Antisense oligonucleotide targeting hepatic Serum Amyloid A limits the progression of angiotensin II-induced abdominal aortic aneurysm formation.

BACKGROUND AND AIMS: Obesity increases the risk for abdominal aortic aneurysms (AAA) in humans and enhances angiotensin II (AngII)-induced AAA formation in C57BL/6 mice. We reported that deficiency of Serum Amyloid A (SAA) significantly reduces AngII-induced inflammation and AAA in both hyperlipidemic apoE-deficient and obese C57BL/6 mice. The aim of this study is to investigate whether SAA plays a role in the progression of early AAA in obese C57BL/6 mice. METHODS: Male C57BL/6J mice were fed a high-fat diet (60% kcal as fat) throughout the study. After 4 months of diet, the mice were infused with AngII until the end of the study. Mice with at least a 25% increase in the luminal diameter of the abdominal aorta after 4 weeks of AngII infusion were stratified into 2 groups. The first group received a control antisense oligonucleotide (Ctr ASO), and the second group received ASO that suppresses SAA (SAA-ASO) until the end of the study. RESULTS: Plasma SAA levels were significantly reduced by the SAA ASO treatment. While mice that received the control ASO had continued aortic dilation throughout the AngII infusion periods, the mice that received SAA-ASO had a significant reduction in the progression of aortic dilation, which was associated with significant reductions in matrix metalloprotease activities, decreased macrophage infiltration and decreased elastin breaks in the abdominal aortas. CONCLUSIONS: We demonstrate for the first time that suppression of SAA protects obese C57BL/6 mice from the progression of AngII-induced AAA. Suppression of SAA may be a therapeutic approach to limit AAA progression.

Deficiency of Acute-Phase Serum Amyloid A Exacerbates Sepsis-Induced Mortality and Lung Injury in Mice.

Serum amyloid A (SAA) is a family of proteins, the plasma levels of which may increase >1000-fold in acute inflammatory states. We investigated the role of SAA in sepsis using mice deficient in all three acute-phase SAA isoforms (SAA-TKO). SAA deficiency significantly increased mortality rates in the three experimental sepsis mouse models: cecal ligation and puncture (CLP), cecal slurry (CS) injection, and lipopolysaccharide (LPS) treatments. SAA-TKO mice had exacerbated lung pathology compared to wild-type (WT) mice after CLP. A bulk RNA sequencing performed on lung tissues excised 24 h after CLP indicated significant enrichment in the expression of genes associated with chemokine production, chemokine and cytokine-mediated signaling, neutrophil chemotaxis, and neutrophil migration in SAA-TKO compared to WT mice. Consistently, myeloperoxidase activity and neutrophil counts were significantly increased in the lungs of septic SAA-TKO mice compared to WT mice. The in vitro treatment of HL-60, neutrophil-like cells, with SAA or SAA bound to a high-density lipoprotein (SAA-HDL), significantly decreased cellular transmigration through laminin-coated membranes compared to untreated cells. Thus, SAA potentially prevents neutrophil transmigration into injured lungs, thus reducing exacerbated tissue injury and mortality. In conclusion, we demonstrate for the first time that endogenous SAA plays a protective role in sepsis, including ameliorating lung injury.

Antisense Oligonucleotide Targeting Hepatic Serum Amyloid A Limits the Progression of Angiotensin II-Induced Abdominal Aortic Aneurysm Formation.

OBJECTIVE: Obesity increases the risk for abdominal aortic aneurysms (AAA) in humans and enhances angiotensin II (AngII)-induced AAA formation in C57BL/6 mice. Obesity is also associated with increases in serum amyloid A (SAA). We previously reported that deficiency of SAA significantly reduces AngII-induced inflammation and AAA in both hyperlipidemic apoE-deficient and obese C57BL/6 mice. In this study, we investigated whether SAA plays a role in the progression of early AAA in obese C57BL/6 mice. APPROACH AND RESULTS: Male C57BL/6J mice were fed a high-fat diet (60% kcal as fat) throughout the study. After 4 months of diet, the mice were infused with angiotensin II (AngII) until the end of the study. Mice with at least a 25% increase in the luminal diameter of the abdominal aorta after 4 weeks of AngII infusion were stratified into 2 groups. The first group received a control antisense oligonucleotide (Ctr ASO), and the second group received ASO that suppresses SAA (SAA-ASO) until the end of the study. Plasma SAA levels were significantly reduced by the SAA ASO treatment. While mice that received the control ASO had continued aortic dilation throughout the AngII infusion periods, the mice that received SAA-ASO had a significant reduction in the progression of aortic dilation, which was associated with significant reductions in matrix metalloprotease activities, decreased macrophage infiltration and decreased elastin breaks in the abdominal aortas. CONCLUSION: We demonstrate for the first time that suppression of SAA protects obese C57BL/6 mice from the progression of AngII-induced AAA. Suppression of SAA may be a therapeutic approach to limit AAA progression.

Serum amyloid A augments the atherogenic effects of cholesteryl ester transfer protein.

Serum amyloid A (SAA) is predictive of CVD in humans and causes atherosclerosis in mice. SAA has many proatherogenic effects in vitro. However, HDL, the major carrier of SAA in the circulation, masks these effects. The remodeling of HDL by cholesteryl ester transfer protein (CETP) liberates SAA restoring its proinflammatory activity. Here, we investigated whether deficiency of SAA suppresses the previously described proatherogenic effect of CETP. ApoE-/- mice and apoE-/- mice deficient in the three acute-phase isoforms of SAA (SAA1.1, SAA2.1, and SAA3; "apoE-/- SAA-TKO") with and without adeno-associated virus-mediated expression of CETP were studied. There was no effect of CETP expression or SAA genotype on plasma lipids or inflammatory markers. Atherosclerotic lesion area in the aortic arch of apoE-/- mice was 5.9 ± 1.2%; CETP expression significantly increased atherosclerosis in apoE-/- mice (13.1 ± 2.2%). However, atherosclerotic lesion area in the aortic arch of apoE-/- SAA-TKO mice (5.1 ± 1.1%) was not significantly increased by CETP expression (6.2 ± 0.9%). The increased atherosclerosis in apoE-/- mice expressing CETP was associated with markedly increased SAA immunostaining in aortic root sections. Thus, SAA augments the atherogenic effects of CETP, which suggests that inhibiting CETP may be of particular benefit in patients with high SAA.

Serum Amyloid A is not obligatory for high-fat, high-sucrose, cholesterol-fed diet-induced obesity and its metabolic and inflammatory complications.

Several studies in the past have reported positive correlations between circulating Serum amyloid A (SAA) levels and obesity. However, based on limited number of studies involving appropriate mouse models, the role of SAA in the development of obesity and obesity-related metabolic consequences has not been established. Accordingly, herein, we have examined the role of SAA in the development of obesity and its associated metabolic complications in vivo using mice deficient for all three inducible forms of SAA: SAA1.1, SAA2.1 and SAA3 (TKO). Male and female mice were rendered obese by feeding a high fat, high sucrose diet with added cholesterol (HFHSC) and control mice were fed rodent chow diet. Here, we show that the deletion of SAA does not affect diet-induced obesity, hepatic lipid metabolism or adipose tissue inflammation. However, there was a modest effect on glucose metabolism. The results of this study confirm previous findings that SAA levels are elevated in adipose tissues as well as in the circulation in diet-induced obese mice. However, the three acute phase SAAs do not play a causative role in the development of obesity or obesity-associated adipose tissue inflammation and dyslipidemia.

Adipocyte-Derived Serum Amyloid A Promotes Angiotensin II-Induced Abdominal Aortic Aneurysms in Obese C57BL/6J Mice.

BACKGROUND: Obesity increases the risk for human abdominal aortic aneurysms (AAAs) and enhances Ang II (angiotensin II)-induced AAA formation in C57BL/6J mice. Obesity is also associated with increases in perivascular fat that expresses proinflammatory markers including SAA (serum amyloid A). We previously reported that deficiency of SAA significantly reduces Ang II-induced inflammation and AAA in hyperlipidemic apoE-deficient mice. In this study. we investigated whether adipose tissue-derived SAA plays a role in Ang II-induced AAA in obese C57BL/6J mice. METHODS: The development of AAA was compared between male C57BL/6J mice (wild type), C57BL/6J mice lacking SAA1.1, SAA2.1, and SAA3 (TKO); and TKO mice harboring a doxycycline-inducible, adipocyte-specific SAA1.1 transgene (TKO-Tgfat; SAA expressed only in fat). All mice were fed an obesogenic diet and doxycycline to induce SAA transgene expression and infused with Ang II to induce AAA. RESULTS: In response to Ang II infusion, SAA expression was significantly increased in perivascular fat of obese C57BL/6J mice. Maximal luminal diameters of the abdominal aorta were determined by ultrasound before and after Ang II infusion, which indicated a significant increase in aortic luminal diameters in wild type and TKO-TGfat mice but not in TKO mice. Adipocyte-specific SAA expression was associated with MMP (matrix metalloproteinase) activity and macrophage infiltration in abdominal aortas of Ang II-infused obese mice. CONCLUSIONS: We demonstrate for the first time that SAA deficiency protects obese C57BL/6J mice from Ang II-induced AAA. SAA expression only in adipocytes is sufficient to cause AAA in obese mice infused with Ang II.

The dual role of group V secretory phospholipase A2 in pancreatic ß-cells.

PURPOSE: Group X (GX) and group V (GV) secretory phospholipase A2 (sPLA2) potently release arachidonic acid (AA) from the plasma membrane of intact cells. We previously demonstrated that GX sPLA2 negatively regulates glucose-stimulated insulin secretion (GSIS) by a prostaglandin E2 (PGE2)-dependent mechanism. In this study we investigated whether GV sPLA2 similarly regulates GSIS. METHODS: GSIS and pancreatic islet-size were assessed in wild-type (WT) and GV sPLA2-knock out (GV KO) mice. GSIS was also assessed ex vivo in isolated islets and in vitro using MIN6 pancreatic beta cell lines with or without GV sPLA2 overexpression or silencing. RESULTS: GSIS was significantly decreased in islets isolated from GV KO mice compared to WT mice and in MIN6 cells with siRNA-mediated GV sPLA2 suppression. MIN6 cells overexpressing GV sPLA2 (MIN6-GV) showed a significant increase in GSIS compared to control cells. Though the amount of AA released into the media by MIN6-GV cells was significantly higher, PGE2 production was not enhanced or cAMP content decreased compared to control MIN6 cells. Surprisingly, GV KO mice exhibited a significant increase in plasma insulin levels following i.p. injection of glucose compared to WT mice. This increase in GSIS in GV KO mice was associated with a significant increase in pancreatic islet size and number of proliferating cells in ß-islets compared to WT mice. CONCLUSIONS: Deficiency of GV sPLA2 results in diminished GSIS in isolated pancreatic beta-cells. However, the reduced GSIS in islets lacking GV sPLA2 appears to be compensated by increased islet mass in GV KO mice.