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The surface of vascular endothelial cells (ECs) is covered by a protective negatively charged layer known as the endothelial glycocalyx. Herein, we hypothesized its transport barrier and mechanosensory role in transmural water flux and low-density lipoprotein (LDL) transport in an isolated rat abdominal aorta perfused under 85 mmHg and 20 dyn/cm 2 ex vivo. The endothelial glycocalyx was digested by hyaluronidase (HAase) from bovine tests. Water infiltration velocity ( V ) was measured by a graduated pipette. LDL coverage and mean maximum infiltration distance (MMID) in the vessel wall were quantified by confocal laser scanning microscopy. EC apoptosis was determined by the terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) technique, and leaky junction rates were evaluated by electron microscopy. The results showed that a 42% degradation of the endothelial glycocalyx by HAase treatment increased w ) was measured by a graduated pipette. LDL coverage and mean maximum infiltration distance (MMID) in the vessel wall were quantified by confocal laser scanning microscopy. EC apoptosis was determined by the terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) technique, and leaky junction rates were evaluated by electron microscopy. The results showed that a 42% degradation of the endothelial glycocalyx by HAase treatment increased V , which cannot be inhibited by HAase treatment. Four hour-shear application increased about fourfolds of LDL coverage, whereas exerted no significant effects on its MMID, EC apoptosis, and the leaky junctions. On the contrary, 24-h shear exposure has no significant effects on LDL coverage, whereas increased 2.74-folds of MMID and about 53% of EC apoptotic rates that could be inhibited by HAase treatment. These results suggest endothelial glycocalyx acts as a transport barrier by decreasing water and LDL transport, as well as a mechanosensor of shear to regulate EC apoptosis, thus affecting leaky junctions and regulating LDL transport into the vessel wall. w A 42% degradation of the endothelial glycocalyx by hyaluronidase of the isolated rat abdominal aorta facilitated water and LDL transport across the vessel wall, suggesting endothelial glycocalyx as a transport barrier. A 24-h shear exposure increased LDL mean maximum infiltration distance, and enhanced EC apoptosis, which could be both inhibited by hyaluronidase treatment, suggesting endothelial glycocalyx may also act as a mechanosensor of shear to regulate EC apoptosis, thus affecting leaky junctions and regulating LDL transport.V w , which cannot be inhibited by HAase treatment. Four hour-shear application increased about fourfolds of LDL coverage, whereas exerted no significant effects on its MMID, EC apoptosis, and the leaky junctions. On the contrary, 24-h shear exposure has no significant effects on LDL coverage, whereas increased 2.74-folds of MMID and about 53% of EC apoptotic rates that could be inhibited by HAase treatment. These results suggest endothelial glycocalyx acts as a transport barrier by decreasing water and LDL transport, as well as a mechanosensor of shear to regulate EC apoptosis, thus affecting leaky junctions and regulating LDL transport into the vessel wall. NEW & NOTEWORTHY A 42% degradation of the endothelial glycocalyx by hyaluronidase of the isolated rat abdominal aorta facilitated water and LDL transport across the vessel wall, suggesting endothelial glycocalyx as a transport barrier. A 24-h shear exposure increased LDL mean maximum infiltration distance, and enhanced EC apoptosis, which could be both inhibited by hyaluronidase treatment, suggesting endothelial glycocalyx may also act as a mechanosensor of shear to regulate EC apoptosis, thus affecting leaky junctions and regulating LDL transport.

Hypercholesterolemia- and atherosclerosis-caused vasomotor property dysfunction may be involved in many clinic manifestations of atherosclerosis, including angina, acute myocardial infarction, and sudden cardiac death. However, its underlying mechanism is not clear. The endothelial glycocalyx is a protective surface layer on the endothelial cells, serving as a molecular sieve, cell adhesion modulator, and mechanosensor for blood flow. In the present study, we demonstrated by confocal microscopy in Sprague-Dawley (SD) male rats fed a 12-wk high-cholesterol diet (HC) compared with the normal diet (NC) that the dimension of the endothelial glycocalyx reduced significantly in both the common carotid artery (2.89 ± 0.41 µm and 3.25 ± 0.44 μm, respectively) and the internal sinus region (2.35 ± 0.07 µm and 3.46 ± 0.86 μm, respectively). Furthermore, we showed by real-time PCR that this dimension modification of endothelial glycocalyx may be attributed to a significant downregulation of heparan sulfate proteoglycan (HSPG)-related genes, including syndecan-3, glypican-1, and EXT1, not resulting from an enhanced shedding of sulfated glycosaminoglycans (sGAGs) from the vessel wall to the plasma. Meanwhile, the mean contraction and relaxation forces of the common carotid artery with responses to norepinephrine (NE) and acetylcholine (ACh) decreased ~0.34- and 0.13-fold, respectively, accompanied by a lower level of nitric oxide (NO) release. These findings suggest that the atherogenic high cholesterol diet diminished endothelial glycocalyx and disturbed the local NO release, thus contributing to the impaired vasomotor properties of the vessel. NEW & NOTEWORTHY Twelve-week high-cholesterol (HC) diet reduces the thickness of the endothelial glycocalyx in Sprague-Dawley (SD) male rats, which is mainly attributed to a downregulation of heparan sulfate proteoglycan-related genes (syndecan-3, glypican-1, EXT1), not resulting from an enhanced shedding of sulfated glycosaminoglycans (sGAGs) into the plasma. HC-diminished glycocalyx may disturb its mechanotransduction of local shear stress, lower nitric oxide (NO) release, and impair vasomotor responses to norepinephrine (NE) and acetylcholine (ACh).

The importance of vascular cell glycocalyx in mechanotransduction has been demonstrated by many studies. The simulated microgravity induced a region-dependent adaptation of arterial glycocalyx including its thickness, coverage, and gene expression in conduit arteries of tail-suspended rats has been reported in our previous studies. Herein, we extended this line of research by quantifying the mRNA levels of three nitric oxide synthase (NOSI, NOSII, and NOSIII) and evaluating the apoptotic rates of endothelial cells (ECs) and smooth muscle cells (SMCs) in the common carotid artery, abdominal aorta, and femoral artery of 3 week tail-suspended rats. Results indicated that the tail suspension of rats induced about 0.36, 0.22, and 0.33 fold down-regulation of NOSI, NOSII, and NOSIII in the abdominal aorta, while 3.21, and 3.48 fold up-regulation of NOSII and NOSIII in the carotid artery and no significant effects on three NOS isoforms in the femoral artery. Moreover, the apoptosis of ECs and SMCs were significantly inhibited in both carotid artery and abdominal aorta, while enhanced in the femoral artery of the tail-suspended rats. A linear positive correlation exists between the normalized coverage of the glycocalyx and the normalized NOSI and NOSIII mRNA levels. These results indicated that the redistribution of haemodynamics in the conduit arteries of 3 week tail-suspended rats regulated the glycocalyx, NOS expression, and vascular cell apoptosis in a region-dependent manner, contributing to the final vascular remodelling under simulated microgravity condition., (© 2019 John Wiley & Sons Australia, Ltd.)

Cancer is a malignant tumor that threatens the health of human beings, and has become the leading cause of death in urban and rural residents in China. The glycocalyx is a layer of multifunctional glycans that covers the surfaces of a variety of cells, including vascular endothelial cells, smooth muscle cells, stem cells, epithelial, osteocytes, as well as cancer cells. The glycosylation and syndecan of cancer cell glycocalyx are unique. However, heparan sulfate (HS), hyaluronic acid (HA), and syndecan are all closely associated with the processes of cancer progression, including cell migration and metastasis, tumor cell adhesion, tumorigenesis, and tumor growth. The possible underlying mechanisms may be the interruption of its barrier function, its radical role in growth factor storage, signaling, and mechanotransduction. In the later sections, we discuss glycocalyx targeting therapeutic approaches reported in animal and clinical experiments. The study concludes that cancer cells' glycocalyx and its role in cancer progression are beginning to be known by more groups, and future studies should pay more attention to its mechanotransduction of interstitial flow-induced shear stress, seeking promising therapeutic targets with less toxicity but more specificity.

Vascular remodeling induced by hemodynamic stimuli contributes to the pathophysiology of cardiovascular diseases. The importance of vascular cells (endothelial cells and smooth muscle cells) glycocalyx in the mechanotransduction of flow-induced shear stress at the cellular and molecular levels has been demonstrated over the past decade. However, its potential mechanotransduction role in vascular remodeling has triggered little attention. In the present study, a home-made apparatus was used to expose the rat abdominal aorta to sterile, flow or no flow, normal-pressure or high-pressure conditions for 4 days. The histomophometric, cellular, and molecular analysis of vessels were performed. The results showed that after exposing the vessels in the flow and high-pressure condition, the apoptotic rate, the cell number, and the RNA level of contractile marker gene smooth muscle 22 of smooth muscle cells were significantly increased, whereas the expression of nitric oxide synthase, α-smooth muscle actin, smoothelin, and calponion showed no significant differences compared with the flow and normal-pressure groups. Moreover, the histomophometric analysis of vascular walls suggested a remodeling induced by flow and high-pressure loading consistent with the classic hypertensive aortic phenotype, which is characterized by a thicker and more rigid vascular wall as well as increased aortic diameter. However, those phenomena were totally abolished after compromising the integrity of glycocalyx by the treatment of vessels with hyaluronidase, which provided evidence of the important mechanotransduction role of the vascular cells glycocalyx in vascular remodeling induced by hemodynamic stimuli., (© 2018 American Heart Association, Inc.)

The vascular smooth muscle cells (VSMCs) are exposed to interstitial flow induced shear stress that may be sensed by the surface glycocalyx, a surface layer composed primarily of proteoglycans and glycoproteins, to mediate cell contraction during the myogenic response. We, therefore, attempted to elucidate the signal pathway of the glycocalyx mechanotransduction in shear stress regulated SMC contraction. Human umbilical vein SMCs (HUVSMCs) deprived of serum for 3-4 days were exposed to a step increase (0 to 20 dyn/cm 2 ) in shear stress in a parallel plate flow chamber, and reduction in the cell area was quantified as contraction. The expressions of Rho kinase (ROCK) and its downstream signal molecules, the myosin-binding subunit of myosin phosphatase (MYPT) and the myosin light chain 2 (MLC2), were evaluated. Results showed that the exposure of HUVSMCs to shear stress for 30 min induced cell contraction significantly, which was accompanied by ROCK1 up-regulation, re-distribution, as well as MYPT1 and MLC activation. However, these shear induced phenomenon could be completely abolished by heparinase III or Y-27632 pre-treatment. These results indicate shear stress induced VSMC contraction was mediated by cell surface glycocalyx via a ROCK-MLC phosphatase (MLCP) pathway, providing evidence of the glycocalyx mechanotransduction in myogenic response., Competing Interests: The authors declare no competing financial interests.

We hypothesized that diminished endothelial glycocalyx (GCX) at atherosclerotic lesion-prone sites accelerates flow-dependent concentration polarization of low-density lipoproteins (LDLs) at the luminal surface, and in turn contributes to vulnerability of these sites to atherosclerosis. A parallel plate flow chamber was applied to expose cultured endothelial monolayers to three different levels of shear stress (3, 12, 20 dyn/cm(2)). Heparinase III (Hep.III) was employed to degrade heparan sulfate proteoglycans selectively and 3-(N-morpholino) propanesulfonic acid-buffered physiological salt solutions (MOPS-PSS) were used at either normal ionic strength (Normal-MOPS), low ionic strength (LO-MOPS) or high ionic strength (HI-MOPS) to modify the effective charge density of the endothelial GCX. Water filtration velocity (V(w)) across the endothelial monolayer, the luminal concentration of LDLs (C(w)) and the uptake of LDLs by endothelial cells were measured and compared among the following five groups of cells: (1) Control; (2) Hep.III treatment; (3) LO-MOPS; (4) Normal-MOPS; and (5) HI-MOPS. The results obtained substantiated the aforementioned hypothesis and demonstrated that compositional or charge density modification of the endothelial GCX facilitated water filtration across the endothelium, enhanced the accumulation of LDLs on the luminal surface and increased the uptake of LDLs by endothelial cells, therefore contributing to atherogenesis.

Vascular remodeling induced by hemodynamic stimuli contributes to the pathophysiology of cardiovascular diseases. The importance of vascular cells (endothelial cells and smooth muscle cells) glycocalyx in the mechanotransduction of flow-induced shear stress at the cellular and molecular levels has been demonstrated over the past decade. However, its potential mechanotransduction role in vascular remodeling has triggered little attention. In the present study, a home-made apparatus was used to expose the rat abdominal aorta to sterile, flow or no flow, normal-pressure or high-pressure conditions for 4 days. The histomophometric, cellular, and molecular analysis of vessels were performed. The results showed that after exposing the vessels in the flow and high-pressure condition, the apoptotic rate, the cell number, and the RNA level of contractile marker gene smooth muscle 22 of smooth muscle cells were significantly increased, whereas the expression of nitric oxide synthase, α-smooth muscle actin, smoothelin, and calponion showed no significant differences compared with the flow and normal-pressure groups. Moreover, the histomophometric analysis of vascular walls suggested a remodeling induced by flow and high-pressure loading consistent with the classic hypertensive aortic phenotype, which is characterized by a thicker and more rigid vascular wall as well as increased aortic diameter. However, those phenomena were totally abolished after compromising the integrity of glycocalyx by the treatment of vessels with hyaluronidase, which provided evidence of the important mechanotransduction role of the vascular cells glycocalyx in vascular remodeling induced by hemodynamic stimuli. [ABSTRACT FROM AUTHOR]

Atherosclerosis preferentially occurs at regions in arterial branching, curvature, and stenosis, which may be explained by the geometric predilection of low-density lipoprotein (LDL) concentration polarization that has been investigated in major arteries in previous studies. Whether this also happens in arterioles remains unknown. Herein, a radially non-uniform distribution of LDL particles and a heterogeneous endothelial glycocalyx layer in the mouse ear arterioles, as shown by fluorescein isothiocyanate labeled wheat germ agglutinin (WGA-FITC), were successfully observed by a non-invasive two-photon laser-scanning microscopy (TPLSM) technique. The stagnant film theory was applied as the fitting function to evaluate LDL concentration polarization in arterioles. The concentration polarization rate (CPR, the ratio of the number of polarized cases to that of total cases) in the inner walls of curved and branched arterioles was 22% and 31% higher than the outer counterparts, respectively. Results from the binary logistic regression and multiple linear regression analysis showed that endothelial glycocalyx thickness increases CPR and the thickness of the concentration polarization layer (CPL). Flow field computation indicates no obvious disturbances or vortex in modeled arterioles with different geometries and the mean wall shear stress is about 7.7–9.0 Pa. These findings suggest a geometric predilection of LDL concentration polarization in arterioles for the first time, and the existence of an endothelial glycocalyx, acting together with a relatively high wall shear stress in arterioles, may explain to some extent why atherosclerosis rarely occurs in these regions. [Display omitted] • Arteriolar LDL concentration polarization shows a geometric predilection. • Arteriolar high WSS counteracts LDL concentration polarization. • Glycocalyx enhances LDL concentration polarization yet reduces its wall deposition. • Glycocalyx and high WSS may explain the absence of atherosclerosis in arterioles. [ABSTRACT FROM AUTHOR]