bluuebunny/crossref_metadata_2025_split huggingface dataset:
The reactive dopamine metabolite 3,4‐Dihydroxyphenylacetaldehyde (DOPAL) is a potent inducer of collagen secretion in human cardiac fibroblasts
Authors: Monroe T. Blake , Anderson Ethan J. 
Published: November 2023

Spontaneous generation of reactive sugar‐ and lipid‐derived aldehydes occurs in vivo as a byproduct of oxidative stress, and these aldehydes form protein adducts via electrophilic attack on cysteine and lysine residues. Catecholaldehydes are biogenic aldehydes formed as products of catecholamine metabolism by monoamine oxidase (MAO). MAO catalyzes the oxidative deamination of dopamine to generate 3,4‐dihydroxyphenylacetaldehyde (DOPAL), a reactive molecule that has been shown to be neurotoxic stemming from the reactivity of both catechol and aldehyde groups. Our lab has recently reported that diabetic patients have higher content and activity of MAO in atrial myocardium as compared with age‐matched nondiabetic patients, and that MAO activity in this tissue is associated with postoperative atrial fibrillation. Increased collagen deposition in the heart (i.e., fibrosis) is a well‐known risk factor for arrhythmias, and is also associated with diabetes. Here, we tested the hypothesis that DOPAL‐adducts induce production and secretion of collagen in human cardiac fibroblasts. Fibroblasts were isolated and cultured from right atrial appendage samples obtained from patients during during cardiac surgery. Cells were then treated with bovine serum albumin (BSA) conjugated with DOPAL, N(6)‐Carboxymethyllysine (CML, an advanced glycation end product), and 4‐hydroxynonenal (HNE, a n6 polyunsaturated fatty acid‐derived aldehyde). Collagen type I and III were measured in the media via immunoblot. Collagen content in the media increased with CML‐, HNE‐ and DOPAL‐BSA (P<0.05), as compared with BSA alone (control). Surprisingly, treatment with DOPAL increased type I and type III collagen secretion by 11‐fold and 32‐fold, respectively, compared to vehicle‐treated cells (P<0.05), an effect that was >5‐fold greater than with either CML or HNE. These findings suggest that reactive aldehydes, particularly catecholaldehydes, may significantly contribute to fibrosis in the heart via increased collagen secretion. Further, they provide evidence that compounds with aldehyde‐scavenging capacity may have therapeutic value in mitigating cardiac fibrosis.Support or Funding InformationR01HL122863‐01
ETB Receptor Dysfunction Leads to Exaggerated Apoptosis and Inflammasome Priming in Renal Outer Medulla during Type 1 Diabetes
Authors: De Miguel Carmen , Liu Xiaofen , Pollock David M. , Pollock Jennifer S. 
Published: November 2023

Endothelin‐1 (ET‐1), a potent vasoactive peptide that acts through two receptors (ETA and ETB), has been implicated as a causative factor in animal models of diabetes‐induced kidney injury and is upregulated in patients with diabetic nephropathy. Renal infiltration of immune cells and increased production of inflammatory markers play a critical role in the pathogenesis of diabetic kidney disease. ET‐1 exerts pro‐inflammatory actions in the diabetic kidney; however, the mechanism(s) by which ET‐1 mediates these effects are unclear. We hypothesized that the ETB receptor mediates renal protection during type 1 diabetes (T1D)‐induced apoptosis and inflammasome activation. Diabetes was induced in ETB deficient (ETB def) and transgenic (TG) control rats by i.v. injection of streptozotocin (STZ). After 10 wk of T1D, rats were terminated, kidneys were isolated and renal apoptosis was detected by TUNEL assay. Loss of ETB receptor function led to exaggerated apoptosis in kidneys of diabetic ETB def rats, especially in the outer medulla, an area of the kidney particularly rich in ETB receptors. Inflammasomes have been recently implicated in activation of apoptotic pathways. Analysis of the outer medulla by RT‐PCR array demonstrated that T1D induces a 4‐fold upregulation of NLRP5 (n=3/group; p<0.05) and a 3‐fold increase of IL‐1β expression (n=3/group; p<0.05) in ETB def rats when compared to diabetic TG controls, while NLRP3 was not different between genotypes. In addition, the negative regulator of the inflammasome PSTPIP1 was decreased in outer medulla of diabetic ETB def rats, further suggesting an activation of inflammasome signaling pathways in this part of the kidney. Recently, endoplasmic reticulum (ER) stress has been identified as an inducer of inflammasome activation; thus, we determined whether diabetic ETB def rats had an exaggerated ER stress response in the outer medulla. Outer medullary RNA expression of GRP78, ATF‐4, ATF‐6, s‐XBP‐1, CHOP and caspase‐12 was not different between genotypes in T1D. In conclusion, absence of functional ETB receptors in diabetes is associated to priming of inflammasomes and exaggerated apoptosis in the renal outer medulla. These results highlight the protective role of ETB receptors against the development of diabetic kidney damage, especially in the renal outer medulla.Support or Funding InformationFunded by NIH T32 DK007545 to CDM and P01 HL95499 to DMP and JSP.
Fructose Induces Metabolic Syndrome Associated with a Marked Inflammatory State
Authors: Santos Fernando , Moreira Edson Dias , De Angelis Katia , Irigoyen Maria Claudia 
Published: November 2023

Owing to controversial effects of fructose consumption, this protocol aims to identify possible changes on inflammatory profile due chronic fructose exposure. Male Wistar rats (weighing 150g at the beginning) were divided in two experimental groups,: Control group (CG), followed for 90 days and fructose group (FG), treated with 10% of fructose in drink water for 90 days Blood pressure was monitored in both groups by DSI® telemetry sensor.. Each animal was allocated individually under controlled environment (temperature: 20–22°C, 12h‐12h light‐dark cycle and soundproof room) with free access to food and water. It was observed that fructose was able to induce metabolic syndrome (MS) after 90 days, evidenced by increased abdominal adipose tissue (FG 12.3±0.6g vs. CG 6.2±0.8g), systolic arterial pressure (FG 146±5.2mmHg vs. CG 119±3.2mmHg), glycaemia (FG 98±1.3mg/dl vs. 81±2.0mg/dl) and triglycerides (FG 161±5.0mg/dl vs. CG 112±3.2mg/dl). The obesity and higher levels of triglycerides caused lipid deposition on kidney of FG. These factors were associated with increase in inflammatory markers evaluated by immunohistochemistry (IL‐6: 4.3±0.7% and NFκB: 0.6±0.1%) when compared to CG (IL‐6: 0.8±0.2% and NFκB: 0.2±0.03%). Furthermore, fructose promoted alteration in cardiac inflammatory profile. There were increase in NFκB (FG 19.1±4.3% vs. CG 6.3±1.2%) and TNFα (FG 25.1±1.4ρg/mg vs. CG 17.1±2.2ρg/mg). These results were associated with an increased action of sympathetic nervous system probably due to fructose consumption, since sympathetic modulation (FG 10.3±1.2mmHg2 vs. CG 4.2±0.3mmHg) evaluated by spectral analyses was increased in the fructose –treated animals. In addition an increase of 74% on expression of cardiac β1 adrenergic receptor was detected in FG. We concluded that chronic consumption of fructose was able to induce MS and to promote a proinflammatory profile dependent on increased lipid deposition and sympathetic modulation.Support or Funding InformationFAPESP 2013/23418‐0
A Genome‐Spanning Map of Chromatin Structure near the Renin Proximal Promoter
Authors: Stodola Timothy , Liu Pengyuan , Liu Yong , Greene Andrew , Liang Mingyu 
Published: November 2023

Emerging evidence suggests the expression of a gene can be regulated by interactions between the gene and genomic regions distant in sequence or on a different chromosome. Chromatin conformation capture with sequencing (3C‐seq) allows high resolution mapping of chromatin structure and enables the determination of all genomic regions that interact with a region of interest. The renin‐angiotensin system (RAS) is one of the major regulatory mechanisms essential for maintaining cardiovascular homeostasis through actions on the kidney and throughout the cardiovascular system. The goal of the current study was to expand our understanding of renin regulation by identifying potential trans‐regulatory elements of renin more than a few kilobases from the renin promoter, using 3C‐seq in isolated SS/JrHrdMcwi cardiac microvascular endothelial cells. Three biological replicates of ten million cells were fixed, digested with EcoRI, ligated, and then decrosslinked. Secondary digestion with NlaIII followed by a second ligation allowed a bait sequence, at the renin promoter, to surround an unknown captured sequence. A sequencing library was prepared using PCR with primers specific to the EcoRI and NlaIII cuts sites at the renin promoter to identify the captured sequences. Sequence reads of captured segments were mapped to rat genome (Rn 5) divided into fragments cut by EcoRI (fragends). The read counts were binarized and Z‐scores calculated for each fragend based on a window surrounding that fragend relative to background. Regions with Z‐score above a threshold Z‐score with false discovery rate ≤ 0.01 in three biological replicates were considered significant interactions. We found 18 genome‐spanning regions consistently interacting with the renin proximal promoter. Interactions were found on Chr13, the chromosome where Ren is located, as well as chromosomes 1, 2, 3, 7, 9, 12, 14 and 15. Quantitative PCR in 6 new 3C samples were used to validate identified interactions and negative control regions not expected to interact. These findings suggest renin expression might be regulated by chromatin conformation involving several chromatin regions located throughout the genome.Support or Funding InformationNIH P01HL116264‐04
Angiotensin converting enzyme enhances the oxidative response and bactericidal activity of neutrophils
Authors: Bernstein Kenneth E , Khan Zakir , Shen Xiao Z , Bernstein Ellen A , Giani Jorge F , Eriguchi Masahiro , Zhao Tuantuan V , Gonzalez‐Villalobos Romer A , Fuchs Sebastien , Liu George Y 
Published: November 2023

We studied the role of angiotensin converting enzyme (ACE) in neutrophils. ACE knockout (KO) mice or mice treated with an ACE inhibitor were more susceptible to bacterial infection by methicillin‐resistant S. aureus (MRSA). Specifically, KO mice have larger skin lesions (4‐fold) and more bacteria per lesion (3.3‐fold) compared to wild type (WT). WT mice transplanted with ACE KO bone marrow are also significantly more susceptible than WT mice transplanted with WT bone marrow. In contrast, mice over expressing ACE in neutrophils (NeuACE mice) have increased resistance to MRSA (smaller lesions, less bacteria), and better in vitro killing of MRSA, P. aeruginosa, and K. pneumoniae. ACE overexpression increased neutrophil production of reactive oxygen species (ROS) following MRSA challenge, an effect independent of the angiotensin II AT1 receptor. Specifically, as compared to WT mice, there was a marked increase of SOD inhibitable superoxide generation (greater than 2‐fold, p<0.0005) in NeuACE neutrophils following MRSA infection. These functional studies and the increased association of cytosolic subunits of NADPH oxidase (p47‐phox, p67‐phox) with membrane gp91‐phox indicate that ACE directly affects NADPH oxidase activity. Myloperoxidase activity in neutrophils was equivalent to that of WT mice. Increased ROS generation mediates the enhanced bacterial resistance of NeuACE mice as the enhanced resistance is lost with DPI inhibition or with ACE inhibitors. NeuACE granulocytes also have increased neutrophil extracellular trap formation (NET) and IL‐1β release in response to MRSA. In a mouse model of chemotherapy‐induced neutrophil depletion, transfusion of ACE overexpressing neutrophils was superior to WT neutrophils in treating MRSA infection. These data indicate a previously unknown function of ACE in neutrophil anti‐bacterial defenses, and suggest caution in the treatment of certain individuals with ACE inhibitors. ACE overexpression in neutrophils may be useful in boosting the immune response to antibiotic resistance bacterial infection.Support or Funding InformationNIH grants R01DK098382 (KEB), R21AI114965 (KEB), and R03DK101592 (RAGV).
Aldosterone‐regulated miRNAs and their target genes in mouse cortical collecting duct mpkCCD cells
Authors: Kwon Tae‐Hwan , Park Eui‐Jung , Jung Hyun Jun , Choi Hyo‐Jung 
Published: November 2023

BackgroundMature microRNA (miRNA) is a modulator in the post‐transcriptional regulation. The present study aimed 1) to profile the aldosterone‐regulated miRNAs in mouse cortical collecting duct cells (mpkCCDc14 cells); 2) to identify the main signaling pathways where putative target genes of identified miRNAs are enriched; and 3) to determine the roles of selected miRNAs and target genes in aldosterone‐mediated action.MethodsMicroarray chip assay (Affymetrix GeneChip miRNA 4.0 array) was done in the mpkCCDc14 cells in the absence or the presence of aldosterone treatment (10−6 M) for 3 d. The candidate miRNAs were selected by 1) more or less than 30% of significant fold‐changes (Protocol 1), or 2) differential expression analysis carried out using the R package ‘bridge’ (Bayesian Robust Inference for Differential Gene Expression. R package version 1.34.0., Protocol 2). Both the putative target genes of identified miRNAs and miRNAs‐enriched pathways were analyzed by using DIANA‐mirPath program.ResultsTwenty‐nine miRNAs were significantly up‐regulated (> 1.3‐fold) and 27 miRNAs were markedly down‐regulated (< 0.7‐fold) in mpkCCDc14 cells after aldosterone treatment (Protocol 1). Moreover, 5 up‐regulated and 7 down‐regulated miRNAs (> 1.2‐fold and < 0.8‐fold) were selected with high posterior probabilities (> 0.95, Protocol 2). Putative target genes of identified miRNAs were associated with 55 KEGG pathways (Protocol 1) and 29 KEGG pathways (Protocol 2) by DIANA‐mirPath program analysis. Among them, Wnt signaling pathway was particularly the most highly ranked. Seven up‐regulated and 8 down‐regulated mature miRNAs were enriched in the Wnt signaling pathways. Their quantitative changes were further examined by qPCR, revealing that miR‐130b‐3p, miR‐34c‐5p and miR‐146a‐5p were significantly changed. Six target genes (Rock1, skp1a, Tbl1xr1, Ppp2re, Wnt2b, Plcb1) of miR‐130b‐3p, 6 target genes (Lef1, Camk2b, Tbl1xr1, Prickle1, Ppp2r5a, Daam1) of miR‐34c‐5p and 4 target genes (Ppp3r2, Cxxc4, Smad4, Nfat5) of miR‐146a‐5p were identified.ConclusionAldosterone induces significant changes of miRNAs expression in mpkCCDc14 cells, and Wnt signaling is the most highly ranked pathway where identified miRNAs are enriched. The changes of target genes and corresponding protein expression may potentially be involved in the aldosterone‐mediated renal fibrosis, which need to be further examined.
Dasatinib, a non‐receptor tyrosine kinase inhibitor, induces aquaporin‐2 membrane accumulation in a phospho‐serine 256 independent manner.
Authors: Bouley Richard , Cheung Pui Wen , Terlouw Abby , Brown Dennis 
Published: November 2023

The cAMP‐sensitive water channel (AQP2) plays an important role in water regulation in the renal collecting ducts. Upon stimulation by vasopressin (VP), AQP2 is phosphorylated at the c‐terminus on serine 256 (pS256). This event leads to AQP2 accumulation in the plasma membrane. Other serine residues including S264 and S269 are also phosphorylated, but phosphorylation of S256 is thought to be essential to AQP2 membrane accumulation, and has to precede phosphorylation of other serine residues. Here we show that by inhibiting the proto‐oncogene tyrosine‐protein kinase Src using dasatinib, we can induce AQP2 membrane accumulation in a S256 independent but S269 dependent manner.Immunostaining using anti‐AQP2 antibody shows that dasatinib increases AQP2 membrane accumulation in LLC‐AQP2 cells. This result was confirmed with other Src inhibitors, bosutinib and KX2‐391. Furthermore, dasatinib also increases apical membrane AQP2 accumulation in collecting duct principal cells in our in situ kidney slice model. Dasatinib does not increase cAMP or PKA activity, and western blot analysis using specific phospho‐AQP2 antibodies showed that, in contrast to VP, dasatinib does not increase phosphorylation of S256 or de‐phosphorylate S261, but does increase the phosphorylation of S269. To confirm the importance of pS269 in dasatinib‐induced AQP2 membrane accumulation, we expressed an AQP2 S269A mutant in the LLC‐PK1 cells. We find that while VP induces AQP2 membrane accumulation in S269A mutant cells, dasatinib does not, suggesting the importance of pS269 for the effect of dasatinib.Src inhibition by dasatinib reduces endocytosis of rhodamine‐conjugated dextran, and rhodamine‐tagged transferrin accumulates at the cell surface and is not internalized. Immunocytochemistry shows an accumulation of clathrin at the plasma membrane in cells and tissues treated with dasatinib. Taken together, these results suggest that dasatinib reduces endocytosis by inhibiting clathrin‐mediated endocytosis.In conclusion, Src inhibition by dasatinib induces AQP2 membrane accumulation in a serine 256 independent, serine 269 dependent manner, and it inhibits clathrin‐mediated endocytosis. This study improves our understanding of the diversity of mechanisms that regulate water reabsorption in the kidneys, and allows us to differentially modulate the phosphorylation of essential AQP2 serine residues to understand their physiological functions.Support or Funding InformationThis work was supported by NIH grant DK096586 (D. Brown). P. W. Cheung was supported by T32 grant 5T32DK007540‐29
Linking epigenetic regulation of short chain fatty acid receptors, gut microbiome and hydrogen sulfide in the hypertensive kidney
Authors: Weber Gregory , Pushpakumar Sathnur , Sen Utpal 
Published: November 2023

Hypertension is one of the leading causes of chronic kidney disease. Recent studies have highlighted the importance of gut microbiota on physiological processes, such as metabolism and immunity, through production of short chain fatty acids (SCFA), which are linked with blood pressure (BP) regulation. Moreover, host epigenetic regulators, such as microRNAs, can directly affect the microbiome, and therefore blood pressure. However, the precise epigenetic regulation of BP is poorly understood. The gaseous molecule, hydrogen sulfide (H2S) is involved in regulating several physiological processes, however, its role and influence on the host gut microbiome and on renal function remains to be elucidated. In this study, we sought to determine the roles of host microRNAs in regulating short chain fatty acid receptors in the hypertensive kidney and whether H2S donor, GYY4137 (GYY), could reverse the effects to mitigate renal dysfunction. C57BL6/J wild‐type mice were treated without or with Ang‐II (1000ng/kg−1/min−1) and GYY (133μM/kg−1/d−1) for 4 weeks. Quantitative PCR, Western blot, immunofluorescence assays, metagenomic sequencing, and glomerular filtration rate were measured. Results indicated that increased expression of miR‐329 and miR‐132 in hypertensive mice were reduced by GYY treatment. SCFA receptors Gpr41 and Gpr43 are expressed in the kidney and are also predicted targets of miR‐329 and miR‐132, respectively. The mRNA and protein expression of the above receptors was reduced in Ang‐II treated mice while animals treated with Ang‐II and GYY showed normalized expression. miR‐129 was induced by GYY supplementation and the predicted SCFA receptor target Olfr78 was decreased in the normal kidney with the opposite effect in hypertensive kidney. Immunofluorescence analysis showed decreased signal of Gpr41 and Gpr43 in the blood vessel and tubules in Ang‐II mice and were normalized in mice treated with GYY. Metagenomic sequencing showed H2S supplementation affected the host gut microbiome composition and improved renal function. In conclusion, our data suggests that microRNAs can regulate SCFA receptors in the kidney and hydrogen sulfide supplementation has a positive effect on renal function through the gut microbiome and SCFA receptors.Support or Funding InformationDK‐104653 and HL‐104103 to U.S.,15SDG25840013 to S.P.
Exaggerated Salt‐Sensitive Hypertension in the ALMS1 (Alstrom syndrome 1) Knockout Rat
Authors: King‐Medina Keyona N. , Jaykumar Ankita , Ortiz Pablo 
Published: November 2023

We recently found that a protein named ALMS1 (Alstrom syndrome 1) is expressed in the kidney thick ascending limb (TAL) where it mediates endocytosis of the renal Na/K/2Cl cotransporter termed NKCC2. To study the role of ALMS1 in renal physiology we generated ALMS1 knockout (KO) rats in a Dahl salt‐sensitive genetic background via zinc‐finger nuclease targeting. We previously found that the amount of NKCC2 in the apical surface is higher in the TALs from ALMS1 KO rats compared to WT‐ Salt‐Sensitive (SS) rats. In order to determine the role of NKCC2 on blood pressure (BP) we utilized both noninvasive tail cuff measurements and invasive radio‐telemetry to study the effects of dietary sodium (.22% Na chow and 4% Na chow) on the systolic blood pressure (SBP) of the ALMS1 KO rats and WT‐SS rats. We hypothesized that deletion of the ALMS1 gene will increase SBP and enhance salt‐sensitivity of BP, in part due to higher NKCC2‐mediated Na reabsorption. First, we used tail cuff measurements to obtain the SBP in both groups of rats. We found that with normal Na intake (0.22% Na chow), the ALMS1 KO rats had a higher SBP than the WT‐SS rats (KO: 136±3 and WT‐SS: 125±3 mmHg, p=0.0461). To further explore the salt‐sensitive response of these rats we utilized radio‐telemetry monitoring for 4 weeks. We found the ALMS1 KO rats to have a higher baseline SBP on .22% Na chow compared to the WT‐SS rats (KO: 145±2 and WT‐SS: 134±1 mmHg, p=0.0009). After 2 weeks of high Na intake (4% Na chow) the SBP in ALMS1 KO rats increased to 181±1 mmHg, a 35±3 mmHg increase, whereas the SBP increased to 159±2 mmHg in WT‐SS rats, a 25±1 mmHg increase (p<0.01 vs ALMS1 KO). Thus, the SBP was higher in ALMS1 KO rats fed a high salt diet (p=0.0001). We then explored the involvement of NKCC2 in the hypertension observed in ALMS1 KO rats. A daily dose of the loop diuretic bumetanide (3mg/kg), an NKCC2 inhibitor, decreased the SBP in both groups (KO: −23±4 and WT: −30±6 mmHg, p=0.40). After 7 days of treatment with bumetanide, the SBP was normalized only in the WT‐SS rats while the SBP remained elevated in the ALMS1 KO rats (KO: 131±3 and WT: 115±4 mmHg, p<0.025). Therefore, we conclude that the ALMS1 KO rats have a higher salt‐sensitivity of BP than the WT‐ (Dahl) SS rats and this in part is mediated by enhanced renal sodium reabsorption in the TAL. Furthermore, our data suggest that additional mechanisms are likely involved in the hypertension observed in ALMS1 KO rats fed a high salt diet.Support or Funding InformationNIH F31 Diversity Grant Number: 5F31HL123285‐02