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Isomaltulose is a low glycemic and insulinaemic carbohydrate increasingly used as an alternative sweetener in commercial beverages. While isomaltulose beverages can improve hydration status compared to sucrose-based beverages, it remains unclear if ingestion of an isomaltulose beverage prior to exercise in the heat may improve plasma volume (PV) and thermoregulatory responses.Twelve endurance-trained men consumed a 1L carbohydrate beverage containing either 6.5%-sucrose (SUC) or 6.5%-isomaltulose (ISO) 60 min prior to 5 successive, 15-min bouts of moderate-intensity (60% of their pre-determined maximum oxygen uptake) in the heat (32 °C, 50% relative humidity), each separated by a 5 min rest. A 6th bout was performed, wherein the participant adjusted running speed to maximize distance covered within the 15-min period. The change (Δ) in PV, heart rate (HR), body core (rectal and gastrointestinal) and skin temperatures, and whole-body sweat loss were assessed during each exercise bout.Ingestion of ISO induced a higher ΔPV at 4th bout only (P 0.001) and lower HR (P = 0.032, main effect of beverage) during exercise compared to those of SUC. Body core and skin temperatures and whole-body sweat loss did not differ between conditions (all P ≥ 0.192, interaction effect). Running distance covered in final exercise bout tended to increase (~ 5%) in ISO versus SUC (P = 0.057, d = 0.64).Relative to a sucrose-based beverage, ISO ingestion prior to exercise in the heat reduced cardiovascular strain by preserving PV and attenuating HR, albeit with no corresponding benefit on thermoregulatory function. The former response may facilitate improvements in exercise performance.

It is known that nitric oxide (NO) is a gas and synthesized from l-arginine by the NO synthase (NOS) in vascular endothelial cells. The diffused NO activates the guanosine monophosphate, which initiates a series of intracellular events, leading to physiological response such as vasodilation. There are three different types of NOS, namely endothelial constitutive NOS (ecNOS), neuronal NOS (nNOS), and cytokine-inducible NOS (iNOS). The ecNOS and nNOS are expressed constitutively at low levels and can be activated rapidly by an increase in cytoplasmic calcium ions. In contrast, the iNOS is induced when macrophages are activated by cytokine, resulting in the induction of pathophysiological effects. Lymph flow is known to stimulate the release of NO from lymphatic endothelial cells (LEC) and then produce the relaxation of lymphatic smooth muscle cells. The NO also plays a key role in the control of lymphatic pump activity

Mesenteric lymph has received attention for its important roles in nutrient absorption and mucosal immunity. It contains cytokines and immune cells that control the immune responses. The aim of this study to evaluate the effects of cocoa butter intake on rat mesenteric lymph flow, cell density, and the IL-1β, IL-6 and IL-10 levels in the lymph. Cocoa butter intake increased the lymph flow, cell density and the cytokine levels in the lymph. Clodronate released IL-1β, IL-6 and IL-10 and slightly accelerated the cocoa butter intake-mediated release of IL-6, but not that of IL-1β and IL-10. A Toll-like receptor (TLR) 2 and 4 inhibitor significantly reduced the linoleic acid-mediated releases of IL-1β, IL-6 and IL-10 in the medium of isolated cells from lamina propria of jejunal villi. Our findings suggest that cocoa butter intake regulates gut immunity through the release of IL-1β, IL-6 and IL-10 via a negative feedback mechanism.

Isomaltulose is a low glycemic and insulinaemic carbohydrate available as a constituent in sports drink. However, it remains unclear whether postexercise rehydration achieved by isomaltulose drink ingestion alone differs as compared to other carbohydrates. Thirteen young men performed intermittent exercise in the heat (35 °C and relative humidity 40%) to induce a state of hypohydration as defined by a 2% loss in body mass. Thereafter, participants were rehydrated by ingesting drinks equal to the volume of body mass loss with either a mixture of 3.25% glucose and 3.25% fructose, 6.5% sucrose (SUC), or 6.5% isomaltulose (ISO) within the first 30 min of a 3-h recovery. The change in plasma volume (ΔPV) from pre-exercise baseline, blood glucose, and plasma insulin concentration were assessed every 30-min. ΔPV was lower in ISO as compared to SUC until 90 min of the recovery (all P ≤ 0.038) with no difference thereafter (all P ≥ 0.391). The ΔPV were paralleled by concomitant changes in blood glucose levels that were greater in ISO as compared to other drinks after 90 min of the recovery (all P ≤ 0.035). Plasma insulin secretion, which potentially enhances renal sodium reabsorption and fluid retention, did not differ between the trials (interaction, P = 0.653). ISO induced a greater net fluid volume retention as compared to SUC (P = 0.010). We showed that rehydration with an isomaltulose drink following exercise-heat stress induces comparable recovery of PV and a greater net fluid retention as compared to other drinks, albeit this response is delayed. The delayed water transport along with glucose absorption may modulate this response. This trial was registered in 25th Sep 2019 at https://www.umin.ac.jp/ as UMIN000038099. (249/250)

We previously demonstrated that water intake increased mesenteric lymph flow and the total flux of IL-22 in rat jejunum. The drained water and the higher permeability of albumin in the jejunal microcirculation contributed to increase the lymph flow and IL-22 transport via the activation of great bulk flow in the jejunal villi. To address the effects of water intake-mediated great bulk flow-dependent mechanical force on jejunal physiological function and immunological regulation of innate lymphoid cells (ILC)-3, we examined the effects of shear stress stimulation on cultured rat myofibroblast cells. Next, we investigated the effects of water intake on podoplanin and IL-22 expressions in cultured human intestinal epithelial cells and rat in vivo jejunal preparations, respectively. Shear stress stimulation of the myofibroblast cells induced ATP release via an activation of cell surface F1/F0 ATP synthase. ATP produced podoplanin expression in the intestinal epithelial cells. Water intake accelerated immunohistochemical expressions of podoplanin and IL-22 in the interepithelial layers and lamina propria of the jejunum. ATP dose-dependently increased IL-22 mRNA expression in ILC-3, which are housed in the lamina propria. Water intake also increased immunohistochemical and mRNA expressions of ecto-nucleoside triphosphate diphosphohydrolases 2 and 5 in jejunal villi. In conclusion, water intake-mediated shear stress stimulation-dependent ATP release from myofibroblast cells maintains higher tissue colloid osmotic pressure in the jejunal microcirculation through podoplanin upregulation in the interepithelial layers. ATP induces IL-22 mRNA expression in ILC-3 in jejunal villi, which may contribute to regulation of mucosal immunity in small intestine.NEW & NOTEWORTHY We investigated effects of shear stress stimulation on cultured myofibroblast cells and water intake on podoplanin and IL-22 expressions in rat jejunal villi. The stimulation induced ATP release from the cells. Water intake accelerated podoplanin and IL-22 expression levels. ATP increased IL-22 mRNA expression in innate lymphoid cells (ILC)-3. Hence, water intake maintains higher osmotic pressure in the jejunal villi through ATP release and podoplanin upregulation. Water intake may regulate the mucosal immunity.

The higher permeability of the venules in jejunal microcirculation to albumin contributes to the increased mesenteric lymph formation. Recently, we demonstrated that water intake induced serotonin release from enterochromaffin cells in rat jejunum, serotonin of which circulated through the portal vein into blood circulation and then increased the mesenteric lymph formation. The mode of action of serotonin remains unclear. Therefore, we aimed to clarify the mechanisms involved in the regulation of the jejunal lymph formation with permeant albumin in in vivo rat experiments. We investigated the effects of intravenous administration of serotonin or water intake on the jejunal-originated lymph volume and the concentration of albumin in the lymph in the presence or absence of L-NAME. The effects of intravenous administration of L-NAME, nicardipine, A23187, and ML-7 on the lymph formation with permeant albumin were also evaluated. Serotonin or water intake significantly increased the mesenteric lymph volume with permeant albumin in the jejunal microcirculation. The serotonin- and water intake-mediated responses were significantly reduced by the pretreatment with intravenous administration of L-NAME. Intravenous administration of L-NAME itself also decreased significantly the jejunal lymph formation. Administration of A23187 and ML-7 significantly reduced the jejunal lymph formation with permeant albumin. In contrast, administration of nicardipine significantly increased the lymph formation. In conclusion, portal venous blood flow- or serotonin-mediated NO release from venular endothelial cells plays physiologically key roles in the lymph formation in rat jejunum via the extrusion of calcium ions and inactivation of MLCK in endothelial cells.

In this review, with our current studies we demonstrated medical evidence that water and food intake are useful for IL-22-related mucosal immunity-dependent maintenance of health care. The traditional Japanese health care practices recommend daily consumption of suitable volume of water. However, immunological mechanisms that support of the traditional practices are still unsolved. We focused on type 3 innate lymphoid cells (ILC3s), because the ILC3s are mainly housed in the lamina propria of the jejunum. IL-22 released from the ILC3 is transported through mesenteric lymph in collaboration with the albumin-mediated movement of consumed water. Thus, water intake-mediated upregulation of IL-22-dependent mucosal immunity contributes to the traditional Japanese health care practices. We also reviewed current studies that food intake-mediated increase in VIP-dependent neuronal activity in the small intestine and the food intake included with tryptophan-derived metabolites may accelerate the IL-22 in ILC3s-dependent mucosal immunity and then contribute in keeping health care.

The traditional Japanese health care custom recommends that a suitable volume of water is consumed. However, physiological and immunological mechanisms in support of this practice are unknown. Therefore, we conducted rat and rabbit in vivo experiments to investigate the effects of intragastric administration of distilled water on the jejunal-originated lymph flow and the concentrations and total flux of cells, albumin, long-chain fatty acids, and innate lymphoid cell 3 (ILC-3)-secreted interleukin-22 (IL-22) through mesenteric lymph vessels. The distribution and activity of ILC-3 in rat small intestine by water intake were evaluated using flow cytometry and RT-PCR. The intragastric administration of distilled water caused significant increases in rat mesenteric lymph flow and in the total flux of cells, albumin, long-chain fatty acids, and IL-22 through the lymph vessels. Intravenously injected Evans blue dye was rapidly transported into rabbit mesenteric lymph vessel and cisterna chyli. The distribution of ILC-3 and the expression of IL-22 mRNA were maximal in the lamina propria cells of the rat jejunum. No significant presence of ILC-3 in the lymph was observed in the control and under water intake conditions. In conclusion, the absorbed water in the jejunum is transported through mesenteric lymph vessels. The higher permeability of albumin in the jejunal microcirculation may play key roles in the transport of consumed water and the reservoir and transporter of long-chain fatty acids. Water intake also accelerates the transfer of IL-22 to the mesenteric lymph, which may contribute, in part, to maintaining and promoting the innate immunity in the body. NEW & NOTEWORTHY The higher permeability of albumin-mediated transport of water-soluble substances in mesenteric lymph vessels of the jejunum may have a large impact on the classic concept suggesting that water-soluble small molecules travel to the liver via the portal vein. ILC-3 is mainly housed in the lamina propria of the jejunum, especially its upper part. IL-22 released from the ILC-3 is also transported through mesenteric lymph in collaboration with the albumin-mediated movement of consumed water.

The present study aims to investigate the roles of water intake in serotonin production and release in rat jejunum. We evaluated the changes in concentrations of serotonin in the portal vein and mesenteric lymph vessel induced by the intragastric administration of distilled water. The density of granules in enterochromaffin cells and the immunoreactivity of serotonin in the jejunal villi were investigated before and after water intake. The effects of intravenous administration of serotonin and/or ketanserin on mesenteric lymph flow and concentrations of albumin and IL-22 in the lymph were also addressed. Water intake increased serotonin concentration in the portal vein, but not in the mesenteric lymph vessel. The flux of serotonin through the portal vein was significantly larger than that through the mesenteric lymph vessel. Water intake decreased the density of granules in the enterochromaffin cells and increased the immunoreactivity of serotonin in the jejunal villi. The intravenous administration of serotonin increased significantly mesenteric lymph flow and the concentrations of albumin and IL-22; both were significantly reduced by the intravenous pretreatment with ketanserin. We showed that serotonin released from enterochromaffin cells by water intake was mainly transported through the portal vein. Additionally, serotonin in blood was found to increase mesenteric lymph formation with permeant albumin in the jejunal villi via the activation of 5-HT2 receptor.

The present study aims to investigate the roles of water intake in serotonin production and release in rat jejunum. We evaluated the changes in concentrations of serotonin in the portal vein and mesenteric lymph vessel induced by the intragastric administration of distilled water. The density of granules in enterochromaffin cells and the immunoreactivity of serotonin in the jejunal villi were investigated before and after water intake. The effects of intravenous administration of serotonin and/or ketanserin on mesenteric lymph flow and concentrations of albumin and IL-22 in the lymph were also addressed. Water intake increased serotonin concentration in the portal vein, but not in the mesenteric lymph vessel. The flux of serotonin through the portal vein was significantly larger than that through the mesenteric lymph vessel. Water intake decreased the density of granules in the enterochromaffin cells and increased the immunoreactivity of serotonin in the jejunal villi. The intravenous administration of serotonin increased significantly mesenteric lymph flow and the concentrations of albumin and IL-22; both were significantly reduced by the intravenous pretreatment with ketanserin. We showed that serotonin released from enterochromaffin cells by water intake was mainly transported through the portal vein. Additionally, serotonin in blood was found to increase mesenteric lymph formation with permeant albumin in the jejunal villi via the activation of 5-HT

Isomaltulose is a low glycemic and insulinaemic carbohydrate now used as an alternative sweetener in beverages. However, it remains unclear if hydration status may be impacted differently with the consumption of beverages containing isomaltulose as compared to sucrose, a common beverage sweetener. Thirteen young adults (7 women) consumed 1 L of a carbohydrate beverage (with low electrolyte content) containing either 6.5%-sucrose, 6.5%-isomaltulose, or water within a 15 min period. For each beverage, beverage hydration index (BHI, a composite measure of fluid balance after consuming a test beverage relative to water) was calculated from urine volume produced over a 3 h period following ingestion of the carbohydrate beverages relative to water. The change in plasma volume (ΔPV), blood glucose, and lactate concentrations were assessed every 30 min post-beverage consumption. Isomaltulose ingestion attenuated urine production as compared to water and sucrose (P ≤ 0.005) over the 3 h post-ingestion period. However, no differences were observed between sucrose and water (P = 0.055). BHI was 1.53 ± 0.44 for isomaltulose (P ≤ 0.022 vs. sucrose and water) and 1.20±0.29 for sucrose (P = 0.210 vs. water). A transient reduction in ΔPV was observed following the ingestion of the isomaltulose beverage (at 30 min, P = 0.007 vs. sucrose). Thereafter, no differences in ΔPV between beverages were measured. Increases in blood glucose and lactate, indices of absorption and utility of glucose, were delayed in the isomaltulose as compared to sucrose beverage. In summary, we demonstrated a greater BHI with a carbohydrate-electrolyte beverage containing isomaltulose as compared to sucrose. This may in part be attributed to a delayed absorption of isomaltulose reducing diuresis.

Background: Previous animal studies have shown that intragastric administration of water can accelerate mesenteric lymph flow. Similarly, human studies have shown that abdominal breathing can induc...

Knowledge of the mechanisms by which aging affects contracting lymphatic vessels remains incomplete; therefore, the functional role of histamine in the reaction of aged lymphatic vessels to increases in flow remains unknown.We measured and analyzed parameters of lymphatic contractility in isolated and pressurized rat mesenteric lymphatic vessels (MLVs) obtained from 9- and 24-month Fischer-344 rats under control conditions and after pharmacological blockade of nitric oxide (NO) by Nω-Nitro-L-arginine methyl ester hydrochloride (L-NAME, 100 μM) or/and blockade of histamine production by α-methyl-DL-histidine dihydrochloride (α-MHD, 10 μM). We also quantitatively compared results of immunohistochemical labeling of the histamine-producing enzyme, histidine decarboxylase (HDC) in adult and aged MLVs. Our data provide the first demonstration of an increased functional role of histamine as an endothelial-derived relaxing factor in aged MLVs, which appears in parallel with the abolished role of NO in the reactions of these lymph vessels to increases in flow. In addition, we found an increased expression of HDC in endothelium of aged MLVs.Our findings provide the basis for better understanding of the processes of aging in lymphatic vessels and for setting new important directions for investigations of the aging-associated disturbances in lymph flow and the immune response.

New findings What is the central question of this study? What are the effects of isomaltulose, an ingredient in carbohydrate-electrolyte beverages to maintain glycaemia and attenuate the risk of dehydration during exercise heat stress, on postexercise rehydration and physiological heat loss responses? What is the main finding and its importance? Consumption of a 6.5% isomaltulose-electrolyte beverage following exercise heat stress restored hydration following a 2 h recovery as compared to a 2% solution or water only. While the 6.5% isomaltulose-electrolytes increased plasma volume and plasma osmolality, which are known to modulate postexercise heat loss, sweating and cutaneous vascular responses did not differ between conditions. Consequently, ingestion beverages containing 6.5% isomaltulose-electrolytes enhanced postexercise rehydration without affecting heat loss responses. Abstract Isomaltulose is a disaccharide carbohydrate widely used during exercise to maintain glycaemia and hydration. We investigated the effects of ingesting a beverage containing isomaltulose and electrolytes on postexercise hydration state and physiological heat loss responses. In a randomized, single-blind cross-over design, 10 young healthy men were hypohydrated by performing up to three 30 min successive moderate-intensity (50% heart rate reserve) bouts of cycling, each separated by 10 min, while wearing a water-perfusion suit heated to 45°C. The protocol continued until a 2% reduction in body mass was achieved. Thereafter, participants performed a final 15 min moderate-intensity exercise bout followed by a 2 h recovery. Following cessation of exercise, participants ingested a beverage consisting of (i) water only (Water), (ii) 2% isomaltulose (CHO-2%), or (iii) 6.5% isomaltulose (CHO-6.5%) equal to the volume of 2% body mass loss within the first 30 min of the recovery. Changes in plasma volume (ΔPV) after fluid ingestion were greater for CHO-6.5% compared with CHO-2% (120 min postexercise) and Water (90 and 120 min) (all P ≤ 0.040). Plasma osmolality remained elevated with CHO-6.5% compared with consumption of the other beverages at 30 and 90 min postexercise (all P ≤ 0.050). Urine output tended to be reduced with CHO-6.5% compared to other fluid conditions (main effect, P = 0.069). Rectal and mean skin temperatures, chest sweat rate and cutaneous perfusion did not differ between conditions (all P > 0.05). In conclusion, compared with CHO-2% and Water, consuming a beverage consisting of CHO-6.5% and electrolytes during recovery under heat stress enhances PV recovery without modulating physiological heat loss responses.

Isomaltulose is a low glycemic and insulinemic carbohydrate available as a constituent of sports drinks. However, it remains unclear whether thermoregulatory responses (sweating and cutaneous vasodilation) after isomaltulose drink ingestion differ from those of sucrose and water during exercise in a hot environment. Ten young healthy males consumed 10% sucrose, 10% isomaltulose, or water drinks. Thirty-five minutes after ingestion, they cycled for fifteen minutes at 75% peak oxygen uptake in a hot environment (30 °C, 40% relative humidity). Sucrose ingestion induced greater blood glucose concentration and insulin secretion at the pre-exercise state, compared with isomaltulose and/or water trials, with no differences during exercise in blood glucose. Change in plasma volume did not differ between the three trials throughout the experiment, but both sucrose and isomaltulose ingestions similarly increased plasma osmolality, as compared with water (main beverage effect, p = 0.040)—a key response that potentially delays the onset of heat loss responses. However, core temperature thresholds and slopes for heat loss responses were not different between the trials during exercise. These results suggest that ingestion of isomaltulose beverages induces low glycemic and insulinemic states before exercise but does not alter thermoregulatory responses during exercise in a hot environment, compared with sucrose or water.

To confirm our previous study that abdominal respiration has induced hemodilution in human subjects, we performed in-vivo experiments involving anesthetized rabbits. Fifteen 6- to 7-month-old male Japanese white rabbits were used in the animal experiments. Anesthesia was maintained with 2.5%-3.0% isoflurane under N

Objectives The knowledge of the basic principles of lymphatic function, still remains, to a large degree, rudimentary and will require significant research efforts. Recent studies of the physiology of the MLVs suggested the presence of an EDRF other than NO. In this study, we tested the hypothesis that lymphatic endothelium-derived histamine relaxes MLVs. Methods We measured and analyzed parameters of lymphatic contractility in isolated and pressurized rat MLVs under control conditions and after pharmacological blockade of NO by l-NAME (100 μM) or/and histamine production by α-MHD (10 μM). Effectiveness of α-MHD was confirmed immunohistochemically. We also used immunohistochemical labeling and Western blot analysis of the histamine-producing enzyme, HDC. In addition, we blocked HDC protein expression in MLVs by transient transfection with vivo-morpholino oligos. Results We found that only combined pharmacological blockade of NO and histamine production completely eliminates flow-dependent relaxation of lymphatic vessels, thus confirming a role for histamine as an EDRF in MLVs. We also confirmed the presence of HDC and histamine inside lymphatic endothelial cells. Conclusions This study supports a role for histamine as an EDRF in MLVs.

Until now, there has been no tool available to provide lymphatic researchers the ability to perform experiments in tissue explants containing lymphatic vessels under tissue position- and lymphatic lumen-controlled conditions.In this article we provide technical details and description of the method of using the newly developed and implemented the position- and lymphatic lumen-controlled tissue chambers to study live lymphatic vessels and surrounding tissues ex vivo. In this study, we, for the first time, performed detailed comparative analysis of the contractile and pumping activity of rat mesenteric lymphatic vessels (MLVs) situated within tissue explants mounted in new tissue chambers and isolated, cannulated, and pressurized rat MLVs maintained in isolated vessel setups. We found no significant differences of the effects of both transmural pressure- and wall shear stress sensitivities of MLVs in tissue chambers and isolated MLVs.We conclude that this new experimental tool, a position- and lymphatic lumen-controlled tissue chamber, allows precise investigation of lymphatic function of MLVs interacting with elements of the tissue microenvironment. This method provides an important new set of experimental tools to investigate lymphatic function.

OBJECTIVE We studied the effects of S1P on the diameter and spontaneous contraction of murine iliac collecting lymph vessels. METHODS The isolated lymph vessel was cannulated with two glass micropipettes and then pressurized to 4 cmH(2) O at the intraluminal pressure. The changes in lymph vessel diameter were measured using a custom-made diameter-detection device. Immunohistochemical studies were also performed to confirm S1P receptors on the lymph vessels. RESULTS S1P (10(-7) M) had no significant effect on the frequency or amplitude of the lymph vessels' spontaneous contractions. In contrast, S1P (10(-8) -10(-6) M) produced a concentration-related reduction in lymph vessel diameter (tonic contraction). Pretreatment with 10(-4) M l-NAME or 10(-5) M aspirin had no significant effect on the S1P-induced tonic contraction of the lymph vessels. To evaluate the intracellular signal transduction pathway responsible for the S1P-induced tonic contractions and their Ca(2+) -dependence, we investigated the effects of JTE013, VPC23019, U-73122, xestospongin C, and nifedipine on the S1P-induced tonic contractions. All of these inhibitors except VPC23019 and nifedipine significantly reduced the S1P-induced tonic contractions. S1P (5x10(-7) M) also induced significant tonic contractions in the lymph vessels that had been superfused with high K(+) Krebs-bicarbonate solution or Ca(2+) -free high K(+) Krebs solution containing 1 mM EGTA. S1P2 receptors were immunohistochemically detected in the lymph vessels. CONCLUSION These findings suggest that neither endogenous NO nor prostaglandins are involved in the S1P-induced tonic contraction of lymph vessels, which is mainly caused by Ca(2+) release from intracellular Ca(2+) stores through the activation of S1P2 and 1,4,5 IP(3) receptors.

We studied the effects of PDGF-BB on changes in the diameters of murine lymph vessels with or without intact endothelium. PDGF-BB induced dilation of the lymph vessels with endothelium. Pretreatment with l-NAME or removal of the endothelium caused a significant attenuation in the PDGF-BB-induced dilation. PDGF-BB also produced dose-related reduction of the diameters of the lymph vessels without endothelium. To evaluate intracellular signal transduction and Ca(2+) -dependence of the PDGF-BB-induced tonic contraction, we investigated the effects of imatinib, GW5074 (an inhibitor of Raf-1 kinase), U-73122 (an inhibitor of phospholipase C), and xestospongin C on the PDGF-BB-induced reduction responses. All of these inhibitors caused a significant attenuation in the PDGF-BB-induced reduction response that was significantly decreased by treatment with Ca(2+) -free Krebs-bicarbonate solution or nifedipine. Higher concentrations of PDGF-BB produced a marked reduction of lymph vessel diameter within both high K(+) Krebs-bicarbonate solution and Ca(2+) -free high K(+) Krebs solution containing 1mM EGTA. These findings suggest that PDGF-BB induced endothelium-dependent NO-mediated relaxation of lymphatic smooth muscles in murine lymph vessels. PDGF receptor β-mediated tonic contraction of the muscles through increased Ca(2+) influx through the membrane and the release of membrane-bound and intracellular Ca(2+) .

The role of lymphatic vessels is to transport fluid, soluble molecules, and immune cells to the draining lymph nodes. Here, we analyze how the aging process affects the functionality of the lymphatic collectors and the dynamics of lymph flow. Ultrastructural, biochemical, and proteomic analysis indicates a loss of matrix proteins, and smooth muscle cells in aged collectors resulting in a decrease in contraction frequency, systolic lymph flow velocity, and pumping activity, as measured in vivo in lymphatic collectors. Functionally, this impairment also translated into a reduced ability for in vivo bacterial transport as determined by time‐lapse microscopy. Ultrastructural and proteomic analysis also indicates a decrease in the thickness of the endothelial cell glycocalyx and loss of gap junction proteins in aged lymph collectors. Redox proteomic analysis mapped an aging‐related increase in the glycation and carboxylation of lymphatic's endothelial cell and matrix proteins. Functionally, these modifications translate into apparent hyperpermeability of the lymphatics with pathogen escaping from the collectors into the surrounding tissue and a decreased ability to control tissue fluid homeostasis. Altogether, our data provide a mechanistic analysis of how the anatomical and biochemical changes, occurring in aged lymphatic vessels, compromise lymph flow, tissue fluid homeostasis, and pathogen transport., Aging Cell, 14 (4), ISSN:1474-9718, ISSN:1474-9728, ISSN:1474-9726

We studied the effects of PDGF-BB on changes in the diameters of murine lymph vessels with or without intact endothelium. PDGF-BB induced dilation of the lymph vessels with endothelium. Pretreatment with l-NAME or removal of the endothelium caused a significant attenuation in the PDGF-BB-induced dilation. PDGF-BB also produced dose-related reduction of the diameters of the lymph vessels without endothelium. To evaluate intracellular signal transduction and Ca(2+) -dependence of the PDGF-BB-induced tonic contraction, we investigated the effects of imatinib, GW5074 (an inhibitor of Raf-1 kinase), U-73122 (an inhibitor of phospholipase C), and xestospongin C on the PDGF-BB-induced reduction responses. All of these inhibitors caused a significant attenuation in the PDGF-BB-induced reduction response that was significantly decreased by treatment with Ca(2+) -free Krebs-bicarbonate solution or nifedipine. Higher concentrations of PDGF-BB produced a marked reduction of lymph vessel diameter within both high K(+) Krebs-bicarbonate solution and Ca(2+) -free high K(+) Krebs solution containing 1mM EGTA. These findings suggest that PDGF-BB induced endothelium-dependent NO-mediated relaxation of lymphatic smooth muscles in murine lymph vessels. PDGF receptor β-mediated tonic contraction of the muscles through increased Ca(2+) influx through the membrane and the release of membrane-bound and intracellular Ca(2+) .

Herein we analyze how the aging process affects the structure and functionality of the lymphatic collectors (LCs) with reference to their ability to maintain pathogen clearance. Ultrastructural, biochemical and proteomic analysis indicated a loss of extracellular matrix proteins, an increase in protein oxidative modifications as well as activation of nuclear factor-κB signaling as sign of “low grade” inflammation in aged LCs. This resulted in a decrease in contractile and pumping activity of LCs, as measured in vivo. Functionally, this impairment also translated into a reduced ability for in vivo bacterial transport as determined by time-lapse microscopy. Ultrastructural and proteomic analysis also indicated a decrease in the thickness of the endothelial cell glycocalyx and loss of gap-junction proteins in aged LCs. Redox proteomic analysis mapped an aging-related increase in the glycation and carboxylation of endothelial cell glycocalyx structural proteins. Functionally, these modifications translated into higher ability of the pathogen to escape from aged LCs into the surrounding tissue. Altogether, our analysis mapped the complexity of the aging-related anatomical, biochemical and functional changes in LCs. The decreased ability to transport bacteria to the draining nodes, associated with increased bacterial escape in the surrounding tissue can contribute to the decreased ability of the immune system to clear pathogens in the elderly, as observed in immunosenescence.