Interaction between cytokines and inflammatory cells in islet dysfunction, insulin resistance and vascular disease

Abstract

Inflammation is an established pathogenic player in insulin resistance, islet demise and atherosclerosis. The complex interactions between cytokines, immune cells and affected tissues result in sustained inflammation in diabetes and atherosclerosis. 12- and 15-lipoxygenase (LO), such as 12/15-LO, produces a variety of metabolites through peroxidation of fatty acids and potentially contributes to the complex molecular crosstalk at the site of inflammation. 12- and 15-LO pathways are frequently activated in tissues affected by diabetes and atherosclerosis including adipose tissue (AT), islets and the vasculature. Moreover, mice with whole body and tissue-specific knockout of 12/15-LO are protected against insulin resistance, hyperglycaemia and atherosclerosis supporting functional contribution of 12- and 15-LO pathways in diabetes and atherosclerosis. Recently, it has emerged that there is a temporal regulation of the particular isoforms of 12- and 15-LO in human AT and islets during the development of type 1 and type 2 diabetes and obesity. Analyses of tissues affected by diabetes and atherosclerosis also implied the roles of interleukin (IL)-12 and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-1 (NOX-1) in islets and IL-17A in atherosclerosis. Future studies should aim to test the efficacy of inhibitions of these mediators for treatment of diabetes and atherosclerosis.

Keywords: 12-lipoxygenase; 15-lipoxygenase; NADPH oxidase-1; interleukin-12; interleukin-17A.

Figure 1. Generation of lipid metabolites by 12- and 15-lipoxygenase

A: arachidonic acid, ω-6 polyunsaturated fatty acid (20:4) is a substrate, 12- and 15-lipoxygenase (LO) produce 12(S)-hydroperoxyeicosatetraenoic acid (H(p)ETE) and 15(S)-H(p)ETE through oxidation at carbon-12 (9^th carbon from ω tail) and carbon-15 (6^th carbon from ω tail) respectively. H(p)ETEs are unstable intermediate metabolites and quickly converted to hydroxyeicosatetraenoic acids (HETEs). B: When docosahexaenoic acid (DHA), ω-3 polyunsaturated fatty acid (22:6) is a substrate, 12-LO activity produces 14(S)-hydroperoxy-docosahexaenoic-acid (H(p)DHA), followed by the formation of 14(S)-hydroxy-docosahexaenoic-acid (HDHA) through catalysis on carbon-14 (9^th carbon from ω tail). 17(S)-H(p)DHA followed by 17(S)-HDHA will be produced from DHA through catalysis on carbon-17 (6^th carbon from ω tail) by 15-LO activity. Different isoforms of 12- and 15-LO (Table 1) possess variable degree of combination of 12- and 15-LO activities. Also, 12- and 15-LO are active toward wide array of fatty acids 4. Here, the major metabolites relevant to the review are shown for clarity.

Figure 2. Variation in adipose tissue expression of leukocyte-12/15-LO and platelet-12-LO and lipid metabolite profile indb/dbmice

A: Phenotype of db/db mice (gray bar) and heterozygous +/db controls (black bar) at 5, 8 and 10 weeks of age (n=7-10 mice/group); at week 10 the db/db mice have full expression of obesity and are severely glucose intolerant; they also show a decline in the average islet number. B: Longitudinal expression of leukocyte-12/15-LO (L-12-LO) and platelet-12-LO (P-12-LO) in adipose tissue, isolated adipocytes and stromal vascular fraction (SVF); the two LO isoforms are abundantly expressed in SVF and show different patterns of expression. L-12-LO is increased early and diminishes with the decline in the metabolic phenotype; P-12-LO increases in parallel with the decline in the glucose tolerance and with beginning of islet loss. Gene expression was measured by real-time PCR using SYBR green primers and 18S as a housekeeping control (n=7-10/group). db/db mice in gray bars and controls in black bars. C: Lipid metabolite profile in the SVF of db/db mice (gray bar) and controls (black bar) at 8 and 10 weeks of age. Lipids were measured using LC-MS/MS on a Kinetec C18 column after extraction with methyl-tert-butyl- ether; chiral analysis was performed on the same system using a Chiralpack AD-LH column (n=7-10 mice/group); Data are mean ± s.e.m. Statistical analysis was performed using one-way ANOVA. HETE = hydroxyeicosatetraenoic acid; HDHA = hydroxy-docosahexanoic-acid.

Figure 3. Lipid profiling of ALOX12 and ALOX15 metabolites in subcutaneous (SC) and omental (OM) human adipose tissue

Paired biopsies were collected during bariatric surgery from 12 morbidly obese subjects (BMI= 44.3±3.8). Total adipose tissue was subjected to lipid extraction using methanol/NaOH under reducing conditions and the lipids were measured by LC-ESI-MS/MS using a Triplequad instrument Agilent 6460/1200SL equipped with a Phenomenex Kinetex Column. Statistical analysis was performed using paired Student's t-test. HETE = hydroxyeicosatetraenoic acid; HDHA: hydroxy-docosahexanoic- acid; HODE = 13-hydroxy-octadecadienoic acid.

Figure 4. Crosstalk between 12- and 15-lipoxygenase, cytokines, oxidative stress, and endoplasmic reticulum stress in cytokine-induced β-cell demise and dysfunction

It has been well established that the combination of cytokines including TNFα, IL-1β, and INFγ results in β-cell demise and dysfunction. 12- and 15-lipoxygenase (LO) plays a role in the process through its crosstalk with cytokines, oxidative stress, and endoplasmic reticulum (ER) stress pathways. 12/15-LO plays the major role in mouse islets, while 12(S)-LO is important in human islets. Both are collectively represented as 12-LO in the figure. The cytokine mixture upregulates 12-LO and increases its proinflammatory metabolite 12(S)-HETE. 12(S)-HETE stimulates production of IL-12, activates p38-MAPK, and induces NADPH oxidase -1 (NOX-1) in islets. Although not directly shown in islets, there is evidence implicating the upregulation of ER stress by 12(S)-HETE in islets. 12-LO also increases IL-12 directly. Once induced, IL-12 increases the production of cytokines in islets and creates a feed-forward loop to amplify islet inflammation. The induction of NOX-1 will increase reactive oxygen species (ROS) in islets, cause oxidative stress, induce CCL2 expression in islets, and further augment islet inflammation. The cytokine mixture also causes ER stress directly or through the activation of 12-LO, which is shown as the increase in Bip, Xbp1, spliced form of Xbp1 (Xbp1s), and CHOP. Importantly, ER stress itself will activate 12-LO, creating another feed-forward loop. Insulin secretion will be impaired as a result of cytokine induction, ROS production and ER stress. Also, the activation of p38-MAPK, oxidative stress, and ER stress results in apoptosis. Therefore, 12-LO plays a significant role in amplifying cytokine effects by creating positive feedback loops at multiple pathways including oxidative stress, ER stress, and inflammatory responses.