Development of the Interosseous Muscles of the Human Hand: Morphological and Functional Aspects of the Terminal Insertion.

INTRODUCTION: To date, there have been no studies conducted on the development of interosseous muscles (IO) in the human hand. This study aimed to investigate the development of these muscles in order to clarify their terminal insertions and their relationship with the metacarpophalangeal joints. METHODS: Serial sections of 25 human specimens (9 embryos and 16 fetuses) between the 7th and 14th weeks of development, sourced from the Collection of the Department of Anatomy and Embryology at UCM Faculty of Medicine, were analyzed bilaterally using a conventional optical microscope. RESULTS: Our findings revealed that, during the 7th week of development, the metacarpophalangeal interzone mesenchyme extended into the extensor apparatus of the fingers. Furthermore, we observed that the joint capsule and the tendon of the IO derive from the articular interzone mesenchyme. By the end of the 7th week, corresponding to Carnegie stage 21, the myotendinous junction appeared, initiating cavitation of the metacarpophalangeal joint. During the fetal period, the terminal insertions of the IO were identified: both the dorsal interosseous (DI) and palmar interosseous (PI) muscles insert into the metacarpophalangeal joint capsule and establish a connection with the volar plate located at the base of the proximal phalanx and the extensor apparatus. Some muscle fibers also attach to the joint capsule at the level of the proximal synovial cul-de-sac. The functional implications of these findings are discussed within this work. CONCLUSION: This study provides the first detailed description of the development of the interosseous muscles in the human hand.

Connexin-45 is expressed in mouse lymphatic endothelium and required for lymphatic valve function.

The expression and functional relevance of the gap junction molecule connexin-45 (Cx45; GJC1) in lymphatic endothelium were not previously known. We found that Cx45 was expressed widely in the endothelium of murine lymphatics, in both valve and nonvalve regions. Cell-specific deletion of Cx45, driven by a constitutive Cre line (Lyve1-Cre) or an inducible Cre line (Prox1-CreERT2), compromised the function of lymphatic valves, as assessed by physiological tests (back leak and closure) of isolated, single-valve vessel segments. The defects were comparable to those previously reported for loss of Cx43, and as with Cx43, deletion of Cx45 resulted in shortening or increased asymmetry of lymphatic valve leaflets, providing an explanation for the compromised valve function. In contrast with Cx43, lymphatic endothelial cell-specific (LEC-specific) deletion of Cx45 did not alter the number of valves in mesenteric or dermal lymphatic networks or the expression patterns of the canonical valve-associated proteins PROX1, ITGA9, or CLAUDIN5. Constitutive deletion of Cx45 from LECs resulted in increased backflow of injected tracer in popliteal networks in vivo and compromised the integrity of the LEC permeability barrier in a subset of collecting vessels. These findings provide evidence for an unexpected role of Cx45 in the development and maintenance of lymphatic valves.

Acute Metabolic Stress Induces Lymphatic Dysfunction Through KATP Channel Activation.

Lymphatic dysfunction is an underlying component of multiple metabolic diseases, including diabetes, obesity, and metabolic syndrome. We investigated the roles of KATP channels in lymphatic contractile dysfunction in response to acute metabolic stress induced by inhibition of the mitochondrial electron transport chain. Ex vivo popliteal lymphatic vessels from mice were exposed to the electron transport chain inhibitors antimycin A and rotenone, or the oxidative phosphorylation inhibitor/protonophore, CCCP. Each inhibitor led to a significant reduction in the frequency of spontaneous lymphatic contractions and calculated pump flow, without a significant change in contraction amplitude. Contraction frequency was restored by the KATP channel inhibitor, glibenclamide. Lymphatic vessels from mice with global Kir6.1 deficiency or expressing a smooth muscle-specific dominant negative Kir6.1 channel were resistant to inhibition. Antimycin A inhibited the spontaneous action potentials generated in lymphatic muscle and this effect was reversed by glibenclamide, confirming the role of KATP channels. Antimycin A, but not rotenone or CCCP, increased dihydrorhodamine fluorescence in lymphatic muscle, indicating ROS production. Pretreatment with tiron or catalase prevented the effect of antimycin A on wild-type lymphatic vessels, consistent with its action being mediated by ROS. Our results support the conclusion that KATP channels in lymphatic muscle can be directly activated by reduced mitochondrial ATP production or ROS generation, consequent to acute metabolic stress, leading to contractile dysfunction through inhibition of the ionic pacemaker controlling spontaneous lymphatic contractions. We propose that a similar activation of KATP channels contributes to lymphatic dysfunction in metabolic disease.

Association of Adverse Clinical Outcomes With Peri-Infarct Ischemia Detected by Stress Cardiac Magnetic Imaging.

BACKGROUND: Early invasive revascularization guided by moderate to severe ischemia did not improve outcomes over medical therapy alone, underlying the need to identify high-risk patients for a more effective invasive referral. CMR could determine the myocardial extent and matching locations of ischemia and infarction. OBJECTIVES: This study sought to investigate if CMR peri-infarct ischemia is associated with adverse events incremental to known risk markers. METHODS: Consecutive patients were included in an expanded cohort of the multicenter SPINS (Stress CMR Perfusion Imaging in the United States) study. Peri-infarct ischemia was defined by the presence of any ischemic segment neighboring an infarcted segment by late gadolinium enhancement imaging. Primary outcome events included acute myocardial infarction and cardiovascular death, whereas secondary events included any primary events, hospitalization for unstable angina, heart failure hospitalization, and late coronary artery bypass surgery. RESULTS: Among 3,915 patients (age: 61.0 ± 12.9 years; 54.7% male), ischemia, infarct, and peri-infarct ischemia were present in 752 (19.2%), 1,123 (28.8%), and 382 (9.8%) patients, respectively. At 5.3 years (Q1-Q3: 3.9-7.2 years) of median follow-up, primary and secondary events occurred in 406 (10.4%) and 745 (19.0%) patients, respectively. Peri-infarct ischemia was the strongest multivariable predictor for primary and secondary events (HRadjusted: 1.72 [95% CI: 1.23-2.41] and 1.71 [95% CI: 1.32-2.20], respectively; both P < 0.001), adjusted for clinical risk factors, left ventricular function, ischemia extent, and infarct size. The presence of peri-infarct ischemia portended to a >6-fold increased annualized primary event rate compared to those with no infarct and ischemia (6.5% vs 0.9%). CONCLUSIONS: Peri-infarct ischemia is a novel and robust prognostic marker of adverse cardiovascular events.

TRPV4-Expressing Tissue-Resident Macrophages Regulate the Function of Collecting Lymphatic Vessels via Thromboxane A2 Receptors in Lymphatic Muscle Cells.

Rationale: TRPV4 channels are critical regulators of blood vascular function and have been shown to be dysregulated in many disease conditions in association with inflammation and tissue fibrosis. These are key features in the pathophysiology of lymphatic system diseases, including lymphedema and lipedema; however, the role of TRPV4 channels in the lymphatic system remains largely unexplored. TRPV4 channels are calcium permeable, non-selective cation channels that are activated by diverse stimuli, including shear stress, stretch, temperature, and cell metabolites, which may regulate lymphatic contractile function. Objective: To characterize the expression of TRPV4 channels in collecting lymphatic vessels and to determine the extent to which these channels regulate the contractile function of lymphatics. Methods and Results: Pressure myography on intact, isolated, and cannulated lymphatic vessels showed that pharmacological activation of TRPV4 channels with GSK1016790A (GSK101) led to contractile dysregulation. The response to GSK101 was multiphasic and included, 1) initial robust constriction that was sustained for ≥1 minute and in some instances remained for ≥4 minutes; and 2) subsequent vasodilation and partial or complete inhibition of lymphatic contractions associated with release of nitric oxide. The functional response to activation of TRPV4 channels displayed differences across lymphatics from four anatomical regions, but these differences were consistent across different species (mouse, rat, and non-human primate). Importantly, similar responses were observed following activation of TRPV4 channels in arterioles. The initial and sustained constriction was prevented with the COX inhibitor, indomethacin. We generated a controlled and spatially defined single-cell RNA sequencing (scRNAseq) dataset from intact and microdissected collecting lymphatic vessels. Our data uncovered a subset of macrophages displaying the highest expression of Trpv4 compared to other cell types within and surrounding the lymphatic vessel wall. These macrophages displayed a transcriptomic profile consistent with that of tissue-resident macrophages (TRMs), including differential expression of Lyve1 , Cd163 , Folr2 , Mrc1 , Ccl8 , Apoe , Cd209f , Cd209d , and Cd209g ; and at least half of these macrophages also expressed Timd4. This subset of macrophages also highly expressed Txa2s , which encodes the thromboxane A2 (TXA2) synthase. Inhibition of TXA2 receptors (TXA2Rs) prevented TRPV4-mediated contractile dysregulation. TXA2R activation on LMCs caused an increase in mobilization of calcium from intracellular stores through Ip3 receptors which promoted store operated calcium entry and vasoconstriction. Conclusions: Clinical studies have linked cancer-related lymphedema with an increased infiltration of macrophages. While these macrophages have known anti-inflammatory and pro-lymphangiogenic roles, as well as promote tissue repair, our results point to detrimental effects to the pumping capacity of collecting lymphatic vessels mediated by activation of TRPV4 channels in macrophages. Pharmacological targeting of TRPV4 channels in LYVE1-expressing macrophages or pharmacological targeting of TXA2Rs may offer novel therapeutic strategies to improve lymphatic pumping function and lymph transport in lymphedema.

Gold(III) Auracycles Featuring C(sp3)-Au-C(sp2) Bonds: Synthesis and Mechanistic Insights into the Cycloauration Step.

The direct auration of arenes is a key step in numerous gold-catalyzed reactions. Although reported more than 100 years ago, understanding of its underlying mechanism has been hampered by the difficulties in the isolation of relevant intermediates given the propensity of gold(III) species to undergo reductive elimination. Here, we report the synthesis and isolation of a new family of intriguing zwitterionic [C(sp3)^C(sp2)]-auracyclopentanes, as well as of their alkyl-gold(III) precursors and demonstrate their value as mechanistic probes to study the C(sp2)-Au bond-forming event. Experimental investigations employing Kinetic Isotope Effects (KIE), Hammett plot, and Eyring analysis provided important insights into the formation of the auracycle. The data suggest a SEAr mechanism wherein the slowest step might be the π-coordination between the arene and the gold(III) center, en route to the Wheland intermediate. We also show that these auracyclopentanes can work as catalysts in several gold-promoted transformations.

Loss of anoctamin 1 reveals a subtle role for BK channels in lymphatic muscle action potentials.

Ca2+ signalling plays a crucial role in determining lymphatic muscle cell excitability and contractility through its interaction with the Ca2+-activated Cl- channel anoctamin 1 (ANO1). In contrast, the large-conductance (BK) Ca2+-activated K+ channel (KCa) and other KCa channels have prominent vasodilatory actions by hyperpolarizing vascular smooth muscle cells. Here, we assessed the expression and contribution of the KCa family to mouse and rat lymphatic collecting vessel contractile function. The BK channel was the only KCa channel consistently expressed in fluorescence-activated cell sorting-purified mouse lymphatic muscle cell lymphatic muscle cells. We used a pharmacological inhibitor of BK channels, iberiotoxin, and small-conductance Ca2+-activated K+ channels, apamin, to inhibit KCa channels acutely in ex vivo isobaric myography experiments and intracellular membrane potential recordings. In basal conditions, BK channel inhibition had little to no effect on either mouse inguinal-axillary lymphatic vessel (MIALV) or rat mesenteric lymphatic vessel contractions or action potentials (APs). We also tested BK channel inhibition under loss of ANO1 either by genetic ablation (Myh11CreERT2-Ano1 fl/fl, Ano1ismKO) or by pharmacological inhibition with Ani9. In both Ano1ismKO MIALVs and Ani9-pretreated MIALVs, inhibition of BK channels increased contraction amplitude, increased peak AP and broadened the peak of the AP spike. In rat mesenteric lymphatic vessels, BK channel inhibition also abolished the characteristic post-spike notch, which was exaggerated with ANO1 inhibition, and significantly increased the peak potential and broadened the AP spike. We conclude that BK channels are present and functional on mouse and rat lymphatic muscle cells but are otherwise masked by the dominance of ANO1. KEY POINTS: Mouse and rat lymphatic muscle cells express functional BK channels. BK channels make little contribution to either rat or mouse lymphatic collecting vessel contractile function in basal conditions across a physiological pressure range. ANO1 limits the peak membrane potential achieved in the action potential and sets a plateau potential limiting the voltage-dependent activation of BK. BK channels are activated when ANO1 is absent or blocked and slightly impair contractile strength by reducing the peak membrane potential achieved in the action potential spike and accelerating the post-spike repolarization.

Fibrinogen in mice cerebral microvessels induces blood-brain barrier dysregulation with aging via a dynamin-related protein 1-dependent pathway.

We previously reported evidence that oxidative stress during aging leads to adverse protein profile changes of brain cortical microvessels (MVs: end arterioles, capillaries, and venules) that affect mRNA/protein stability, basement membrane integrity, and ATP synthesis capacity in mice. As an extension of our previous study, we also found that proteins which comprise the blood-brain barrier (BBB) and regulate mitochondrial quality control were also significantly decreased in the mice's cortical MVs with aging. Interestingly, the neuroinflammatory protein fibrinogen (Fgn) was increased in mice brain MVs, which corresponds with clinical reports indicating that the plasma Fgn concentration increased progressively with aging. In this study, protein-protein interaction network analysis indicated that high expression of Fgn is linked with downregulated expression of both BBB- and mitochondrial fission/fusion-related proteins in mice cortical MVs with aging. To investigate the mechanism of Fgn action, we observed that 2 mg/mL or higher concentration of human plasma Fgn changed cell morphology, induced cytotoxicity, and increased BBB permeability in primary human brain microvascular endothelial cells (HBMECs). The BBB tight junction proteins were significantly decreased with increasing concentration of human plasma Fgn in primary HBMECs. Similarly, the expression of phosphorylated dynamin-related protein 1 (pDRP1) and other mitochondrial fission/fusion-related proteins were also significantly reduced in Fgn-treated HBMECs. Interestingly, DRP1 knockdown by shRNA(h) resulted in the reduction of both BBB- and mitochondrial fission/fusion-related proteins in HBMECs. Our results suggest that elevated Fgn downregulates DRP1, leading to mitochondrial-dependent endothelial and BBB dysfunction in the brain microvasculature.

An Unexpected Cause of Abdominal Discomfort: Non-pancreatic Mesenteric Pseudocyst.

Mesenteric cysts (MCs), rare entities of embryologic origin, predominantly affect the small bowel's mesentery. The clinical manifestations of MCs often lack specificity, which complicates diagnosis. Given their rarity, detailed reporting of MC cases is essential to enhance understanding and improve treatment strategies. We present a case of a 45-year-old male who presented to the emergency department with a one-month history of abdominal pain in the umbilical region, postprandial fullness, progressive decrease in food intake, 12 kg weight loss, and increased abdominal girth. Computed tomography (CT) imaging revealed a well-defined mass in the jejunoileal area. During the exploratory laparotomy, we identified and excised a fibrotic mass on the mesentery of the ileal jejunum, which was not adherent to the intestines. We discharged the patient with no complications following an uneventful four-day observational period. Histopathological examination, including immunohistochemical staining, confirmed the lesion as a non-pancreatic mesenteric pseudocyst. On the follow-up visit, the patient reported no complications. This case report underscores the solitary, multilocular nature of the jejunoileal MC, distinct for its serosanguineous fluid content. In conclusion, this case highlights the diagnostic challenge of MCs and illustrates the potential for successful management with a timely and multidisciplinary approach.

Potentiating glymphatic drainage minimizes post-traumatic cerebral oedema.

Cerebral oedema is associated with morbidity and mortality after traumatic brain injury (TBI)1. Noradrenaline levels are increased after TBI2-4, and the amplitude of the increase in noradrenaline predicts both the extent of injury5 and the likelihood of mortality6. Glymphatic impairment is both a feature of and a contributor to brain injury7,8, but its relationship with the injury-associated surge in noradrenaline is unclear. Here we report that acute post-traumatic oedema results from a suppression of glymphatic and lymphatic fluid flow that occurs in response to excessive systemic release of noradrenaline. This post-TBI adrenergic storm was associated with reduced contractility of cervical lymphatic vessels, consistent with diminished return of glymphatic and lymphatic fluid to the systemic circulation. Accordingly, pan-adrenergic receptor inhibition normalized central venous pressure and partly restored glymphatic and cervical lymphatic flow in a mouse model of TBI, and these actions led to substantially reduced brain oedema and improved functional outcomes. Furthermore, post-traumatic inhibition of adrenergic signalling boosted lymphatic export of cellular debris from the traumatic lesion, substantially reducing secondary inflammation and accumulation of phosphorylated tau. These observations suggest that targeting the noradrenergic control of central glymphatic flow may offer a therapeutic approach for treating acute TBI.

IP3R1 underlies diastolic ANO1 activation and pressure-dependent chronotropy in lymphatic collecting vessels.

Pressure-dependent chronotropy of murine lymphatic collecting vessels relies on the activation of the Ca2+-activated chloride channel encoded by Anoctamin 1 (Ano1) in lymphatic muscle cells. Genetic ablation or pharmacological inhibition of ANO1 results in a significant reduction in basal contraction frequency and essentially complete loss of pressure-dependent frequency modulation by decreasing the rate of the diastolic depolarization phase of the ionic pacemaker in lymphatic muscle cells (LMCs). Oscillating Ca2+ release from sarcoendoplasmic reticulum Ca2+ channels has been hypothesized to drive ANO1 activity during diastole, but the source of Ca2+ for ANO1 activation in smooth muscle remains unclear. Here, we investigated the role of the inositol triphosphate receptor 1 (Itpr1; Ip3r1) in this process using pressure myography, Ca2+ imaging, and membrane potential recordings in LMCs of ex vivo pressurized inguinal-axillary lymphatic vessels from control or Myh11CreERT2;Ip3r1fl/fl (Ip3r1ismKO) mice. Ip3r1ismKO vessels had significant reductions in contraction frequency and tone but an increased contraction amplitude. Membrane potential recordings from LMCs of Ip3r1ismKO vessels revealed a depressed diastolic depolarization rate and an elongation of the plateau phase of the action potential (AP). Ca2+ imaging of LMCs using the genetically encoded Ca2+ sensor GCaMP6f demonstrated an elongation of the Ca2+ flash associated with an AP-driven contraction. Critically, diastolic subcellular Ca2+ transients were absent in LMCs of Ip3r1ismKO mice, demonstrating the necessity of IP3R1 activity in controlling ANO1-mediated diastolic depolarization. These findings indicate a critical role for IP3R1 in lymphatic vessel pressure-dependent chronotropy and contractile regulation.

Adult Diaphorina citri Biocontrol Using Hirsutella citriformis Strains and Gum Formulations.

Hirsutella citriformis Speare is the only entomopathogenic fungus that has been applied to control the hemipteran Diaphorina citri Kuwayama. However, the use of available commercial products under field conditions is limited due to conidia's shelf life and short environmental persistence. We have previously reported the citrus psyllid D. citri adults' biocontrol potential using H. citriformis strains. The aim of the present study was to evaluate different formulations based on H. citriformis (OP-Hir-3, OP-Hir-10, and OP-Hir-12 strains) conidia and gums as additives to improve D. citri adults' biocontrol, under laboratory, greenhouse, and field conditions, using Hirsutella gums as conidia stabilizers to improve their viability under environmental drought conditions and as insecticide. Laboratory bioassay results showed that the highest (p < 0.05) D. citri mortality was achieved using FOP-Hir-10GH (63.5%), followed by the Hirsutella gum control (42.2%). Under greenhouse conditions, adults' mortality reached up to 84.6% with FOP-Hir-12 and 49.0% with Hirsutella gum. In addition, we applied H. citriformis formulations under field conditions in a commercial citrus grove located in Tecomán, Colima, México, at 21.5 °C and 73.3% relative humidity (RH) in March and 25.7 °C and 72.5% RH in October 2022 and observed 67.3% and 94.0% mortality of D. citri adults, respectively. Hirsutella gum alone showed significant insecticidal activity against D. citri adults. In conclusion, this study demonstrated that Hirsutella gum functioned as additive to H. citriformis conidia formulations, improving D. citri adults' mortality and showing potential for this pest biocontrol in citrus orchards.

Electric field stimulation unmasks a subtle role for T-type calcium channels in regulating lymphatic contraction.

We previously identified two isoforms of T-type, voltage-gated calcium (Cav3) channels (Cav3.1, Cav3.2) that are functionally expressed in murine lymphatic muscle cells; however, contractile tests of lymphatic vessels from single and double Cav3 knock-out (DKO) mice, exhibited nearly identical parameters of spontaneous twitch contractions as wild-type (WT) vessels, suggesting that Cav3 channels play no significant role. Here, we considered the possibility that the contribution of Cav3 channels might be too subtle to detect in standard contraction analyses. We compared the sensitivity of lymphatic vessels from WT and Cav3 DKO mice to the L-type calcium channel (Cav1.2) inhibitor nifedipine and found that the latter vessels were significantly more sensitive to inhibition, suggesting that the contribution of Cav3 channels might normally be masked by Cav1.2 channel activity. We hypothesized that shifting the resting membrane potential (Vm) of lymphatic muscle to a more negative voltage might enhance the contribution of Cav3 channels. Because even slight hyperpolarization is known to completely silence spontaneous contractions, we devised a method to evoke nerve-independent, twitch contractions from mouse lymphatic vessels using single, short pulses of electric field stimulation (EFS). TTX was present throughout to block the potential contributions of voltage-gated Na+ channels in perivascular nerves and lymphatic muscle. In WT vessels, EFS evoked single contractions that were comparable in amplitude and degree of entrainment to those occurring spontaneously. When Cav1.2 channels were blocked or deleted, only small residual EFS-evoked contractions (~ 5% of normal amplitude) were present. These residual, EFS-evoked contractions were enhanced (to 10-15%) by the KATP channel activator pinacidil (PIN) but were absent in Cav3 DKO vessels. Our results point to a subtle contribution of Cav3 channels to lymphatic contractions that can be unmasked in the absence of Cav1.2 channel activity and when the resting Vm is more hyperpolarized than normal.

Postnatal development of extracellular matrix and vascular function in small arteries of the rat.

Introduction: Vascular extracellular matrix (ECM) is dominated by elastic fibers (elastin with fibrillin-rich microfibrils) and collagens. Current understanding of ECM protein development largely comes from studies of conduit vessels (e.g., aorta) while resistance vessel data are sparse. With an emphasis on elastin, we examined whether changes in postnatal expression of arteriolar wall ECM would correlate with development of local vasoregulatory mechanisms such as the myogenic response and endothelium-dependent dilation. Methods: Rat cerebral and mesenteric arteries were isolated at ages 3, 7, 11, 14, 19 days, 2 months, and 2 years. Using qPCR mRNA expression patterns were examined for elastin, collagen types I, II, III, IV, fibrillin-1, and -2, lysyl oxidase (LOX), and transglutaminase 2. Results: Elastin, LOX and fibrillar collagens I and III mRNA peaked at day 11-14 in both vasculatures before declining at later time-points. 3D confocal imaging for elastin showed continuous remodeling in the adventitia and the internal elastic lamina for both cerebral and mesenteric vessels. Myogenic responsiveness in cannulated cerebral arteries was detectable at day 3 with constriction shifted to higher intraluminal pressures by day 19. Myogenic responsiveness of mesenteric vessels appeared fully developed by day 3. Functional studies were performed to investigate developmental changes in endothelial-dependent dilation. Endothelial-dependent dilation to acetylcholine was less at day 3 compared to day 19 and at day 3 lacked an endothelial-derived hyperpolarizing factor component that was evident at day 19. Conclusion: Collectively, in the rat small artery structural remodeling and aspects of functional control continue to develop in the immediate postnatal period.

Electric field stimulation unmasks a subtle role for T-type calcium channels in regulating lymphatic contraction.

We previously identified two isoforms of T-type, voltage-gated calcium (Ca v 3) channels (Ca v 3.1, Ca v 3.2) that are functionally expressed in murine lymphatic muscle cells; however, contractile tests of lymphatic vessels from single and double Ca v 3 knock-out (DKO) mice, exhibited nearly identical parameters of spontaneous twitch contractions as wild-type (WT) vessels, suggesting that Ca v 3 channels play no significant role. Here, we considered the possibility that the contribution of Ca v 3 channels might be too subtle to detect in standard contraction analyses. We compared the sensitivity of lymphatic vessels from WT and Ca v 3 DKO mice to the L-type calcium channel (Ca v 1.2) inhibitor nifedipine and found that the latter vessels were significantly more sensitive to inhibition, suggesting that the contribution of Ca v 3 channels might normally be masked by Ca v 1.2 channel activity. We hypothesized that shifting the resting membrane potential (Vm) of lymphatic muscle to a more negative voltage might enhance the contribution of Ca v 3 channels. Because even slight hyperpolarization is known to completely silence spontaneous contractions, we devised a method to evoke nerve-independent, twitch contractions from mouse lymphatic vessels using single, short pulses of electric field stimulation (EFS). TTX was present throughout to block the potential contributions of voltage-gated Na + channels in perivascular nerves and lymphatic muscle. In WT vessels, EFS evoked single contractions that were comparable in amplitude and degree of entrainment to those occurring spontaneously. When Ca v 1.2 channels were blocked or deleted, only small residual EFS-evoked contractions (~ 5% of normal amplitude) were present. These residual, EFS-evoked contractions were enhanced (to 10-15%) by the K ATP channel activator pinacidil (PIN) but were absent in Ca v 3 DKO vessels. Our results point to a subtle contribution of Ca v 3 channels to lymphatic contractions that can be unmasked in the absence of Ca v 1.2 channel activity and when the resting Vm is more hyperpolarized than normal.

Lymphatic contractile dysfunction in mouse models of Cantú Syndrome with KATP channel gain-of-function.

Cantú Syndrome (CS) is an autosomal dominant disorder caused by gain-of-function (GoF) mutations in the Kir6.1 and SUR2 subunits of KATP channels. KATP overactivity results in a chronic reduction in arterial tone and hypotension, leading to other systemic cardiovascular complications. However, the underlying mechanism of lymphedema, developed by >50% of CS patients, is unknown. We investigated whether lymphatic contractile dysfunction occurs in mice expressing CS mutations in Kir6.1 (Kir6.1[V65M]) or SUR2 (SUR2[A478V], SUR2[R1154Q]). Pressure myograph tests of contractile function of popliteal lymphatic vessels over the physiological pressure range revealed significantly impaired contractile strength and reduced frequency of spontaneous contractions at all pressures in heterozygous Kir6.1[V65M] vessels, compared to control littermates. Contractile dysfunction of intact popliteal lymphatics in vivo was confirmed using near-infrared fluorescence microscopy. Homozygous SUR2[A478V] vessels exhibited profound contractile dysfunction ex vivo, but heterozygous SUR2[A478V] vessels showed essentially normal contractile function. However, further investigation of vessels from all three GoF mouse strains revealed significant disruption in contraction wave entrainment, decreased conduction speed and distance, multiple pacemaker sites, and reversing wave direction. Tests of 2-valve lymphatic vessels forced to pump against an adverse pressure gradient revealed that all CS-associated genotypes were essentially incapable of pumping under an imposed outflow load. Our results show that varying degrees of lymphatic contractile dysfunction occur in proportion to the degree of molecular GoF in Kir6.1 or SUR2. This is the first example of lymphatic contractile dysfunction caused by a smooth muscle ion channel mutation and potentially explains the susceptibility of CS patients to lymphedema.

Circulating Plasma Exosomal Proteins of Either SHIV-Infected Rhesus Macaque or HIV-Infected Patient Indicates a Link to Neuropathogenesis.

Despite the suppression of human immunodeficiency virus (HIV) replication by combined antiretroviral therapy (cART), 50-60% of HIV-infected patients suffer from HIV-associated neurocognitive disorders (HAND). Studies are uncovering the role of extracellular vesicles (EVs), especially exosomes, in the central nervous system (CNS) due to HIV infection. We investigated links among circulating plasma exosomal (crExo) proteins and neuropathogenesis in simian/human immunodeficiency virus (SHIV)-infected rhesus macaques (RM) and HIV-infected and cART treated patients (Patient-Exo). Isolated EVs from SHIV-infected (SHIV-Exo) and uninfected (CTL-Exo) RM were predominantly exosomes (particle size < 150 nm). Proteomic analysis quantified 5654 proteins, of which 236 proteins (~4%) were significantly, differentially expressed (DE) between SHIV-/CTL-Exo. Interestingly, different CNS cell specific markers were abundantly expressed in crExo. Proteins involved in latent viral reactivation, neuroinflammation, neuropathology-associated interactive as well as signaling molecules were expressed at significantly higher levels in SHIV-Exo than CTL-Exo. However, proteins involved in mitochondrial biogenesis, ATP production, autophagy, endocytosis, exocytosis, and cytoskeleton organization were significantly less expressed in SHIV-Exo than CTL-Exo. Interestingly, proteins involved in oxidative stress, mitochondrial biogenesis, ATP production, and autophagy were significantly downregulated in primary human brain microvascular endothelial cells exposed with HIV+/cART+ Patient-Exo. We showed that Patient-Exo significantly increased blood-brain barrier permeability, possibly due to loss of platelet endothelial cell adhesion molecule-1 protein and actin cytoskeleton structure. Our novel findings suggest that circulating exosomal proteins expressed CNS cell markers-possibly associated with viral reactivation and neuropathogenesis-that may elucidate the etiology of HAND.

Sex-Specific Stress Perfusion Cardiac Magnetic Resonance Imaging in Suspected Ischemic Heart Disease: Insights From SPINS Retrospective Registry.

BACKGROUND: Cardiovascular disease (CVD) remains the leading cause of mortality in women, but current noninvasive cardiac imaging techniques have sex-specific limitations. OBJECTIVES: In this study, the authors sought to investigate the effect of sex on the prognostic utility and downstream invasive revascularization and costs of stress perfusion cardiac magnetic resonance (CMR) for suspected CVD. METHODS: Sex-specific prognostic performance was evaluated in a 2,349-patient multicenter SPINS (Stress CMR Perfusion Imaging in the United States [SPINS] Study) Registry. The primary outcome measure was a composite of cardiovascular death and nonfatal myocardial infarction; secondary outcomes were hospitalization for unstable angina or heart failure, and late unplanned coronary artery bypass grafting. RESULTS: SPINS included 1,104 women (47% of cohort); women had higher prevalence of chest pain (62% vs 50%; P < 0.0001) but lower use of medical therapies. At the 5.4-year median follow-up, women with normal stress CMR had a low annualized rate of primary composite outcome similar to men (0.54%/y vs 0.75%/y, respectively; P = NS). In contrast, women with abnormal CMR were at higher risk for both primary (3.74%/y vs 0.54%/y; P < 0.0001) and secondary (9.8%/y vs 1.6%/y; P < 0.0001) outcomes compared with women with normal CMR. Abnormal stress CMR was an independent predictor for the primary (HR: 2.64 [95% CI: 1.20-5.90]; P = 0.02) and secondary (HR: 2.09 [95% CI: 1.43-3.08]; P < 0.0001) outcome measures. There was no effect modification for sex. Women had lower rates of invasive coronary angiography (3.6% vs 7.3%; P = 0.0001) and downstream costs ($114 vs $171; P = 0.001) at 90 days following CMR. There was no effect of sex on diagnostic image quality. CONCLUSIONS: Stress CMR demonstrated excellent prognostic performance with lower rates of invasive coronary angiography referral in women. Stress CMR should be considered as a first-line noninvasive imaging tool for the evaluation of women. (Stress CMR Perfusion Imaging in the United States [SPINS] Study [SPINS]; NCT03192891).

Titanium versus zirconia complete arch implant-supported fixed prostheses: A comparison of plaque accumulation.

STATEMENT OF PROBLEM: Complete arch implant-supported zirconia prostheses appear to have less plaque accumulation than titanium prostheses, but a comparison of the materials and the possible influence on the adjacent soft tissue is lacking. PURPOSE: The purpose of this clinical study was to compare the plaque accumulation and soft-tissue inflammation of complete arch implant-supported fixed maxillary prostheses fabricated with either a titanium framework or monolithic zirconia. MATERIAL AND METHODS: Twenty participants with a complete arch implant-supported fixed maxillary prosthesis were enrolled in the study. The participants were divided into 2 groups according to the prosthesis material, titanium (Ti) or zirconia (Zir). The prosthesis had to have been in function for at least 6 months, and participants were examined during at least 3 maintenance appointments at 3-month intervals. Clinical information collected in each appointment included standardized photographs to record the Plaque Area Index (PAI) of the intaglio surface of the prosthesis; clinical parameters including modified Plaque Index (mPI), modified Bleeding Index (mBI), implant mobility (MOB), probing depths ≥5 mm (PD), suppuration (SUP), keratinized tissue band ≥2 mm (KT), and an intraoral photograph of the maxillary arch without the prosthesis to evaluate the redness of the soft tissues. RESULTS: MOB was not present at any implant at any time point. SUP could not be analyzed because it was an infrequent finding. Both groups exhibited significant increases in mBI over time. No significant differences were observed for PD between the groups at any time point. Implants in the Ti group had significantly higher KT values than those in the Zir group; levels remained constant over time for both groups. Zirconia prostheses had slightly lower PAI levels than Ti prostheses. The PAI in the Zir group significantly decreased over time (P=.035); in the Ti group, they remained constant (P=.45). Higher PAI levels were correlated with increased levels of erythema; both groups had a significant decrease in erythema values over time (P=.04). CONCLUSIONS: Zirconia complete arch implant-supported fixed maxillary prostheses displayed a significant decrease in plaque accumulation in individuals who had received periodic maintenance and oral hygiene instructions. Ti prostheses had significantly higher plaque levels than zirconia prostheses at all time points, which was not reduced by maintenance and oral hygiene measures. The present study suggests that patients receiving zirconia prostheses respond well to plaque control measures, while plaque control for those with titanium prostheses may be more challenging.