Modifiable Host Factors for the Prevention and Treatment of COVID-19: Diet and Lifestyle/Diet and Lifestyle Factors in the Prevention of COVID-19.

Many studies have shown that the immune system requires adequate nutrition to work at an optimal level. Not only do optimized nutritional strategies support the immune system, but they also reduce chronic inflammation. Nutritional supplements that are recommended for patients with critical illnesses are thought to also be effective for the coronavirus disease 2019 (COVID-19) patients in the intensive care unit. Some studies have recommended fresh fruits and vegetables, soy, nuts, and antioxidants, such as omega-3 fatty acids, to improve immune system activity. Although nutritional status is considered to be an important prognostic factor for patients with COVID-19, there is to date no sufficient evidence that optimal nutritional therapies can be beneficial for these patients. Some have argued that the COVID-19 pandemic is a good opportunity to test the effectiveness of nutritional intervention for infectious diseases. Many researchers have suggested that testing the proposed nutritional approaches for infectious diseases in the context of a pandemic would be highly informative. The authors of other review papers concluded that it is important to have a diet based on fresh foods, such as fruits, vegetables, whole grains, low-fat dairy products, and healthy fats (i.e., olive oil and fish oil), and to limit the intake of sugary drinks as well as high-calorie and high-salt foods. In this review, we discuss the clinical significance of functional food ingredients as complementary therapies potentially beneficial for the prevention or treatment of COVID-19. We believe that our review will be helpful to plan and deploy future studies to conclude these potentials against COVID-19, but also to new infectious diseases that may arise in the future.

Coreceptor functions of cell surface heparan sulfate proteoglycans.

Receptor-ligand interactions play an important role in many biological processes by triggering specific cellular responses. These interactions are frequently regulated by coreceptors that facilitate, alter, or inhibit signaling. Coreceptors work in parallel with other specific and accessory molecules to coordinate receptor-ligand interactions. Cell surface heparan sulfate proteoglycans (HSPGs) function as unique coreceptors because they can bind to many ligands and receptors through their HS and core protein motifs. Cell surface HSPGs are typically expressed in abundance of the signaling receptors and, thus, are capable of mediating the initial binding of ligands to the cell surface. HSPG coreceptors do not possess kinase domains or intrinsic enzyme activities and, for the most part, binding to cell surface HSPGs does not directly stimulate intracellular signaling. Because of these features, cell surface HSPGs primarily function as coreceptors for many receptor-ligand interactions. Given that cell surface HSPGs are widely conserved, they likely serve fundamental functions to preserve basic physiological processes. Indeed, cell surface HSPGs can support specific cellular interactions with growth factors, morphogens, chemokines, extracellular matrix (ECM) components, and microbial pathogens and their secreted virulence factors. Through these interactions, HSPG coreceptors regulate cell adhesion, proliferation, migration, and differentiation, and impact the onset, progression, and outcome of pathophysiological processes, such as development, tissue repair, inflammation, infection, and tumorigenesis. This review seeks to provide an overview of the various mechanisms of how cell surface HSPGs function as coreceptors.

Role of HSPGs in Systemic Bacterial Infections.

Heparan sulfate proteoglycans (HSPGs) are at the forefront of host-microbe interactions. Cell surface HSPGs are thought to promote infection as attachment and internalization receptors for many bacterial pathogens and as soluble inhibitors of host immunity when released from the cell surface by ectodomain shedding. However, the importance of HSPG-pathogen interactions in vivo has yet to be clearly established. Here we describe several representative methods to study the role of HSPGs in systemic bacterial infections, such as bacteremia and sepsis. The overall experimental strategy is to use mouse models to establish the physiological significance of HSPGs, to determine the identity of HSPGs that specifically promote infection, and to define key structural features of HSPGs that enhance bacterial virulence in systemic infections.

Syndecan-1 limits the progression of liver injury and promotes liver repair in acetaminophen-induced liver injury in mice.

Accidental or intentional misuse of acetaminophen (APAP) is the leading cause of acute liver failure in the Western world. Although mechanisms that trigger APAP-induced liver injury (AILI) are well known, those that halt the progression of APAP liver disease and facilitate liver recovery are less understood. Heparan sulfate proteoglycans (HSPGs) bind to and regulate various tissue injury factors through their heparan sulfate (HS) chains, but the importance of HSPGs in liver injury in vivo remains unknown. Here, we examined the role of syndecan-1, the major cell-surface HSPG of hepatocytes, in AILI. Ablation of syndecan-1 in mice led to unopposed progression of liver injury upon APAP overdose. However, direct APAP hepatoxicity and liver injury at early times post-APAP overdose were unaffected by syndecan-1, suggesting that syndecan-1 influences later mechanisms that lead to liver repair. The exuberant liver injury phenotypes in syndecan-1 null (Sdc1-/- ) mice were traced to a deficiency in protein kinase B (Akt) activation in hepatocytes, which led to a delayed increase in glycogen synthase kinase-3ß (GSK-3ß)-mediated hepatocyte apoptosis. Inhibition of Akt worsened, whereas inhibition of GSK-3ß and caspases protected mice from AILI. Moreover, administration of purified syndecan-1, HS, or engineered heparan compounds containing 2-O-sulfate groups rescued Sdc1-/- mice from AILI by potentiating Akt signaling and inhibiting GSK-3ß-mediated apoptosis in hepatocytes. In addition, HS showed a significantly prolonged therapeutic efficacy as compared to N-acetylcysteine. CONCLUSION: These results demonstrate that 2-O-sulfated domains in syndecan-1 HS halt disease progression and promote liver repair by enhancing hepatocyte survival in AILI. We propose that syndecan-1 is a critical endogenous factor that controls the balance between prosurvival signaling and apoptosis in hepatocytes in APAP liver disease. (Hepatology 2017;66:1601-1615).

EXTL3 mutations cause skeletal dysplasia, immune deficiency, and developmental delay.

We studied three patients with severe skeletal dysplasia, T cell immunodeficiency, and developmental delay. Whole-exome sequencing revealed homozygous missense mutations affecting exostosin-like 3 (EXTL3), a glycosyltransferase involved in heparan sulfate (HS) biosynthesis. Patient-derived fibroblasts showed abnormal HS composition and altered fibroblast growth factor 2 signaling, which was rescued by overexpression of wild-type EXTL3 cDNA. Interleukin-2-mediated STAT5 phosphorylation in patients' lymphocytes was markedly reduced. Interbreeding of the extl3-mutant zebrafish (box) with Tg(rag2:green fluorescent protein) transgenic zebrafish revealed defective thymopoiesis, which was rescued by injection of wild-type human EXTL3 RNA. Targeted differentiation of patient-derived induced pluripotent stem cells showed a reduced expansion of lymphohematopoietic progenitor cells and defects of thymic epithelial progenitor cell differentiation. These data identify EXTL3 mutations as a novel cause of severe immune deficiency with skeletal dysplasia and developmental delay and underline a crucial role of HS in thymopoiesis and skeletal and brain development.

Pentraxin 3 as a biomarker for acute coronary syndrome: comparison with biomarkers for cardiac damage.

BACKGROUND: Pentraxin 3 (PTX3) is increased in circulating blood during the acute stage of acute coronary syndrome (ACS). Therefore, we compared diagnostic values of PTX3 for ACS with those of biomarkers for myocardial damage, such as troponin T (TnT) and heart-type fatty acid binding protein (H-FABP). METHODS AND RESULTS: Patients (n = 87) undergoing coronary angiography (CAG), consisting of 16 ACS and 71 non-ACS patients were enrolled. Non-ACS consists of 12 patients with normal CAG, 30 stable angina pectoris (SAP) patients controlled by medical treatment, and 29 SAP patients who required elective coronary revascularization (percutaneous coronary intervention or coronary artery bypass graft). Age, gender, or prevalence of diabetes, hypertension, or smoking was not significantly different between ACS and non-ACS groups. Serum total, high-density lipoprotein, or low-density lipoprotein cholesterol, or triglyceride levels were not significantly different between ACS and non-ACS. PTX3 levels were not significantly correlated with lipid profiles or different between those with and without conventional risk factors. Circulating PTX3, TnT, and H-FABP levels were significantly higher in ACS than non-ACS. In receiver-operating characteristic (ROC) curves, area under the curve (AUC) values for PTX3, TnT and H-FABP were 0.920, 0.674, and 0.690, respectively. ROC curves of PTX3 (AUC: 0.901), TnT (AUC: 0.731), and H-FABP (AUC: 0.633) for ST-elevation ACS were similar to those for whole ACS. In a TnT-negative subgroup, the AUC values of PTX3 and H-FABP for ACS were 0.981 and 0.489, respectively. CONCLUSIONS: PTX3 is a sensitive and specific biomarker for the diagnosis of ACS, and shows additional diagnostic values when measured in combination with TnT.

Soluble lectin-like oxidized LDL receptor-1 (sLOX-1) as a sensitive and specific biomarker for acute coronary syndrome--comparison with other biomarkers.

BACKGROUND: Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) appears to be involved in atherosclerotic plaque vulnerability and rupture. Circulating soluble LOX-1 (sLOX-1) levels are dramatically elevated in patients with acute coronary syndrome (ACS), and its diagnostic sensitivity and specificity is superior to high-sensitivity C-reactive protein (hs-CRP). In this study, we have compared the diagnostic value of sLOX-1 for ACS with those of troponin T (TnT) and heart-type fatty acid binding protein (H-FABP). METHODS: One hundred and seven patients who underwent coronary angiography (CAG), including 18 ACS and 89 non-ACS patients were enrolled. Peripheral blood samples were obtained during the emergent or elective CAG. The non-ACS group consisted of 30 patients with normal CAG, 30 stable angina pectoris patients controlled by medical treatment, and 29 patients with stable angina who required elective coronary revascularization (percutaneous coronary intervention or coronary artery bypass graft). RESULTS: Age, gender, lipid profiles, or prevalence of diabetes, smoking, or hypertension were not significantly different between ACS and non-ACS. These factors did not significantly affect blood sLOX-1 levels. Circulating sLOX-1, TnT, and H-FABP levels were significantly higher in ACS than non-ACS. Area under the curve (AUC) values of the receiver-operating characteristic curves were 0.948, 0.704, and 0.691 for sLOX-1, TnT, and H-FABP, respectively. In a TnT-negative (<0.03 ng/mL) subgroup, the AUC values for sLOX-1 and H-FABP were 0.848 and 0.476, respectively. CONCLUSION: Circulating sLOX-1 is a more sensitive and specific biomarker for ACS than TnT and H-FABP, and provides additional diagnostic values when measured in combination with TnT.

Soluble lectin-like oxidized low-density lipoprotein receptor-1 predicts prognosis after acute coronary syndrome--a pilot study.

BACKGROUND: Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) is implicated in atherosclerotic plaque vulnerability. It is shed, in part, by proteases and released as soluble LOX-1 (sLOX-1), which is a specific and sensitive biomarker of acute coronary syndrome (ACS). The present study explored if sLOX-1 can also predict prognosis after ACS. METHODS AND RESULTS: ACS patients undergoing emergency percutaneous coronary intervention and measurement of circulating sLOX-1 were followed (median: 896 days). Among 94 patients, 13 had ACS recurrence or died (re-ACS/death group). None of age, sex, lipid profile or prevalence of diabetes, smoking or hypertension was significantly different between the re-ACS/death group and the event-free survival group. Circulating sLOX-1 levels, but not those of high-sensitivity C-reactive protein (hs-CRP) or troponin T (TnT), were significantly (P<0.005) higher in the re-ACS/death group than in the event-free survival group. Kaplan-Meier survival curves showed that ACS patients with sLOX-1 values in the highest quartile or tertile had more frequent and earlier ACS recurrence or death. Receiver-operating characteristic curves for prediction of re-ACS or death showed higher sensitivity and specificity for sLOX-1 (area under the curve for sLOX-1, hs-CRP and TnT: 0.764, 0.658 and 0.524, respectively). CONCLUSIONS: Circulating sLOX-1, a diagnostic biomarker of ACS, also predicts ACS recurrence or death.

Molecular and cellular mechanisms of ectodomain shedding.

The extracellular domain of several membrane-anchored proteins is released from the cell surface as soluble proteins through a regulated proteolytic mechanism called ectodomain shedding. Cells use ectodomain shedding to actively regulate the expression and function of surface molecules, and modulate a wide variety of cellular and physiological processes. Ectodomain shedding rapidly converts membrane-associated proteins into soluble effectors and, at the same time, rapidly reduces the level of cell surface expression. For some proteins, ectodomain shedding is also a prerequisite for intramembrane proteolysis, which liberates the cytoplasmic domain of the affected molecule and associated signaling factors to regulate transcription. Ectodomain shedding is a process that is highly regulated by specific agonists, antagonists, and intracellular signaling pathways. Moreover, only about 2% of cell surface proteins are released from the surface by ectodomain shedding, indicating that cells selectively shed their protein ectodomains. This review will describe the molecular and cellular mechanisms of ectodomain shedding, and discuss its major functions in lung development and disease.

Generation of monoclonal antibodies against a soluble form of lectin-like oxidized low-density lipoprotein receptor-1 and development of a sensitive chemiluminescent enzyme immunoassay.

Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1), expressed prominently in atherosclerotic lesions, is cleaved and released as a soluble LOX-1 (sLOX-1), which is a specific biomarker to diagnose acute coronary syndrome (ACS) at an early stage. Although sLOX-1 levels in patient's blood were successfully measured with our previously established enzyme-linked immunosorbent assay (ELISA), the assay was not sensitive enough to detect normal serum levels of sLOX-1 in healthy human subjects. We therefore developed sensitive and specific monoclonal antibodies (mAbs) against sLOX-1 in order to establish a more sensitive immunoassay. Mice were immunized with recombinant human LOX-1 extracellular domain. mAbs were subsequently generated by standard myeloma cell fusion techniques with a novel screening method using time-resolved fluorescence immunoassay. Using two anti-human sLOX-1 mAbs and alkaline phosphatase as a label, a sandwich chemiluminescent enzyme immunoassay (CLEIA) was developed. In total, nine mAbs were obtained. The dissociation constant (K(d)) values of these mAbs for sLOX-1 were 0.12-1.32 nM. Characteristics of these mAbs were estimated and the best combination for CLEIA was selected. The newly established CLEIA could determine sLOX-1 levels as low as 8 pg/mL, and thus, was sensitive enough to measure serum sLOX-1 levels in normal human subjects and to evaluate subtle differences. Values for sLOX-1 measured by monoclonal CLEIA and polyclonal ELISA were highly correlated (r(2)=0.7594, p<0.0001). Area under the curve values of the receiver-operating characteristic curves in detecting ACS were 0.948 and 0.978 for monoclonal CLEIA and polyclonal ELISA, respectively. Thus, a more sensitive sLOX-1 CLEIA was established using newly developed mAbs against sLOX-1. In addition to its advantage in early diagnosis of ACS, this assay may also be useful in predicting cardiovascular disease risk in disease-free subjects.

Circulating soluble SR-PSOX/CXCL16 as a biomarker for acute coronary syndrome -comparison with high-sensitivity C-reactive protein.

AIM: Diagnostic values of soluble SR-PSOX/CXCL16 (sSR-PSOX/CXCL16), a receptor for atherogenic oxidized LDL and a membrane-anchored chemokine for CXCR6-positive lymphocytes, for acute coronary syndrome (ACS) were evaluated.. METHODS: We examined 106 patients undergoing coronary angiography (CAG); 17 patients with ACS and 89 patients without ACS (non-ACS) including stable angina. Circulating sSR-SPOX/CXCL16 was measured in peripheral venous blood by sandwich ELISA. RESULTS: Age, gender, prevalence of diabetes or hypertension, and serum lipid profiles were not significantly different between ACS and non-ACS. Presence or absence of risk factors, such as diabetes, smoking and hypertension, did not significantly affect circulating sSR-PSOX/CXCL16 levels. Circulating sSR-PSOX/CXCL16 levels were significantly lower in ACS than non-ACS (median: 3.05 versus 3.36 ng/mL, p<0.02). Lipid profiles, high-sensitivity C-reactive protein (hs-CRP), cardiac troponin T (TnT), and soluble LOX-1 (sLOX-1) showed no significant correlation with sSR-PSOX/CXCL16. Receiver-operating characteristic (ROC) curves for ACS detection indicate higher sensitivity and specificity for sSR-PSOX/CXCL16 than hs-CRP. In the TnT-negative and sLOX-1-negative subpopulation, sSR-PSOX/CXCL16 showed similar sensitivity and specificity for ACS; however, hs-CRP showed less sensitivity and specificity for ACS when compared with the whole population. CONCLUSION: sSR-PSOX/CXCL16 is a biomarker for ACS, which would provide additional diagnostic information besides TnT and sLOX-1.

Syndecan-1 shedding facilitates the resolution of neutrophilic inflammation by removing sequestered CXC chemokines.

Heparan sulfate binds to and regulates many inflammatory mediators in vitro, suggesting that it serves an important role in directing the progression and outcome of inflammatory responses in vivo. Here, we evaluated the role of syndecan-1, a major heparan sulfate proteoglycan, in modulating multiorgan host injury responses in murine endotoxemia. The extent of systemic inflammation was similar between endotoxemic syndecan-1-null and wild-type mice. However, high levels of CXC chemokines (KC and MIP-2), particularly at later times after LPS, were specifically sustained in multiple organs in syndecan-1-null mice and associated with exaggerated neutrophilic inflammation, organ damage, and lethality. Syndecan-1 shedding was activated in several organs of endotoxemic wild-type mice, and this associated closely with the removal of tissue-bound CXC chemokines and resolution of accumulated neutrophils. Moreover, administration of a shedding inhibitor exacerbated disease by impeding the removal of CXC chemokines and neutrophils, whereas administration of heparan sulfate inhibited the accumulation of CXC chemokines and neutrophils in tissues and attenuated multiorgan injury and lethality. These data show that syndecan-1 shedding is a critical endogenous mechanism that facilitates the resolution of neutrophilic inflammation by aiding the clearance of proinflammatory chemokines in a heparan sulfate-dependent manner.

Interleukin 18 stimulates release of soluble lectin-like oxidized LDL receptor-1 (sLOX-1).

Lectin-like oxidized LDL receptor-1 (LOX-1) appears to play crucial roles in atherosclerotic plaque rupture. We previously reported that circulating soluble LOX-1 (sLOX-1) levels are elevated in acute coronary syndrome (ACS) and that sLOX-1 can be a specific and sensitive biomarker for ACS. A proinflammatory cytokine interleukin 18 (IL-18) and its receptor are prominently expressed in atherosclerotic plaques. In addition, circulating IL-18 levels were reported to be high in ACS. In this study, we have examined if IL-18 can stimulate shedding of LOX-1 and subsequent release of sLOX-1. After transfection with LOX-1 cDNA, HEK-293T cells were incubated with or without IL-18. Cell-conditioned media and total cell lysates were subjected to immunoblot analyses with an anti-LOX-1 monoclonal antibody. In addition, ADAM10 cDNA, ADAM10 siRNA or control vector were also co-transfected into HEK-293T cells, and the cell-conditioned media and total cell lysates were subjected to LOX-1 immunoblotting after treatment with or without IL-18. The cell-conditioned medium/total cell lysate ratios in the amounts of LOX-1 or sLOX-1 were determined as sLOX-1 cleavage ratios. IL-18 (10-100ng/mL) stimulation increased the sLOX-1 cleavage by 3-4-fold in a concentration- and time-dependent manner. ADAM10 overexpression alone similarly enhanced the sLOX-1 cleavage. ADAM10 inhibition by ADAM10 siRNA transfection significantly suppressed IL-18-induced sLOX-1 cleavage. IL-18 similarly enhanced sLOX-1 cleavage in TNF-alpha-activated cultured endothelial cells, as well as LOX-1 transgenic mice in vivo. IL-18 appears one of the stimuli that enhance sLOX-1 release in ACS and ADAM10 may be involved in this process.

Syndecan-1 ectodomain shedding is regulated by the small GTPase Rab5.

The ectodomain shedding of syndecan-1, a major cell surface heparan sulfate proteoglycan, modulates molecular and cellular processes central to the pathogenesis of inflammatory diseases. Syndecan-1 shedding is a highly regulated process in which outside-in signaling accelerates the proteolytic cleavage of syndecan-1 ectodomains at the cell surface. Several extracellular agonists that induce syndecan-1 shedding and metalloproteinases that cleave syndecan-1 ectodomains have been identified, but the intracellular mechanisms that regulate syndecan-1 shedding are largely unknown. Here we examined the role of the syndecan-1 cytoplasmic domain in the regulation of agonist-induced syndecan-1 shedding. Our results showed that the syndecan-1 cytoplasmic domain is essential because mutation of invariant cytoplasmic Tyr residues abrogates ectodomain shedding, but not because it is Tyr phosphorylated upon shedding stimulation. Instead, our data showed that the syndecan-1 cytoplasmic domain binds to Rab5, a small GTPase that regulates intracellular trafficking and signaling events, and this interaction controls the onset of syndecan-1 shedding. Syndecan-1 cytoplasmic domain bound specifically to Rab5 and preferentially to inactive GDP-Rab5 over active GTP-Rab5, and shedding stimulation induced the dissociation of Rab5 from the syndecan-1 cytoplasmic domain. Moreover, the expression of dominant-negative Rab5, unable to exchange GDP for GTP, interfered with the agonist-induced dissociation of Rab5 from the syndecan-1 cytoplasmic domain and significantly inhibited syndecan-1 shedding induced by several distinct agonists. Based on these data, we propose that Rab5 is a critical regulator of syndecan-1 shedding that serves as an on-off molecular switch through its alternation between the GDP-bound and GTP-bound forms.

Syndecan-1 is an in vivo suppressor of Gram-positive toxic shock.

Heparan sulfate proteoglycans bind to and regulate many inflammatory mediators in vitro, suggesting that they serve an important role in influencing inflammatory responses in vivo. Here we evaluated the role of syndecan-1, a major heparan sulfate proteoglycan, in modulating inflammatory responses in Gram-positive toxic shock, a systemic disease that is a significant cause of morbidity and mortality. Syndecan-1-null and wild-type mice were injected intraperitoneally with staphylococcal enterotoxin B, a pyrogenic superantigen, and their inflammatory responses were assessed. Syndecan-1-null mice showed significantly increased liver injury, vascular permeability, and death in response to staphylococcal enterotoxin B challenge compared with wild-type mice. Although serum levels of systemic IL-2 and IFNgamma were similar between the two backgrounds, those of TNFalpha and IL-6 were significantly increased in syndecan-1-null mice undergoing Gram-positive toxic shock. Furthermore, syndecan-1-null mice challenged with staphylococcal enterotoxin B showed enhanced T cell accumulation in tissues, whereas immunodepletion of T cells protected syndecan-1-null mice from the magnified systemic cytokine storm, inflammatory tissue injury, and death. Importantly, syndecan-1 shedding was induced in wild-type mice injected with staphylococcal enterotoxin B, and the administration of heparan sulfate, but not syndecan-1 core protein, rescued syndecan-1-null mice from lethal toxic shock by suppressing the production of TNFalpha and IL-6, and attenuating inflammatory tissue injury. Altogether, these data suggest that syndecan-1 shedding is a key endogenous mechanism that protects the host from Gram-positive toxic shock by inhibiting the dysregulation and amplification of the inflammatory response.

Molecular and cellular mechanisms of syndecans in tissue injury and inflammation.

The syndecan family of heparan sulfate proteoglycans is expressed on the surface of all adherent cells. Syndecans interact with a wide variety of molecules, including growth factors, cytokines, proteinases, adhesion receptors and extracellular matrix components, through their heparan sulfate chains. Recent studies indicate that these interactions not only regulate key events in development and homeostasis, but also key mechanisms of the host inflammatory response. This review will focus on the molecular and cellular aspects of how syndecans modulate tissue injury and inflammation, and how syndecans affect the outcome of inflammatory diseases in vivo.

Mulberry leaf aqueous fractions inhibit TNF-alpha-induced nuclear factor kappaB (NF-kappaB) activation and lectin-like oxidized LDL receptor-1 (LOX-1) expression in vascular endothelial cells.

Mulberry (Morus Alba L., family Moraceae) leaf extracts have various biological effects including inhibition of oxidative modification of low-density lipoprotein (LDL), which is the major cause of atherosclerosis. Endothelial dysfunction elicited by oxidized LDL (Ox-LDL) has been implicated in atherogenesis. Lectin-like Ox-LDL receptor-1 (LOX-1), a cell-surface receptor for atherogenic Ox-LDL, appears to mediate Ox-LDL-induced inflammation, which may be crucial in atherogenesis. Previous studies revealed that expression of LOX-1 is highly inducible by proinflammatory stimuli, including tumor necrosis factor-alpha (TNF-alpha), lipopolysaccharide (LPS), and transforming growth factor-beta (TGF-beta). Therefore, we examined whether mulberry leaf aqueous fractions inhibit LOX-1 expression induced by proinflammatory stimuli. Pretreatment of cultured bovine aortic endothelial cells (BAECs) with mulberry leaf aqueous fractions inhibited TNF-alpha- and LPS-induced expression of LOX-1 at both protein and mRNA levels in a time- and concentration-dependent manner. In contrast, mulberry leaf aqueous fractions did not affect TGF-beta-induced LOX-1 expression. Furthermore, mulberry leaf aqueous fractions inhibited TNF-alpha-induced activation of nuclear factor-kappaB (NF-kappaB) and phosphorylation of inhibitory factor of NF-kappaB-alpha (IkappaB-alpha) in a time- and concentration-dependent fashion. Thus, mulberry leaf aqueous fractions suppress TNF-alpha- and LPS-induced LOX-1 gene expression, by inhibiting NF-kappaB activation.

Syndecan-1 expression in epithelial cells is induced by transforming growth factor beta through a PKA-dependent pathway.

Syndecans comprise a major family of cell surface heparan sulfate proteoglycans (HSPGs). Syndecans bind and modulate a wide variety of biological molecules through their heparan sulfate (HS) moiety. Although all syndecans contain the ligand binding HS chains, they likely perform specific functions in vivo because their temporal and spatial expression patterns are different. However, how syndecan expression is regulated has yet to be clearly defined. In this study, we examined how syndecan-1 expression is regulated in epithelial cells. Our results showed that among several bioactive agents tested, only forskolin and three isoforms of TGFbeta (TGFbeta1-TGFbeta3) significantly induced syndecan-1, but not syndecan-4, expression on various epithelial cells. Steady-state syndecan-1 mRNA was not increased by TGFbeta treatment and cycloheximide did not inhibit syndecan-1 induction by TGFbeta, indicating that TGFbeta induces syndecan-1 in a post-translational manner. However, TGFbeta induction of syndecan-1 was inhibited by transient expression of a dominant-negative construct of protein kinase A (PKA) and by specific inhibitors of PKA. Further (i) syndecan-1 cytoplasmic domains were Ser-phosphorylated when cells were treated with TGFbeta and this was inhibited by a PKA inhibitor, (ii) PKA was co-immunoprecipitated from cell lysates by anti-syndecan-1 antibodies, (iii) PKA phosphorylated recombinant syndecan-1 cytoplasmic domains in vitro, and (iv) expression of a syndecan-1 construct with its invariant Ser(286) replaced with a Gly was not induced by TGFbeta. Together, these findings define a regulatory mechanism where TGFbeta signals through PKA to phosphorylate the syndecan-1 cytoplasmic domain and increases syndecan-1 expression on epithelial cells.

Serum soluble lectin-like oxidized low-density lipoprotein receptor-1 levels are elevated in acute coronary syndrome: a novel marker for early diagnosis.

BACKGROUND: Markers of cardiac injury, including troponin-T (TnT), are used to diagnose acute coronary syndrome (ACS); however, markers for plaque instability may be more useful for diagnosing ACS at the earliest stage. Lectin-like oxidized LDL receptor-1 (LOX-1) appears to play crucial roles in the pathogenesis of atherosclerotic plaque rupture and ACS onset. LOX-1 is released in part as soluble LOX-1 (sLOX-1) by proteolytic cleavage. METHODS AND RESULTS: We examined serum sLOX-1 levels in 521 patients, consisting of 427 consecutive patients undergoing coronary angiography, including 80 ACS patients, 173 symptomatic coronary heart disease patients, 122 patients with significant coronary stenosis without ischemia, and 52 patients without apparent coronary atherosclerosis plus 34 patients with noncardiac acute illness and 60 patients with noncardiac chronic illness. Time-dependent changes in sLOX-1 and TnT levels were analyzed in an additional 40 ACS patients. Serum sLOX-1 levels were significantly higher in ACS than the other groups and were associated with ACS as shown by multivariable logistic regression analyses. Given a cutoff value of 1.0 ng/mL, sLOX-1 can discriminate ACS from other groups with 81% and 75% of sensitivity and specificity, respectively. sLOX-1 can also discriminate ACS without ST elevation or abnormal Q waves and ACS without TnT elevation from non-ACS with 91% and 83% of sensitivity, respectively. Peak values of sLOX-1 in ACS were observed earlier than those of TnT. CONCLUSIONS: sLOX-1 appears to be a useful marker for early diagnosis of ACS.