBB)or microglial immune cells of the brain as they relate to the vasculature. To investigate microglial and vascular responses to intranasal NNK in vivo, we combined the use of a transgenic mouse (CX3CR-1GFP) with intravital neuroimaging. We show that NNK given intranasally over acute (4-7days) and chronic (12 weeks) exposures, leads to disruption of the BBB and heterogenous microglia activation. Machine learning algorithm was employed to segment microglial soma from processes, and analyses performed to evaluate morphometry and associations with vasculature. In vivo microglia activation was found e heterogeneous; with ameboid activated microglia localized to the vasculature. An increase in vessel-associated microglia compared to PBS controls was found. Pronounced effect of NNK on vasculature included vasoconstriction, vasodilation, and microhemorrhage. In chronic treatment, these effects were sustained and, in some cases, exacerbated. Both neuronal and DNA damage was found; interestingly the later confined to cells other than microglia indicating microglia may be acting in a productive phagocytic function (quantified in vivo). The transgenic 3xTg-AD (Alzheimer’s disease) mouse model was used to investigate the vascular and microglia effects of NNK during ongoing disease progression, and marked increase in vascular events, similar to CX3CR-1 GFP were found. Lipofuscin autofluorescence was observed in intravital imaging for both PBS and NNK-treated groups and showed overall increase in lipofuscin deposition with NNK treatment over 4 days vs. PBS. Vessel leakage was also increased with NNK. Large-scale microscopy of optically cleared mouse lungs revealed an increase of internal red blood cell deposition in NNK-treated groups and distorted alveolar structure that was not observed in PBS-treated groups. Taken together, this work suggests an immunomodulatory effect of NNK marked by highly vessel-localized microglia activity and vascular dysfunction that can produce long-term pathophysiology that could contribute to or exacerbate ongoing or developing disease processes. This work lays the groundwork for studies examining the potential role of other components in cigarettes or e-cigs as it relates to neurological disorders as well as studies examining the multiorgan effects (lung-brain) of NNK.

Name thesis:degree_name

Neuroscience (Doctoral)

Discipline thesis:degree_discipline

Neuroscience

Grantor

The University of Texas Medical Branch at Galveston

Year dc:date.issued

2023

Author dc:creator

• Ochoa, Lorenzo Francisco 09/06/1986-

Advisor dc:contributor.advisor

• Vargas, Gracie (grvargas@utmb.edu)

Committee members dc:contributor.committeemember

• Motamedi, Massoud
• Bucchieri, Fabio
• Ameredes, Bill
• Taglialatela, Giulio

Language dc:language.iso

English

Handle dc:identifier.uri

https://hdl.handle.net/2152.3/12493

OAI identifier oai:identifier

oai:utmb-ir.tdl.org:2152.3/12493