Nardilysin (N-arginine dibasic convertase; NRDc), a metalloendopeptidase, was identified in previous work by Nishi and colleagues as a potent activator for ectodomain shedding of several membrane proteins, including HB-EGF, TNF-alpha and APP, as well as some molecules involved in metabolic syndrome, including TNF-alpha receptor and leptin receptor. Ectodomain shedding is a post-translational modification that releases the extracellular domain of membrane-anchored proteins through proteolysis. NRDc, which has widespread expression throughout the body, potentiates catalytic activity of ADAM proteases via direct interaction.

Nishi presented data from their current in vivo examination of the biologic roles of NRDc in NRDc knockout (NRDcKO). About 70% of the mice died within 48 hours, despite the expected birth rate. Survivors, although healthy, had growth retardation, particularly in puberty, and decreased body temperature.

Findings on CT scanning at 3 months included:

• Less adipose tissue mass in NRDcKO mice (about 48% vs 100% in wild-type; WT).
• Less visceral fat in NRDcKO mice at about 20% compared to 100% in WT mice.

Other remarkable findings at 3 months included:

• Decreased size of adipocytes in white adipose tissue of NRDcKO versus WT mice.
• Lower serum insulin levels in NRDcKO mice vs WT mice before and after glucose injection, despite normal glucose tolerance.
• Enhanced insulin sensitivity on insulin tolerance testing in NRDcKO vs WT mice.
• No significant difference in total cholesterol between groups (about 70 mg/dL).
• Significantly lower triglycerides in NRDcKO vs WT mice, about 40 mg/dL vs 100 mg/dL.
• Hypotension in the NRDcKO, with systolic blood pressure about 25% lower than WT mice and diastolic about 35% less than WT mice.
• No significant difference in food intake between groups.
• Higher oxygen consumption over 24 hours (30% on average) in NRDcKO vs WT mice, particularly during the day.

Oxygen consumption is a measure of total expenditure, thus the higher oxygen consumption in the knockout mice may explain the reduced energy storage as fat accumulation.

Body temperature was lower at about 36 degrees Celsius in knockout mice compared to about 38 degrees in wild-type mice. Further, a cold challenge test revealed that the WT mice maintained their body temperature for 3 hours when exposed to 4 degrees Celsius, while body temperature was below 15 degrees Celsius in the knockout mice. This indicates NRDc is essential for adaptive thermogenesis.

Hyperactivity was revealed in the NRDcKO (1200 movement counts/hour, vs 800 in WT mice). This may partially explain the higher energy expenditure in the knockout mice.

Nishi stated these data indicate the essential roles of NRDc in energy metabolism. NRDc may be a novel therapeutic target for the treatment of metabolic syndrome, as the characteristics of the NRDcKO mice (decreased visceral fat content, low triglycerides, low blood pressure, enhanced insulin sensitivity, increased energy expenditure, hypothermia, hyperactivity) is the exact opposite of the characteristics of metabolic syndrome.

Production of plasminogen activator inhibitor-1 (PAI-1) is increased in vitro by insulin, IL-1, IL-6, and TNF-alpha, which are implicated in metabolic syndrome (MetS), as shown by previous work by Furumoto and colleagues at Hokkaido University Graduate School of Medicine. The effects of PAI-1 in MetS are not established, although its role in development of atherogenesis and cardiovascular disease has been established. Work by this group examining the role of PAI-1 on atherosclerotic vascular changes in two animal models was presented.

Studies in two mice models

In genetically obese mice, characterized by insulin resistance, hyperinsulinemia, obesity, and hypertension, this group found increased production of PAI-1 in coronary walls, PAI-1 mRNA in medial vascular walls, and at 20 weeks increased (nearly double) perivascular fibrosis and increased wall thickness.

In insulin resistance substrate-1 knockout mice (IRS-1KO), characterized by insulin resistance, hyperinsulinemia, and hypertension, they found at 20 weeks perivascular fibrosis and increased coronary wall thickness and overexpression of  PAI-1 protein and messenger levels in coronary arteries. These data suggest that atherosclerotic vascular changes in an animal model of MetS are accompanied by increasing PAI-1 expression in vascular walls.

PAI-1 in clinical setting

In the clinical setting, PAI-1levels indicate the clustering of clinical risk factors. Brachial femoral medial diameter (FMD), an initial marker of atherosclerotic vascular changes, has been shown to be significantly decreased in persons with MetS and hypertension. Brachial FMD is negatively correlated with brachial wall stress and PAI-1 activity is positively correlated with wall stress, suggesting that maintenance of normal wall stress may protect endothelial function. Thus, PAI-1 can also indicate atherosclerotic vascular changes in persons with MetS.

PAI-1: principal adipocytokine, new marker of MetS

In the obese mouse model, this group showed that PAI-1 is significantly increased, while it is significantly decreased in the IRS-1KO mice.

In the IRS-1KO mice, plasma PAI-1 is overexpressed in adipose tissues. This may be a result of the significant decrease in adipose tissue weight (by about one-half), especially in visceral fat, in IRS-1KO mice.

Insulin and TNF-alpha can increase PAI-1 levels alone and synergistically. In the setting of obesity, a synergistic effect is seen where an increase in adipose tissue leads to an increase in insulin and TNF-alpha which leads to an increase in PAI-1. Thus, in MetS, levels of PAI-1 can easily indicate adipose tissue levels and PAI-1 can cause vascular changes in MetS.

Medical treatment of PAI-1 in MetS

Work by Furumoto and colleagues showed that the ACE inhibitor temocapril decreased fibrotic changes in the vascular lumen and wall of obese mice. Temocapril also decreased PAI-1 levels by about one-third in the vascular wall, as measured by quantitative imaging, in obese animals. Also in obese animals, temocapril reduced perivascular fibrosis. A significant reduction in plasma PAI-1 levels was found with ACE inhibitors and angiotensin receptor blockers.  Thus, PAI-1 guided medication for MetS may be effective to prevent the development of atherosclerotic vascular changes, stated Furumoto.

PAI-1 modulates intravascular thrombosis and thrombolysis and breaks down the extracellular matrix and regulates cell adhesion and migration. In conclusion, PAI-1 indicates the clustering of risk factors and is an initiator insulin resistance, hyperinsulinemia, and early vascular damage, and reflects the amount of visceral fat. RAS inhibition can improve atherosclerotic vascular changes by decreasing PAI-1 production in the setting of metabolic syndrome.