American journal of physiology. Renal physiology最新文献

Mechanosensitive neurons are important sensors of bladder distention, but their role in urologic disease remains unclear. Our current knowledge about how disease alters bladder sensation comes from studies that focus primarily on peptidergic nociceptors, leaving our understanding of neuropeptide-negative mechanoreceptors incomplete. In this study, we found that a substantial proportion of neurofilament heavy (NFH)-positive A fibers innervating the bladder were calcitonin gene-related peptide (CGRP)-negative, potentially representing uncharacterized mechanoreceptors. We then identified two genetic strategies that label mechanoreceptors in mouse skin and confirmed that they likewise label bladder afferents. Cre-mediated tdTomato reporter expression driven by tyrosine kinase B (TrkB), which labels A-delta mechanoreceptors in skin, successfully labeled bladder nerve terminals. The majority of TrkB bladder afferents were CGRP-negative and NFH-positive, with more characteristic staining patterns seen at the level of the cell body. The Ret proto-oncogene also produced robust labeling of bladder afferents, where colocalization with CGRP and NFH was consistent with multiple afferent subtypes. Because TrkB labeling was more specific for putative mechanoreceptors, we directly tested the role of TrkB neurons in bladder mechanosensation in vivo. Using an intersectional genetic strategy, we selectively ablated TrkB afferents and measured bladder responses to mechanical distention using anesthetized cystometry. Compared to controls, mice with ablated TrkB afferents required higher distention pressure to elicit voids. Interestingly, after ablation, distention also increased the frequency of non-voiding contractions, a poorly understood phenotype of several urologic diseases. These genetic strategies comprise critical new tools to advance the study of mechanoreceptors in bladder function and urologic disease pathophysiology.

SGLT2 inhibitors (SGLT2i) and GLP1 receptor (GLP1R) agonists have kidney protective effects. To better understand their molecular effects, RNA sequencing was performed in SGLT2-positive proximal tubule segments isolated by immunostaining-guided laser capture microdissection. Male adult DBA wildtype (WT) and littermate diabetic Akita mice ± Sglt1 knockout (Sglt1-KO) were given vehicle or SGLT2i dapagliflozin (dapa; 10mg/kg diet) for 2 weeks, and other Akita mice received GLP1R agonist semaglutide (sema; 3nmol/[kg body weight*day], s.c.). Dapa (254±11mg/dL) and Sglt1-KO (367±11mg/dL) but not sema (407±44mg/dL) significantly reduced hyperglycemia in Akita mice (480±33mg/dL). The 20,748 detected annotated protein-coding genes included robust enrichment of S1-segment marker genes. Akita showed 198 (~1%) differentially expressed genes vs. WT (DEGs; adjusted p<0.1) including downregulation of anionic transport, unsaturated fatty acid and carboxylic acid metabolism. Dapa changed only 2 genes in WT but restored 43% of DEGs in Akita, including upregulation of lipid metabolic pathway, carboxylic acid metabolism and organic anion transport. In Akita, sema restored ~10% of DEGs, and Sglt1-KO and dapa were synergistic (restored ~61%) possibly involving additive blood glucose effects (193±15mg/dl). Targeted analysis of transporters and channels (t-test p<0.05) revealed that ~10% of 526 detectable transporters and channels were downregulated by Akita, with ~60% restored by dapa. Dapa, dapa+Sglt1-KO and sema also altered Akita-insensitive genes. Among DEGs in Akita, ~30% were unresponsive to any treatment, indicating potential new targets. In conclusion, SGLT2i restored transcription for multiple metabolic pathways and transporters in SGLT2-positive proximal tubule segments in diabetic mice, with a smaller effect also observed for GLP1R agonism.

Renin is crucial for blood pressure regulation and electrolyte balance, and its expressing cells arise from Foxd1+ stromal progenitors. However, factors guiding these progenitors toward renin-secreting cell fate remain unclear. Tcf21, a basic helix-loop-helix (bHLH) transcription factor, is essential in kidney development. Utilizing Foxd1^Cre/+;Tcf21^f/f and Ren1^dCre/+;Tcf21^f/f mouse models, we investigated the role of Tcf21 in the differentiation of Foxd1+ progenitor cells into juxtaglomerular (JG) cells. Immunostaining and in-situ hybridization demonstrated fewer renin-positive areas and altered renal arterial morphology, including the afferent arteriole, in Foxd1^Cre/+;Tcf21^f/f kidneys compared to controls, indicating Tcf21's critical role in the emergence of renin-expressing cells. However, Tcf21 inactivation in renin-expressing cells (Ren1^dCre/+;Tcf21^f/f) did not recapitulate this phenotype, suggesting Tcf21 is dispensable once renin cell identity is established. Using an integrated analysis of single-cell RNA sequencing (scRNA-seq) and single-cell assay for transposase-accessible chromatin sequencing (scATAC-seq) on GFP+ cells (stromal lineage) from E12, E18, P5, and P30 Foxd1^Cre/+;Rosa26^mTmG control kidneys, we analyzed the temporal dynamics of Tcf21 expression in cells comprising the JG lineage (n=2,054). A pseudotime trajectory analysis revealed that Tcf21 expression is highest in metanephric mesenchyme and stromal cells at early developmental stages (E12), with a decline in expression as cells mature into renin-expressing JG cells. Motif enrichment analyses supported Tcf21's significant involvement in early kidney development. These findings underscore the critical role of Tcf21 in Foxd1+ cell differentiation into JG cells during early stages of kidney development, offering insights into the molecular mechanisms governing JG cell differentiation and highlight Tcf21's pivotal role in kidney development.

Mucin 1 (or MUC1) is a heterodimeric transmembrane glycoprotein expressed on the apical surface of polarized epithelial cells in several tissues including the kidney. Recent studies have revealed several novel roles for MUC1 in the kidney, potentially including bacterial infection, mineral balance, and genetic interstitial kidney disease, even though MUC1 levels are reduced not only in the kidney but in all tissues due to MUC1 mutations. A careful localization of MUC1 in discrete segments of the nephron is a first step in understanding the multiple functional roles of MUC1 in the kidney. Most published reports of MUC1 expression to date have been largely confined to cultured cells, tumor tissues, selective nephron segments of experimental rodents, and very few studies have been performed using human kidney tissues. Given the rising attention to the role of MUC1 in differing components of renal physiology, we carefully examined the kidney distribution of MUC1 in both human and mouse kidney sections using well-defined markers for different nephron segments or cell types. We further extended our investigation to include sections of early stages of mouse kidney development and upon injury in humans. We included staining for MUC1 in urothelial cells, the highly specialized epithelial cells lining the renal pelvis and bladder. These data implicate a role for MUC1 in antimicrobial defense. Our study provides the groundwork to test the physiological relevance of MUC1 in the urinary tract.

Here, we review key events in the accrual of knowledge about the cortical thick ascending limb (CTAL) of the kidney, starting with its initial characterization by Maurice Burg in 1973. Burg's work showed that the CTAL actively reabsorbs NaCl and that, because its water permeability is virtually zero, it can lower the luminal NaCl concentration to a 'static head' level well below blood levels. This process is central to the kidney's ability to excrete a dilute urine in states of high water intake. Following Burg's original observations, Greger and Schlatter, working in the 1980's, identified the membrane transport processes responsible for transepithelial NaCl transport in the CTAL. In the 1990's, several investigators identified the key transporter genes and proteins at a molecular level by cDNA cloning. The successful completion of human and mouse genome sequencing projects at the turn of the century, led to development of transcriptomic and proteomic methodologies that allowed identification of complete transcriptomes and proteomes of CTAL cells. Knowledge accrual was enhanced by the development of differential equation-based models of transport in the CTAL in the 2010's. Here we used a simplified mathematical model of NaCl ('salt'), urea and water transport in the CTAL to address three key questions about CTAL function: 1) What is the mechanism of Burg's 'static head' phenomenon? 2) How does the kidney compensate for the very short length of the CTALs of juxtamedullary nephrons? 3) Which of the three isoforms of the apical Na-K-2Cl cotransporter (NKCC2) dominates functionally in the CTAL?

Chronic kidney disease (CKD) is associated with an increased risk of cardiovascular disease (CVD). Despite the entry of sodium glucose cotransporter 2 (SGLT2) inhibitors, CKD persists as a medical challenge. Proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibition reduces low-density lipoprotein (LDL)-cholesterol, a major risk factor of CVD. Interestingly, studies indicate that PCSK9 inhibition decreases proteinuria in kidney disease, complementing the reduced CVD risk. This study aimed to validate obese ZSF1 rats as a model for the renoprotective effects of PCSK9 and SGLT2 inhibition using alirocumab and empagliflozin for 15 weeks. Obese rats revealed a significant reduction in measured glomerular filtration rate (mGFR) and increased urine albumin/creatinine ratio (UACR) during follow-up compared to lean controls. Alirocumab treatment resulted in a decline in mGFR and increased UACR compared to vehicle-treated obese rats. Immunohistochemistry showed increased fibrosis and inflammation in kidney tissue from obese rats treated with empagliflozin or alirocumab, whereas hepatic cholesterol and triglyceride levels were lowered compared to vehicle-treated obese rats. While alirocumab lowered circulating free cholesterol levels throughout the treatment period, certain cholesteryl esters were increased at the end of the study, resulting in no overall difference in total cholesterol levels in the alirocumab group. Correspondingly, only a trend toward increased hepatic LDL-receptor levels was observed. In conclusion, these findings suggest that alirocumab treatment aggravates kidney dysfunction in obese ZSF1 rats. Moreover, in contrast to the renoprotective properties of empagliflozin observed in CKD patients, empagliflozin did not ameliorate kidney disease progression in the obese ZSF1 rat.

The maintenance of fluid and electrolyte homeostasis by the kidney requires proper folding and trafficking of ion channels and transporters in kidney epithelia. Each of these processes requires a specific subset of a diverse class of proteins termed molecular chaperones. One such chaperone is GRP170, which is an Hsp70-like, endoplasmic reticulum (ER)-localized chaperone that plays roles in protein quality control and protein folding in the ER. We previously determined that loss of GRP170 in the mouse nephron leads to hypovolemia, electrolyte imbalance, and rapid weight loss. In addition, GRP170-deficient mice develop an AKI-like phenotype, typified by tubular injury, elevation of kidney injury markers, and induction of the unfolded protein response (UPR). By using an inducible GRP170 knockout cellular model, we confirmed that GRP170 depletion induces the UPR, triggers apoptosis, and disrupts protein homeostasis. Based on these data, we hypothesized that UPR induction underlies hyponatremia and volume depletion in these rodents, and that these and other phenotypes might be rectified by sodium supplementation. To test this hypothesis, control and GRP170 tubule-specific knockout mice were provided a diet containing 8% sodium chloride. We discovered that sodium supplementation improved electrolyte imbalance and kidney injury markers in a sex-specific manner but was unable to restore weight or tubule integrity. These results are consistent with UPR induction contributing to the kidney injury phenotype in the nephron-specific GR170 knockout model and indicate that GRP170 function in kidney epithelia is essential to both maintain electrolyte balance and ER homeostasis.

Epithelial sodium channel (ENaC) represents a major route of Na⁺ reabsorption in the aldosterone-sensitive distal nephron where the bulk of ENaC activity is considered to occur in the cortical collecting duct (CCD). Relatively, ENaC activity in the medulla, especially the inner medulla, is often neglected. (Pro)renin receptor (PRR), also termed as ATP6AP2, a newly characterized member of the renin-angiotensin system (RAS), has emerged as an important regulator of ENaC in the distal nephron. The ENaC regulatory action of PRR is largely mediated by the 28 kDa soluble PRR (sPRR). Although all three subunits of ENaC are under the control of aldosterone, sPRR only mediates the upregulation of α-ENaC but not the other two subunits. Furthermore, sPRR-dependent regulation of α-ENaC only occur in the renal inner medulla but not the cortex. sPRR also rapidly upregulates ENaC activity via Nox4-derived H₂O₂. Overall, sPRR has emerged as an important regulator of renal medullary Na⁺ reabsorption in the context of overactivation of the renin-angiotensin-aldosterone system (RAAS).

Natriuretic peptide receptor-A (NPR-A) is the principal receptor for the natriuretic peptides atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP). Targeted deletion of NPR-A in mouse glomerular podocytes significantly enhances renal injury in vivo in the DOCA-salt experimental model. It was therefore hypothesized that natriuretic peptides exert a direct protective effect on glomerular barrier integrity through activation of NPR-A and modulation of gene expression patterns in podocytes. Green fluorescence-positive podocytes from mice with a conditional deletion of Npr1 encoding NPR-A were isolated by fluorescence-activated cell sorting (FACS). Differentially expressed genes (DEGs) in podocytes were identified by RNA sequencing of podocytes from wild-type and NPR-A-deleted mice. Enrichment analysis was performed on the DEGs using Gene Ontology (GO) terms. Identified transcripts were validated by real-time PCR and ELISA of cultured isolated human and mouse glomeruli. In addition, the effect of natriuretic peptides on podocyte migration was investigated by measuring the outgrowth of podocytes from cultured glomeruli. A total of 158 DEGs were identified with 81 downregulated DEGs and 77 upregulated DEGs in Npr1-deficient podocytes. Among the downregulated genes were protein S and semaphorin 3G, which are known to have protective effects in podocytes. Protein S was also expressed in and secreted from isolated human glomeruli. GO enrichment analysis revealed that the upregulated DEGs in NPR-A deficient podocytes were associated with cell migration and motility. In line, BNP significantly decreased podocyte outgrowth from cultured glomeruli. In conclusion, endogenous levels of natriuretic peptides in mice support baseline protective pathways at glomerular podocytes such as protein S and suppress podocyte migration.NEW & NOTEWORTHY A combination of fluorescence-activated cell sorting and RNA sequencing identified 158 changed gene products in adult mouse kidneys with and without podocyte-specific deletion of the natriuretic peptide receptor A. Downregulated products included protein S and semaphorin 3G, both with proven renoprotective impact, whereas upregulated products were related to mobility of podocytes. Protein S was produced and released from human and murine isolated glomeruli, and atrial natriuretic peptide (ANP) led to decreased migration of podocytes.

Renal denervation (RDN) has been used for treating resistant hypertension. A few recent studies have shown vagal innervation of kidneys causing confusion. This study aimed to provide anatomical and functional evidence for renal autonomic innervation. Experiments were performed in male Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR). Pseudorabies virus (PRV) in the paraventricular nucleus and rostral ventrolateral medulla was prevented by bilateral RDN, but not subdiaphragmatic vagotomy. PRV did not appear in the dorsal motor nucleus of the vagus and nucleus tractus solitarii 72 h after renal injection of PRV. Adrenergic fibers were approximately seven times more than cholinergic fibers in the main renal artery (MRA) and its first (1RA) and second grade (2RA) branches. Adrenergic fibers in 1RA were more than those in MRA and 2RA. Tyrosine hydroxylase immunoreactivity in these arteries was higher in SHR than in WKY. Norepinephrine (NE) increased and α-receptor antagonist reduced vascular ring tension of renal arteries. The effect of NE was greater in 1RA and 2RA than in MRA, which was prevented by α-receptor antagonist. Acetylcholine (ACh) or blockage of β-receptors, M receptors, or N receptors had no significant effects on vascular ring tension and the effect of NE. Renal blood flow was reduced by electrical stimulation of renal nerves but not affected by stimulation of the subdiaphragmatic vagus. These results provide anatomical and functional evidence that kidneys are innervated and renal blood flow is regulated by renal sympathetic nerves rather than the vagus. Renal vasoconstriction is regulated by NE and adrenergic fibers rather than ACh or cholinergic fibers in WKY and SHR.NEW & NOTEWORTHY Kidneys are innervated by renal nerves rather than the vagus. Adrenergic fibers in renal arteries are about seven times more than cholinergic fibers. Renal vasoconstriction is regulated by norepinephrine and adrenergic fibers rather than acetylcholine or cholinergic fibers. Renal blood flow is regulated by renal sympathetic nerves and is not affected by the vagus. These findings provide anatomical and functional evidence for renal autonomic innervation in normotensive and hypertensive rats.