Plasma junctional adhesion molecule C levels are associated with the presence and severity of coronary artery disease.

BACKGROUND: Junctional adhesion molecule C (JAM-C) is a novel cell adhesion molecule that belongs to the immunoglobulin superfamily. Previous studies have demonstrated the up-regulation of JAM-C in atherosclerotic vessels in human and in spontaneous early lesions of apoe-/- mice. However, insufficient research is currently available on the association of plasma JAM-C levels with the presence and severity of coronary artery disease (CAD). OBJECTIVES: To explore the relationship between plasma JAM-C levels and CAD. DESIGN AND METHODS: Plasma JAM-C levels were examined in 226 patients who underwent coronary angiography. Unadjusted and adjusted associations were assessed using logistic regression models. ROC curves were generated to examine the predictive performance of JAM-C. C-statistics, continuous net reclassification improvement (NRI) and integrated discrimination improvement (IDI) were obtained to assess the incremental predictive value of JAM-C. RESULTS: Plasma JAM-C levels were significantly higher in patients with CAD and high GS. Multivariate logistic regression analysis showed that JAM-C was independent predictor for the presence and severity of CAD [adjusted OR (95% CI): 2.04(1.28-3.26) and 2.81 (2.02-3.91), respectively]. The optimal cutoff value of plasma JAM-C levels for predicting the presence and severity of CAD was 98.26 pg/ml and 122.48 pg/ml, respectively. Adding JAM-C to the baseline model improved the global performance of the model [C-statistic increased from 0.853 to 0.872, p = 0.171; continuous NRI (95% CI): 0.522 (0.242-0.802), p < 0.001; IDI (95% CI): 0.042 (0.009-0.076), p = 0.014]. CONCLUSIONS: Our data showed that plasma JAM-C levels are associated with the presence and severity of CAD, suggesting that JAM-C may be a useful marker for the prevention and management of CAD.

ADAM10 facilitates rapid neural stem cell cycling and proper positioning within the subventricular zone niche via JAMC/RAP1Gap signaling.

The mechanisms that regulate neural stem cell (NSC) lineage progression and maintain NSCs within different domains of the adult neural stem cell niche, the subventricular zone are not well defined. Quiescent NSCs are arranged at the apical ventricular wall, while mitotically activated NSCs are found in the basal, vascular region of the subventricular zone. Here, we found that ADAM10 (a disintegrin and metalloproteinase 10) is essential in NSC association with the ventricular wall, and via this adhesion to the apical domain, ADAM10 regulates the switch from quiescent and undifferentiated NSC to an actively proliferative and differentiating cell state. Processing of JAMC (junctional adhesion molecule C) by ADAM10 increases Rap1GAP activity. This molecular machinery promotes NSC transit from the apical to the basal compartment and subsequent lineage progression. Understanding the molecular mechanisms responsible for regulating the proper positioning of NSCs within the subventricular zone niche and lineage progression of NSCs could provide new targets for drug development to enhance the regenerative properties of neural tissue.

Use of Proximity Labeling in Cardiovascular Research.

Protein-protein interactions are of paramount importance in regulating normal cardiac physiology. Methodologies to elucidate these interactions in vivo have been limited. Recently, proximity-dependent biotinylation, with the use of BioID, TurboID, and ascorbate peroxidase, has been developed to uncover cellular neighborhoods and novel protein-protein interactions. These cutting-edge techniques have enabled the identification of subcellular localizations of specific proteins and the neighbors or interacting proteins within these subcellular regions. In contrast to classic methods such as affinity purification and subcellular fractionation, these techniques add covalently bound tags in living cells, such that spatial relationships and interaction networks are not disrupted. Recently, these methodologies have been used to identify novel protein-protein interactions relevant to the cardiovascular system. In this review, we discuss the development and current use of proximity biotin-labeling for cardiovascular research.

Role of Junctional Adhesion Molecule-C in the Regulation of Inner Endothelial Blood-Retinal Barrier Function.

Although JAM-C is abundantly expressed in the retinae and upregulated in choroidal neovascularization (CNV), it remains thus far poorly understood whether it plays a role in the blood-retinal barrier, which is critical to maintain the normal functions of the eye. Here, we report that JAM-C is highly expressed in retinal capillary endothelial cells (RCECs), and VEGF or PDGF-C treatment induced JAM-C translocation from the cytoplasm to the cytomembrane. Moreover, JAM-C knockdown in RCECs inhibited the adhesion and transmigration of macrophages from wet age-related macular degeneration (wAMD) patients to and through RCECs, whereas JAM-C overexpression in RCECs increased the adhesion and transmigration of macrophages from both wAMD patients and healthy controls. Importantly, the JAM-C overexpression-induced transmigration of macrophages from wAMD patients was abolished by the administration of the protein kinase C (PKC) inhibitor GF109203X. Of note, we found that the serum levels of soluble JAM-C were more than twofold higher in wAMD patients than in healthy controls. Mechanistically, we show that JAM-C overexpression or knockdown in RCECs decreased or increased cytosolic Ca2+ concentrations, respectively. Our findings suggest that the dynamic translocation of JAM-C induced by vasoactive molecules might be one of the mechanisms underlying inner endothelial BRB malfunction, and inhibition of JAM-C or PKC in RCECs may help maintain the normal function of the inner BRB. In addition, increased serum soluble JAM-C levels might serve as a molecular marker for wAMD, and modulating JAM-C activity may have potential therapeutic value for the treatment of BRB malfunction-related ocular diseases.

Targeting junctional adhesion molecule-C ameliorates sepsis-induced acute lung injury by decreasing CXCR4+ aged neutrophils.

Sepsis is a severe inflammatory condition associated with high mortality. Transmigration of neutrophils into tissues increases their lifespan to promote deleterious function. Junctional adhesion molecule-C (JAM-C) plays a pivotal role in neutrophil transmigration into tissues. We aim to study the role of JAM-C on the aging of neutrophils to cause sepsis-induced acute lung injury (ALI). Sepsis was induced in C57BL/6J mice by cecal ligation and puncture (CLP) and JAM-C expression in serum was assessed. Bone marrow-derived neutrophils (BMDN) were treated with recombinant mouse JAM-C (rmJAM-C) ex vivo and their viability was assessed. CLP-operated animals were administrated with either isotype IgG or anti-JAM-C Ab at a concentration of 3 mg/kg and after 20 h, aged neutrophils (CXCR4+ ) were assessed in blood and lungs and correlated with systemic injury and inflammatory markers. Soluble JAM-C level in serum was up-regulated during sepsis. Treatment with rmJAM-C inhibited BMDN apoptosis, thereby increasing their lifespan. CLP increased the frequencies of CXCR4+ neutrophils in blood and lungs, while treatment with anti-JAM-C Ab significantly reduced the frequencies of CXCR4+ aged neutrophils. Treatment with anti-JAM-C Ab significantly reduced systemic injury markers (alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase) as well as systemic and lung inflammatory cytokines (IL-6 and IL-1ß) and chemokine (macrophage inflammatory protein-2). The blockade of JAM-C improved lung histology and reduced neutrophil contents in lungs of septic mice. Thus, reduction of the pro-inflammatory aged neutrophils by blockade of JAM-C has a novel therapeutic potential in sepsis-induced ALI.

Neuronal Expression of Junctional Adhesion Molecule-C is Essential for Retinal Thickness and Photoreceptor Survival.

BACKGROUND: Photoreceptor cell death is a key pathology of retinal degeneration diseases. To date, the molecular mechanisms for this pathological process remain largely unclear. Junctional adhesion molecule-c (Jam-c) has been shown to play important roles in different biological events. However, its effect on retinal neuronal cells is unknown. OBJECTIVE: To determine the effect of Jam-c on adult mouse eyes, particularly, on retinal structure, vasculature and photoreceptor cells, in order to explore potential important target molecules for ocular diseases. METHODS: Jam-c global knockout mice, endothelial-specific and neuronal-specific Jam-c conditional knockout mice using Tie2-Cre and Nestin-Cre mice respectively were used in this study. Mouse eyes were harvested from the different groups and eye size examined. Cryosections of the eyes were made and stained with Hematoxylin and Eosin (H&E) and the thicknesses of retinal layers measured. Retinal blood vessels and cone and rod photoreceptors were analyzed using isolectin B4, peanut agglutinin and rhodopsin as markers respectively. In vivo Jam-c knockdown in mouse eyes was performed by intravitreal injection of Jam-c shRNA. Jam-c expression in the retinae was quantified by real-time PCR. RESULTS: Global Jam-c gene deletion in mice resulted in smaller eyes and decreased the diameters of lens and iris. Jam-c-/- mice display marked thinning of the outer nuclear layer (ONL), less numbers of photoreceptor cells, and abnormal retinal vasculature. Importantly, neuronal-specific Jam-c deletion led to similar phenotype, whereas no obvious defect was observed in endothelial-specific Jam-c knockout mice. Moreover, Jam-c knockdown by shRNA also decreased ONL thickness and photoreceptor numbers. CONCLUSION: We found that Jam-c is critically required for the normal size and retinal structure. Particularly, Jam-c plays important roles in maintaining the normal retinal thickness, vasculature and photoreceptor numbers. Jam-c thus may therefore have important roles in various ocular diseases.

Dachengqi Decoction Attenuates Intestinal Vascular Endothelial Injury in Severe Acute Pancreatitis in Vitro and in Vivo.

BACKGROUND/AIMS: Dachengqi decoction (DCQD) is a well-known traditional Chinese herbal drug with strong anti-inflammatory effects. Angiopoietin-1 (Ang-1) plays a vital role in maintaining the stability and integrity of the vascular wall and prevents vascular leakage due to inflammatory mediators. Our previous work found that DCQD protects against pancreatic injury in rats with severe acute pancreatitis (SAP). This study aims to investigate the effects of DCQD on intestinal endothelial damage in both damaged human umbilical vein endothelial cells (HUVECs) and SAP rats. METHODS: HUVECs were randomly divided into four groups: control group, TNF-α group, TNF-α plus Ang-1 group (Ang-1 group), and TNF-α plus DCQD group (DCQD group). Cells were incubated for 6 h, 12 h, and 24 h, before collection. The treatment concentration of DCQD was decided based on a Cell Counting Kit-8 (CCK-8) assay. The monolayer permeability of the HUVECs was assessed by measuring the transendothelial electrical resistance (TEER). Apoptosis was analyzed by flow cytometry. mRNA and protein expression of aquaporin 1 (AQP-1), matrix metalloproteinase 9 (MMP9), and junctional adhesion molecule-C (JAM-C) was evaluated by RT-PCR, immunocytofluorescence, and western blot. Forty male Sprague-Dawley rats were randomized into a control group, SAP group, SAP plus Ang-1 group (Ang-1 group), and SAP plus DCQD group (DCQD group). SAP was induced by intraperitoneal injection of cerulein and lipopolysaccharide (LPS), while the control group received 0.9% saline solution. Evans blue was injected through the penile vein and the rats were then sacrificed 12 h after modeling. Levels of serum amylase, TNF-α, IL-1ß, IL-2, and IL-6 were determined by using ELISA. Intestinal tissue was analysed by histology, and capillary permeability in the tissues was evaluated by Evans blue extravasation assay. Protein and mRNA expression of AQP-1, MMP9, and JAM-C were assessed by immunohistofluorescence, western blot, and RT-PCR. RESULTS: DCQD reduced the permeability of HUVEC induced by TNF-α in vitro. Furthermore, DCQD altered the mRNA and protein levels of JAM-C, MMP9, and AQP-1 in HUVECs after TNF-α induction. SAP intestinal injury induced by cerulein combined with lipopolysaccharides was concomitant with increased expression of JAM-C and MMP9, and reduced expression of AQP-1 in intestinal tissue. Pretreatment with DCQD attenuated SAP intestinal injury and lowered the levels of serum amylase, TNF-α, IL-1ß, IL-2, and IL-6 effectively. Our study demonstrated that DCQD decreased the expression of JAM-C and MMP9 and increased the expression of AQP-1 both in vitro and in vivo. CONCLUSION: DCQD can reduce capillary endothelial damage in acute pancreatitis-associated intestinal injury and the mechanism may be associated with the regulation of endothelial barrier function-associated proteins AQP-1, MMP9, and JAM-C.

Blocking junctional adhesion molecule C promotes the recovery of cisplatin-induced acute kidney injury.

BACKGROUND/AIMS: Recent findings have demonstrated the occurrence of neutrophil transendothelial migration in the reverse direction (reverse TEM) and that endothelial junctional adhesion molecule C (JAM-C) is a negative regulator of reverse TEM. In this study, we tested the effects of a JAM-C blocking antibody on the resolution of kidney injuries and inflammation in a mouse model of cisplatin-induced acute kidney injury (AKI). METHODS: Cisplatin was administered via intraperitoneal injection. A JAM-C blocking antibody or a control immunoglobulin G was administered intraperitoneal at 1.5 mg/kg, with the injection being delayed until day 4 following cisplatin administration to restrict the effect of antibodies on recovery. RESULTS: After cisplatin injection, serum creatinine and histologic injuries peaked on day 4. Treatment with a JAM-C blocking antibody on days 4 and 5 promoted the functional and histologic recovery of cisplatin-induced AKI on days 5 and 6. Facilitating recovery with a JAM-C blocking antibody correlated with significantly increased circulating intercellular adhesion molecule 1+ Tamm-Horsfall protein+ neutrophils and significantly decreased renal neutrophil infiltration, indicating that facilitating reverse the TEM of neutrophils from the kidney to the peripheral circulation partially mediated the resolution of inflammation and recovery. CONCLUSIONS: These results demonstrated that reverse TEM is involved in the resolution of neutrophilic inflammation in cisplatin-induced AKI and that JAM-C is an important regulator of this process.

JAM-C maintains VEGR2 expression to promote retinal pigment epithelium cell survival under oxidative stress.

Junctional adhesion molecule-C (JAM-C) has been shown to play critical roles during development and in immune responses. However, its role in adult eyes under oxidative stress remains poorly understood. Here, we report that JAM-C is abundantly expressed in adult mouse retinae and choroids in vivo and in cultured retinal pigment epithelium (RPE) and photoreceptor cells in vitro. Importantly, both JAM-C expression and its membrane localisation are downregulated by H2O2-induced oxidative stress. Under H2O2-induced oxidative stress, JAM-C is critically required for the survival of human RPE cells. Indeed, loss of JAM-C by siRNA knockdown decreased RPE cell survival. Mechanistically, we show that JAM-C is required to maintain VEGFR2 expression in RPE cells, and VEGFR2 plays an important role in keeping the RPE cells viable since overexpression of VEGFR2 partially restored impaired RPE survival caused by JAM-C knockdown and increased RPE survival. We further show that JAM-C regulates VEGFR2 expression and, in turn, modulates p38 phosphorylation. Together, our data demonstrate that JAM-C plays an important role in maintaining VEGR2 expression to promote RPE cell survival under oxidative stress. Given the vital importance of RPE in the eye, approaches that can modulate JAM-C expression may have therapeutic values in treating diseases with impaired RPE survival.

Blocking junctional adhesion molecule C promotes the recovery of cisplatin-induced acute kidney injury

BACKGROUND/AIMS: Recent findings have demonstrated the occurrence of neutrophil transendothelial migration in the reverse direction (reverse TEM) and that endothelial junctional adhesion molecule C (JAM-C) is a negative regulator of reverse TEM. In this study, we tested the effects of a JAM-C blocking antibody on the resolution of kidney injuries and inflammation in a mouse model of cisplatin-induced acute kidney injury (AKI). METHODS: Cisplatin was administered via intraperitoneal injection. A JAM-C blocking antibody or a control immunoglobulin G was administered intraperitoneal at 1.5 mg/kg, with the injection being delayed until day 4 following cisplatin administration to restrict the effect of antibodies on recovery. RESULTS: After cisplatin injection, serum creatinine and histologic injuries peaked on day 4. Treatment with a JAM-C blocking antibody on days 4 and 5 promoted the functional and histologic recovery of cisplatin-induced AKI on days 5 and 6. Facilitating recovery with a JAM-C blocking antibody correlated with significantly increased circulating intercellular adhesion molecule 1+ Tamm-Horsfall protein+ neutrophils and significantly decreased renal neutrophil infiltration, indicating that facilitating reverse the TEM of neutrophils from the kidney to the peripheral circulation partially mediated the resolution of inflammation and recovery. CONCLUSIONS: These results demonstrated that reverse TEM is involved in the resolution of neutrophilic inflammation in cisplatin-induced AKI and that JAM-C is an important regulator of this process.