American Journal of Physiology-Regulatory, Integrative and Comparative Physiology

BackgroundHydrogen sulfide (H2S) is an important endothelial-derived vasodilator, but the signaling mechanism remains incompletely understood. We previously demonstrated that H2S-mediated vasodilation requires transient receptor potential vanilloid type 4 (TRPV4) channels. Because H2S has been reported to enhance heme oxygenase (HO) activity and HO-derived carbon monoxide (CO) regulates endothelial signaling, we hypothesized that H2S-mediated vasodilation requires HO-2-derived CO. MethodsPressure myography was performed in isolated rat mesenteric arteries to determine the contribution of HO, TRPV4, eBK, and SK/IK channels to H2S-mediated vasodilation. HO-2 sulfhydration was assessed using a maleimide assay, and spatial association among HO-2 and TRPV4 was examined using proximity ligation assays in human aortic endothelial cells. ResultsH2S Selicited concentration-dependent vasodilation that was abolished by HO inhibition. Repletion of CO restored H2S-mediated vasodilation in the presence of HO inhibition. CO-mediated vasodilation was abolished by TRPV4 and SK/IK inhibition but was unaffected by eBK inhibition. H2S increased HO-2 sulfhydration and enhanced HO activity. In endothelial cells, HO-2 and TRPV4 exhibited close spatial association. ConclusionsThese findings support a model in which H2S stimulates HO-2-derived CO production, leading to TRPV4-dependent endothelial signaling, SK/IK activation, and vasodilation. Together, the data support the existence of an endothelial HO-2/TRPV4/SK/IK signaling domain that contributes to H2S-mediated vascular reactivity.

Arginine vasopressin (AVP) increases excitability of myometrial smooth muscle cells (MSMCs) through Gq-coupled AVP receptors. Although excitability requires membrane depolarization, the mechanisms linking AVP receptor activation to membrane depolarization and Ca{superscript 2} signaling are incompletely understood. Here, we show that AVPR1 is the predominant AVP receptor in primary MSMCs. In Xenopus oocytes, AVP signals through AVPR1 to inhibit SLO2.1-mediated potassium currents, reducing current amplitude to approximately 60% of control currents. Consistent with suppression of a hyperpolarizing conductance, AVP depolarized a myometrial cell line (hTERT-HM) and increased intracellular Ca{superscript 2} signaling. Analysis of Ca{superscript 2} dynamics revealed that the initial Ca{superscript 2} peak was largely preserved under conditions limiting extracellular Ca{superscript 2} entry, consistent with intracellular store release. Conversely, the oscillatory phase depended on extracellular Ca{superscript 2} influx and was reduced by SLO2.1 knockdown. Together, these findings support a model in which AVP preferentially signals through AVPR1A to inhibit SLO2.1, depolarize myometrial cells, enhance VDCC-dependent Ca{superscript 2} entry, and promote excitability, enhancing conditions for uterine contraction.

The urinary bladder functions to store and release urine, yet how the sensation of bladder fullness is conveyed and perceived to the central nervous system is not understood. During bladder filling, the detrusor smooth muscle (DSM) generates phasic contractions, resulting in pressure fluctuations within the bladder. These transient pressure events drive bursts of afferent nerve activity, yet the underlying mechanism leading to rhythmic contractions remains unclear. Here, we examined the role of Gq protein-coupled receptor (GqPCR) activity on DSM excitability and contractility. Using ex vivo pressurized urinary bladder preparations and sharp microelectrode experiments on bladder strips from mice, we evaluated whole bladder transient pressure events, whole bladder DSM Ca2+ activity, and membrane potential in bladder strips. We found that global inhibition of urinary bladder GqPCR activity with YM-254890 abates phasic contractility and transient pressure events through a reduction in DSM Ca2+ activity and propagation of Ca2+ waves. Further, we found inhibition of GqPCR significantly hyperpolarizes DSM, reducing action potentials and decreasing excitability, and activation of protein kinase C restores membrane potential to baseline levels. These findings highlight that GqPCR activity mediates DSM excitability and contractility in such a way as to result in phasic detrusor contractions and transient pressure events.

Cerebrospinal fluid (CSF) provides a specialized extracellular environment for the central nervous system, which is predominantly produced by the choroid plexus, a highly vascularized epithelial structure whose ion transport processes are fundamental to CSF secretion, composition, and homeostasis. The mechanisms of Na+ entry into choroid plexus epithelial cells (CPECs) from the interstitial side remain disputed. The slc4a10 gene product encoding the Na+-dependent Cl-/HCO3- exchanger, Ncbe, was suggested as a key transport mechanism based on its impact on the cell's Na+-dependent regulation of intracellular pH and its basolateral membrane expression. The current study was undertaken to directly assess the contribution of Ncbe to the Na+ uptake into CPECs. Intracellular Na+ was recorded by fluorometry using the Na+ probe Sodium Binding Fluorescent Indicator in clusters of CPECs with access to both the luminal and basolateral membranes. Removal of extracellular Na+ reduced the apparent ex vivo intracellular [Na+] to ~5 mM from a baseline of ~43 mM in the absence of CO2/HCO3- and ~54 mM in the presence of CO2/HCO3-. Flame photometry estimated the intracellular [Na+] ex vivo to ~28 mM. The CO2/HCO3--dependent rate of [Na+] recovery amounted to ~53% of the total recovery rate upon re-addition of Na+. Experiments with access to only the luminal membrane show a [Na+] recovery of a similar rate as observed in the absence of CO2/HCO3- in the clusters. The CO2/HCO3--independent [Na+] recovery was inhibited to ~50% by the NKCC1 inhibitor bumetanide and to ~30% by the TRPv4 inhibitor RN1734. NHE contributed to a minor extent to the CO2/HCO3--independent transport. The HCO3- transport inhibitor DIDS, however, inhibited the total [Na+] recovery rate to ~50%, indicating a role for Ncbe rather than NBCn1 in the cellular [Na+] recovery. Indeed, docking of DIDS into Ncbe and NBCn1 indicated that both proteins can accommodate the binding of DIDS. However, the orientation of the DIDS poses in Ncbe suggests a binding mode more similar to that found in the Anion Exchangers (SLC4A1-3), which seems to accommodate the covalent-type docking more than NBCn1. The Ncbe inhibition by DIDS was supported by the rate of [Na+] recovery that was significantly higher in CPECs from Ncbe-wt than Ncbe-ko mice in the presence of CO2/HCO3-. As both NKCC1 and TRPv4 are localized to the luminal membrane, the findings collectively suggest that Ncbe is the most prominent mechanism for Na+ entry into CPECs expressed at the basolateral side. We suggest Ncbe as the rate-limiting mechanism in the vectorial Na+ transport driving CSF secretion.

Nitrogenous waste excretion is essential for all developmental stages of fish. Embryonic fish excrete urea, transitioning to cutaneous and later branchial ammonia excretion. In zebrafish, ammonia excretion involves rhesus glycoproteins Rhbg and Rhcgb in keratinocytes and ionocytes, but the developmental moment they appear in the gill remains unclear. Potential redundancy between Rhbg and Rhcgb in ammonia excretion is also not fully investigated, nor is the difference in response to low pH. We hypothesized that rhesus glycoproteins are partially redundant, and that they differ in their response to low pH as ammonia excretion enables ionocytes to exchange Na+ and H+ (Rh-NHE-metabolon). We predicted that a loss of rhbg or rhcgb induces compensatory responses. We characterized the transition from urea to branchial ammonia excretion from 0 to 8 days-post fertilization (dpf) and the response to pH 5.0 on the expression and localization of rhesus glycoproteins in control zebrafish and rhbg or rhcgb-crispants. Effects of high external ammonia (HEA, 500 M NH4Cl) and 10 mM HEPES-buffering were further characterized in rhcgb-crispants. Rhag and Rhbg appeared in the gill at 5 dpf, while Rhcgb appeared at 6 dpf. A loss of rhbg or rhcgb did not impact baseline N-excretion, illustrating that zebrafish can maintain ammonia excretion without the full complement of rhesus glycoproteins. We observed no compensatory increase in rhesus glycoproteins, but expression of the transporter hippocampus-abundant transcript 1b increased. HEA-exposed rhcgb-crispants switched to urea as primary nitrogen waste. Together, these findings underline the plasticity of the larval in dealing with nitrogenous waste.

Objective: Circadian rhythms are frequently disrupted in patients in the intensive care unit (ICU), potentially worsening clinical outcomes. Continuous enteral nutrition throughout the day and night is common in the ICU, but eliminates feeding-fasting cycles that serve as important timing cues for the circadian system. The objective of this study was to determine the effect of providing enteral nutrition in a cyclic daytime pattern, compared with continuous administration, on circadian rhythmicity in critically ill patients in the ICU. Design: Single-center randomized controlled trial Setting: Mixed medical-surgical tertiary intensive care unit in the Netherlands Patients: Adult ICU patients ([≥]18 yr) receiving enteral nutrition. Intervention: Patients were randomized to receive either continuous, or cyclic daytime enteral feeding (08:00-20:00), initiated from the start of nutritional support. Measurements and Main Results: Sixty-two ICU patients were enrolled, of whom 51 were included in the per-protocol analysis. While the amplitude of the 24-hour rhythm in core body temperature did not differ significantly between the cyclic daytime and continuous feeding groups (0.17 [interquartile range: 0.09-0.24] vs. 0.20 [0.13-0.30], p=0.182), the 24-hour rhythm in heart rate was enhanced in patients receiving cyclic daytime feeding, as reflected by significantly higher amplitudes and more synchronized peak times. No significant differences in 24-hour rhythmicity were observed between groups for the other vital signs or melatonin. Conclusions: Our findings suggest that cyclic daytime feeding may strengthen circadian rhythms in critically ill patients. Further studies are warranted to evaluate its impact on clinical outcomes.

Globally, hypercholesterolemia affects over 20% of the population; and while many studies have examined its impact on cardiovascular health, little is known about its effects on the lymphatic system. In mice, hypercholesterolemia has been linked to multiple aspects of lymphatic dysfunction; and a recent study demonstrated that cholesterol depletion by cyclodextrins promoted lymphatic vessel regeneration and restored lymphatic drainage in mouse models of lymphedema. Collecting lymphatic vessels rely on the spontaneous and highly entrained contractions of lymphatic muscle cells (LMCs) and competent unidirectional on-way valves to propel lymph forward. Critical to lymphatic pacemaking and contractility is the proper functioning of ion channels, which are known to be modulated by the cholesterol content in the plasma membrane. Therefore, we sought to understand the role cholesterol plays in regulating lymphatic contractility. The effects of cholesterol depletion by the cyclodextrins M{beta}CD and HP{beta}CD were assessed in cannulated and pressurized inguinal-axillary collecting lymphatic vessels (CLVs) from C57BL6/J (WT) mice. Noteworthy, studies have shown that HP{beta}CD is safe for human use, and in fact, it is commonly used as a drug excipient. Acute treatment with both cyclodextrins significantly increased the pumping capacity of CLVs, as demonstrated by the increased contraction amplitudes by [~]50{+/-}12% and calculated fluid volume displacement by each contraction by [~]35{+/-}11%. Calcium imaging demonstrated that HP{beta}CD increased the amplitude and duration of the large Cav1.2-mediated calcium events (termed calcium flashes. In contrast, cholesterol supplementation by incubation with BODIPY-cholesterol, which presumably incorporates cholesterol into the cell membrane, significantly impaired the contractile activity of CLVs compared to controls by decreasing contraction amplitude (control: 42{+/-}2 {micro}m versus BODIPY-cholesterol: 20{+/-}7{micro}m) and calculated fluid volume displacement (control: 9.2{+/-}3.9nL versus BODIPY cholesterol: 3.3{+/-}1.2nL) which were significantly restored with subsequent cholesterol depletion using HP{beta}CD (amplitude: 36{+/-}11{micro}m, volume displacement: 5.5{+/-}2.4nL). Similarly, treatment with HP{beta}CD significantly improved the contractile capacity of dysfunctional CLVs isolated from hypercholesterolemic ApoEKO mice. In conclusion, changes to cell membrane cholesterol content acutely and significantly altered CLV contractility with depletion improving contractility associated with recruitment of voltage-gated Cav1.2 channels in lymphatic muscle cells (LMCs). Future studies from our lab will determine whether pharmacological depletion of membrane cholesterol can be therapeutic strategy to improve and/or restore lymphatic contractile function in secondary lymphedema, including obesity/hypercholesterolemia-induced and cancer-related lymphedemas.

BackgroundHypertension, the persistent elevation of blood pressure (BP), is characterized by chronic low-grade inflammation and systemic cytokine release. Circulating cytokines contribute to the development of hypertension and end-organ damage. However, the specific immune profile associated with the progression of hypertension remains unclear. We hypothesize that a plasma cytokine signature reflects early BP changes in older adults. MethodsSeventy participants aged 57-81 years were categorized as normotensive (n = 17), elevated BP (n = 10), or hypertensive (n = 43) based on 24-hour ambulatory BP monitoring and antihypertensive treatment status. Plasma IL-1{beta}, IL-6, IL-10, IL-17A, IL-21, IL-22, IL-23, and TNF- were quantified using immunoassays. Partial Pearson correlations adjusted for demographic and biochemical covariates were used to assess associations between cytokines, BP, and cytokine-cytokine networks. ResultsIn untreated hypertensive individuals, plasma IL-23 was positively correlated with 24-hour diastolic BP. Antihypertensive treatment was associated with reduced IL-17A concentrations, which are negatively associated with 24-hour systolic BP. In the elevated BP group, IL-21 concentrations were higher than in normotensive individuals. To further characterize the cytokine signature, cytokine-cytokine correlations were examined. IL-23 and IL-17A were positively correlated with most interleukins, whereas TNF- showed few associations. IL-1{beta} exhibited strong correlations with both IL-23 and IL-17A, particularly in untreated participants. ConclusionIL-23 and IL-17A are associated with BP status and are broadly interconnected with other inflammatory cytokines, highlighting the potential importance of the IL-23/IL-17A axis in the hypertension of development. Early alterations in IL-21 in elevated BP may reflect immune changes that precede the onset of hypertension.

Tear fluid is amenable to non-invasive and repeated collection, making it a practical specimen for evaluating exercise-related physiological responses. However, the immediate proteome-wide alterations in tear fluid following acute exercise have not been characterised. In this study, we performed quantitative proteomic profiling of paired tear samples from healthy female participants before and immediately after a single exercise session using data-independent acquisition liquid chromatography-tandem mass spectrometry (DIA-LC-MS/MS). Among the 3,173 identified proteins, 744 were significantly altered post-exercise, of which 484 were up-regulated and 260 down-regulated. Functional enrichment analysis revealed that up-regulated proteins were predominantly associated with translation and ribosome biogenesis, whereas down-regulated proteins were involved in glycan metabolism, lysosomal processing, and extracellular matrix organisation. Collectively, these findings indicate that acute exercise elicits a rapid and coordinated reconfiguration of the tear proteome. This investigation provides a molecular basis for understanding exercise-mediated modulation of tear composition and ocular surface homeostasis.

Environmental temperature strongly influences muscle contractile mechanics and locomotor performance in ectotherms, yet animals routinely develop across a range of temperatures while maintaining effective movement. We tested the hypothesis that developmental temperature induces compensatory changes in the intrinsic mechanical properties of the muscles that power the fast-start escape response in larval zebrafish (Danio rerio). Larvae were reared at 25{degrees}C, 28{degrees}C, or 32{degrees}C, and contractile properties of intact tail myotomal muscles were measured across experimental temperatures. Acute changes in experimental temperature strongly affected twitch kinetics, particularly relaxation rate (Q10 = 2.1), resulting in substantial changes in twitch duration. In contrast, rearing temperature produced adaptive changes that opposed these acute thermal effects. At a common experimental temperature, muscles from cold-reared larvae exhibited faster intrinsic relaxation and greater force production during shortening at a physiologically relevant velocity, whereas warm-reared larvae showed slower relaxation and reduced shortening force. As a result, twitch kinetics were largely normalized when measurements were made at each group's rearing temperature, reducing the apparent thermal sensitivity of relaxation rate (Q10 = 1.1). To identify molecular correlates of these functional adaptations, we performed label-free quantitative LCMS proteomic analysis. Cold rearing increased the abundance of Sarco/Endoplasmic Reticulum Calcium-ATPase (SERCA) proteins, driven primarily by elevated atp2a1 expression, while warm rearing reduced the abundance of the major parvalbumin isoforms pvalb1 and pvalb2. These changes implicate remodeling of intracellular calcium handling as a mechanism underlying thermal compensation of muscle function. Together, our results demonstrate that developmental temperature modifies the intrinsic mechanical properties of larval zebrafish muscle in ways that counteract the direct effects of environmental temperature, thereby preserving the timing and power-generating capacity required for fast-start escape performance.

Background: The placenta has a unique transcriptomic profile, including microRNAs that are secreted into maternal circulation throughout pregnancy. MicroRNAs are small, non-coding RNA that post-transcriptionally regulate gene expression. Spontaneous preterm birth (sPTB) is associated with substantial differences in both placental pathophysiology and placental gene expression compared to term birth. We aimed to generate microRNA signatures of sPTB and map them to target genes using a microRNA-mRNA network. Methods: This study was conducted within the Conditions Affecting Neurocognitive Development and Learning in Early childhood (CANDLE) study. Placental samples were collected at delivery, and RNA was isolated for mRNA and microRNA sequencing. To investigate sPTB, this study excluded placental samples of participants with iatrogenic indications for PTB or induced labor. We examined differences in microRNA expression in participants who delivered before 37 weeks (N=35) compared to term participants (N=404) in a series of covariate-adjusted linear regression models. We used paired placental microRNA and mRNA expression data from this cohort to validate associations between computationally predicted microRNA-mRNA pairs and establish a microRNA-mRNA network. Results: Expression of 7 microRNAs were increased in sPTB (FDR<0.05) and were inversely correlated with sPTB-associated genes involved in immune signaling. Expression of 12 microRNAs were decreased in sPTB, including 4 members of the maternally expressed chromosome 14 microRNA cluster (miR-376a-3p, miR-376c-3p, miR-377-3p, and miR-381-3p). These microRNAs were predicted to negatively regulate oxidative phosphorylation genes that were increased in sPTB. The associations between miR-376c-3p and miR-377-3p and oxidative phosphorylation were confirmed in microRNA knockdown experiments. Conclusions: This study highlights potential biological mechanisms by which placental microRNA dysfunction might contribute to sPTB and highlights putative sPTB biomarkers that may be detectable in maternal circulation.

Gut-brain communication has emerged as a rapidly expanding field of research, with recent electrophysiological studies revealing rhythmic gut-brain coupling between gastric activity and brain oscillations in humans. Gut motility is a key determinant of gastrointestinal function, but it remains unclear whether individual differences in gut motility reflected by weekly bowel movements (e.g., defecation frequency) are associated with differences in gut-brain coupling. Here, we address this question by examining women with self-reported daily bowel movements (N = 38) and women with less frequent bowel movements (N = 38). We recorded simultaneous electroencephalography (EEG) and electrogastrography (EGG) at fasting state, performed cognitive assessments, and analysed faecal short-chain fatty acids (SCFAs) as markers of colonic fermentation. In a subset of participants, EEG-EGG coupling was assessed twice over an interval of at least eight weeks to assess test-retest reliability. In this group, EEG-EGG coupling showed moderate test-retest reliability (Intraclass Correlation Coefficient (ICC) = 0.50). When comparing the two groups of women, the phase-amplitude coupling (PAC) analysis between EEG and EGG signals revealed a significantly stronger gut-brain coupling in women with daily bowel movements compared to women with less frequent bowel movements (p = 0.03). We additionally found that women with daily bowel movements made less errors in the cognitive tasks and had higher levels of faecal SCFAs. A path analysis suggested that bowel movements significantly affect gut-brain phase-amplitude coupling through faecal SCFAs. However, neither faecal SCFAs nor phase-amplitude coupling significantly predicted cognitive performance, suggesting the existence of alternative pathways for the association between bowel movements and cognitive performance. Together, our findings suggest that the strength of gut-brain coupling is associated with bowel movements and cognitive performance, making EEG-EGG coupling a promising marker of human gut-brain interactions.

Elephant seals are remarkable breath-hold divers, capable of remaining submerged for up to two hours during diving bouts. These dives entail repeated, extreme hypoxia/reoxygenation events that would induce severe lipid peroxidation and tissue dysfunction in most mammals. Here, we show that primary vascular endothelial cells derived from elephant seals possess an intrinsic resistance to lipid peroxidation. Comparative transcriptomic and lipidomic profiling across seal, human, and sheep cells identified ferroptosis - an iron-dependent, lipid peroxidation-driven cell death pathway - as uniquely regulated in seal cells following hydroperoxide exposure. Mechanistically, seal cells exhibit robust baseline expression of acyl-CoA synthetase long-chain family member 3 (ACSL3), alongside rapid, seal-specific induction of the sole mammalian iron exporter, ferroportin (SLC40A1). Functional validation using genetic and pharmacological approaches revealed that seal cells are naturally enriched in monounsaturated fatty acids and triglycerides and utilize lipid droplet biogenesis and active iron export as dual protective axes to evade lipid peroxidation. Together, these findings show that elephant seal cells employ a coordinated cytoprotective network of lipid remodeling and iron handling to withstand the severe challenges of deep diving. SIGNIFICANCE STATEMENTDeep-diving marine mammals repeatedly experience extreme hypoxia-reoxygenation events that would induce severe oxidative damage in most terrestrial mammals. However, vascular cells derived from seals naturally resist lipid peroxidation, a major driver of ischemia-reperfusion injury. Here, we show that elephant seal endothelial cells evade lipid peroxidation through two complementary mechanisms: lipid droplets that sequester peroxidation-prone phospholipids, and rapid iron export that limits lipid peroxide formation. These findings reveal naturally evolved cellular strategies that protect against vascular oxidative stress, offering new insights into physiological resilience against ischemia-reperfusion injury.

Modulation of renal nerves to improve blood pressure (BP) control has become a topic of intense investigation over the last 10-15 years. Given that renal innervation is composed of mixed nerve fibers containing both afferent (sensory) and efferent (sympathetic) fibers, subsequent preclinical studies have been investigating their respective roles in hypertension pathobiology in different genetic and salt-sensitive rat models. Here we set out to investigate how renal afferent and efferent nerves regulate hypertension development in the chronic mild stress model (CMS). We show that in male CMS rats, ablation of afferent renal nerves (ARDNx) and all renal nerves (TRDNx) resulted in similar BP (104 {+/-} 2 mmHg vs. 101 {+/-} 3 mmHg, respectively), both reduced compared to the SHAM group (118 {+/-} 1 mmHg, p = 0.003 and p < 0.001, respectively) arguing for a prominent role of afferent renal nerves in CMS hypertension. Additionally, we show a reduction of vasopressin (AVP) V1b but not V1a receptor abundance in ARDNx CMS males but not females, suggesting that afferent renal nerves are involved in increase in BP via V1b AVP receptor. We additionally show that despite normal BP, female CMS rats display increased renal sympathetic nerve activity (RSNA; 2.39 {+/-} 0.23 bursts/beat vs. 1.44 {+/-} 0.12 bursts/beat, p < 0.005) measured directly with implanted telemetry in conscious rats over one week and aortic stiffness, as evidenced by increased aortic pulse wave velocity (173.2 {+/-} 50.9 mm/s vs. - 10.7 {+/-} 54.6 mm/s in controls, p = 0.0393). NEW & NOTEWORTHYWe show that renal denervation mitigates the rise in blood pressure (BP) in a model that is not genetic nor diet-dependent, the chronic mild stress model (CMS). Specifically, we demonstrate the role of afferent, rather than efferent, renal nerves in mediating the rise in BP in male CMS rats. Finally, we report that renal sympathetic nerve activity, but not BP, is elevated in female CMS rats measured by telemetry over seven days in conscious rats.

Objectives: Menopausal symptoms are heterogeneous and commonly assessed by questionnaires. We explored serum two-dimensional gel electrophoresis (2-DE) protein spots associated with menopausal symptom burden. Methods: This exploratory cross-sectional study included 27 women aged 45-55 years. A total of 550 matched serum 2-DE spots were quantified. A frequency-adjusted symptom burden score was calculated as the sum of severity x frequency products across 10 symptoms. Spots were screened using Spearman rank correlation with Benjamini-Hochberg false discovery rate (FDR) adjustment, followed by qualitative image review. Spots #285 and #636 were prioritized for vasomotor and psychological domain analyses. Results: The median age was 51.0 years; 13 participants were menstruating and 14 were amenorrheic. The median overall symptom burden score was 45.0 (interquartile range, 6.5-58.5) and was inversely correlated with spots #285 and #636. Spot #285 was inversely correlated with vasomotor symptom score, including inverse correlations in both menstrual-status groups. Spot #636 was inversely correlated with psychological symptom score overall, with a stronger descriptive correlation among menstruating participants. Neither candidate remained significant after FDR adjustment. Conclusions: Spots #285 and #636 are hypothesis-generating candidates requiring molecular identification, analytical validation, multiplicity-aware confirmation, and independent replication.

Introduction: Central regulation of energy homeostasis is essential for balancing lipid storage and reproductive investment; however, the hypothalamic factors governing this trade-off remain incompletely defined in birds. Neurosecretory protein GM (NPGM), an 83-amino acid hypothalamic factor, was investigated for its role in energy allocation during sexual maturation in Japanese quail (Coturnix japonica). Methods: Male and female quails at the onset of sexual maturation received chronic intracerebroventricular administration of NPGM for 13 days via osmotic pumps, during which their body mass, food intake, and water intake were monitored daily. At the endpoint, peripheral tissue and muscle masses, serum metabolite levels (glucose, fatty acids, triglycerides, testosterone, and 17{beta}-estradiol), hepatic triglyceride content, and gene expression profiles of hypothalamic feeding/reproductive genes and hepatic/adipose lipid metabolic genes were evaluated. Results: NPGM increased subcutaneous and abdominal fat in both sexes and was associated with suppressed gonadal maturation, as indicated by reduced testicular mass relative to body mass and lower testosterone levels in males, as well as a trend toward reduced ovarian mass and lower 17{beta}-estradiol levels in females. Sex-dependent metabolic phenotypes emerged: males exhibited increased body mass gain, hyperphagia, elevated water intake, enlarged liver, pancreas, and heart, higher serum and hepatic triglyceride levels, increased hepatic SCD1 expression, and reduced hepatic CGI-58, PPAR{gamma}, SLC2A2, and CD36. In contrast, females showed fat accumulation without hyperphagia or hepatic triglyceride elevation, accompanied by reduced hepatic VTG2 and APOV1 and decreased adipose ATGL, LPL, and FATP. Hypothalamic AGRP expression decreased in males, whereas both NPY and AGRP decreased in females. Discussion: These findings demonstrate that central NPGM shifts energy allocation from reproduction toward lipid storage through sex-dependent endocrine and metabolic mechanisms, identifying NPGM as a neuroendocrine regulator of energy allocation during sexual maturation in Japanese quails.

Objective: Lymphatic dysfunction has been implicated in exacerbating fibrosis in numerous diseases, yet the role of the lymphatic system in laryngotracheal injury has not previously been explored. This study aims to evaluate lymphatic vascular remodeling in a murine model of laryngotracheal stenosis (LTS) and determine how pharmacologic blockade of lymphangiogenesis impacts airway healing after mucosal injury. Methods: LTS was induced in C57BL6 mice using an established chemomechanical injury model. Lymphatic density was quantified using LYVE-1 immunohistochemistry. Mice were treated with the VEGFR-3-selective tyrosine kinase inhibitor SAR131675 to block lymphangiogenesis after injury. Outcomes assessed included survival, histopathology, immunohistochemistry, and Evans blue dye vascular leakage. Results: Laryngotracheal injury induced a substantial increase in subepithelial lymphatic vessel density concomitant with fibrotic remodeling. Pharmacologic inhibition of VEGFR-3 signaling with SAR131675 abrogated this lymphangiogenic response and resulted in markedly increased mortality, impaired epithelial repair with obstructive sloughing, increased edema, and persistent histopathologic evidence of tissue injury. A qualitative increase in pathologic fibrocellular remodeling was also observed, though with no measurable difference in lamina propria thickness. Conclusion: These findings establish lymphatic remodeling as an essential component of successful airway repair following mucosal injury. Lymphatic dysfunction is a common feature of known risk factors for LTS including diabetes, obesity, and prematurity, and can be exacerbated by positive pressure ventilation. Disruption of the lymphangiogenic response to airway injury may lead to stasis of pro-inflammatory factors that result in chronic inflammation, maladaptive remodeling, and pathologic tissue changes. The lymphatic vasculature is a viable target for future mechanistic study and potential therapeutic intervention following airway injury.

Children of undernourished women have impaired pre- and postnatal growth. Undernourished women and children have a high incidence of environmental enteric dysfunction (EED), an enteropathy characterized by gut barrier dysfunction and systemic inflammation. Here, we employ gnotobiotic mice to compare the effects of bacterial consortia cultured from the duodenal microbiota of Bangladeshi women with EED and their healthy counterparts. Female mice harboring the EED-derived consortium exhibited fetal and placental growth restriction. Transcriptomic and proteomic analyses disclosed pronounced effects of the EED-derived consortium on the decidual component of the maternal-fetal interface involving tissue-resident uterine natural killer (uNK) cells and disruption of TGF-{beta} signaling between uNK and decidual stromal cells. Co-housing mice with EED and healthy consortia ameliorated these effects, disclosing bacterial targets to improve prenatal development.

We identified Sleep Bursts (SBs), as a novel phenomenon of brief (1-2 sec), often periodic bursts in cranial forces occurring during human sleep. Our goal was to characterize SBs in normal subjects, then compare SB in a cohort of subjects with neurodegenerative disease (NDD). We recorded 32 cognitively healthy subjects (23 -87 years) and 13 subjects with NDD (51 - 84 years). SBs occurred in all 45 subjects. SBs occurred at 0.57 SB/min (once per 105 seconds) in controls and 0.40 SB/min (once per 150 seconds) in NDD (p = 0.0043). SB occurred with equal rates across all sleep stages in both groups. When occurring periodically, SBs had modal intervals (3.75 bursts/min (0.0625 Hz) - 2.67 bursts/min (0.044 Hz)). EEG power increased in the delta range 1-2 seconds before and following the SB. EEG delta power during a SB was significantly lower in all NDD subjects across sleep stages compared to controls. The relatively low frequency of SB events and synchronization with EEG power has no parallel in human sleep; we hypothesize that SBs may represent a brain-generated pulsatile component of brain glymphatic drainage.

Dietary iron deficiency in the absence of anemia (IDNA) affects numerous people worldwide, with a wide range of negative effects on brain functioning and cognition. Although studies employing electroencephalography (EEG) have revealed a number of negative effects of IDNA in both the time- and frequency domains, to date there have been no attempts to characterize the effects of IDNA on the temporal dynamics of whole brain interactions. To address this issue, we applied multiscale entropy (MSE) analysis to resting-state EEG data collected from IDNA (n = 21) and iron sufficient (IS, n = 21) women. The MSE analysis on this data revealed that entropy was higher overall for the IS than the IDNA group, with significant differences appearing primarily at longer time scales and under right frontal and left and right parietal electrodes. These results suggest that IDNA may negatively affect long-distance interactions among brain regions and that this could conceivably be a source of diminished cognitive function and neural resilience in IDNA.