*3.1. In Physiology*

HS are loaded with biological roles, as illustrated in Figure 2A. Acting as mediators in a multitude of regulatory mechanisms, ranging from embryonic development to ECM assembly and regulation of cell signalling [6,47,48]. HS interact with a plethora of molecular partners, including soluble proteins (growth factors, morphogens and chemokines), ECM proteins, bioactive fragments and membrane receptors, such as integrins and receptor tyrosine kinase (RTKs). HS chains also promote pathogen attachment and invasion of specific tissues by binding to numerous microorganisms, including viruses, bacteria, parasites and fungi [2,5,49–51]. Moreover, HSPGs are expressed in all main organ systems having essential roles in several biological activities like metabolism regulation, transcellular transport, cellular communication, ECM support and modulation. The classical role attributed to cell surface HSPGs was to assist as signalling co-receptor for growth factors activity, allowing a correct presentation to their cognate receptors and helping to stabilise gradients, to control the range of signalling and to protect the proteins against degradation [1]. However, it has been increasingly accepted that besides these co-receptor functions, HSPGs stand alone as key regulators of cell behaviour [52].

During embryonic development, HSPGs modulate the morphogen gradients distribution and other extracellular ligands signalling involved in the formation of the different tissue architectures [53]. In this light, the particular interaction of HS with the Hedgehog signalling pathway is very important to a proper embryonic development [54]. Similarly, HSPGs, being the most abundant PGs in basal lamina and cell surface of skeletal muscle, have been shown to regulate fibroblast growth factor (FGF)**,** Wnt and bone morphogenetic protein pathways, fundamental for the development of skeletal structures [55].

More recently, it was revealed that SDCs can regulate calcium channels of the TRPC (transient receptor potential canonical) family, with functional consequences on the actin cytoskeleton, cell adhesion, junctions and migration. Moreover, this interaction was suggested to be evolutionary conserved and relevant for the progression of some diseases [48,56].

HSPGs are also important modulators of metabolism, as illustrated by their role in the liver mediated clearance of triglyceride-rich lipoproteins [57]. Additionally, several SDCs and GPCs have been implicated in the uptake of different forms of lipoproteins [58–60].

Given the many essential cellular and developmental processes in which HS and HSPGs are involved, it is expected that modifications in HSPG expression and structure contribute to a dysregulation in function and lead to pathological scenarios, such as cancer [12,17]. In Section 4, we address several cancer cellular features that are regulated by changes in expression, glycosylation and sulfation profiles of HSPGs, which in turn translate into cancer progression.

**Figure 2.** Overview of heparan sulfate proteoglycans functions. (**A**) HSPGs roles in cellular activities, (**B**) HSPGs aberrant expression and functional implications in cancer and (**C**) HSPGs biomedical potential as a biomarker and as a therapeutic target regarding cancer improved diagnosis and prognosis.
