14-3-3ζ: A suppressor of inflammatory arthritis.

Inflammatory arthritis (IA) is a common disease that affects millions of individuals worldwide. Proinflammatory events during IA pathogenesis are well studied; however, loss of protective immunity remains underexplored. Earlier, we reported that 14-3-3zeta (ζ) has a role in T-cell polarization and interleukin (IL)-17A signal transduction. Here, we demonstrate that 14-3-3ζ knockout (KO) rats develop early-onset severe arthritis in two independent models of IA, pristane-induced arthritis and collagen-induced arthritis. Arthritic 14-3-3ζ KO animals showed an increase in bone loss and immune cell infiltration in synovial joints. Induction of arthritis coincided with the loss of anti-14-3-3ζ antibodies; however, rescue experiments to supplement the 14-3-3ζ antibody by passive immunization did not suppress arthritis. Instead, 14-3-3ζ immunization during the presymptomatic phase resulted in significant suppression of arthritis in both wild-type and 14-3-3ζ KO animals. Mechanistically, 14-3-3ζ KO rats exhibited elevated inflammatory gene signatures at the messenger RNA and protein levels, particularly for IL-1ß. Furthermore, the immunization with recombinant 14-3-3ζ protein suppressed IL-1ß levels, significantly increased anti-14-3-3ζ antibody levels and collagen production, and preserved bone quality. The 14-3-3ζ protein increased collagen expression in primary rat mesenchymal cells. Together, our findings indicate that 14-3-3ζ causes immune suppression and extracellular remodeling, which lead to a previously unrecognized IA-suppressive function.

Genetically engineered Lactobacillus paracasei rescues colonic angiotensin converting enzyme 2 (ACE2) and attenuates hypertension in female Ace2 knock out rats.

Engineered gut microbiota represents a new frontier in medicine, in part serving as a vehicle for the delivery of therapeutic biologics to treat a range of host conditions. The gut microbiota plays a significant role in blood pressure regulation; thus, manipulation of gut microbiota is a promising avenue for hypertension treatment. In this study, we tested the potential of Lactobacillus paracasei, genetically engineered to produce and deliver human angiotensin converting enzyme 2 (Lacto-hACE2), to regulate blood pressure in a rat model of hypertension with genetic ablation of endogenous Ace2 (Ace2-/- and Ace2-/y). Our findings reveal a sex-specific reduction in blood pressure in female (Ace2-/-) but not male (Ace2-/y) rats following colonization with the Lacto-hACE2. This beneficial effect of lowering blood pressure was aligned with a specific reduction in colonic angiotensin II, but not renal angiotensin II, suggesting the importance of colonic Ace2 in the regulation of blood pressure. We conclude that this approach of targeting the colon with engineered bacteria for delivery of ACE2 represents a promising new paradigm in the development of antihypertensive therapeutics.

Beyond the gastrointestinal tract: oral and sex-specific skin microbiota are associated with hypertension in rats with genetic disparities.

Current knowledge of the link between microbiota and hypertension is limited to the gut. Besides the gut, oral cavity and skin are other locations where sodium chloride (NaCl) is in direct contact with microbiota. Although oral nitrate-reducing bacteria generate nitric oxide, which leads to vasodilation and lowering of blood pressure (BP), the skin excretes sodium via sweat glands and is an important site for sodium and BP homeostasis. However, knowledge on the contributions of oral and skin microbiota to BP regulation, is limited. Therefore, the current study was conducted to compare the tripartite relationship between site, sex, and genetic effects on the composition of oral, skin, and gut microbiota impacting hypertension. Microbiota were profiled from the oral cavity, skin, and feces of both male and female hypertensive Dahl salt-sensitive (S) and congenic rats with genomic substitutions on rat chromosomes (RNO) 1, 5, 9, and 10, demonstrating disparate BP effects. Sex-specific differences in ß-diversity were observed only in skin microbiota. The most abundant taxa of the oral and skin microbiota were Actinobacteria and Cyanobacteria, respectively. Oral Actinobacteria were inversely associated with BP. Although the abundance of oral Actinobacteria was upregulated by the BP locus on RNO10 in both sexes, depletion of skin Cyanobacteria decreased the protection from hypertension in the RNO5 female, but not male, congenic strain. In conclusion, to our knowledge this is the first study to identify specific microbiota in sites other than gut as contributors to BP regulation. Notably, both oral Actinobacteria and skin Cyanobacteria were beneficial for lowering BP.

Low-dose 1,3-butanediol reverses age-associated vascular dysfunction independent of ketone body ß-hydroxybutyrate.

With an aging global population, identifying novel therapeutics are necessary to increase longevity and decrease the deterioration of essential end organs such as the vasculature. Secondary alcohol, 1,3-butanediol (1,3-BD), is commonly administered to stimulate the biosynthesis of the most abundant ketone body ß-hydroxybutyrate (ßHB), in lieu of nutrient deprivation. However, suprapharmacological concentrations of 1,3-BD are necessary to significantly increase systemic ßHB, and 1,3-BD per se can cause vasodilation at nanomolar concentrations. Therefore, we hypothesized that 1,3-BD could be a novel antiaging therapeutic, independent of ßHB biosynthesis. To test this hypothesis, we administered a low-dose (5%) 1,3-BD to young and old Wistar-Kyoto (WKY) rats via drinking water for 4 wk and measured indices of vascular function and metabolism posttreatment. We observed that low-dose 1,3-BD was sufficient to reverse age-associated endothelial-dependent and -independent dysfunction, and this was not associated with increased ßHB bioavailability. Further analysis of the direct vasodilator mechanisms of 1,3-BD revealed that it is predominantly an endothelium-dependent vasodilator through activation of potassium channels and nitric oxide synthase. In summary, we report that 1,3-BD, at a concentration that does not stimulate ßHB biosynthesis, could be a nutraceutical that can reverse the age-associated decline in vascular function. These results emphasize that 1,3-BD has multiple, concentration-dependent mechanisms of action. Therefore, we suggest alternative approaches to study the physiological and cardiovascular effects of ßHB.NEW & NOTEWORTHY 1,3-Butanediol (1,3-BD) is often administered to stimulate the biosynthesis of the most abundant ketone body, ß-hydroxybutyrate (ßHB), and its purported salubrious effects. Here, we report that a low dose of 1,3-BD (5%) is sufficient to reverse age-associated vascular dysfunction, independent of ßHB. Therefore, low-dose 1,3-BD could be a novel therapeutic to increase blood flow and improve the quality of life in the elderly.

Reconstitution of the host holobiont in germ-free born male rats acutely increases bone growth and affects marrow cellular content.

Integration of microbiota in a host begins at birth and progresses during adolescence, forming a multidirectional system of physiological interactions. Here, we present an instantaneous effect of natural, bacterial gut colonization on the acceleration of longitudinal and radial bone growth in germ-free born, 7-wk-old male rats. Changes in bone mass and structure were analyzed after 10 days following the onset of colonization through cohousing with conventional rats and revealed unprecedented acceleration of bone accrual in cortical and trabecular compartments, increased bone tissue mineral density, improved proliferation and hypertrophy of growth plate chondrocytes, bone lengthening, and preferential deposition of periosteal bone in the tibia diaphysis. In addition, the number of small in size adipocytes increased, whereas the number of megakaryocytes decreased, in the bone marrow of conventionalized germ-free rats indicating that not only bone mass but also bone marrow environment is under control of gut microbiota signaling. The changes in bone status paralleled with a positive shift in microbiota composition toward short-chain fatty acids (SCFA)-producing microbes and a considerable increase in cecal SCFA concentrations, specifically butyrate. Furthermore, reconstitution of the host holobiont increased hepatic expression of IGF-1 and its circulating levels. Elevated serum levels of 25-hydroxy vitamin D and alkaline phosphatase pointed toward an active process of bone formation. The acute stimulatory effect on bone growth occurred independently of body mass increase. Overall, the presented model of conventionalized germ-free rats could be used to study microbiota-based therapeutics for combatting dysbiosis-related bone disorders.

Deep transcriptomic profiling of Dahl salt-sensitive rat kidneys with mutant form of Resp18.

Expression of Regulated endocrine specific protein 18 (Resp18) is localized in numerous tissues and cell types; however, its exact cellular function is unknown. We previously showed that targeted disruption of the Resp18 locus in the Dahl SS (SS) rat (Resp18mutant) results in higher blood pressure (BP), increased renal fibrosis, increased urinary protein excretion, and decreased mean survival time following a chronic (6 weeks) 2% high salt (HS) diet compared with the SS rat. Based on this prominent renal injury phenotype, we hypothesized that targeted disruption of Resp18 in the SS rat promotes an early onset hypertensive-signaling event through altered signatures of the renal transcriptome in response to HS. To test this hypothesis, both SS and Resp18mutant rats were exposed to a 7-day 2% HS diet and BP was recorded by radiotelemetry. After a 7-day exposure to the HS diet, systolic BP was significantly increased in the Resp18mutant rat compared with the SS rat throughout the circadian cycle. Therefore, we sought to investigate the renal transcriptomic response to HS in the Resp18mutant rat. Using RNA sequencing, Resp18mutant rats showed a differential expression of 25 renal genes, including upregulation of Ren. Upregulation of renal Ren and other differentially expressed genes were confirmed via qRT-PCR. Moreover, circulating renin activity was significantly higher in the Resp18mutant rat compared with the WT SS rat after 7 days on HS. Collectively, these observations demonstrate that disruption of the Resp18 gene in the SS rat is associated with an altered renal transcriptomics signature as an early response to salt load.

Physiologic, Metabolic, and Toxicologic Profile of 1,3-Butanediol.

Ketone bodies are essential energy substrates in the absence of exogenous nutrients, and more recently, they have been suggested to prevent disease and improve longevity. ß-hydroxybutyrate (ßHB) is the most abundant ketone body. The secondary alcohol, 1,3-butanediol (1,3-BD), is commonly administered to raise ßHB bioavailability in vivo and in the absence of nutrient deprivation. However, the concentration of 1,3-BD that yields a systemic concentration of ßHB similar to that observed after a 24-hour fast has yet to be determined. To evaluate this knowledge gap, we administered 5%, 10%, or 20% 1,3-BD via the drinking water to adult, male Wistar-Kyoto rats for four weeks. In addition to systemic and excreted ßHB concentration, physiologic, metabolic, and toxicologic parameters were measured. We report that only 20% 1,3-BD significantly elevates the systemic and urinary concentrations of ßHB. Rats treated with 20% 1,3-BD had a rapid and sustained reduction in body mass. All concentrations of 1,3-BD decreased food consumption, but only the 20% concentration decreased fluid consumption. Urine volume, red blood cell count, and hematocrit suggested dehydration in the 10% and 20% 1,3-BD-treated rats. Finally, 20% 1,3-BD-treated rats presented with indicators of metabolic acidosis and sinusoidal dilation, but no evidence of fatty liver or hepatotoxicity. In summary, we report that 20% 1,3-BD, but not 5% or 10%, produces a systemic concentration of ßHB similar to that observed after a 24-hour fast. However, this concentration is associated with deleterious side effects such as body mass loss, dehydration, metabolic acidosis, and sinusoidal dilation. SIGNIFICANCE STATEMENT: 1,3-Butanediol (1,3-BD) is often administered to stimulate the biosynthesis of the most abundant ketone body, ß-hydroxybutyrate (ßHB), and its purported salubrious effects. This article reports that suprapharmacological concentrations of 1,3-BD are necessary to yield a systemic concentration of ßHB similar to that observed after a 24-hour fast, and this is associated with undesirable side effects. On the other hand, low concentrations of 1,3-BD were better tolerated and may improve health independent of its conversion into ßHB.

Vertical selection for nuclear and mitochondrial genomes shapes gut microbiota and modifies risks for complex diseases.

Here we postulate that the heritability of complex disease traits previously ascribed solely to the inheritance of the nuclear and mitochondrial genomes is broadened to encompass a third component of the holobiome, the microbiome. To test this, we expanded on the selectively bred low capacity runner/high capacity runner (LCR/HCR) rat exercise model system into four distinct rat holobiont model frameworks including matched and mismatched host nuclear and mitochondrial genomes. Vertical selection of varying nuclear and mitochondrial genomes resulted in differential acquisition of the microbiome within each of these holobiont models. Polygenic disease risk of these novel models were assessed and subsequently correlated with patterns of acquisition and contributions of their microbiomes in controlled laboratory settings. Nuclear-mitochondrial-microbiotal interactions were not for exercise as a reporter of health, but significantly noted for increased adiposity, increased blood pressure, compromised cardiac function, and loss of long-term memory as reporters of disease susceptibility. These findings provide evidence for coselection of the microbiome with nuclear and mitochondrial genomes as an important feature impacting the heritability of complex diseases.

Positional cloning of quantitative trait nucleotides for blood pressure and cardiac QT-interval by targeted CRISPR/Cas9 editing of a novel long non-coding RNA.

Multiple GWAS studies have reported strong association of cardiac QT-interval to a region on HSA17. Interestingly, a rat locus homologous to this region is also linked to QT-intervals. The high resolution positional mapping study located the rat QT-interval locus to a <42.5kb region on RNO10. This region contained no variants in protein-coding sequences, but a prominent contiguous 19bp indel polymorphism was noted within a novel predicted long non-coding RNA (lncRNA), which we named as Rffl-lnc1. To assess the candidacy of this novel lncRNA on QT-interval, targeted CRISPR/Cas9 based genome-engineering approaches were applied on the rat strains used to map this locus. Targeted disruption of the rat Rffl-lnc1 locus caused aberrant, short QT-intervals and elevated blood pressure. Further, to specifically examine the significance of the 19bp polymorphism within the Rffl-lnc1 locus, a CRISPR/Cas9 based targeted knock-in rescue model was constructed by inserting the 19bp into the strain which contained the deletion polymorphism. The knock-in alleles successfully rescued the aberrant QT-interval and blood pressure phenotypes. Further studies revealed that the 19bp polymorphism was necessary and sufficient to recapitulate the phenotypic effect of the previously mapped <42.5kb rat locus. To our knowledge, this study is the first demonstration of a combination of both CRISPR/Cas9 based targeted disruption as well as CRISPR/Cas9 based targeted knock-in rescue approaches applied for a mammalian positional cloning study, which defines the quantitative trait nucleotides (QTNs) within a rat long non-coding RNA as being important for the pleiotropic regulation of both cardiac QT-intervals and blood pressure.

Targeted disruption of regulated endocrine-specific protein ( Resp18) in Dahl SS/Mcw rats aggravates salt-induced hypertension and renal injury.

Hypertension is a classic example of a complex polygenic trait, impacted by quantitative trait loci (QTL) containing candidate genes thought to be responsible for blood pressure (BP) control in mammals. One such mapped locus is on rat chromosome 9, wherein the proof for a positional candidate gene, regulated endocrine-specific protein-18 ( Resp18) is currently inadequate. To ascertain the status of Resp18 as a BP QTL, a custom targeted gene disruption model of Resp18 was developed on the Dahl salt-sensitive (SS) background. As a result of this zinc-finger nuclease (ZFN)-mediated disruption, a 7 bp deletion occurred within exon 3 of the Resp18 locus. Targeted disruption of Resp18 gene locus in SS rats decreases its gene expression in both heart and kidney tissues regardless of their dietary salt level. Under a high-salt dietary regimen, both systolic and diastolic BP of Resp18mutant rats were significantly increased compared with SS rats. Resp18mutant rats demonstrated increased renal damage, as evidenced by higher proteinuria and increased renal fibrosis compared with SS rats. Furthermore, under a high-salt diet regimen, the mean survival time of Resp18mutant rats was significantly reduced compared with SS rats. These findings serve as evidence in support of Resp18 as a gene associated with the development of hypertension and renal disease.

Microbiotal-Host Interactions and Hypertension.

Hypertension, or elevated blood pressure (BP), has been extensively researched over decades and clearly demonstrated to be caused due to a combination of host genetic and environmental factors. Although much research remains to be conducted to pin-point the precise genetic elements on the host genome that control BP, new lines of evidence are emerging to indicate that, besides the host genome, the genomes of all indigenous commensal micro-organisms, collectively referred to as the microbial metagenome or microbiome, are important, but largely understudied, determinants of BP. Unlike the rigid host genome, the microbiome or the "second genome" can be altered by diet or microbiotal transplantation in the host. This possibility is attractive from the perspective of exploiting the microbiotal composition for clinical management of inherited hypertension. Thus, focusing on the limited current literature supporting a role for the microbiome in BP regulation, this review highlights the need to further explore the role of the co-existence of host and the microbiota as an organized biological unit called the "holobiont" in the context of BP regulation.

Targeted disruption of Cd40 in a genetically hypertensive rat model attenuates renal fibrosis and proteinuria, independent of blood pressure.

High blood pressure is a common cause of chronic kidney disease. Because CD40, a member of the tumor necrosis factor receptor family, has been linked to the progression of kidney disease in ischemic nephropathy, we studied the role of Cd40 in the development of hypertensive renal disease. The Cd40 gene was mutated in the Dahl S genetically hypertensive rat with renal disease by targeted-gene disruption using zinc-finger nuclease technology. These rats were then given low (0.3%) and high (2%) salt diets and compared. The resultant Cd40 mutants had significantly reduced levels of both urinary protein excretion (41.8 ± 3.1 mg/24 h vs. 103.7 ± 4.3 mg/24 h) and plasma creatinine (0.36 ± 0.05 mg/dl vs. 1.15 ± 0.19 mg/dl), with significantly higher creatinine clearance compared with the control S rats (3.04 ± 0.48 ml/min vs. 0.93 ± 0.15 ml/min), indicating renoprotection was conferred by mutation of the Cd40 locus. Furthermore, the Cd40 mutants had a significant attenuation in renal fibrosis, which persisted on the high salt diet. However, there was no difference in systolic blood pressure between the control and Cd40 mutant rats. Thus, these data serve as the first evidence for a direct link between Cd40 and hypertensive nephropathy. Hence, renal fibrosis is one of the underlying mechanisms by which Cd40 plays a crucial role in the development of hypertensive renal disease.

High-resolution mapping of a novel rat blood pressure locus on chromosome 9 to a region containing the Spp2 gene and colocalization of a QTL for bone mass.

Through linkage analysis of the Dahl salt-sensitive (S) rat and the spontaneously hypertensive rat (SHR), a blood pressure (BP) quantitative trait locus (QTL) was previously located on rat chromosome 9. Subsequent substitution mapping studies of this QTL revealed multiple BP QTLs within the originally identified logarithm of odds plot by linkage analysis. The focus of this study was on a 14.39 Mb region, the distal portion of which remained unmapped in our previous studies. High-resolution substitution mapping for a BP QTL in the setting of a high-salt diet indicated that an SHR-derived congenic segment of 787.9 kb containing the gene secreted phosphoprotein-2 (Spp2) lowered BP and urinary protein excretion. A nonsynonymous G/T polymorphism in the Spp2 gene was detected between the S and S.SHR congenic rats. A survey of 45 strains showed that the T allele was rare, being detected only in some substrains of SHR and WKY. Protein modeling prediction through SWISSPROT indicated that the predicted protein product of this variant was significantly altered. Importantly, in addition to improved cardiovascular and renal function, high salt-fed congenic animals carrying the SHR T variant of Spp2 had significantly lower bone mass and altered bone microarchitecture. Total bone volume and volume of trabecular bone, cortical thickness, and degree of mineralization of cortical bone were all significantly reduced in congenic rats. Our study points to opposing effects of a congenic segment containing the prioritized candidate gene Spp2 on BP and bone mass.

Evidence for a link between gut microbiota and hypertension in the Dahl rat.

The gut microbiota plays a critical role in maintaining physiological homeostasis. This study was designed to evaluate whether gut microbial composition affects hypertension. 16S rRNA genes obtained from cecal samples of Dahl salt-sensitive (S) and Dahl salt-resistant (R) rats were sequenced. Bacteria of the phylum Bacteroidetes were higher in the S rats compared with the R rats. Furthermore, the family S24-7 of the phylum Bacteroidetes and the family Veillonellaceae of the phylum Firmicutes were higher in the S rats compared with the R rats. Analyses of the various phylogenetic groups of cecal microbiota revealed significant differences between S and R rats. Both strains were maintained on a high-salt diet, administered antibiotics for ablation of microbiota, transplanted with S or R rat cecal contents, and monitored for blood pressure (BP). Systolic BP of the R rats remained unaltered irrespective of S or R rat cecal transplantation. Surprisingly, compared with the S rats given S rat cecal content, systolic BP of the S rats given a single bolus of cecal content from R rats was consistently and significantly elevated during the rest of their life, and they had a shorter lifespan. A lower level of fecal bacteria of the family Veillonellaceae and increased plasma acetate and heptanoate were features associated with the increased BP observed in the S rats given R rat microbiota compared with the S rats given S rat microbiota. These data demonstrate a link between microbial content and BP regulation and, because the S and R rats differ in their genomic composition, provide the necessary basis to further examine the relationship between the host genome and microbiome in the context of BP regulation in the Dahl rats.

Conjugated bile acids are nutritionally re-programmable antihypertensive metabolites.

BACKGROUND: Hypertension is the largest risk factor affecting global mortality. Despite available medications, uncontrolled hypertension is on the rise, whereby there is an urgent need to develop novel and sustainable therapeutics. Because gut microbiota is now recognized as an important entity in blood pressure regulation, one such new avenue is to target the gut-liver axis wherein metabolites are transacted via host-microbiota interactions. Knowledge on which metabolites within the gut-liver axis regulate blood pressure is largely unknown. METHOD: To address this, we analyzed bile acid profiles of human, hypertensive and germ-free rat models and report that conjugated bile acids are inversely correlated with blood pressure in humans and rats. RESULTS: Notably intervening with taurine or tauro-cholic acid rescued bile acid conjugation and reduced blood pressure in hypertensive rats. Subsequently, untargeted metabolomics uncovered altered energy metabolism following conjugation of bile acids as a mechanism alleviating high blood pressure. CONCLUSION: Together this work reveals conjugated bile acids as nutritionally re-programmable anti-hypertensive metabolites.

Repurposing a Drug Targeting Inflammatory Bowel Disease for Lowering Hypertension.

Background The gut and gut microbiota, which were previously neglected in blood pressure regulation, are becoming increasingly recognized as factors contributing to hypertension. Diseases affecting the gut such as inflammatory bowel disease (IBD) present with aberrant energy metabolism of colonic epithelium and gut dysbiosis, both of which are also mechanisms contributing to hypertension. We reasoned that current measures to remedy deficits in colonic energy metabolism and dysbiosis in IBD could also ameliorate hypertension. Among them, 5-aminosalicylic acid (5-ASA; mesalamine) is a PPARγ (peroxisome proliferator-activated receptor gamma) agonist. It attenuates IBD by a dual mechanism of selectively enhancing colonic epithelial cell energy metabolism and ameliorating gut dysbiosis. Methods and Results A total of 2 groups of 11- to 12-week-old male, hypertensive, Dahl salt-sensitive (S) rats were gavaged with (n=10) or without (n=10) 5-aminosalicylic acid (150 mg/kg) for 4 weeks. Rats receiving 5-aminosalicylic acid treatment had a lower mean blood pressure than controls (145±3 mm Hg versus 153±4 mm Hg; P<0.0001). This reduction in blood pressure was accompanied by increased activity of PPARγ, increased expression of energy metabolism-related genes, and lowering of the Firmicutes/Bacteroidetes ratio in the colon, the reduction of which is a marker for the correction of gut dysbiosis. Furthermore, these data were consistent with the American Gut Project wherein the Firmicutes/Bacteroidetes ratio of non-IBD (n=611) patients was significantly lower than patients with IBD (n=631). Conclusions 5-Aminosalicylic acid could be repurposed for hypertension by specifically enhancing the gut energy metabolism and correction of microbiota dysbiosis.

Identification of a Gut Commensal That Compromises the Blood Pressure-Lowering Effect of Ester Angiotensin-Converting Enzyme Inhibitors.

BACKGROUND: Despite the availability of various classes of antihypertensive medications, a large proportion of hypertensive individuals remain resistant to treatments. The reason for what contributes to low efficacy of antihypertensive medications in these individuals is elusive. The knowledge that gut microbiota is involved in pathophysiology of hypertension and drug metabolism led us to hypothesize that gut microbiota catabolize antihypertensive medications and compromised their blood pressure (BP)-lowering effects. METHODS AND RESULTS: To test this hypothesis, we examined the BP responses to a representative ACE (angiotensin-converting enzyme) inhibitor quinapril in spontaneously hypertensive rats (SHR) with or without antibiotics. BP-lowering effect of quinapril was more pronounced in the SHR+antibiotics, indicating that gut microbiota of SHR lowered the antihypertensive effect of quinapril. Depletion of gut microbiota in the SHR+antibiotics was associated with decreased gut microbial catabolism of quinapril as well as significant reduction in the bacterial genus Coprococcus. C. comes, an anaerobic species of Coprococcus, harbored esterase activity and catabolized the ester quinapril in vitro. Co-administration of quinapril with C. comes reduced the antihypertensive effect of quinapril in the SHR. Importantly, C. comes selectively reduced the antihypertensive effects of ester ramipril but not nonester lisinopril. CONCLUSIONS: Our study revealed a previously unrecognized mechanism by which human commensal C. comes catabolizes ester ACE inhibitors in the gut and lowers its antihypertensive effect.

Selective IgA Deficiency in Spontaneously Hypertensive Rats With Gut Dysbiosis.

BACKGROUND: The spontaneously hypertensive rat (SHR) is extensively used to study hypertension. Gut microbiota dysbiosis is a notable feature in SHR for reasons unknown. Immunoglobulin A (IgA) is a major host factor required for gut microbiota homeostasis. We hypothesized that inadequate IgA contributes to gut microbiota dysbiosis in SHR. METHODS: IgA was measured in feces, cecum, serum, liver, gut-associated lymphoid tissue, and milk from SHR and Wistar Kyoto rats. IgA regulatory factors like IgM, IgG, and pIgR (polymeric immunoglobulin receptor) were analyzed. IgA and IgG antibodies and blood pressure (BP) were measured before and after administrating a bacterial antigen (ie, flagellin). RESULTS: Compared with Wistar Kyoto rats, SHR displayed remarkably near-deficient IgA levels accompanied by compensatory increases in serum IgM and IgG and gut-liver pIgR expression. Inadequate milk IgA in SHR emphasized this immune defect stemmed from the neonatal stage. Reduced IgA+ B cells in circulation and Peyer patches indicated a possible reason for the lower IgA in SHR. Noteworthy, a genetic insufficiency was unlikely because administering flagellin to SHR induced anti-flagellin IgA antibodies. This immune response surprisingly accelerated hypertension development in SHR, suggesting IgA quiescence may help maintain lower BP. CONCLUSIONS: This study is the first to reveal IgA deficiency in SHR as one host factor associated with gut microbiota dysbiosis and invigorates future research to determine the pathophysiological role of IgA in hypertension.

Ciliary polycystin-2 is a mechanosensitive calcium channel involved in nitric oxide signaling cascades.

Cardiovascular complications such as hypertension are a continuous concern in patients with autosomal dominant polycystic kidney disease (ADPKD). The PKD2 encoding for polycystin-2 is mutated in approximately 15% of ADPKD patients. Here, we show that polycystin-2 is localized to the cilia of mouse and human vascular endothelial cells. We demonstrate that the normal expression level and localization of polycystin-2 to cilia is required for the endothelial cilia to sense fluid shear stress through a complex biochemical cascade, involving calcium, calmodulin, Akt/PKB, and protein kinase C. In response to fluid shear stress, mouse endothelial cells with knockdown or knockout of Pkd2 lose the ability to generate nitric oxide (NO). Consistent with mouse data, endothelial cells generated from ADPKD patients do not show polycystin-2 in the cilia and are unable to sense fluid flow. In the isolated artery, we further show that ciliary polycystin-2 responds specifically to shear stress and not to mechanical stretch, a pressurized biomechanical force that involves purinergic receptor activation. We propose a new role for polycystin-2 in transmitting extracellular shear stress to intracellular NO biosynthesis. Thus, aberrant expression or localization of polycystin-2 to cilia could promote high blood pressure because of inability to synthesize NO in response to an increase in shear stress (blood flow).

Ketone body ß-hydroxybutyrate is an autophagy-dependent vasodilator.

Autophagy has long been associated with longevity, and it is well established that autophagy reverts and prevents vascular deterioration associated with aging and cardiovascular diseases. Currently, our understanding of how autophagy benefits the vasculature is centered on the premise that reduced autophagy leads to the accumulation of cellular debris, resulting in inflammation and oxidative stress, which are then reversed by reconstitution or upregulation of autophagic activity. Evolutionarily, autophagy also functions to mobilize endogenous nutrients in response to starvation. Therefore, we hypothesized that the biosynthesis of the most physiologically abundant ketone body, ß-hydroxybutyrate (ßHB), would be autophagy dependent and exert vasodilatory effects via its canonical receptor, Gpr109a. To the best of our knowledge, we have revealed for the first time that the biosynthesis of ßHB can be impaired by preventing autophagy. Subsequently, ßHB caused potent vasodilation via potassium channels but not Gpr109a. Finally, we observed that chronic consumption of a high-salt diet negatively regulates both ßHB biosynthesis and hepatic autophagy and that reconstitution of ßHB bioavailability prevents high-salt diet-induced endothelial dysfunction. In summary, this work offers an alternative mechanism to the antiinflammatory and antioxidative stress hypothesis of autophagy-dependent vasculoprotection. Furthermore, it reveals a direct mechanism by which ketogenic interventions (e.g., intermittent fasting) improve vascular health.