TRPV4 channels' dominant role in the temperature modulation of intrinsic contractility and lymph flow of rat diaphragmatic lymphatics.

The lymphatic system drains and propels lymph by extrinsic and intrinsic mechanisms. Intrinsic propulsion depends upon spontaneous rhythmic contractions of lymphatic muscles in the vessel walls and is critically affected by changes in the surrounding tissue like osmolarity and temperature. Lymphatics of the diaphragm display a steep change in contraction frequency in response to changes in temperature, and this, in turn, affects lymph flow. In the present work, we demonstrated in an ex vivo diaphragmatic tissue rat model that diaphragmatic lymphatics express transient receptor potential channels of the vanilloid 4 subfamily (TRPV4) and that their blockade by both the nonselective antagonist Ruthenium Red and the selective antagonist HC-067047 abolished the response of lymphatics to temperature changes. Moreover, the selective activation of TRPV4 channels by means of GSK1016790A mirrored the behavior of vessels exposed to increasing temperatures, pointing out the critical role played by these channels in sensing the temperature of the lymphatic vessels' environment and thus inducing a change in contraction frequency and lymph flow.NEW & NOTEWORTHY The present work addresses the putative receptor system that enables diaphragmatic lymphatics to change intrinsic contraction frequency and thus lymph flow according to the changes in temperature of the surrounding environment, showing that this role can be sustained by TRPV4 channels alone.

Nitric oxide regulates homeoprotein OTX1 and OTX2 expression in the rat myenteric plexus after intestinal ischemia-reperfusion injury.

Neuronal and inducible nitric oxide synthase (nNOS and iNOS) play a protective and damaging role, respectively, on the intestinal neuromuscular function after ischemia-reperfusion (I/R) injury. To uncover the molecular pathways underlying this dichotomy we investigated their possible correlation with the orthodenticle homeobox proteins OTX1 and OTX2 in the rat small intestine myenteric plexus after in vivo I/R. Homeobox genes are fundamental for the regulation of the gut wall homeostasis both during development and in pathological conditions (inflammation, cancer). I/R injury was induced by temporary clamping the superior mesenteric artery under anesthesia, followed by 24 and 48 h of reperfusion. At 48 h after I/R intestinal transit decreased and was further reduced by Nω-propyl-l-arginine hydrochloride (NPLA), a nNOS-selective inhibitor. By contrast this parameter was restored to control values by 1400W, an iNOS-selective inhibitor. In longitudinal muscle myenteric plexus (LMMP) preparations, iNOS, OTX1, and OTX2 mRNA and protein levels increased at 24 and 48 h after I/R. At both time periods, the number of iNOS- and OTX-immunopositive myenteric neurons increased. nNOS mRNA, protein levels, and neurons were unchanged. In LMMPs, OTX1 and OTX2 mRNA and protein upregulation was reduced by 1400W and NPLA, respectively. In myenteric ganglia, OTX1 and OTX2 staining was superimposed with that of iNOS and nNOS, respectively. Thus in myenteric ganglia iNOS- and nNOS-derived NO may promote OTX1 and OTX2 upregulation, respectively. We hypothesize that the neurodamaging and neuroprotective roles of iNOS and nNOS during I/R injury in the gut may involve corresponding activation of molecular pathways downstream of OTX1 and OTX2.NEW & NOTEWORTHY Intestinal ischemia-reperfusion (I/R) injury induces relevant alterations in myenteric neurons leading to dismotility. Nitrergic neurons seem to be selectively involved. In the present study the inference that both neuronal and inducible nitric oxide synthase (nNOS and iNOS) expressing myenteric neurons may undergo important changes sustaining derangements of motor function is reinforced. In addition, we provide data to suggest that NO produced by iNOS and nNOS regulates the expression of the vital transcription factors orthodenticle homeobox protein 1 and 2 during an I/R damage.

Temperature-dependent modulation of regional lymphatic contraction frequency and flow.

Lymph drainage and propulsion are sustained by an extrinsic mechanism, based on mechanical forces acting from the surrounding tissues against the wall of lymphatic vessels, and by an intrinsic mechanism attributable to active spontaneous contractions of the lymphatic vessel muscle. Despite being heterogeneous, the mechanisms underlying the generation of spontaneous contractions share a common biochemical nature and are thus modulated by temperature. In this study, we challenged excised tissues from rat diaphragm and hindpaw, endowed with spontaneously contracting lymphatic vessels, to temperatures from 24°C (hindpaw) or 33°C (diaphragmatic vessels) to 40°C while measuring lymphatic contraction frequency (fc) and amplitude. Both vessel populations displayed a sigmoidal relationship between fc and temperature, each centered around the average temperature of surrounding tissue (36.7 diaphragmatic and 32.1 hindpaw lymphatics). Although the slope factor of the sigmoidal fit to the fc change of hindpaw vessels was 2.3°C·cycles-1·min-1, a value within the normal range displayed by simple biochemical reactions, the slope factor of the diaphragmatic lymphatics was 0.62°C·cycles-1·min-1, suggesting the added involvement of temperature-sensing mechanisms. Lymph flow calculated as a function of temperature confirmed the relationship observed on fc data alone and showed that none of the two lymphatic vessel populations would be able to adapt to the optimal working temperature of the other tissue district. This poses a novel question whether lymphatic vessels might not adapt their function to accommodate the change if exposed to a surrounding temperature, which is different from their normal condition.NEW & NOTEWORTHY This study demonstrates to what extent lymphatic vessel intrinsic contractility and lymph flow are modulated by temperature and that this modulation is dependent on the body district that the vessels belong to, suggesting a possible functional misbehavior should lymphatic vessels be exposed to a chronically different temperature.

Lymph flow pattern in pleural diaphragmatic lymphatics during intrinsic and extrinsic isotonic contraction.

Peripheral rat diaphragmatic lymphatic vessels, endowed with intrinsic spontaneous contractility, were in vivo filled with fluorescent dextrans and microspheres and subsequently studied ex vivo in excised diaphragmatic samples. Changes in diameter and lymph velocity were detected, in a vessel segment, during spontaneous lymphatic smooth muscle contraction and upon activation, through electrical whole-field stimulation, of diaphragmatic skeletal muscle fibers. During intrinsic contraction lymph flowed both forward and backward, with a net forward propulsion of 14.1 ± 2.9 µm at an average net forward speed of 18.0 ± 3.6 µm/s. Each skeletal muscle contraction sustained a net forward-lymph displacement of 441.9 ± 159.2 µm at an average velocity of 339.9 ± 122.7 µm/s, values significantly higher than those documented during spontaneous contraction. The flow velocity profile was parabolic during both spontaneous and skeletal muscle contraction, and the shear stress calculated at the vessel wall at the highest instantaneous velocity never exceeded 0.25 dyne/cm(2). Therefore, we propose that the synchronous contraction of diaphragmatic skeletal muscle fibers recruited at every inspiratory act dramatically enhances diaphragmatic lymph propulsion, whereas the spontaneous lymphatic contractility might, at least in the diaphragm, be essential in organizing the pattern of flow redistribution within the diaphragmatic lymphatic circuit. Moreover, the very low shear stress values observed in diaphragmatic lymphatics suggest that, in contrast with other contractile lymphatic networks, a likely interplay between intrinsic and extrinsic mechanisms be based on a mechanical and/or electrical connection rather than on nitric oxide release.

Hyperpolarization-activated cyclic nucleotide-gated channels in peripheral diaphragmatic lymphatics.

Diaphragmatic lymphatic function is mainly sustained by pressure changes in the tissue and serosal cavities during cardiorespiratory cycles. The most peripheral diaphragmatic lymphatics are equipped with muscle cells (LMCs), which exhibit spontaneous contraction, whose molecular machinery is still undetermined. Hypothesizing that spontaneous contraction might involve hyperpolarization-activated cyclic nucleotide-gated (HCN) channels in lymphatic LMCs, diaphragmatic specimens, including spontaneously contracting lymphatics, were excised from 33 anesthetized rats, moved to a perfusion chamber containing HEPES-Tyrode's solution, and treated with HCN channels inhibitors cesium chloride (CsCl), ivabradine, and ZD-7288. Compared with control, exposure to 10 mM CsCl reduced (-65%, n = 13, P < 0.01) the contraction frequency (FL) and increased end-diastolic diameter (DL-d, +7.3%, P < 0.01) without changes in end-systolic diameter (DL-s). Ivabradine (300 µM) abolished contraction and increased DL-d (-14%, n = 10, P < 0.01) or caused an incomplete inhibition of FL (n = 3, P < 0.01), leaving DL-d and DL-s unaltered. ZD-7288 (200 µM) completely (n = 12, P < 0.01) abolished FL, while DL-d decreased to 90.9 ± 2.7% of control. HCN gene expression and immunostaining confirmed the presence of HCN1-4 channel isoforms, likely arranged in different configurations, in LMCs. Hence, all together, data suggest that HCN channels might play an important role in affecting contraction frequency of LMCs.

Diaphragmatic lymphatic vessel behavior during local skeletal muscle contraction.

The mechanism through which the stresses developed in the diaphragmatic tissue during skeletal muscle contraction sustain local lymphatic function was studied in 10 deeply anesthetized, tracheotomized adult Wistar rats whose diaphragm was exposed after thoracotomy. To evaluate the direct effect of skeletal muscle contraction on the hydraulic intraluminal lymphatic pressures (Plymph) and lymphatic vessel geometry, the maximal contraction of diaphragmatic fibers adjacent to a lymphatic vessel was elicited by injection of 9.2 nl of 1 M KCl solution among diaphragmatic fibers while Plymph was recorded through micropuncture and vessel geometry via stereomicroscopy video recording. In lymphatics oriented perpendicularly to the longitudinal axis of muscle fibers and located at <300 µm from KCl injection, vessel diameter at maximal skeletal muscle contraction (Dmc) decreased to 61.3 ± 1.4% of the precontraction value [resting diameter (Drest)]; however, if injection was at >900 µm from the vessel, Dmc enlarged to 131.1 ± 2.3% of Drest. In vessels parallel to muscle fibers, Dmc increased to 122.8 ± 2.9% of Drest. During contraction, Plymph decreased as much as 22.5 ± 2.6 cmH2O in all submesothelial superficial vessels, whereas it increased by 10.7 ± 5.1 cmH2O in deeper vessels running perpendicular to contracting muscle fibers. Hence, the three-dimensional arrangement of the diaphragmatic lymphatic network seems to be finalized to efficiently exploit the stresses exerted by muscle fibers during the contracting inspiratory phase to promote lymph formation in superficial submesothelial lymphatics and its further propulsion in deeper intramuscular vessels.

Regional lung tissue changes with mechanical ventilation and fluid load.

PURPOSE: To investigate the regional gravity-dependent impact of mechanical ventilation and fluid overload on lung extracellular matrix (ECM) in healthy lungs. MATERIALS AND METHODS: The glycosaminoglycans (GAGs) composition of the ventral and dorsal lung parenchyma was determined in anesthetized supine healthy rats mechanically ventilated for 4 hours in air: (a) at low (∼7.5 mL/kg) or high (∼ 23 mL /kg) tidal volume (V(T)) and 0 cmH2O positive end-expiratory pressure (PEEP); (b) at low or high V(T) at 5 cmH2O PEEP and (c) with or without 7 mL /(kg·h) intravenous saline infusion. RESULTS: Mechanical ventilation degraded lung ECM, with alveolar septa thinning and structural GAGs disorganization. Low V(T) ventilation was associated with significant tissue structure changes in both ventral and dorsal lung regions, while high VT mainly affected the dependent ones. PEEP decreased ECM injury mainly in the ventral lung regions, although it did not prevent matrix fragmentation and washout at high V(T). Intravascular fluid load increased lung damage prevalently in the ventral lung regions. CONCLUSION: Mechanical ventilation and fluid load may cause additive injuries in healthy lungs, mainly in ventral regions.

COVID-19-Associated Multisystem Inflammatory Syndrome in Children and Cardiovascular Autonomic Control: A Prospective Cohort Study Nine Months after SARS-CoV-2 Infection.

Background: Multisystem Inflammatory Syndrome in Children (MIS-C) has emerged as a severe pediatric complication during the SARS-CoV-2 pandemic, with potential long-term cardiovascular repercussions. We hypothesized that heart rate and blood pressure control at rest and during postural maneuvers in MIS-C patients, months after the remission of the inflammatory syndrome, may reveal long-term autonomic dysfunctions. Methods: We assessed 17 MIS-C patients (13 males; 11.9 ± 2.6 years, m ± SD) 9 months after acute infection and 18 age- (12.5 ± 2.1 years) and sex- (13 males) matched controls. Heart rate and blood pressure variability, baroreflex function, and hemodynamic parameters were analyzed in supine and standing postures. Results: MIS-C patients exhibited reduced heart rate variability, particularly in parasympathetic parameters during standing (pNN50+: 6.1 ± 6.4% in controls, 2.5 ± 3.9% in MIS-C; RMSSD: 34 ± 19 ms in controls, 21 ± 14 ms in MIS-C, p < 0.05), with no interaction between case and posture. Blood pressure variability and baroreflex sensitivity did not differ between groups except for the high-frequency power in systolic blood pressure (3.3 ± 1.2 mmHg2 in controls, 1.8 ± 1.2 mmHg2 in MIS-C, p < 0.05). The MIS-C group also showed lower diastolic pressure-time indices (DPTI) and systolic pressure-time indices (SPTI), particularly in standing (DPTI: 36.2 ± 9.4 mmHg·s in controls, 29.4 ± 6.2 mmHg·s in MIS-C; SPTI: 26.5 ± 4.3 mmHg·s in controls, 23.9 ± 2.4 mmHg·s in MIS-C, p < 0.05). Conclusions: Altered cardiovascular autonomic control may persist in MIS-C patients with, however, compensatory mechanisms that may help maintain cardiovascular homeostasis during light autonomic challenges, such as postural maneuvers. These results highlight the importance of assessing long-term cardiovascular autonomic control in children with MIS-C to possibly identify residual cardiovascular risks and inform targeted interventions and rehabilitation protocols.

Spontaneous activity in peripheral diaphragmatic lymphatic loops.

The spontaneous contractility of FITC-dextran-filled lymphatics at the periphery of the pleural diaphragm was documented for the first time "in vivo" in anesthetized Wistar rats. We found that lymphatic segments could be divided into four phenotypes: 1) active, displaying rhythmic spontaneous contractions (51.8% of 197 analyzed sites); 2) stretch-activated, whose contraction was triggered by passive distension of the vessel lumen (4.1%); 3) passive, which displayed a completely passive distension (4.5%); and 4) inert, whose diameter never changed over time (39.6%). Smooth muscle actin was detected by immunofluorescence and confocal microscopy in the vessel walls of active but also of inert sites, albeit with a very different structure within the vessel wall. Indeed, while in active segments, actin was arranged in a dense mesh completely surrounding the lumen, in inert segments actin decorated the vessels wall in sparse longitudinal strips. When located nearby along the same lymphatic loop, active, stretch-activated, and passive sites were always recruited in temporal sequence starting from the active contraction. The time delay was ∼0.35 s between active and stretch-activated and 0.54 s between stretch-activated and passive segments, promoting a uniform lymph flux of ∼150/200 pl/min. We conclude that, unlike more central diaphragmatic lymphatic vessels, loops located at the extreme diaphragmatic periphery do require an intrinsic pumping mechanism to propel lymph centripetally, and that such an active lymph propulsion is attained by means of a complex interplay among sites whose properties differ but are indeed able to organize lymph flux in an ordered fashion.

Fluid Osmolarity Modulates the Rate of Spontaneous Contraction of Lymphatic Vessels and Lymph Flow by Means of a Cooperation between TRPV and VRAC Channels.

Lymphatic vessels are capable of sustaining lymph formation and propulsion via an intrinsic mechanism based on the spontaneous contraction of the lymphatic muscle in the wall of lymphatic collectors. Exposure to a hyper- or hypo-osmolar environment can deeply affect the intrinsic contraction rate and therefore alter lymph flow. In this work, we aimed at defining the putative receptors underlying such a response. Functional experiments were conducted in ex vivo rat diaphragmatic specimens containing spontaneously contracting lymphatic vessels that were exposed to either hyper- or hypo-osmolar solutions. Lymphatics were challenged with blockers to TRPV4, TRPV1, and VRAC channels, known to respond to changes in osmolarity and/or cell swelling and expressed by lymphatic vessels. Results show that the normal response to a hyperosmolar environment is a steady decrease in the contraction rate and lymph flow and can be prevented by blocking TRPV1 channels with capsazepine. The response to a hyposmolar environment consists of an early phase of an increase in the contraction rate, followed by a decrease. The early phase is abolished by blocking VRACs with DCPIB, while blocking TRPV4 mainly resulted in a delay of the early response. Overall, our data suggest that the cooperation of the three channels can shape the response of lymphatic vessels in terms of contraction frequency and lymph flow, with a prominent role of TRPV1 and VRACs.

Draining the Pleural Space: Lymphatic Vessels Facing the Most Challenging Task.

Lymphatic vessels exploit the mechanical stresses of their surroundings together with intrinsic rhythmic contractions to drain lymph from interstitial spaces and serosal cavities to eventually empty into the blood venous stream. This task is more difficult when the liquid to be drained has a very subatmospheric pressure, as it occurs in the pleural cavity. This peculiar space must maintain a very low fluid volume at negative hydraulic pressure in order to guarantee a proper mechanical coupling between the chest wall and lungs. To better understand the potential for liquid drainage, the key parameter to be considered is the difference in hydraulic pressure between the pleural space and the lymphatic lumen. In this review we collected old and new findings from in vivo direct measurements of hydraulic pressures in anaesthetized animals with the aim to better frame the complex physiology of diaphragmatic and intercostal lymphatics which drain liquid from the pleural cavity.

Effects of speed, agility and quickness training programme on cognitive and physical performance in preadolescent soccer players.

The aim of the present study was to investigate the effect of a short-term (4 weeks) non-soccer-specific training programme based on speed, agility and quickness (SAQ) and a soccer-specific training programme based on small-sided games (SSG) on cognitive and physical performance in preadolescent soccer players. Twenty-one participants were randomly assigned to SAQ group (n = 11) or SSG group (n = 10). They were tested pre and post interventions on physical (5 m sprint, 20 m sprint and sprint with turns of 90°) and cognitive (inhibitory control by means of the Flanker task and perceptual speed by means of the visual search task) performances. Although no significant time x group interactions were observed, the main effect of time was significant for cognitive performance and 5 m and 20 m sprint, showing improvements after both SAQ and SSG. These findings highlight that 4 weeks of SAQ training programme induced comparable improvements in cognitive and physical performance with respect to a soccer-specific training programme based on SSG in preadolescent soccer players. Non-sport-specific activities targeting speed, agility and quickness combined with cognitive engagement (i.e., SAQ) should be useful strategies as soccer-specific activities to be included within a soccer training programme for promoting both physical and cognitive domain in preadolescent individuals.

Lymphatic anatomy and biomechanics.

Lymph formation is driven by hydraulic pressure gradients developing between the interstitial tissue and the lumen of initial lymphatics. While in vessels equipped with lymphatic smooth muscle cells these gradients are determined by well-synchronized spontaneous contractions of vessel segments, initial lymphatics devoid of smooth muscles rely on tissue motion to form lymph and propel it along the network. Lymphatics supplying highly moving tissues, such as skeletal muscle, diaphragm or thoracic tissues, undergo cyclic compression and expansion of their lumen imposed by local stresses arising in the tissue as a consequence of cardiac and respiratory activities. Active muscle contraction and not passive tissue displacement is required to support an efficient lymphatic drainage, as suggested by the fact that the respiratory activity promotes lymph formation during spontaneous, but not mechanical ventilation. The mechanical properties of the lymphatic wall and of the surrounding tissue also play an important role in lymphatic function. Modelling of stress distribution in the lymphatic wall suggests that compliant vessels behave as reservoirs accommodating absorbed interstitial fluid, while lymphatics with stiffer walls, taking advantage of a more efficient transmission of tissue stresses to the lymphatic lumen, propel fluid through the lumen of the lymphatic circuit.

Interplay between Gut Lymphatic Vessels and Microbiota.

Lymphatic vessels play a distinctive role in draining fluid, molecules and even cells from interstitial and serosal spaces back to the blood circulation. Lymph vessels of the gut, and especially those located in the villi (called lacteals), not only serve this primary function, but are also responsible for the transport of lipid moieties absorbed by the intestinal mucosa and serve as a second line of defence against possible bacterial infections. Here, we briefly review the current knowledge of the general mechanisms allowing lymph drainage and propulsion and will focus on the most recent findings on the mutual relationship between lacteals and intestinal microbiota.

Tissue contribution to the mechanical features of diaphragmatic initial lymphatics.

The role of the mechanical properties of the initial lymphatic wall and of the surrounding tissue in supporting lymph formation and/or progression was studied in six anaesthetized, neuromuscularly blocked and mechanically ventilated rats. After mid-sternal thoracotomy, submesothelial initial lymphatics were identified on the pleural diaphragmatic surface through stereomicroscopy. An 'in vivo' lymphatic segment was prepared by securing two surgical threads around the vessel at a distance of ∼2.5 mm leaving the vessel in place. Two glass micropipettes were inserted into the lumen, one for intraluminar injections of 4.6 nl saline boluses and one for hydraulic pressure (Plymph) recording. The compliance of the vessel wall (Clymph) was calculated as the slope of the plot describing the change in segment volume as a function of the post-injection Plymph changes. Two superficial lymphatic vessel populations with a significantly different Clymph (6.7 ± 1.6 and 1.5 ± 0.4 nl mmHg−1 (mean ± S.E.M.), P < 0.001) were identified. In seven additional rats, the average elastic modulus of diaphragmatic tissue strips was determined by uniaxial tension tests to be 1.7 ± 0.3 MPa. Clymph calculated for an initial lymphatic completely surrounded by isotropic tissue was 0.068 nl mmHg−1, i.e. two orders of magnitude lower than in submesothelial lymphatics. Modelling of stress distribution in the lymphatic wall suggests that compliant vessels may act as reservoirs accommodating large absorbed fluid volumes, while lymphatics with stiffer walls serve to propel fluid through the lumen of the lymphatic vessel by taking advantage of the more efficient mechanical transmission of tissue stresses to the lymphatic lumen.

The contribution of cationic conductances to the potential of rod photoreceptors.

The contribution of cationic conductances in shaping the rod photovoltage was studied in light adapted cells recorded under whole-cell voltage- or current-clamp conditions. Depolarising current steps (of size comparable to the light-regulated current) produced monotonic responses when the prepulse holding potential (V (h)) was -40 mV (i.e. corresponding to the membrane potential in the dark). At V (h) = -60 mV (simulating the steady-state response to an intense background of light) current injections <35 pA (mimicking a light decrement) produced instead an initial depolarisation that declined to a plateau, and voltage transiently overshot V (h) at the stimulus offset. Current steps >40 pA produced a steady depolarisation to approximately -16 mV at both V (h). The difference between the responses at the two V (h) was primarily generated by the slow delayed-rectifier-like K(+) current (I (Kx)), which therefore strongly affects both the photoresponse rising and falling phase. The steady voltage observed at both V (h) in response to large current injections was instead generated by Ca-activated K(+) channels (I (KCa)), as previously found. Both I (Kx) and I (KCa) oppose the cation influx, occurring at the light stimulus offset through the cGMP-gated channels and the voltage-activated Ca(2+) channels (I (Ca)). This avoids that the cation influx could erratically depolarise the rod past its normal resting value, thus allowing a reliable dim stimuli detection, without slowing down the photovoltage recovery kinetics. The latter kinetics was instead accelerated by the hyperpolarisation-activated, non-selective current (I (h)) and I (Ca). Blockade of all K(+) currents with external TEA unmasked a I (Ca)-dependent regenerative behaviour.

Lymphatic Vessels and Their Surroundings: How Local Physical Factors Affect Lymph Flow.

Lymphatic vessels drain and propel lymph by exploiting external forces that surrounding tissues exert upon vessel walls (extrinsic mechanism) and by using active, rhythmic contractions of lymphatic muscle cells embedded in the vessel wall of collecting lymphatics (intrinsic mechanism). The latter mechanism is the major source of the hydraulic pressure gradient where scant extrinsic forces are generated in the microenvironment surrounding lymphatic vessels. It is mainly involved in generating pressure gradients between the interstitial spaces and the vessel lumen and between adjacent lymphatic vessels segments. Intrinsic pumping can very rapidly adapt to ambient physical stimuli such as hydraulic pressure, lymph flow-derived shear stress, fluid osmolarity, and temperature. This adaptation induces a variable lymph flow, which can precisely follow the local tissue state in terms of fluid and solutes removal. Several cellular systems are known to be sensitive to osmolarity, temperature, stretch, and shear stress, and some of them have been found either in lymphatic endothelial cells or lymphatic muscle. In this review, we will focus on how known physical stimuli affect intrinsic contractility and thus lymph flow and describe the most likely cellular mechanisms that mediate this phenomenon.

Acute Exposure of Collecting Lymphatic Vessels to Low-Density Lipoproteins Increases Both Contraction Frequency and Lymph Flow: An In Vivo Mechanical Insight.

Background: Lymphatic vessels drain fluids and solutes from interstitial spaces and serosal cavities. Among the solutes, low-density lipoproteins (LDL) are drained and can be detected in peripheral lymph, where they have been reported to exert a modulatory action on lymphatic vessels intrinsic contraction rate. In the present work, we investigated lymphatic vessel mechanical properties (contraction frequency and amplitude) that may be modulated by LDL application and the consequence on lymph flow. Methods and Results: Human-derived LDL were resuspended in phosphate-buffered saline (PBS) and microinjected in the interstitial space surrounding spontaneously contracting lymphatic vessels of the rat diaphragm, in vivo. Vessels' contraction rate and diameter were measured in control conditions (PBS) and after LDL injection. Lymph flow (Jlymph) was computed from contraction rate and diameter change. In some animals, after the recording procedure, diaphragmatic tissue samples were excised and immunostained with antilymphatic muscle (LM) actin to investigate the correlation between LM signal level and contraction amplitude. Data indicate a positive, saturating correlation between the abundance of LM actin and contraction amplitude, and LDL microinjection caused an acute increase in contraction frequency (+126%), a reduction of contraction amplitude to 75% of that obtained after PBS injection, and a +63% increase in Jlymph. Conclusions: From our in vivo analysis of the mechanical parameters affected by LDL, Jlymph was increased by a predominant effect on the contraction rate rather than amplitude, suggesting that the still elusive messaging system might be linked to the pacemaker sites.

Method for Studying the Physical Effect of Extracellular Matrix on Voltage-Dependent Ion Channel Gating.

Divalent cations can change the actual electrical potential at the outer surface of the plasma membrane. They do so by the so-called Gouy-Chapman-Stern effect which is due to the electrical "masking" that certain ions, especially divalents, can exert onto the electrically negative charged polar heads of the membrane phospholipids.Chondroitin sulfates can chelate free calcium ions to a different extent based on the spatial arrangement of their sulfate groups and can thus alter the actual availability of screening divalent ions at the outer membrane surface.Voltage-dependent ion channels sense the actual potential difference between the two sides of the plasma membrane and are thus exquisite and extremely sensitive "devices" able to react to changes in the electrical potential across the membrane.Hence, by recording the shift in the activation curve of well-known voltage-dependent ionic channels it will be possible to study the physical effect of ECM chondroitin sulfates on membrane conductances.

Gait screening of a population of young, healthy athletes by means of a portable, low-cost device unveils hidden left-right asymmetries in both quadriceps and anterior cruciate ligament forces.

OBJECTIVE: The present study reports the on-field screening of a population of young soccer players in the pursuit of alterations in gait using a portable and low-cost gait analysis system composed of a Wii Balance Board and a webcam. RESULTS: Recordings of motion of the lower extremities along with vertical ground reaction force (GRF) were used to quantify coefficients of symmetry for the overall GRF and the forces exerted by the quadriceps femori and acting on the anterior cruciate ligament (ACL). Data show that, in face of a quite homogeneous symmetry of GRF during left and right stance phases of gait, quadriceps and ACL exert and are subjected to left-right asymmetrical forces that might prelude, especially in young athletes, later alterations of gait.