The Endothelial Glycocalyx as a Target of Excess Soluble Fms-like Tyrosine Kinase-1.

Soluble fms-like tyrosine kinase-1 (sFlt-1) is a secreted protein that binds heparan sulfate expressed on the endothelial glycocalyx (eGC). In this paper we analyze how excess sFlt-1 causes conformational changes in the eGC, leading to monocyte adhesion, a key event triggering vascular dysfunction. In vitro exposure of primary human umbilical vein endothelial cells to excess sFlt-1 decreased eGC height and increased stiffness as determined by atomic force microscopy (AFM). Yet, structural loss of the eGC components was not observed, as indicated by Ulex europaeus agglutinin I and wheat germ agglutinin staining. Moreover, the conformation observed under excess sFlt-1, a collapsed eGC, is flat and stiff with unchanged coverage and sustained content. Functionally, this conformation increased the endothelial adhesiveness to THP-1 monocytes by about 35%. Heparin blocked all these effects, but the vascular endothelial growth factor did not. In vivo administration of sFlt-1 in mice also resulted in the collapse of the eGC in isolated aorta analyzed ex vivo by AFM. Our findings show that excess sFlt-1 causes the collapse of the eGC and favors leukocyte adhesion. This study provides an additional mechanism of action by which sFlt-1 may cause endothelial dysfunction and injury.

NCS-1 is a regulator of calcium signaling in health and disease.

Neuronal Calcium Sensor-1 (NCS-1) is a highly conserved calcium binding protein which contributes to the maintenance of intracellular calcium homeostasis and regulation of calcium-dependent signaling pathways. It is involved in a variety of physiological cell functions, including exocytosis, regulation of calcium permeable channels, neuroplasticity and response to neuronal damage. Over the past 30 years, continuing investigation of cellular functions of NCS-1 and associated disease states have highlighted its function in the pathophysiology of several disorders and as a therapeutic target. Among the diseases that were found to be associated with NCS-1 are neurological disorders such as bipolar disease and non-neurological conditions such as breast cancer. Furthermore, alteration of NCS-1 expression is associated with substance abuse disorders and severe side effects of chemotherapeutic agents. The objective of this article is to summarize the current body of evidence describing NCS-1 and its interactions on a molecular and cellular scale, as well as describing macroscopic implications in physiology and medicine. Particular attention is paid to the role of NCS-1 in development and prevention of chemotherapy induced peripheral neuropathy (CIPN).

Neuronal Calcium Sensor 1 Has Two Variants with Distinct Calcium Binding Characteristics.

Neuronal calcium sensor-1 (NCS-1 Var1) is a calcium-binding protein expressed in most tissues. We examined a poorly characterized variant of NCS-1 (Var2), identified only in humans where the N-terminal 22 amino acid residues of native NCS-1(MGKSNSKLKPEVVEELTRKTY) were replaced with 4 different residues (MATI). Because alterations in the level of expression of NCS-1 Var1 and the expression of NCS-1 variants have been correlated with several neurological diseases, the relative expression and functional role of NCS-1 Var2 was examined. We found that NCS-1 Var2 mRNA levels are not found in mouse tissues and are expressed at levels ~1000-fold lower than NCS-1 Var1 in three different human cell lines (SHSY5Y, HEK293, MB231). Protein expression of both variants was only identified in cell lines overexpressing exogenous NCS-1 Var2. The calcium binding affinity is ~100 times weaker in purified NCS-1 Var2 than NCS-1 Var1. Because truncation of NCS-1 Var1 has been linked to functional changes in neurons, we determined whether the differing properties of the NCS-1 variants could potentially contribute to the altered cell function. In contrast to previous reports showing that overexpression of NCS-1 Var1 increases calcium-dependent processes, functional differences in cells overexpressing NCS-1 Var2 were undetectable in assays for cell growth, cell death and drug (paclitaxel) potency. Our results suggest that NCS-1 Var1 is the primary functional version of NCS-1.