This study aims to investigate the relationship between oral glutamate administration and the progression of atherosclerosis in ApoE⁻/⁻ mice that have been fed a high-fat diet (HFD). Additionally, the study seeks to identify proteins associated with this process.
Atherosclerosis was induced in ApoE⁻/⁻ mice through a high-fat diet for a period of three months. The mice were assigned to four distinct groups:
To assess atherosclerotic lesions, Lillie's oil red O staining and histological analysis were utilized. Furthermore, proteomics profiling based on LC-MS/MS was conducted to identify proteins with varying expressions, with validation achieved through Western blotting and quantitative PCR (qPCR).
The administration of glutamate alone (in the Glu group) resulted in minimal plaque formation. However, the combination of the high-fat diet and glutamate (in the AS-GLU group) was linked to significantly larger atherosclerotic plaque areas compared to those on the high-fat diet alone (AS group). Proteomic analysis revealed that the proteins RAPGEF1, OPN, and MYL7 exhibited increased expression in the AS-GLU group when compared to the AS group. These associations were corroborated through validation techniques like Western blotting and qPCR.
This study provides evidence that the combination of oral glutamate administration with a high-fat diet in ApoE⁻/⁻ mice is associated with aggravated atherosclerosis and elevated expression of RAPGEF1, OPN, and MYL7. These proteins may act as candidate molecules linked to glutamate exposure, though their precise causal role in atherogenesis remains to be determined. Interestingly, serum levels of glutamate did not show significant increases, suggesting that changes in lipid profiles could also play a role in the observed effects.
Atherosclerosis (AS) is a significant inflammatory disease of the vascular system characterized by lipid accumulation, cell infiltration, and plaque formation in arterial walls. As a central contributor to cardiovascular disease, understanding its pathobiology is critical but remains an area of active research, particularly regarding dietary and metabolic influences.
Glutamate, a widely-used dietary additive, has recently drawn attention for its potential pathological effects beyond its traditional role. Epidemiological studies suggest a connection between plasma glutamate levels and cardiovascular risk, alongside subclinical atherosclerosis. However, it's unclear whether this association indicates a direct role during atherogenesis or stems from underlying metabolic disturbances.
In this study, we explored whether exogenous glutamate affects atherosclerosis progression away from systemic metabolic influences, utilizing the ApoE⁻/⁻ mouse model as it simulates hypercholesterolemia and atherosclerotic processes. By administering glutamate while controlling diet types, we aimed to isolate its potential effects on atherosclerosis, independent from endogenous variation. Results indicated that glutamate administration exacerbates AS progression when paired with a high-fat diet, despite not elevating serum glutamate levels substantially, suggesting localized vascular activity rather than systemic impacts.
A total of 40 male ApoE⁻/⁻ mice aged 42 to 48 days were acquired. Initial weights ranged from 19 to 21 grams. The mice were randomly divided into four treatment groups: the AS-GLU group received a high-fat diet and glutamate; the AS group received a high-fat diet with a placebo; the Glu group received a regular diet with glutamate; and the Control group received a regular diet with a placebo. Precise and consistent dosing of glutamate was ensured through oral administration as food intake could vary significantly among HFD-fed mice. All animals underwent euthanasia three months post-treatment, following ethical guidelines, with subsequent isolation and preparation of aortic tissues for analysis.
The aortic tissues underwent standard procedures for fixation, dehydration, and embedding. Histopathological evaluations included Hematoxylin and eosin (HE) staining, Masson trichrome, and Oil Red O to assess cellular architecture, collagen distribution, and lipid accumulation, respectively. Quantitative evaluations were conducted in a double-blinded manner utilizing Image J software to measure lesion areas, ensuring objective results across tested groups. Additionally, immunofluorescence assessments were performed for specific markers to evaluate inflammation and smooth muscle function within the aortal tissues.
This investigation underscores the necessity for further inquiry into the mechanistic implications of glutamate's dietary role concerning atherosclerosis, particularly concerning human health and dietary regulations.