## 3 Umbilical Cord and HUVEC
### Advantages and Disadvantages of the HUVEC Model  
An in vitro model must be accessible, easy to handle, reproducible and in accordance with an appropriate ethical framework. It must also reproduce the pathophysiology of the human disease to be studied as accurately as possible. In this sense, HUVEC more faithfully represents human EC behavior as compared to cell lines [73]. The HUVEC model is physiologically representative of the human vascular endothelium, allowing the study of the physiological and pathological effects of different stimuli both in an isolated form, and in co-culture with other cell types, such as leukocytes and smooth muscle cells [72]. It is also useful to study the effects of novel drugs on human endothelium [74]. Moreover, a recent innovation in gene editing provides the opportunity to study the effects of specific mutations in EC [1; 34]. Most importantly, the HUVEC model has been useful to study a broad array of biological processes and diseases, such as inflammation, apoptosis, preeclampsia, cancer, CVD and regenerative medicine [12; 13; 49; 58].  
Among technical advantages, cultured HUVEC are subject to contact inhibition and limitations in growth density, as cell proliferation is strongly inhibited once the culture has reached confluence [75]. Moreover, because different components of the extracellular matrix can promote the formation of capillary-like structures in HUVEC cultures, they are useful as angiogenesis models, although they do not reflect all the steps of physiological angiogenesis. These models are simple, particularly suitable for in vitro screening of angiostatic molecule activity and can also be used to evaluate the ultrastructure of capillary formation, matrix synthesis of EC or the role of adhesion molecules in the process of tubular morphogenesis. They are also useful to functionally characterize EC lines, to study the putative role of adhesion molecules (such as E-selectin, platelet endothelial cell adhesion molecule [PECAM-1], cadherin-5 and proteases), the synthesis of extracellular proteins and blood vessel maturation [37].  
Because the HUC is fetal and not maternal tissue, gender differences must be considered. Umbilical cord cells are subject to the effect of hormones produced by the fetal gonad in addition to maternal hormones. In fact, various studies have shown gender differences in male and female umbilical cords regarding gene expression, protein expression, cell viability after serum starvation, tube formation capacity, autophagy, intracellular ATP and metabolite levels, oxidative stress and angiogenesis [76; 77; 78]. Sexual dimorphism shown by HUVEC indicates that sex differences exist in prenatal life and are parameter-specific, suggesting that HUVEC of both sexes should be used as a model to increase the quality and the translational value of research. The sex differences observed in HUVEC could be relevant in explaining the adulthood diseases because endothelial dysfunction has a crucial role in the pathogenesis of diabetes mellitus, cardiovascular, neurodegenerative and immune disease [76]. Among possible disadvantages, while it was initially assumed that EC were homogeneous, it is now known that vascular cells show a high degree of heterogeneity, allowing biological adaptation to local needs [80]. In addition, the EC growth rate is much higher in vitro than in vivo; although this can be controlled using inhibitors and growth factors to make them quiescent [49]. Finally, the lifespan of HUVEC is approximately 10 passages, although some authors affirm that these cells lose their primary characteristics and ability to respond to stimuli after passage 6. Although HUVEC can be maintained in culture for up to five months, one of their main disadvantages is that they are not suitable for long-term experiments [75].