Recently, it has been reported that 25% of plasma angiotensinogen (Agt) is derived from fat. Meanwhile, liver-specific Agt knockout (KO) mice have markedly low plasma Agt, which may be due to reduced fat mass. To study the contribution of the fat to plasma Agt, we tested whether increasing fat mass can elevate plasma Agt and blood pressure in liver- Agt KO mice. Epididymal fat mass in liver- Agt KO mice fed a high-fat diet (HFD) was 4.1-fold larger than that in liver- Agt KO mice on a normal-fat diet (NFD). The liver- Agt KO mice on NFD were hypotensive with low levels of plasma Agt (on average, 0.11 vs 2.38 μg/ml). HFD slightly increased plasma Agt (0.17 μg/ml) without increase in blood pressure. To further increase fat mass, liver- Agt KO mice were fed HFD and simultaneously supplemented with low-dose angiotensin II and compared with control mice. Fat mass was comparable between the two groups. However, liver- Agt KO mice had uniformly low plasma Agt (0.09 vs 2.07 μg/ml) and systolic blood pressure (78±12 vs 111±6 mm Hg). In conclusion, adipocyte-derived Agt has essentially no contribution to the plasma concentration and no impact on blood pressure compared to liver-derived Agt.

Angiotensinogen is the precursor of angiotensin II, which is associated with ischemia-reperfusion injury. Angiotensin II reduces liver regeneration after hepatectomy and causes dysfunction and failure of reduced-size liver transplants. However, the regulation of angiotensinogen during liver regeneration is still unclear.

Elevated urinary angiotensinogen (UA) was identified as novel prognostic biomarker capable of predicting chronic kidney disease, and in the present study, we will investigate the diagnostic value of UA in the patients of nephrolithiasis.

Angiotensinogen (AGT) belongs to the serpin superfamily. It acts as the unique substrate of all angiotensin peptides, which generates a spectrum of angiotensin peptides in the renin-angiotensin system and regulates hypertension. This serpin belongs to the multiple member group V2 of the intron encoded vertebrate serpin classification. Despite huge advancements in the understanding of angiotensinogen based on biochemical properties and its roles in the RAS, phylogenetic history of AGT remains forgotten. To date, there is no comprehensive study illustrating the phylogenetic history of AGT. Herein, we investigated phylogenetic traits of AGT gene across vertebrates. Gene structures of AGT gene from selected ray-finned fishes varied in exon I and II with insertions of two novel introns in the core domain for ray-finned fishes at the position 77c and 233c. We that found AGT loci is conserved from lampreys to human and estimated to be older than 500 MY. By comparing AGT protein in 57 vertebrate genomes, we illustrated that the reactive center loop (RCL) of AGT protein became from inhibitory (in lampreys, GTEAKAETVVGIMPI†SMPPT) to non-inhibitory (in human, EREPTESTQQLNKPE†VLEVT) during period of 500 MY. We identified 690 AGT variants by analysis of 1092 human genomes with top three variation classes belongs to SNPs (89.7%), somatic SNVs (5.2%) and deletion (2.9%). There are 32 key residues out of 121 missense variants, which are deleterious for AGT protein, computed by combination of SIFT and PolyPhen V2 methods. These results may have clinical implications for understanding hypertension.

Activation of the renin-angiotensin-aldosterone system (RAAS) is an important mediator of diabetic nephropathy. Urinary angiotensinogen, a novel biomarker of the intrarenal RAAS, is associated with progressive kidney injury. In this study, the authors investigated the determinants of urinary angiotensinogen and its associations with staging of diabetic nephropathy.

Blood pressure is critically controlled by angiotensins, which are vasopressor peptides specifically released by the enzyme renin from the tail of angiotensinogen-a non-inhibitory member of the serpin family of protease inhibitors. Although angiotensinogen has long been regarded as a passive substrate, the crystal structures solved here to 2.1 Å resolution show that the angiotensin cleavage site is inaccessibly buried in its amino-terminal tail. The conformational rearrangement that makes this site accessible for proteolysis is revealed in our 4.4 Å structure of the complex of human angiotensinogen with renin. The co-ordinated changes involved are seen to be critically linked by a conserved but labile disulphide bridge. Here we show that the reduced unbridged form of angiotensinogen is present in the circulation in a near 40:60 ratio with the oxidized sulphydryl-bridged form, which preferentially interacts with receptor-bound renin. We propose that this redox-responsive transition of angiotensinogen to a form that will more effectively release angiotensin at a cellular level contributes to the modulation of blood pressure. Specifically, we demonstrate the oxidative switch of angiotensinogen to its more active sulphydryl-bridged form in the maternal circulation in pre-eclampsia-the hypertensive crisis of pregnancy that threatens the health and survival of both mother and child.

Earlier published studies investigating the association between polymorphisms in the angiotensinogen gene and lung cancer risk showed no consistent results. In this study, we have summarized all currently available data to examine the correlation by meta-analysis. Case-control studies addressing the association being examined were identified through Embase, the Cochrane Library, ISI Web of Science (Web of Knowledge), Google Scholar, PubMed, and CNKI databases. Risk of lung cancer (odds ratio [OR] and 95% confidence interval [CI]) was estimated with the fixed or the random effects model assuming homozygous, allele, heterozygous, dominant, and recessive models for all angiotensinogen polymorphisms. We identified a total of 10 articles in this meta-analysis, including 7 for Leu84Phe, 4 for Ile143Val, and 3 for Leu53Leu. In the meta-analysis of Leu84Phe polymorphism, the homozygous model provided an OR of 1.44 (Phe/Phe vs Ile/Ile: OR = 1.44, 95% CI = 1.04-1.99, P values for heterogeneity test (Q-test) [P(Het)] = 0.382). The significantly increased risk was similarly indicated in the recessive model (Phe/Phe vs Phe/Ile + Ile/Ile: OR = 1.41, 95% CI = 1.02-1.95, P(Het) = 0.381). We also observed a positive association in the Caucasian subgroup. The heterozygous model and the dominant model tested for the Ile143Val polymorphism showed a marginally increased risk (Ile/Val vs Ile/Ile: OR = 1.16, 95% CI = 1.00-1.36, P(Het) = 0.323; Val/Val + Ile/Val vs Ile/Ile: OR = 1.15, 95% CI = 0.99-1.34, P(Het) = 0.253). These data suggest that Leu84Phe and Ile143Val polymorphisms in the angiotensinogen gene may be useful biomarkers for lung cancer in some specific populations.

Renin-angiotensin system (RAS) inhibitors reduce glomerular injury and proteinuria, indicating that angiotensin II (Ang II) is involved in glomerular diseases. Although the local RAS is reported to play an essential role in maintaining local tissue functions, the role of the local RAS in regulating glomerular function is not well evaluated. In this study, we analyzed the glomerular expression of RAS components in nephrotic models and the effect of Ang II receptor blockers (ARB) on the expression of angiotensinogen (AGT).

This study was performed to determine whether immunoreactivity of intrarenal hemeoxygenase-1 and angiotensinogen are increased in IgA nephropathy (IgAN) patients. Hemeoxygenase-1 and angiotensinogen immunoreactivity were determined by immunohistochemistry robot system in renal specimens from 39 patients with IgAN. Normal portions of surgically resected kidney served as controls. IgAN patients showed moderate proteinuria (1.1+/-0.2 g/day); however, the control group did not show any proteinuria. Immunoreactivity of intrarenal hemeoxygenase-1 and angiotensinogen in IgAN were significantly increased compared to normal kidneys (2.42+/-0.42 vs 1.00+/-0.26 for hemeoxygenase-1 and 4.05+/-0.40 vs 1.00+/-0.21 for angiotensinogen, arbitrary unit). Even though these IgAN patients did not show massive renal damage, hemeoxygenase-1 and angiotensinogen immunoreactivity were increased in these patients at this time point. These data suggest that activated intrarenal reactive oxygen species-angiotensinogen axis plays some roles in development of IgAN at the early stage and will provide supportive foundation of effectiveness of the renin-angiotensin system blockade in IgAN.

Prenatal programming results in an increase in blood pressure in adult offspring. We have shown that compared to control adult offspring whose mothers were fed a 20% protein diet, programmed adults whose mothers were fed a 6% protein diet during the last half of pregnancy have an increase in renal sympathetic nerve activity and urinary angiotensinogen/creatinine levels. We hypothesized that the increase in urinary angiotensinogen was mediated by renal sympathetic nerve activity in programmed rats. In this study performed in 3 month old rats, renal denervation resulted in normalization of blood pressure in the 6% programmed group (150 ± 3 Hg in 6% sham vs. 121 ± 4 Hg in 6% denervated, P < 0.001), and a reduction in blood pressure in the 20% group (126 ± 2 Hg 20% sham vs. 113 ± 4 Hg 20% denervated (P < 0.05). We confirm that the intrarenal renin-angiotensin system assessed by urinary angiotensinogen/creatinine is upregulated in offspring of rats fed a 6% protein diet rats compared to 20% controls. To determine if sympathetic nerve activity was mediating the increase in urinary angiotensinogen in programmed rats, we compared denervated to sham-operated control and programmed rats. Renal denervation had no effect on urinary angiotensinogen/creatinine ratio in the 20% group and no effect on the increased urinary angiotensinogen/creatinine ratio found in programmed rats. This study demonstrates that the increase in urinary angiotensinogen in programmed rats is not mediated by renal sympathetic nerve activity.

Enhanced intrarenal renin-angiotensin system (RAS) is implicated in the development and progression of renal injury. To investigate whether angiotensinogen (AGT) expression is involved in glomerular RAS activity and glomerular injury, we examined glomerular AGT expression and its correlation with expression of other RAS components, and levels of glomerular injury in samples from patients with immunoglobulin A nephropathy (IgAN) (23) and minor glomerular abnormalities (MGA) (8). Immunohistochemistry showed that AGT protein was highly expressed by glomerular endothelial cells (GEC) and mesangial cells in nephritic glomeruli of IgAN compared with glomeruli of MGA. Levels of glomerular AGT protein were well correlated with levels of glomerular angiotensin II (ang II), transforming growth factor-beta (TGF-beta), alpha-smooth-muscle actin, glomerular cell number, and glomerulosclerosis score but not with those of glomerular angiotensin-converting enzyme and ang II type 1 receptor. Real-time polymerase chain reaction (RT-PCR) and Western blot analyses using cultured human GEC indicated that ang II upregulated AGT messenger ribonucleic acid (mRNA) and protein expression in a dose- and time-dependent manner. These data suggest that activated glomerular AGT expression is likely involved in elevated local ang II production and, thereby, may contribute to increased TGF-beta production and development of glomerular injury in IgAN. Augmentation of GEC-AGT production with ang II stimulation might drive further glomerular injury in a positive-feedback loop.

Numerous studies have evaluated the association between the angiotensinogen (AGT) T174M polymorphism and ischemic stroke(IS) risk. However, the specific association is still controversial.

The role of intranephron angiotensinogen (AGT) in blood pressure (BP) regulation is not fully understood. Previous studies showed that proximal tubule-specific overexpression of AGT increases BP, whereas proximal tubule-specific deletion of AGT did not alter BP. The latter study may not have completely eliminated nephron AGT production; in addition, BP was only assessed on a normal salt diet. To evaluate this issue in greater detail, we developed mice with inducible nephron-wide AGT deletion. Mice were generated which were hemizygous for the Pax8-rtTA and LC-1 transgenes and homozygous for loxP-flanked AGT alleles to achieve nephron-wide AGT disruption after doxycycline induction. Compared to controls, AGT knockout (KO) mice demonstrated markedly reduced renal AGT immunostaining, mRNA, and protein levels; unexpectedly AGT KO mice had reduced AGT mRNA levels in the liver along with 50% reduction in plasma AGT levels. BP was significantly lower in the AGT KO mice compared to controls fed a normal, low, or high Na(+) intake, with the highest BP reduction on a low Na(+) diet. Regardless of Na(+) intake, AGT KO mice had higher plasma renin concentration (PRC) and markedly reduced urinary AGT levels compared to controls. Following angiotensin-II (Ang-II) infusion, AGT KO mice demonstrated an attenuated hypertensive response despite similar suppression of PRC in the two groups. Taken together, these data suggest that nephron-derived AGT may be involved in Ang-II-dependent hypertension, however, a clear role for nephron-derived AGT in physiological BP regulation remains to be determined.

Angiotensinogen (AGT), the precursor of angiotensin I, is known to be involved in tumor angiogenesis and associated with the pathogenesis of coronary atherosclerosis. This study was undertaken to determine the role played by AGT in endothelial progenitor cells (EPCs) in tumor progression and metastasis. It was found that the number of EPC colonies formed by AGT heterozygous knockout (AGT(+/-)) cells was less than that formed by wild-type (WT) cells, and that the migration and tube formation abilities of AGT(+/-) EPCs were significantly lower than those of WT EPCs. In addition, the gene expressions of vascular endothelial growth factor (VEGF), Flk1, angiopoietin (Ang)-1, Ang-2, Tie-2, stromal derived factor (SDF)-1, C-X-C chemokine receptor type 4 (CXCR4), and of endothelial nitric oxide synthase (eNOS) were suppressed in AGT(+/-) EPCs. Furthermore, the expressions of hypoxia-inducible factor (HIF)-1α and -2α were downregulated in AGT(+/-) early EPCs under hypoxic conditions, suggesting a blunting of response to hypoxia. Moreover, the activation of Akt/eNOS signaling pathways induced by VEGF, epithelial growth factor (EGF), or SDF-1α were suppressed in AGT(+/-) EPCs. In AGT(+/-) mice, the incorporation of EPCs into the tumor vasculature was significantly reduced, and lung tumor growth and melanoma metastasis were attenuated. In conclusion, AGT is required for hypoxia-induced vasculogenesis.

Patients with recurrent glioblastoma achieving response to bevacizumab combined with chemotherapy have clinical improvement and prolonged survival. High gene expression of angiotensinogen (AGT) is associated with a poor bevacizumab response. Because AGT expression is epigenetically regulated, we aimed to investigate whether AGT promoter methylation in tumor tissue predicts response to bevacizumab combination therapy in patients with recurrent glioblastoma. The study included 159 patients with recurrent glioblastoma, treated with bevacizumab combination treatment (training cohort, n = 77; validation cohort, n = 82). All patients could be evaluated for treatment response and biomarkers. DNA methylation of 4 CpG sites in the AGT promoter was measured using pyrosequencing. A model for nonresponse was established using logistic regression analysis. In the training cohort, lower methylation of each of the four CpG sites in the AGT promoter was significantly associated with nonresponse (all P < 0.05). Moreover, the mean methylation level of all four CpG sites was associated with an increased likelihood of not achieving response to bevacizumab combination therapy (twofold decrease: odds ratio = 3.01; 95% confidence interval: 1.41-6.44; P = 0.004). We developed a model for nonresponse in the training cohort, where a threshold of mean AGT promoter methylation levels was set to below 12%. The model could predict bevacizumab nonresponse with 96% specificity. Importantly, this predictor was also significantly associated with nonresponse in the validation cohort (P = 0.037). Taken together, our findings suggest that low AGT promoter methylation in tumor tissue predicts nonresponse to bevacizumab combination treatment in patients with recurrent glioblastoma. We have, thus, established and successfully validated a predictor for nonresponse that can be used to identify patients who will not benefit from bevacizumab combination therapy.

Hypertension is caused by a combination of genetic and environmental factors. Angiotensinogen (AGT) is a component of RAAS, that regulates blood pressure. The human angiotensinogen (hAGT) gene has -6A/-6G polymorphism and -6A variant is associated with human hypertension. In this study, we have investigated the epigenetic regulation of the hAGT. To understand transcriptional regulation of the hAGT, we have made transgenic animals containing -6A. We show that HS affects DNA methylation and modulates transcriptional regulation of this gene in liver and kidney. High salt (HS) increases hAGT gene expression in -6A TG mice. We have observed that the number of CpG sites in the hAGT promoter is decreased after HS treatment. In the liver, seven CpG sites are methylated whereas after HS treatment, only three CpG sites remain methylated. In the kidney, five CpG sites are methylated, whereas after HS treatment, only three CpG sites remain methylated. These results suggest that HS promotes DNA demethylation and increasing AGT gene expression. RT-PCR and immunoblot analysis show that hAGT gene expression is increased by HS. Chip assay has shown that transcription factors bind strongly after HS treatment. RNA-Seq identified differentially expressed genes, novel target genes associated with hypertension, top canonical pathways, upstream regulators. One of the plausible mechanisms for HS induced up-regulation of the hAGT gene is through IL-6/JAK/STAT3/AGT axis.

Nonalcoholic fatty liver disease (NAFLD) is considered as a liver manifestation of metabolic disorders. Previous studies indicate that the renin-angiotensin system (RAS) plays a complex role in NAFLD. As the only precursor of the RAS, decreased angiotensinogen (AGT) profoundly impacts RAS bioactivity. Here, we investigated the role of hepatocyte-derived AGT in liver steatosis. AGT floxed mice (hepAGT+/+) and hepatocyte-specific AGT-deficient mice (hepAGT-/-) were fed a Western diet and a normal laboratory diet for 12 weeks, respectively. Compared with hepAGT+/+ mice, Western diet-fed hepAGT-/- mice gained less body weight with improved insulin sensitivity. The attenuated severity of liver steatosis in hepAGT-/- mice was evidenced by histologic changes and reduced intrahepatic triglycerides. The abundance of SREBP1 and its downstream molecules, acetyl-CoA carboxylase and FASN, was suppressed in hepAGT-/- mice. Furthermore, serum derived from hepAGT+/+ mice stimulated hepatocyte SREBP1 expression, which could be diminished by protein kinase B (Akt)/mammalian target of rapamycin (mTOR) inhibition in vitro. Administration of losartan did not affect diet-induced body weight gain, liver steatosis severity, and hepatic p-Akt, p-mTOR, and SREBP1 protein abundance in hepAGT+/+ mice. These data suggest that attenuation of Western diet-induced liver steatosis in hepAGT-/- mice is associated with the alternation of the Akt/mTOR/SREBP-1c pathway.

The renin-angiotensin cascade is a hormone system that regulates blood pressure and fluid balance. Renin-mediated cleavage of the angiotensin I peptide from the N terminus of angiotensinogen (AGT) is the rate-limiting step of this cascade; however, the detailed molecular mechanism underlying this step is unclear. Here, we solved the crystal structures of glycosylated human AGT (2.30 Å resolution), its encounter complex with renin (2.55 Å), AGT cleaved in its reactive center loop (RCL; 2.97 Å), and spent AGT from which the N-terminal angiotensin peptide was removed (2.63 Å). These structures revealed that AGT undergoes profound conformational changes and binds renin through a tail-into-mouth allosteric mechanism that inserts the N terminus into a pocket equivalent to a hormone-binding site on other serpins. These changes fully extended the N-terminal tail, with the scissile bond for angiotensin release docked in renin's active site. Insertion of the N terminus into this pocket accompanied a complete unwinding of helix H of AGT, which, in turn, formed key interactions with renin in the complementary binding interface. Mutagenesis and kinetic analyses confirmed that renin-mediated production of angiotensin I is controlled by interactions of amino acid residues and glycan components outside renin's active-site cleft. Our findings indicate that AGT adapts unique serpin features for hormone delivery and binds renin through concerted movements in the N-terminal tail and in its main body to modulate angiotensin release. These insights provide a structural basis for the development of agents that attenuate angiotensin release by targeting AGT's hormone binding pocket.

Urinary angiotensinogen (UAGT) is supposed to be a marker of activation of the intrarenal renin- angiotensin system (RAS) system in early diabetic nnephropathy (EDN). Vitamin D has been studied as a negative regulator of the circulating and tissue RAS activity, so its supplementation may prevent the progression of diabetic nephropathy (DN). This study was planned to assess the RAS activation and effect of vitamin D supplementation in EDN progression by estimating the UAGT level.

Sexual difference has been shown in the pathogenesis of chronic kidney disease induced by hypertension. Females are protected from hypertension and related end-organ damage. Augmentation of renal proximal tubular angiotensinogen (AGT) expression can promote intrarenal angiotensin formation and the development of associated hypertension and kidney injury. Female rodents exhibit lower intrarenal AGT levels than males under normal conditions, suggesting that the suppressed intrarenal AGT production by programmed mechanisms in females may provide protection from these diseases. This study was performed to examine whether epigenetic mechanisms serve as repressors of AGT.