Perioperative fluid therapy impairs lymphatic pump function in male rats

Physiological Reports · 2025-06-01 · 1 citations

Because of its life-saving benefits, perioperative IV fluid therapy remains a cornerstone of medical treatment. However, it also induces sustained edemagenic stress. The resulting persistent interstitial edema-excessive fluid accumulation in the interstitium-significantly delays recovery and worsens patient outcomes. Therefore, to gain a detailed understanding of the lymphatic functional consequences of perioperative fluid therapy, this study aimed to test the hypothesis that perioperative IV fluid therapy compromises lymphatic pump function within 3 days after major surgery. Following a midline laparotomy, animals received IV fluid therapy over 48 h during recovery (FLTP). Three days post-surgery, mesenteric lymphatic vessels from FLTP and sham surgery (CTRL) animals were isolated, and lymphatic pump function was assessed in vitro. The transmural pressure-pump flow and circumferential length-wall tension relationships of FLTP vessels were altered-contraction frequency and normalized pump flow and active and passive wall tensions were significantly lower than CTRL. In vessels from another group of animals with surgically produced mesenteric venous hypertension to induce sustained edemagenic stress, only the pressure-pump flow relationship was altered similarly to FLTP. These results demonstrate the detrimental effects of perioperative fluid therapy on lymphatic pumping, which is essential for restoring interstitial fluid pressure and resolving edema and inflammation.

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

The Journal of Physiology · 2024-05-05 · 3 citations

Abstract Ca 2+ signalling plays a crucial role in determining lymphatic muscle cell excitability and contractility through its interaction with the Ca 2+ ‐activated Cl − channel anoctamin 1 (ANO1). In contrast, the large‐conductance (BK) Ca 2+ ‐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 Ca 2+ ‐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 ( Myh11CreERT 2 ‐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. image 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.

Short-chain fatty acids of the intestinal microbiota reduce mesenteric lymphatic pumping

Physiology · 2023-05-01

Short-chain fatty acids (SCFAs; acetate (AC), propionate (PP), and butyrate (BT)) are the primary metabolites of the intestinal microbiota. In addition to being an epithelial energy source, SCFAs exert numerous local effects, including maintenance of the epithelial barrier integrity and protection against inflammation, and are commonly assumed beneficial. It is also well understood that mesenteric lymphatic function is vital for normal gut health; impaired pumping leads to interstitial accumulation of fluid and immune cells, inducing intestinal edema and inflammation. Mesenteric lymphatic vessels are exposed to significantly high concentrations of SCFAs, yet the acute effects of SCFAs on lymphatic function have remained unexplored. Following earlier reports of SCFAs-induced relaxation of vascular smooth and cardiac muscle, in the present study, we tested the hypothesis that AC, PP, and BT reduce lymphatic pumping.Following our recently reported methods, pumping was characterized in vitro using rat mesenteric lymphatic vessels. Briefly, isolated vessels were cannulated, perfused, and bathed with warmed APSS. Following 60-min initial equilibration, in spontaneously contracting vessels, the effects of acute treatment with AC (10 mM), PP (10 mM), BT (10 mM), or AC+PP+BT were determined. In an additional set of vessels, the effects of short-chain fatty acid receptor antagonists (GLPG0974, FFAR2 antagonist, 1 μM and ((R)-3-Hydroxybutyric acid, FFAR3 antagonist, 1 μM) on lymphatic acute responses to AC+PP+BT were determined. Lymphatic responses were characterized using contraction frequency (CF), normalized stroke volume (difference in diastolic and systolic volume normalized to passive volume, NSV), normalized pump flow (CF*NSV, NPF), and diastolic tone (difference in passive and diastolic diameters normalized to passive diameter, DT).Treatment with AC, PP, or BT alone decreased CF and NPF significantly. Accordingly, CF and NPF were also lowered significantly after the combined treatment with the three SCFAs. Nevertheless, both indices were completely restored after FFAR2 and FFAR3 inhibition. Changes in NSV and DT after treatment with AC, PP, or BT, however, were not significant.To our knowledge, the present study is the first to demonstrate adverse chronotropic effects of the three primary SCFAs produced by intestinal microbiota on lymphatic pumping. The lymphatic responses are consistent with reports of reduced heart rate and cardiac contractile indices more than the dilation of blood vessels induced by SCFAs. Further studies are needed to reconcile the apparent conflict between the beneficial effects of SCFAs and the damaging effects of impaired lymphatic pumping. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Interstitial edema

Small Animal Critical Care Medicine · 2023-01-01

Effects of Acetate on Mesenteric Lymphatic Pump Function

The FASEB Journal · 2022-05-01

Pump function of mesenteric lymphatic vessels (MLVs) is essential for active transport of intestinal lymph and lipids from the gut to the venous circulation. MLV pump function is also critical for transport of immune cells in the GI tract, which account for almost 70% of the body’s immune cells. Short‐chain fatty acids (SCFAs)—the metabolites of the intestinal microbiota—are absorbed rapidly across the intestinal epithelial barrier and their concentration reaches 1‐100 mM in the intestinal tissues. There is a growing interest in the effects SCFAs on blood vessels. Blood vessel studies using acetate, the most abundant SCFA in peripheral circulation, reported that vasodilation induced by acetate mediates the blood pressure lowering effects of acetate. Furthermore, nitric oxide (NO) pathway was identified to mediate the acetate‐induced vasodilation. MLVs are exposed to significantly higher SCFA concentrations (as high as those in the intestinal tissues) in the undiluted lymph compared to the blood vessels. However, the effects of SCFAs on lymphatic vessel function have yet to be investigated thoroughly. Therefore, the purpose of the present study was to evaluate our hypothesis that acetate induces lymphatic dilation and decreases lymphatic pump function. MLVs were isolated from male Sprague Dawley rats and cannulated and perfused with warm APSS. Transmural pressure was set to 3 cmH 2 O and MLVs were allowed to equilibrate. In spontaneously contracting vessels, changes in pump function in response to cumulative concentrations of acetate (10 ‐8 to 10 ‐2 M) were determined. In separate MLVs, pump function at three transmural pressures (3, 5 and 7 cmH 2 O; selected randomly) was evaluated before and after incubating the MLVs with acetate (10 mM). Additionally, in another set of MLVs, lymphatic pump function at three transmural pressures (3, 5 and 7 cmH 2 O; selected randomly) was evaluated before and after incubating the MLVs with N(ω)‐nitro‐L‐arginine methyl ester (L‐NAME; 100 µM) and again after incubating the MLVs with L‐NAME (100 µM) + acetate (10 mM). The dose‐response study revealed that acetate led to decrease in lymphatic contraction frequency and calculated active lymph flow in a dose‐dependent manner. IC50 of the dose‐response curve was 1mM and lymphatic pumping was completely abolished at 50 mM concentration. Lymphatic contraction frequency and calculated active lymph flow were significantly decreased at all transmural pressures. Lymphatic stroke volume normalized to passive volume and diastolic tone were not significantly altered by acetate. Blockade of the NO pathway with L‐NAME did not restore the acetate‐induced decrease in lymphatic contraction frequency at all transmural pressures. Consistent with the effects of acetate on the blood vessels, acetate decreased lymphatic pump function in a dose‐dependent manner. However, the findings suggest that the NO pathway, reported to mediate blood vessel responses to acetate, does not mediate lymphatic responses to acetate. Two different pathways mediating acetate‐induced vasodilation in blood and lymphatic vessels may enable regulation of blood or lymphatic vessel function independently.

In Vitro Irradiation of Lymphatic Vessels Enhances Lymphatic Pump Function

The FASEB Journal · 2022-05-01

Active lymphatic pumping is critical for propulsion of lymph against a pressure gradient as well as transport of lipids and immune cells to the central venous circulation. When lymphatic pumping is impaired, diminished lymph flow leads to interstitial edema, immune cell accumulation and sustained inflammation. However, a majority of radiation biology research has primarily focused on the effects of ionizing radiation (IR) on blood vessels. These studies reported that IR induces imbalance between vaso‐dilatory and ‐constrictive pathways and impairs vascular tone. Only a handful of the studies have investigated the acute effects of IR on lymphatic vessels. Furthermore, the studies evaluating IR‐induced changes in lymphatic tone and pumping in vivo are limited because, in addition to the intrinsic contractility of the lymphatic muscle, lymphatic tone and pumping in vivo is modulated by a variety of substances present in the luminal lymph and the surrounding interstitial fluid. Therefore, the goal of the present study was to characterize the effects of IR on lymphatic tone and pumping independent of the confounding effects of IR on the other tissues. Mesenteric lymphatic vessels (MLVs), isolated from male Sprague Dawley rats, were randomly divided into SHAM and IR groups and were placed in chilled APSS. MLVs from the IR group were then irradiated with X‐rays at 10 Gray Dose. Immediately after irradiation, MLVs from both groups were placed in chilled APSS until further use. MLVs were then cannulated and perfused with warm APSS. Lymphatic pumping was assessed in spontaneously contracting vessels. Role of nitric oxide (NO) and cyclooxygenase (COX) pathways was evaluated by assessing pump function in separate MLVs before and after incubation in N(ω)‐nitro‐L‐arginine methyl ester (L‐NAME; 100 µM) and Indomethacin (INDO; 10 µM). Evaluation of the lymphatic pumping in response to changes in transmural pressure revealed significantly increased contraction frequency and calculated active lymph flow in IR MLVs compared to those in SHAM MLVs. However, stroke volume normalized to passive volume and diastolic tone were not significantly different between the IR and SHAM MLVs. The increase in contraction frequency in IR MLVs was partially restored by the blockade of COX pathways with INDO. Blockade of NOS with L‐NAME, however, led to further increase in contraction frequency of IR MLVs, which was significantly higher than that in SHAM MLVs. Early studies investigating the IR effects on lymphatic endothelial cells reported that lymphatic endothelium is resistant to IR. Conflicting findings of the in vivo lymphatic studies have further confounded the insights into the lymphatic responses to IR. It has been widely believed that lymphatic vessels are unaffected by IR, and therefore, the role of the lymphatic system‐ the other half of the circulatory system, in radiation injury has been mostly ignored. However, it is evident from the findings of the present study that lymphatic vessels are indeed affected by IR. Furthermore, IR‐induced increase in contraction frequency observed in the present study suggests that lymphatic acute responses to IR differ from the blood vessel responses.

Prolonged Exposure to Intestinal Edemagenic Stress Decreases Mesenteric Lymphatic Contractility

The FASEB Journal · 2022-05-01

Mesenteric lymphatic vessels (MLVs) are the primary route for removal of excess intestinal interstitial fluid. Lymphatic pumping, the virtue of the intrinsic contractility of lymphatic muscle, is necessary for active lymph transport to the venous circulation and thus resolution of interstitial edema. Acute exposure to edemagenic stress has been reported to increase contractility and pumping of MLVs. However, our recent studies revealed that when exposure to edemagenic stress is prolonged, lymphatic contraction frequency decreases and pump failure ensues. It is unknown how the ability of lymphatic muscle to generate active tension is affected in these maladapted MLVs. Therefore, the purpose of the present study was to evaluate our hypothesis that prolonged exposure to edemagenic stress decreases lymphatic muscle contractility. Male Sprague Dawley rats underwent midline laparotomy with 3‐day exposure to either venous hypertension (VHT; induced by coarctation of superior mesenteric vein), fluid overload (HD; induced by daily IV infusion of 80 ml/kg lactated Ringer’s saline), or neither (SHAM; no additional intervention). In isometric biomechanics studies, MLVs were mounted on a wire myograph and perfused with warm APSS. Vessel rings were incrementally stretched, stimulated with substance P (1 µM) in 60 mM K + ‐APSS (SP K‐APSS), followed by wash out with APSS. This sequence was repeated and the following steady state tensions were recorded at each level of stretch: unstimulated total tension (passive stretch, in APSS), stimulated total tension (SP K‐APSS), and passive tension (Ca ++ ‐free APSS, at the end of the experiment). The circumferential length at the level of stretch yielding optimal delta tension (= stimulated total tension ‐ unstimulated total tension) for each vessel was identified as L max . Additionally, unstimulated active tension (= unstimulated total tension ‐ passive tension) and stimulated active tension (= stimulated total tension ‐ passive tension) were calculated. Vessels from VHT and SHAM groups yielded nearly identical length‐tension relationships. However, compared to SHAM and VHT vessels, HD vessels generated significantly lower tension across all indices: unstimulated total tension, stimulated total tension, passive tension, unstimulated active tension, and stimulated active tension. At optimal stretch (L max ), the delta tension was lower in HD vessels than SHAM or VHT vessels. Consistent with findings of the previous studies, prolonged exposure to intestinal edemagenic stress induced mesenteric lymphatic pump failure in VHT and HD vessels. However, lymphatic muscle contractility was reduced only in HD vessels. Taken together, these findings suggest that peri‐ and post‐oprative fluid therapy in major surgery patients receiving fluid therapy may lead to mesenteric lymphatic maladaptation. The consequential decrease in intestinal lymph drainage would then contribute to delayed resolution of intestinal edema, increased morbidity and extended hospital stays.

Dichotomous effects of in vivo and in vitro ionizing radiation exposure on lymphatic function

AJP Heart and Circulatory Physiology · 2022 · 8 citations

• Biology
• Chemistry
• Cell biology

Earlier studies leading to the common belief that lymphatic vessels are radioresistant either did not characterize lymphatic pumping, deemed necessary for the resolution of edema and inflammation, or did it in vivo. By characterizing pumping in vitro, the present study, for the first time, demonstrated that lymphatic pumping was impaired in vessels irradiated in vivo and enhanced in vessels irradiated in vitro. Furthermore, the pathways implicated in ionizing radiation-induced blood vessel damage did not mediate lymphatic responses.

P193 Audit of cough swab sampling techniques in children with cystic fibrosis

Journal of Cystic Fibrosis · 2021-01-01

Algebraic formulas characterizing an alternative to Guyton’s graphical analysis relevant for heart failure

American Journal of Physiology-Regulatory, Integrative and Comparative Physiology · 2021-02-18 · 1 citations

Although Guyton’s graphical analysis of cardiac output-venous return has become a ubiquitous tool for explaining how circulatory equilibrium emerges from heart-vascular interactions, this classical model relies on a formula for venous return that contains unphysiological assumptions. Furthermore, Guyton’s graphical analysis does not predict pulmonary venous pressure, which is a critical variable for evaluating heart failure patients’ risk of pulmonary edema. Therefore, the purpose of the present work was to use a minimal closed-loop mathematical model to develop an alternative to Guyton’s analysis. Limitations inherent in Guyton’s model were addressed by 1) partitioning the cardiovascular system differently to isolate left ventricular function and lump all blood volumes together, 2) linearizing end-diastolic pressure-volume relationships to obtain algebraic solutions, and 3) treating arterial pressures as constants. This approach yielded three advances. First, variables related to morbidities associated with left ventricular failure were predicted. Second, an algebraic formula predicting left ventricular function was derived in terms of ventricular properties. Third, an algebraic formula predicting flow through the portion of the system isolated from the left ventricle was derived in terms of mechanical properties without neglecting redistribution of blood between systemic and pulmonary circulations. Although complexities were neglected, approximations necessary to obtain algebraic formulas resulted in minimal error, and predicted variables were consistent with reported values.