Vascular endothelial growth factor signaling in obesity and adipose tissue microstructures: A meta-analysis

Introduction::
Obesity is a major public health issue that increases the risk of serious comorbidities. Angiogenesis, the formation of new blood vessels, is a critical process for adipose tissue expansion, which is mediated by the binding of vascular endothelial growth factor (VEGF) to its receptors (VEGFR1 and VEGFR2) and co-receptor (NRP1). While experimental studies have suggested the relationship between VEGF signaling and obesity (1,2), the precise role of VEGF remains unclear. To address this knowledge gap, we propose using systems biology to examine the role of VEGF signaling in obesity.

Although systems biology has been used to examine the role of VEGF in cancer and peripheral vascular disease, there are unique challenges to modeling adipose tissue due to several distinct features, such as: 1) the large proportion of adipocytes not present in previously modeled systems, 2) highly vascularized, yet phenotypically different endothelium in adipose tissue, and the 3) differential VEGFRs levels in adipose tissue. This study analyzes two types of parameters: 1) adipose tissue geometry parameters including: adipocyte size and capillary basement membrane (BM) thickness, and 2) VEGF binding kinetics to VEGFRs and NRP1. Our study aims to fill the knowledge gap on the role of VEGF signaling in obesity, enabling the development of obesity-specific computational models, which could lead to novel therapeutic strategies for obesity-related disorders.




Materials and Methods::
We conducted a literature search through PubMed, Google Scholar, and the University of Washington Libraries for this meta-analysis. Ex vivo studies measuring adipocyte size and capillary BM thickness were included for lean and obese murine species. Here, we calculated the weighted average and standard deviation for each data set. We included in vitro studies that reported equilibrium dissociation constants (Kd) using chip-based (surface plasmon resonance; SPR) and cell-based (radioligand) assays. We excluded studies reporting significantly different Kd values and an enzyme-linked immunosorbent assay. Standard errors or standard deviations were also required, resulting in a total of 34 studies included, as follows (data, n=number of studies): adipocyte size, n=10; capillary BM thickness, n=6; VEGF-A—VEGFR1 binding, n=8; VEGF-A—VEGFR2 binding, n=9; and VEGF-A—NRP1 binding, n=8. We used a non-pooled Student’s t-test to determine the association of the geometric parameters with obesity. The same approach was used to analyze the binding affinities of VEGF to its receptors and to identify statistical differences in cell- and chip-based assays.




Results, Conclusions, and Discussions::
Results and Discussion
Our study analyzed ten experiments and found that adipocyte size in obese mice is significantly larger than in lean mice (Fig. 1; 76±4.3 vs. 45±3.8 µm; p < 0.001), which was expected due to higher lipid content. However, we found that capillary BM thickness in obese mice is similar to that of lean mice (80±17 nm vs. 70±5.2 nm; p = 0.638>0.05), which was unexpected given the high expression of collagen IV, the most abundant BM component, in obese mice (3). Further analysis suggests that age-related factors may play a more significant role in determining BM thickness (4), and the effects of tissue type may need to be considered.

Our study also compared VEGF-A binding affinities measured by cell-based, radioligand assays, and chip-based, surface plasmon resonance (SPR). There was no significant difference in weighted averages for VEGFR1 (39±20 pM vs. 5.9±2.6 pM; p = 0.17 > 0.05), VEGFR2 (483±174 pM vs. 48±16.7 pM; p = 0.08>0.05), and NRP1 (0.27±0.04 nM vs. 2.69±1.96 nM; p = 0.33>0.05) (Fig. 2). These non-significant differences are possibly attributed to large between-study variations. Notably, the binding affinities are affected by interaction conditions including temperature and pH. Despite the absence of statistical significance in our analysis, it's important to acknowledge a noticeable divergence in the measurements obtained from SPR and radioligand assays. Thus, computational modelers should be cautious when selecting values to build models and recognize the potential variations in binding affinities resulting from different binding assays.



Conclusions
Our study provides a comprehensive understanding of the parameters influencing VEGF signaling in obesity through integrative statistical analysis. Adipocyte size in obese mice is significantly larger (about 70%) than in lean mice, but capillary BM thickness shows similarity, potentially due to aging and tissue type. The study also highlights the importance of considering differences in binding affinities measured by cell-based and chip-based assays when building computational models, as these could influence model outcomes. Our research lays a foundation for the development of a computational model for VEGF signaling in obesity, contributing to the understanding of this complex pathway in metabolic disorders.


Acknowledgements (Optional): :
We thank Dr. Vincenza Cifarelli for providing invaluable support through discussions. This material is based upon work supported by the National Science Foundation under Grant No. 1923151. Research reported in this publication was also supported by the National Institute of Heart, Lung, and Blood of the National Institutes of Health under Award Number 7R01HL159946-02. Any opinions, findings, and conclusions or recommendations expressed in this material are those of the author(s) and do not necessarily reflect the views of the National Science Foundation or the National Institutes of Health.




References (Optional): :
1. Claffey KP, Wilkison WO, Spiegelman BM. Vascular endothelial growth factor. Regulation by cell differentiation and activated second messenger pathways. Journal of Biological Chemistry. 1992;267(23).




2. Nishimura S, Manabe I, Nagasaki M, Hosoya Y, Yamashita H, Fujita H, et al. Adipogenesis in obesity requires close interplay between differentiating adipocytes, stromal cells, and blood vessels. Diabetes. 2007;56(6).




3. Chen H jian, Yan X yue, Sun A, Zhang L, Zhang J, Yan Y e. High-Fat-Diet-Induced Extracellular Matrix Deposition Regulates Integrin—FAK Signals in Adipose Tissue to Promote Obesity. Molecular Nutrition & Food Research. 2022;66(7).




4. Creutzfeldt W, Mende D, Willms B, Söling HD. Vascular basement membrane thickness in muscle of spiny mice and activities of glycolysis and gluconeogenesis in the liver of animals with spontaneous and experimental diabetes and of untreated human diabetics. Diabetologia. 1970;6(3).