Sickle cell nephropathy (SCN) significantly shortens the life expectancy of patients with sickle cell disease (SCD). We previously reported that endothelin-1 (ET-1) and endothelin A receptor (ETA) are upregulated in SCN, and ETA antagonism mitigates SCN early in the disease progression in a humanized mouse model of SCD. We hypothesized that endothelium-derived ET-1 mediates the progression of SCN and T cell inflammation in the kidney of SCD mice. To test this hypothesis, we first used allogenic bone marrow transplantation from humanized sickle cell mice (HbSS) into endothelial-derived ET-1 knockout (VEET KO) mice, revealing that endothelial-derived ET-1 mitigates SCN and regulates the renal inflammatory response and T cell infiltration. Second, using young (4-5 mo old) and middle-aged (10-15 mo old) HbSS mice lacking endothelial-specific ET-1 (HbSS-VEET KO), we found a temporal maintenance of glomerular filtration rate, reduced infiltration of T cells to the kidney, and reduced progression of SCN. Furthermore, 2-wk ETA antagonism in middle-aged HbSS mice reduced infiltration of T cells. Finally, flow cytometric analyses revealed blunting of kidney T helper 17 (TH17) cells without a change in kidney T regulatory cells in HbSS-VEET KO mice, suggesting T cell subset-specific regulation by endothelial-derived ET-1 signaling. In vitro studies showed that ETA antagonism directly inhibits TH17 polarization and IL-17A production, suggesting that in established sickle cell disease, the ETA receptor-TH17 cell axis may play a key role in maintenance of fibrosis in SCN. Taken together, these data indicate that endothelial-derived ET-1 mediates the progression of SCN and strengthens the rationale for targeting ET-1 signaling as a new therapeutic approach.

Modern lifestyles increasingly promote unhealthy eating behaviors, contributing to the global rise in cardiovascular-kidney-metabolic diseases (CKMD). Circadian rhythms, anticipatory responses that coordinate biological processes with the 24-h rotation of the earth, are tightly linked to feeding behavior and nutrient metabolism, which are crucial for cardiovascular-kidney-metabolic homeostasis. At the molecular level, core circadian "clock genes" take part in a highly-conserved, regulatory feedback loop that optimizes metabolic processes for the anticipated time of food intake. Among the molecular components of the circadian clock, the transcription factor brain and muscle aryl hydrocarbon receptor nuclear translocator-like 1 (BMAL1) plays a central role in coordinating metabolic functions. Notably, BMAL1 and feeding behavior are at the intersection of circadian biology and metabolism, regulating each other. In this review, we summarize recent insights in metabolic physiology from global and tissue-specific Bmal1 knockout animal models, highlighting how BMAL1 influences CKMD consequences in response to dietary composition and feeding schedules and whether sex is a determinant in these outcomes. We also discuss the potential of molecular clock-targeted therapies for CKMD. Maintaining circadian integrity is central to mitigating CKMD; thus, modulating clock-regulated pathways may offer novel therapeutic strategies.

Time-restricted feeding (TRF), which confines food intake to specific time periods without altering nutrient content or reducing calories, has shown promise in improving cardiometabolic health. This study tested whether a 2-wk TRF intervention during the active (dark) period could reverse long-term effects of a high-fat diet (HFD) on liver mitochondrial function, steatosis, and metabolism in mice. Male C57BL/6J mice were fed either a normal-fat diet (NFD, 10% kcal fat) or an HFD (45% kcal fat) ad libitum for 18 wk, followed by 2 wk of active period TRF. Assessments included whole body metabolism, gene expression, histopathology, plasma lipid levels, and mitochondrial bioenergetic function. Chronic HFD feeding abolished the day-night difference in the respiratory exchange ratio (RER), altered 24-h expression rhythms of clock, lipid, and mitochondrial metabolism genes in the liver, and eliminated diurnal variation in liver mitochondrial bioenergetics. TRF partially restored RER rhythmicity without altering body composition or reducing caloric intake in HFD mice. TRF also reset 24-h expression rhythms in clock and several metabolic genes, normalized liver and plasma triglyceride oscillations, and reduced small droplet macrosteatosis in the livers of HFD mice. Importantly, TRF improved liver mitochondrial respiration and reduced circulating levels of mitochondrial transcription factor A, a mitochondrially-derived damage-associated molecule pattern, indicating reduced mitochondrial injury in HFD mice. These findings suggest that TRF can rapidly reverse HFD-induced disruptions in metabolic and mitochondrial function, offering a promising new nonpharmacologic strategy for improving liver health in obesity-related metabolic disease.

Obesity is a major risk factor for chronic kidney disease. Time-restricted feeding (TRF) shows promise to reduce kidney inflammation in chronic kidney disease. We hypothesized that TRF blunts kidney fibrosis in obese mice by mitigating T cell inflammation. We used a diet-induced obese mouse model fed a high fat diet (DIO, 45% fat) ad libitum for 18 weeks followed by 2 weeks of TRF or ad libitum high fat feeding. We found that TRF reversed kidney fibrosis as well as reduced kidney CD8+ T cells in DIO mice. Our study also revealed that DIO mice had increased kidney CD8+ T cell infiltration from the small intestine that was blunted with TRF. Furthermore, anti-CD8 intervention in DIO showed reduced kidney fibrosis and damage compared to anti-IgG treated DIO mice. Single cell RNA sequencing data revealed that DIO increased, while TRF reduced, the frequency of a specific cluster of CD8+ T cells that featured high expression of exhaustion/activation genes. Spatial analyses showed DIO mice had significant infiltration of PD-1+CD8+ T cells near CD31+ endothelial cells that was diminished by TRF. In conclusion, this study discovered that TRF reverses kidney fibrosis through reducing CD8+ T cell infiltration in obese mice.

Early life stress (ELS) is an independent risk factor for cardiovascular disease, including hypertension. The current study was designed to test the hypothesis that exposure to ELS disrupts autonomic function in adult mice at baseline and in response to an acute behavioral stressor. Using the mouse model of ELS, maternal separation with early weaning (MSEW), we determined the effects of ELS on heart rate (HR), blood pressure (BP), locomotor activity (LMA), HR variability (HRV), and spontaneous baroreceptor sensitivity (sBRS) in adult male mice. Under basal conditions, normally reared (NR) and MSEW mice had similar HR, BP, LMA, and sBRS when characterized at different phases of the circadian cycle. Spectral analysis of HRV indicated that MSEW mice had significantly lower total power and low-frequency (LF) power during the inactive or light period, as well as a loss of diurnal rhythm of LF power compared with NR mice. When subjected to an acute behavioral stress, cage switch stress (CSS), NR and MSEW mice showed similar changes in HR, BP, and LMA. Frequency domain analysis of HRV indicated that total, LF, and high frequency power during CSS and recovery from CSS were all significantly decreased in MSEW mice compared with NR mice. Taken together, these findings indicate that exposure to ELS in mice disrupts autonomic function basally and in response to an acute behavioral stress in a time-of-day dependent manner.NEW & NOTEWORTHY Early life stress is a known risk factor for cardiovascular disease in adulthood. The current study provides evidence for impaired regulation of the autonomic nervous system in adult mice after being exposed to maternal separation with early weaning, an established model of neglect.

Hypertension (HTN), the chronic elevation of blood pressure, accounts for more atherosclerotic cardiovascular disease deaths than any other modifiable risk factor.1 In the arteries, stable blood flow (s-flow) drives healthy, atheroprotective endothelial cell (EC) functions including nitric oxide (NO) production, barrier function, and anti-inflammatory programs via the action of flow-sensitive proteins. We showed that s-flow stimulates Heart-of-Glass 1 (HEG1) protein expression, localization to cell-cell junctions, and secretion from ECs.2 We found that conditional, endothelial cell-specific knockout of (Heg1 ECKO) exacerbates atherosclerosis2, however the mechanism was unknown. Here, we report a new role of HEG1 in controlling EC dysfunction, hypertension and atherosclerosis. We discover a novel mechanism: HEG1 regulates NO bioavailability via a flow-dependent HEG1-eNOS interaction (endothelial nitric oxide synthase, NOS3). Heg1 ECKO develops spontaneous hypertension and severe atherosclerosis, both of which are effectively treated by Angiotensin-Converting Enzyme inhibition (ACEi). UK BioBank and Swedish cohort studies reveal that plasma HEG1 levels are associated with hypertension and cardiovascular disease risk.3,4 Our findings suggest HEG1 may serve as a biomarker to advance personalized therapies for EC dysfunction, hypertension, and atherosclerosis.

Two-thirds of U.S. adults report experiencing early life stress (ELS), such as physical/sexual abuse, neglect, violence exposure, and parental separation or divorce. ELS is associated with an increased risk of cardiovascular disease (CVD) later in life. The CDC reported that 1.9 million cases of CVD could be prevented by preventing ELS, highlighting its profound impact on public health. Studies have also indicated that young adults exposed to ELS suffer from hemodynamic damage, such as increased resting blood pressure and arterial stiffness. In addition, ELS is associated with impaired emotion regulation, resulting in maladaptive cardiovascular responses to psychological stress that increase CVD risk. Parental emotion socialization refers to the process by which children acquire the ability to comprehend and regulate their emotions through interactions with their caregivers and parents. Higher levels of supportive parental emotion socialization in childhood are associated with enhanced emotion regulation and better stress management. Therefore, supportive parental emotion socialization may reduce the impact of ELS on CVD; however, this has not been tested. This study tested the hypothesis that supportive parental emotion socialization in childhood can buffer the effect of ELS on CVD risk factors in adulthood. Participants (Mage=19.58 years) enrolled in Wave 4 of the longitudinal Healthy Passages study reported their exposure to adverse and stressful events using the Adolescent Life Change Event Scale. They also reported how their parents responded to their sadness, fear, and anger during childhood using the supportive parenting subscale of the Emotions as a Child Scale. Approximately 10 years later, participants participated in Wave 5, and their cardiovascular function was measured. The casual and ambulatory blood pressures were collected along with two measures of vascular stiffness (pulse wave velocity (PWV) and augmentation index (AIx75)). Multiple linear regression analyses were conducted to examine the associations among ELS, emotion socialization, and cardiovascular outcomes while controlling for age, gender, body mass index, household income, and race. Interactions between the emotion socialization scales and ELS were used to test whether parental emotion socialization buffers the link between ELS and CVD risk. The sample included 164 young adults (Mage=29.62 years; 64.0% female; 29% White, 71% Black). Greater measures of early life stress were significantly associated with higher AIx75 (β=0.192; p=0.006). Additionally, greater supportive parental socialization was associated with lower PWV (β=-0.203; p=0.009). No significant association was found between ELS and supportive parental emotion socialization and either casual or ambulatory blood pressure. Supportive parental emotion socialization did not moderate the relationship between ELS and cardiovascular functioning (p>0.05). Higher ELS levels predicted higher arterial stiffness in young adulthood, whereas supportive emotional socialization was associated with lower arterial stiffness. These preliminary results indicate that supportive parental emotion socialization may have a beneficial impact on cardiovascular health among individuals who have not experienced ELS. However, supportive parental emotion socialization may not counteract the long-term harmful effects of ELS exposure on cardiovascular health. More research is needed to understand the interactions between ELS and protective mechanisms in cardiovascular disease risk across the lifespan. This work was supported by a NHLBI P01 HL158500 to J.S. Pollock, M.E. Siefert, and S. Mrug and a NHLBI K99 HL165091 to K.M. Kemp. This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

Diet-induced obesity (DIO) is a major risk factor for cardiovascular disease (CVD) and chronic kidney disease (CKD). Previous studies show that pan inhibition of histone deacetylase (HDAC) isoforms may alleviate obesity-related vascular, cardiac, or kidney dysfunction. HDAC1 is a key mediator of endothelial cell (EC) activation, and we previously showed that HDAC1 overexpression in EC reduces nitric oxide production and mediates endothelial dysfunction. However, it is unclear whether EC-specific HDAC1 drives CVD or CKD risk. We hypothesized that EC-specific HDAC1 mediates DIO-related aortic stiffness, cardiac dysfunction, and/or kidney damage. To test this hypothesis, we utilized male tamoxifen-inducible EC-specific HDAC1 (iEC-HDAC1) wildtype (WT) and knockout (KO) mice on a chronic high-fat diet (45% fat) for 20 weeks. On week 17, all mice were treated with tamoxifen for five consecutive days. Immunofluorescence confirmed the specificity of HDAC1 in EC of the iEC-HDAC1 KO and WT DIO mice. iEC-HDAC1 WT DIO male had similar body weight gain to iEC-HDAC1 KO DIO mice (WT (n=9): 12.20±1.74 g, KO (n=18): 11.06±1.37 g, unpaired T-Test, p=0.62). Pulse wave velocity (PWV, measure of aortic stiffness) and heart function were assessed with Vevo 3100 ultrasound technology. iEC-HDAC1 KO DIO mice had significantly reduced PWV compared to iEC-HDAC1 WT DIO mice (WT (n=8): 2.99±0.24 m/s, KO (n=7): 1.89±0.12 m/s; unpaired T-Test, p=0.002). Further, iEC-HDAC1 KO DIO mice had improved cardiac diastolic function compared to iEC-HDAC1 WT DIO mice (WT (n=5): 23.62±3.88 cm/s, KO (n=6): 14.73±3.18 cm/s, unpaired T-test, p=0.11). Ejection fraction and fractional shortening were not different (EF: unpaired T-test, n=5, p=0.92; FS: unpaired T-test, n=5, p=0.91). Kidney damage was assessed by histological staining with Masson’s trichrome blue to identify proximal tubular vacuoles and medullary interstitial fibrosis. An unbiased scorer was utilized to determine the vacuolar or fibrosis score, with 0 being no vacuoles or fibrosis and 2 being maximum vacuoles or fibrosis. We found that iEC-HDAC1 KO DIO mice had reduced proximal tubular vacuoles compared to iEC-HDAC1 WT DIO (WT (n=12): 0.98±0.08, KO (n=14): 0.55±0.12, unpaired T-test, p=0.0118). Also, iEC-HDAC1 KO DIO mice had reduced medullary interstitial fibrosis compared to iEC-HDAC1 WT DIO (WT (n=6): 0.67±0.14, KO (n=13): 0.37±0.05; unpaired T-test, p=0.0228). In summary, iEC-HDAC1 mediates aortic stiffness, cardiac diastolic dysfunction, and kidney damage in a chronic high-fat model of DIO. We conclude that HDAC1, specifically in EC, mediates obesity-related cardiovascular dysfunction and renal damage. NIH R25 DK 115353 - in partnership with: NIH U54 DK126087 UAB Childhood Cystic Kidney Disease Core Center (UAB-CCKDCC); NIH R01: 1R01DK134562-01A1 This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

The term early life stress encompasses traumatic events occurring before the age of 18 years, such as physical abuse, verbal abuse, household dysfunctions, sexual abuse, childhood neglect, child maltreatment, and adverse childhood experiences. Adverse psychological experiences in early life are linked to enduring effects on mental and physical health in adulthood. In this review, we first describe the effects and potential mechanisms of early life stress on the components of the vasculature. Next, we dive into the impact of early life stress on the vasculature across the lifespan through alterations of the epigenetic landscape. Finally, we consolidate the critical gaps in knowledge for focusing future research including the potential for resilience in combatting the impact of early life stress on vascular health.

Excess dietary salt and salt-sensitivity contribute to cardiovascular disease. Distinct T cell phenotypic responses to high salt and hypertension, as well as influences from environmental cues, are not well understood. The aryl hydrocarbon receptor (AhR) is activated by dietary ligands, promoting T cell and systemic homeostasis. We hypothesized that activating AhR supports CD4+ homeostatic functions, such as cytokine production and mobilization, in response to high salt intake while mitigating salt-sensitive hypertension. In the intestinal mucosa, we demonstrate that a high-salt diet (HSD) is a key driving factor, independent of hypertension, in diminishing interleukin 17A (IL-17A) production by CD4+ T (Th17) cells without disrupting circulating cytokines associated with Th17 function. Previous studies suggest that hypertensive patients and individuals on a HSD are deficient in AhR ligands or agonistic metabolites. We found that activating AhR augments Th17 cells during experimental salt-sensitive hypertension. Further, we demonstrate that activating AhR in vitro contributes to sustaining Th17 cells in the setting of excess salt. Using photoconvertible Kikume Green-Red mice, we also revealed that HSD drives CD4+ T cell mobilization. Next, we found that excess salt augments T cell mobilization markers, validating HSD-driven T cell migration. Also, we found that activating AhR mitigates HSD-induced T cell migration markers. Using telemetry in a model of experimental salt-sensitivity, we found that activating AhR prevents the development of salt-sensitive hypertension. Collectively, stimulating AhR through dietary ligands facilitates immunologic and systemic functions amid excess salt intake and restrains the development of salt-sensitive hypertension.

Early life stress (ELS) refers to adverse childhood experiences, including stressors such as parental separation, household dysfunction, and exposure to trauma, and is well-established to be associated with increased risk for cardiovascular disease (CVD) in adulthood. We previously found that aorta from mice exposed to ELS have significantly higher oxidative stress and inflammation. Utilizing immunohistochemical localization, we found that aortic endothelial cells show increased HDAC9 expression in adult MSEW mice compared to NR mice. HDAC9, a class II HDAC, is known to be associated with vascular dysfunction in humans and mice. We hypothesized that ELS exposure induces increased endothelial cell HDAC9 expression and that endothelial cell HDAC9 mediates increased pro-oxidant gene, NOX4, and pro-inflammatory gene VCAM-1, expression in the aorta of mice exposed to ELS. We utilized a mouse model of ELS, Maternal Weaning and Early Separation (MSEW), with normal-reared (NR) mice as controls with tamoxifen-inducible endothelial cell specific HDAC9 wild-type (iEC-HDAC9 WT) and knockout (iEC-HDAC9 KO) mice. Briefly, MSEW mice were separated from their mothers for increasing durations from days 2 to 16 post-birth and weaned on day 17. NR mice remained undisturbed until weaning on day 21. Aorta from adult iEC-HDAC9 WT MSEW mice have significantly greater NOX4 expression compared to iEC-HDAC9 KO MSEW (75.9 ± 18.4 copies/μL, 34.1 ± 9.0 copies/μL, n=9, p=0.0197). Additionally, aorta from iEC-HDAC9 KO MSEW mice had significantly lower VCAM-1 expression than aorta from iEC-HDAC9 WT MSEW mice critical for inflammatory cell migration (78.2 ± 19.2 copies/μL, 218.2 ± 69.8 copies/μL, n=9, p=0.030). These findings suggest that endothelial cell-specific HDAC9 mediates NOX4-driven oxidative stress and VCAM-1 contributing to endothelial dysfunction and inflammation with exposure to ELS. To follow up these findings, we further hypothesized that ELS exposure would promote macrophage priming with increased HDAC9 expression, thereby contributing to ELS-induced inflammation. Bone marrow-derived macrophages (BMDM) from MSEW and NR groups were cultured in media supplemented with macrophage colony stimulating factor for seven days followed by acute lipopolysaccharide stimulation. BMDM from MSEW mice showed significantly increased HDAC9 to the NR group (228.9 ± 143.3 copies/μL, 1.8 ± 0.8 copies/μL, p=0.014). These findings suggest BMDM from MSEW mice are primed for inflammation in contrast to BMDM from NR mice. We conclude that targeting HDAC9, an epigenetic regulator associated with ELS, may alleviate oxidative stress and decrease inflammation for individuals with ELS exposure. NIH P01HL158500 This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

Disrupted feeding and fasting cycles as well as chronic high-fat diet-induced (HFD-induced) obesity are associated with cardiovascular disease risk factors. We designed studies that determined whether 2 weeks of time-restricted feeding (TRF) intervention in mice fed a chronic HFD would reduce cardiovascular disease risk factors. Mice were fed a normal diet (ND;10%fat) ad libitum or HFD (45% fat) for 18 weeks ad libitum to establish diet-induced obesity. ND or HFD mice were continued on ad libitum diet or subjected to TRF (limiting food availability to 12 hours only during the dark phase) during the final 2 weeks of the feeding protocol. TRF improved whole-body metabolic diurnal rhythms without a change in body weight. HFD mice showed reduced blood pressure dipping compared with ND, which was restored by TRF. Further, TRF reduced aortic wall thickness, decreased aortic stiffness, as well as increased kidney tubular brush border integrity, decreased renal medullaryfibrosis, and reduced renal medullaryT cell inflammation in HFD mice. These findings indicate that TRF may be an effective intervention for improving vascular and kidney health in a model of established diet-induced obesity.

Angiotensin (Ang) II-induced hypertension is a common model of hypertension that entails infusing Ang II in rodents beginning during the inactive (lights-on) period. Using this model, investigators found that the immune system, particularly activated T cells, plays an instrumental role in the pathogenesis of hypertension. T cells exhibit time-of-day differences in activation capacity, cell fate, and tissue migration. However, it is unknown whether Ang II-induced hypertension is time-of-day dependent. Thus, we hypothesized that the blood pressure and T cell responses to Ang II infusion are dependent on the time-of-day. Male C57BL/6J mice were randomly implanted with telemetry (DSI, C10) at 9 weeks of age to monitor blood pressure (BP). After 2 weeks recovery, mice were then assigned to receive vehicle or continuous Ang II infusion (490 ng/kg*min, osmotic minipump, Alzet) initiated during the light, inactive period (zeitgeber time (ZT) 0-2) or during the dark, active period (ZT 12-14, minipump implanted under red light). In a separate set of mice, kidneys were dissected for IHC, peripheral blood T cells (PBTs) were harvested (Invitrogen 11443D) for bulk RNA sequencing (Azenta) after 7 days of vehicle or Ang II infusion, and PBTs were harvested on day 14 for assaying T cell activation capacity. Initiation of Ang II infusion at ZT 12-14 resulted in significantly lower systolic blood pressure response compared to initiation of Ang II infusion at ZT 0-2 (p<0.01, n=8/group, Day 7 24-hr BP = 117±11.65 or 135±11.17 mmHg [±SD], respectively). To address the potential time-of-surgery confounder, we implanted programmable peristaltic minipumps (iPrecio) at ZT 0-2 and delayed Ang II infusion by 30 or 42 hours while monitoring BP via telemetry. The interaction between timing of Ang II initiation and blood pressure was independent of time-of-surgery (P=0.0235, n=3/group). There was reduced CD3 + T cell accumulation (p<0.05, n=20,19) on day 7 in the kidneys of mice with Ang II infusion initiated at ZT 12-14 compared to ZT 0-2. BulkRNA sequencing of PBTs led to the identification of 1429 genes (adj.p<0.05) that were significantly different based on the time-of-day irrespective of Ang II infusion, while 233 genes (adj.p<0.05) were differentially expressed based on the time of initiation of Ang II infusion. Initiation of Ang II infusion at ZT 12-14 resulted in attenuated peripheral blood CD8 + IFNγ and IL-17a (P < 0.05, n=6/group) production capacity on day 14 compared to ZT 0-2 infusion. These findings demonstrate that BP and T cell responses to exogenous Ang II infusion are time-of-day dependent. Furthermore, these data indicate that mechanistic studies in rodent models need to consider the time of day when modeling hypertension. NIH KUH PRIME U2C DK133422 TL1 DK139566 (LNB), NIH R01 DK134562 (DMP&JSP). This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

Decades of research has indicated that T cells contribute to the development of hypertension and target organ damage. There is a critical role of spleen-derived T cells in angiotensin II-mediated hypertension and kidney damage in mice. Splenectomy abolished sex differences in the development of hypertension in spontaneously hypertensive rats, suggesting sex differences in the spleen-kidney immune axis. It is well-appreciated that immune cells migrate from different tissues to infiltrate organs. However, whether there are sex differences in T cell migration from the spleen to the kidney remained unexplored. Thus, we hypothesized that male mice would show higher immune cell migration from the spleen to the kidney at baseline and in response to angiotensin II infusion when compared to female mice. To quantify T cell migration in male and female mice, we utilized Kikume Green-Red (KikGR) transgenic mice that ubiquitously express the kikume green protein in all cells throughout the body, and this protein can be stably photoconverted to kikume red with exposure to 405 nm blue light. We specifically photoconverted mouse spleens with 10-minute exposure to blue light and infused angiotensin II via mini-pump implantation (490 ng/kg*min) in half of the photoconverted mice. After 3 days post-photoconversion, all mice were sacrificed, and both kidneys were removed for analysis via spectral flow cytometry to determine spleen-derived infiltrating immune cells (CD45+, KikRed+ cells) as well as spleen-derived T cells (CD45+, KikRed+, CD3+) in the kidney. There were no differences in the immune cell profiles found between left and right kidneys within each sex (p = 0.8 [male kidneys] and 0.8292 [female kidneys], n = 6-10 kidneys per group). At baseline, we found significantly increased spleen-derived immune cells in the kidneys of male compared to female mice (P = 0.0173, n = 3-5 per group). In addition, sex was a significant factor in spleen-derived T cell percentages within the kidney (Two-Way ANOVA, P = 0.029, n = 3-5 mice). In response to 3 days of angiotensin II infusion, spleen-derived immune cells were significantly higher in the kidneys of males compared to females (P = 0.0499, n = 3-5 mice). We conclude that there is reduced immune cell migration to the kidney from the spleen at baseline and in response to angiotensin II infusion in female mice, which may contribute to the lower blood pressure and kidney damage observed in female mice. NIH TL1 DK139566 to LNB, NIH R01 DK134562 to JSP. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

AIM:We aimed to test the hypothesis that a high-salt diet (HS) impairs NO signaling in kidney microvascular endothelial cells through a histone deacetylase 1 (HDAC1)-dependent mechanism. METHODS:Male Sprague Dawley rats were fed normal salt diet (NS; 0.49% NaCl) or HS (4% NaCl) for 2 weeks. NO signaling was assessed by measuring L-NAME induced vasoconstriction of the afferent arteriole using the blood perfused juxtamedullary nephron (JMN) preparation. In this preparation, kidneys were perfused with blood from a donor rat on a matching or different diet to that of the kidney donor. Kidney endothelial cells were isolated with magnetic activated cell sorting and HDAC1 activity was measured. RESULTS:We found HS-induced impaired NO signaling in the afferent arteriole. This was restored by inhibition of HDAC1 with MS-275. Consistent with these findings, HDAC1 activity was increased in kidney endothelial cells. We further found the loss of NO to be dependent upon the diet of the blood donor rather than the diet of the kidney donor and the plasma from HS-fed rats to be sufficient to induce impaired NO signaling. This indicates the presence of a humoral factor we termed plasma-derived endothelial dysfunction mediator (PDEM). Pretreatment with the antioxidants, PEG-SOD and PEG-catalase, as well as the NOS cofactor, tetrahydrobiopterin, restored NO signaling. CONCLUSION:We conclude that HS activates endothelial HDAC1 through PDEM leading to decreased NO signaling. This study provides novel insights into the molecular mechanisms by which a HS decreases renal microvascular endothelial NO signaling.

Diet-induced obesity (DIO) is an epidemic in the United States and globally. Obesity is well known to exacerbate and/or promote kidney disease. Inflammation is a major factor in DIO and kidney disease. Utilizing a 20-week DIO mouse model (45% fat) and a normal diet (ND; 10% fat), our lab previously showed greater kidney medullary interstitial fibrosis as well as increased kidney medullary CD3+ T cells in DIO mice when compared to ND mice. Specifically, we found increased kidney CD8+ T cells, but similar CD4+ T cells, in DIO mice compared to ND mice. We hypothesized that CD8+ T cells in DIO mice are a primary factor in promoting kidney interstitial fibrosis. DIO mice were treated with anti-CD8 antibody or control anti-IgG antibody and ND mice were treated with control anti-IgG antibody for the final two weeks of the DIO protocol (n=7-9). DIO/anti-IgG and DIO/anti-CD8 mice had significantly higher body weight than ND/anti-IgG mice as expected (one-way ANOVA; p<0.0001). Administration of anti-CD8 treatment in DIO mice had null effects on body weight (one-way ANOVA, p=0.509) and kidney weight (one-way ANOVA; p=0.067). Utilizing flow cytometry, we verified that anti-CD8 treatment significantly reduced CD8+ T cells compared to anti-IgG in the kidney (one-way ANOVA; p=0.0077; DIO/anti-IgG vs DIO/anti-CD8, p=0.0066) and in the blood (one-way ANOVA; p=0.0034; DIO/anti-IgG vs DIO/anti-CD8, p=0.0048). Using Picrosirius Red histological staining to assess kidney fibrosis, we found that anti-CD8 treatment significantly blunted kidney medullary fibrosis (one-way ANOVA, p=0.0177, ND/anti-IgG vs DIO/anti-IgG, p=0.0329; DIO/anti-IgG vs DIO/anti-CD8, p=0.0318), whereas glomerular fibrosis was not affected by anti-CD8 treatment in DIO mice (one-way ANOVA, p=0.31). Urine was collected in 12-hour increments and analyzed for markers of kidney damage. All groups of mice showed similar urinary KIM-1 excretion (one-way ANOVA; p=0.62) and protein excretion (one-way ANOVA; p=0.69). DIO mice with anti-CD8 or anti-IgG treatment had significantly increased urinary NGAL excretion compared to ND mice (one-way ANOVA; p=0.0203). In summary, anti-CD8 treatment reduced circulating and kidney CD8+ T cells, as well as kidney medullary interstitial fibrosis but not urinary NGAL excretion. These results indicate that CD8+ T cells mediate kidney medullary interstitial fibrosis in a 20 week DIO mouse model. Funding: R01 DK134562, R25 DK115353, F31HL167626. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Environmental factors play an important role in inflammatory bowel diseases (IBD; Crohn’s disease, [CD], ulcerative colitis [UC]). As part of the Crohn’s & Colitis Challenges 2024 agenda, the Environmental Triggers workgroup summarized the progress made in the field of environmental impact on IBD since the last Challenges cycle in this document. The workgroup identified 4 unmet gaps in this content area pertaining to 4 broad categories: (1) Epidemiology; (2) Exposomics and environmental measurement; (3) Biologic mechanisms; and (4) Interventions and Implementation. Within epidemiology, the biggest unmet gaps were in the study of environmental factors in understudied populations including racial and ethnic minority groups and in populations witnessing rapid rise in disease incidence globally. The workgroup also identified a lack of robust knowledge of how environmental factors may impact difference stages of the disease and for different disease-related end points. Leveraging existing cohorts and targeted new prospective studies were felt to be an important need for the field. The workgroup identified the limitations of traditional questionnaire-based assessment of environmental exposure and placed high priority on the identification of measurable biomarkers that can quantify cross-sectional and longitudinal environmental exposure. This would, in turn, allow for identifying the biologic mechanisms of influence of environmental factors on IBD and understand the heterogeneity in effect of such influences. Finally, the working group emphasized the importance of generating high-quality data on effective environmental modification on an individual and societal level, and the importance of scalable and sustainable methods to deliver such changes.