Sequential MRI Evaluation of Lymphatic Abnormalities over the Course of Fontan Completion.

Purpose To evaluate lymphatic abnormalities before and after Fontan completion using noncontrast lymphatic imaging and relate findings with postoperative outcomes. Materials and Methods This study is a retrospective review of noncontrast T2-weighted lymphatic imaging performed at The Children's Hospital of Philadelphia from June 2012 to February 2023 in patients with single ventricle physiology. All individuals with imaging at both pre-Fontan and Fontan stages were eligible. Lymphatic abnormalities were classified into four types based on severity and location of lymphatic vessels. Classifications were compared between images and related to clinical outcomes such as postoperative drainage and hospitalization, lymphatic complications, heart transplant, and death. Results Forty-three patients (median age, 10 years [IQR, 8-11]; 20 [47%] boys, 23 [53%] girls) were included in the study. Lymphatic abnormalities progressed in 19 individuals after Fontan completion (distribution of lymphatic classifications: type 1, 23; type 2, 11; type 3, 6; type 4, 3 vs type 1, 10; type 2, 18; type 3, 10; type 4, 5; P = .04). Compared with individuals showing no progression of lymphatic abnormalities, those progressing to a high-grade lymphatic classification had longer postoperative drainage (median time, 9 days [IQR, 6-14] vs 17 days [IQR, 10-23]; P = .04) and hospitalization (median time, 13 days [IQR, 9-25] vs 26 days [IQR, 18-30]; P = .03) after Fontan completion and were more likely to develop chylothorax (12% [three of 24] vs 75% [six of eight]; P < .01) and/or protein-losing enteropathy (0% [0 of 24] vs 38% [three of eight]; P < .01) during a median follow-up of 8 years (IQR, 5-9). Progression to any type was not associated with an increased risk of adverse events. Conclusion The study demonstrated that lymphatic structural abnormalities may progress in select individuals with single ventricle physiology after Fontan completion, and progression of abnormalities to a high-grade classification was associated with worse postoperative outcomes. Keywords: Congenital Heart Disease, Glenn, Fontan, Lymphatic Imaging, Cardiovascular MRI Supplemental material is available for this article. Published under a CC BY 4.0 license.

Octreotide improves human lymphatic fluid transport a translational trial.

OBJECTIVES: Chylothorax is a complex condition and many different pharmacological agents have been tried as treatment. Octreotide is used off-label to treat chylothorax, but the efficacy of octreotide remains unclear. A decrease in lymph production is suggested as the mechanism. In this cross-over study, we explore the direct effect of octreotide on human lymphatic drainage. METHODS: Pre-clinical: the effect of octreotide on force generation was assessed during acute and prolonged drug incubation on human lymphatic vessels mounted in a myograph. Clinical: in a double-blinded, randomized, cross-over trial including 16 healthy adults, we administered either octreotide or saline as an intravenous infusion for 2.5 h. Near-infrared fluorescence imaging was used to examine spontaneous lymphatic contractions and lymph pressure in peripheral lymphatic vessels and plethysmography was performed to assess the capillary filtration rate, capillary filtration coefficient and isovolumetric pressures of the lower leg. RESULTS: Pre-clinical: human thoracic duct (n = 12) contraction rate was concentration-dependently stimulated by octreotide with a maximum effect at 10 and 100 nmol/l in the myograph chamber. Clinical: spontaneous lymphatic contractions and lymph pressure evaluated by near-infrared fluorescence did not differ between octreotide or placebo (P = 0.36). Plethysmography revealed similar capillary filtration coefficients (P = 0.057), but almost a doubling of the isovolumetric pressures (P = 0.005) during octreotide infusion. CONCLUSIONS: Octreotide stimulated lymphatic contractility in the pre-clinical setup but did not affect the spontaneous lymphatic contractions or lymph pressure in healthy individuals. Plethysmography revealed a doubling in the isovolumetric pressure. These results suggest that octreotide increases lymphatic drainage capacity in situations with high lymphatic afterload.

Lymphatic Function Decreases over Time in the Arms of Breast Cancer Patients following Treatment.

In patients with breast cancer-related lymphedema, distinct lymphatic patterns and changed lymphatic contractile function have been described, but it is unknown how these characteristics change over time and to what extent they appear before clinical edema is detectable. Recently, we described the lymphatic morphology and function in a cohort of breast cancer patients shortly after radiation therapy (RT). In the current study, we investigate lymphatic function and morphology in the same cohort after 1 year of follow-up. Methods: The study population consisted of 28 breast cancer patients investigated 12 months after adjuvant locoregional RT. Lymphatic contraction frequency (CF), propulsion velocity, and the morphology of lymphatic vessels in the upper extremities were described in vivo using near-infrared fluorescence imaging. Lymphatic stress test was performed using hyperthermia. Results: At 1 year after RT, (n = 28) 46% of the patients presented with lymphatic morphological abnormalities with a degree of dermal backflow and 21% had developed clinical breast cancer-related lymphedema. In the ipsilateral arm, CF was 23% lower than in the contralateral arm (P = 0.04). Since primary examination, CF in the ipsilateral arm decreased by 40% (P = 0.03), whereas no change was observed in the contralateral arm. During hyperthermia, the ipsilateral arms with lymphatic complications were not able to increase CF as the remaining subgroups. Conclusions: Lymphatic function in the ipsilateral arm deteriorated over time after adjuvant breast cancer therapy. Furthermore, the presence of abnormal torturous lymphatic vessels in asymptomatic arms appeared to be associated with weak lymphatic reserve pumping capacity.

Gravity and lymphodynamics.

The lymphatic system is compromised in different groups of patients. To recognize pathology, we must know what is healthy. We use Near-Infrared Fluorescence (NIRF) to assess peripheral lymphatic function in humans. We have shown that external factors such as exercise, hyperthermia, and pharmacological mediators influence the function of peripheral lymphatic vessels. In this study, we explored the impact on lymphatic vessels by the ever-present external factor-gravity. We used NIRF imaging to investigate the lymphatic changes to gravity. Gravity was assessed by changing body position from supine to standing. We extracted following lymphatic functional parameters: lymphatic packet propulsion frequency (contractions/min), velocity (cm/s), and pressure (mmHg). Raw data analysis was performed using a custom-written Labview program. All sequences were analyzed by two observers and interclass correlation scores were calculated. All statistical analysis was performed using RStudio Team (2021). RStudio: Integrated Development Environment for R. RStudio, PBC. Healthy participants (n = 17, 11 males, age 28.1 ± 2.6 years) were included. The lymphatic packet propulsion frequency at baseline was 0.5 ± 0.2 contractions/min and rose within 3 min significantly to a maximum of 1.2 ± 0.5 contractions/min during upright posture and remained significantly higher than the baseline lymphatic packet propulsion frequency after lying down again for up to 6 min. The lymph velocity was 1.5 ± 0.4 cm/s at baseline and changed in both directions and without a specific pattern at different points in time during standing. Lymph pressure was significantly higher while standing (mean increase 9 mmHg, CI: 2-15 mmHg). The ICC scores were 89.8% (85.9%-92.7%), 59.3% (46.6%-69.6%) and 89.4% (79.0%-94.8%) in lymphatic packet propulsion frequency (130 observations), velocity (125 observations), and pressure (30 observations), respectively. The lymphatic system responds within few minutes to gravitational changes by increasing lymphatic packet propulsion frequency and pressure.

Lymphatic Function in the Arms of Breast Cancer Patients-A Prospective Cohort Study.

BACKGROUND: Lymphedema is a highly feared complication of breast cancer treatment, but the underlying complex mechanisms are still unknown. Thus, we investigated the lymphatic morphology and contractility in the lymphatic vessels of arms of high-risk breast cancer patients treated for node-positive early breast cancer. METHODS: In this prospective cohort study 32 women treated for unilateral node-positive breast cancer were enrolled and studied 36 ± 23 days after loco-regional radiotherapy. Near-infrared fluorescence imaging was used to assess morphology and function of the superficial lymphatic vessels. Strain-gauge plethysmography was performed to evaluate the capillary filtration of fluid.Both arms were investigated, with the non-treated arm acting as control. The patients were questioned about the presence of lymphedema yearly and finally 574 ± 118 days after ended radiotherapy. RESULTS: Morphologically, 25% of the treated arms expressed lymphatic vessel abnormalities compared to the control arms (p = 0.0048). No difference in functional parameters (maximal pumping pressure, p = 0.20; contraction frequency, p = 0.63; contraction velocity, p = 0.55) was found between the treated and control arms. Patients who later developed lymphedema had a difference in velocity compared to those who did not develop lymphedema (p = 0.02). The capillary filtration rate was similar between the two arms (p = 0.18). CONCLUSIONS: Peripheral lymphatic vessels were morphologically changed in the ipsilateral arm in 25% of the patients and patients who later developed lymphedema showed an early increase in velocity. Other functional parameters and capillary filtration were unchanged in this early phase. These discrete changes might be early indicators of later development of lymphedema.

Functional lymphatic reserve capacity is depressed in patients with a Fontan circulation.

BACKGROUND: Lymphatic abnormalities play a role in effusions in individuals with a Fontan circulation. Recent results using near-infrared fluorescence imaging disclosed an increased contraction frequency of lymphatic vessels in Fontan patients compared to healthy controls. It is proposed that the elevated lymphatic pumping seen in the Fontan patients is necessary to maintain habitual interstitial fluid balance. Hyperthermia has previously been used as a tool for lymphatic stress test. By increasing fluid filtration in the capillary bed, the lymphatic workload and contraction frequency are increased accordingly. Using near-infrared fluorescence imaging, the lymphatic functional reserve capacity in Fontan patients were explored with a lymphatic stress test. METHODS: Fontan patients (n = 33) were compared to a group of 15 healthy individuals of equal age, weight, and gender. The function of the superficial lymphatic vessels in the lower leg during rest and after inducing hyperthermia was investigated, using near-infrared fluorescence imaging. RESULTS: Baseline values in the Fontan patients showed a 57% higher contraction frequency compared to the healthy controls (0.4 ± 0.3 min-1 vs. 0.3 ± 0.2 min-1 , p = 0.0445). After inducing stress on the lymphatic vessels with hyperthermia the ability to increase contraction frequency was decreased in the Fontan patients compared to the controls (0.6 ± 0.5 min-1 vs. 1.2 ± 0.8 min-1 , p = 0.0102). CONCLUSIONS: Fontan patients had a higher lymphatic contraction frequency during normal circumstances. In the Fontan patients, the hyperthermia response is dampened indicating that the functional lymphatic reserve capacity is depressed. This diminished reserve capacity could be part of the explanation as to why some Fontan patients develop late-onset lymphatic complications.

Diagnosis and Management of Lymphatic Disorders in Congenital Heart Disease.

PURPOSE OF REVIEW: Lymphatic disorders have received an increasing amount of attention over the last decade. Sparked primarily by improved imaging modalities and the dawn of lymphatic interventions, understanding, diagnostics, and treatment of lymphatic complications have undergone considerable improvements. Thus, the current review aims to summarize understanding, diagnostics, and treatment of lymphatic complications in individuals with congenital heart disease. RECENT FINDINGS: The altered hemodynamics of individuals with congenital heart disease has been found to profoundly affect morphology and function of the lymphatic system, rendering this population especially prone to the development of lymphatic complications such as chylous and serous effusions, protein-losing enteropathy and plastic bronchitis. Although improved, a full understanding of the pathophysiology and targeted treatment for lymphatic complications is still wanting. Future research into pharmacological improvement of lymphatic function and continued implementation of lymphatic imaging and interventions may improve knowledge, treatment options, and outcome for affected individuals.

Function of Upper Extremity Human Lymphatics Assessed by Near-Infrared Fluorescence Imaging.

Background: Knowledge of functional parameters that can be used for evaluation of upper extremity lymphatic function is limited. This study aims to evaluate near-infrared fluorescence (NIRF) imaging for evaluation of lymphatic function in the human arm. Methods and Results: Ten healthy male volunteers (age 25.7 ± 1.3 years) were included and examined at two occasions. The lymphatic functional frequency and velocity were examined at baseline, during hyperthermia and after exercise. In addition, the maximum pressure generated by contraction of the lymphatic vessels was estimated. The mean contraction frequency was found to be 0.9 ± 0.4/min, and the mean velocity of the propulsions was 1.1 ± 0.3 cm/s. The average maximal pressure generated by the contraction of the lymphatic vessels was 59 ± 12 mmHg. No significant difference in frequency, velocity, or pumping pressure was found between the two visits (p > 0.05). Local hyperthermia increased contraction frequency significantly, whereas exercise decreased frequency and increased propulsion velocity. Conclusions: The functional lymphatic parameters evaluated by NIRF imaging showed good repeatability with no significant difference between visits. Future examinations should be conducted with standardization of temperature and exercise, as these parameters were shown to alter lymphatic function.

Reduced Lymphatic Function Predisposes to Calcium Channel Blocker Edema: A Randomized Placebo-Controlled Clinical Trial.

Background: The current belief is that the calcium channel blocker (CCB)-induced edema is due to a preferential arterial over venous dilatation leading to increased fluid filtration. We challenged this conviction by measuring the lymphatic removal of interstitial fluid during chronic systemic treatment with the CCB, amlodipine. Lymphatic vessels could potentially be an off-target effect of the drugs and play a role in CCB edema. Methods and Results: Sixteen healthy postmenopausal women completed a 12-week double-blinded randomized placebo-controlled crossover trial. Lymphatic function was assessed by near-infrared fluorescence imaging. The lymphatic function during amlodipine treatment compared with placebo did not show any difference in pumping pressure (53.9 ± 13.9 mmHg vs. 54.7 ± 9.4 mmHg, p = 0.829), contraction frequency (0.4 ± 0.2/min vs. 0.4 ± 0.3/min, p = 0.932), refill time (440 ± 438 seconds vs. 442 ± 419 seconds, p = 0.990), or propagation velocity of lymph packets (18 ± 10 mm/s vs. 15 ± 7 mm/s, p = 0.124). However, the subjects who developed edema during CCB treatment had a 20% lower baseline lymphatic pumping pressure (48.9 ± 4.4 mmHg, n = 7) than the subjects not affected by treatment (59.1 ± 1.2 mmHg, n = 9, p = 0.025). Contraction frequency, refill time, and lymph packet velocity showed no differences in baseline values between the two groups. Conclusion: Our results suggest that CCB does not directly impair lymphatic function. However, our results show that a reduced lymphatic function predisposes to CCB edema, which may explain why some patients develop edema during treatment.

Morphology and Function of the Lymphatic Vasculature in Patients With a Fontan Circulation.

BACKGROUND: The Fontan procedure has revolutionized the treatment of univentricular hearts. However, it is associated with severe complications such as protein-losing enteropathy, plastic bronchitis, and peripheral edema that may involve the lymphatic circulation. We aimed to assess lymphatic function and morphology in patients with a univentricular circulation. METHODS: The functional state of lymphatic vessels in the lower extremities of patients with a Fontan circulation (n=10) was investigated using the novel technique near-infrared fluorescence imaging and compared with an age-, sex-, and weight-matched control group of healthy volunteers (n=10). The lymphatic morphology was described using T2-weighted magnetic resonance imaging, and microvascular permeability was estimated by strain gauge plethysmography. RESULTS: The Fontan patients had 17% lower lymphatic pumping pressure (50±3.1 mm Hg) compared with controls (60±2.8 mm Hg; P=0.0341) and a 62% higher contraction frequency (0.8±0.1 min-1) compared with the healthy controls (0.5±0.1 min-1; P=0.0432). Velocity by which the lymph is moved and refill time after manual emptying of the lymphatic vessels showed no differences between the 2 groups. The thoracic duct was elongated 10% ( P=0.0409) and with an abnormal course in the Fontan patients compared with normal. No difference in microvascular permeability was found between the 2 groups. CONCLUSIONS: Patients with a Fontan circulation have an impaired lymphatic pumping capacity and morphologically changed thoracic duct. Our results indicate a challenged lymphatic vasculature in the Fontan circulation and may play a role in the pathogenesis of the complications that are seen in Fontan patients. CLINICAL TRIAL REGISTRATION: URL: https://www.clinicaltrials.gov . Unique identifier: NCT03379805.

Spontaneous and α-adrenoceptor-induced contractility in human collecting lymphatic vessels require chloride.

Human lymphatic vessels are myogenically active and respond to sympathetic stimulation. The role of various cations in this behavior has recently been investigated, but whether the anion Cl- is essential is unclear. With ethical approval and informed consent, human thoracic duct and mesenteric lymphatic vessels were obtained from surgical patients. Spontaneous or norepinephrine-induced isometric force production from isolated vessels was measured by wire myography; the transmembrane Cl- gradient and Cl- channels were investigated by substitution of extracellular Cl- with the impermeant anion aspartate and inhibition of Cl- transport and channels with the clinical diuretics furosemide and bendroflumethiazide as well as DIDS and 5-nitro-2-(3-phenylpropylamino)benzoic acid. The molecular expression of Ca2+-activated Cl- channels was investigated by RT-PCR, and proteins were localized using immunoreactivity. Spontaneous and norepinephrine-induced contractility in human lymphatic vessels was highly abrogated after Cl- substitution with aspartate. About 100-300 µM DIDS or 5-nitro-2-(3-phenylpropylamino)benzoic acid inhibited spontaneous contractile behavior. Norepinephrine-stimulated tone was furthermore markedly abrogated by 200 µM DIDS. Furosemide lowered only spontaneous constrictions, whereas bendroflumethiazide had nonspecific inhibitory effects. Consistent expression of transmembrane member 16A [TMEM16A (anoctamin-1)] was found in both the thoracic duct and mesenteric lymphatic vessels, and immunoreactivity with different antibodies localized TMEM16A to lymphatic smooth muscle cells and interstitial cells. The significant change in contractile function observed with inhibitors and anion substitution suggests that Cl- movement over the plasma membrane of lymphatic myocytes is integral for spontaneous and α-adrenoceptor-evoked contractility in human collecting lymphatic vessels. Consistent detection and localization of TMEM16A to myocytes suggests that this channel could play a major functional role. NEW & NOTEWORTHY In this study, we report the first observations of Cl- being a critical ionic component of spontaneous and agonist-evoked contractility in human lymphatics. The most consistently expressed Ca2+-activated Cl- channel gene in the human thoracic duct and mesenteric lymphatic vessels appears to be transmembrane member 16A, suggesting that this channel plays a major role.

Spontaneous and Evoked Contractility of Human Intestinal Lymphatic Vessels.

BACKGROUND: Mesenteric lymphatic vessels (MLVs) from various animal species have been intensively studied. We aimed to establish the viability and basic contractile characteristics of human MLVs maintained in vitro and to determine the reactivity of MLVs with norepinephrine (NE) and substance P (SP) and to compare with the thoracic duct (TD). METHODS AND RESULTS: Isolated human lymphatic vessels were mounted on a wire myograph under isometric conditions and tension was recorded. The diameter-tension characteristics for MLVs were generated by stretching the vessels and stimulating with a 125 mM K+ solution containing 10 µM NE. The diameter-tension data generated for MLVs from two separate surgical patient groups were found to be similar: maximum active tension for MLVs occurred when the passive stretch corresponded to a transmural pressure of 22 mmHg. Subsequent experiments on human MLVs were performed by normalization with 22 mmHg as the equivalent target pressure. The majority of MLVs were responders (spontaneous activity and/or reactivity with 10 µM NE or 125 mM K+ solution). Nonresponders (16% of vessel segments) had significantly smaller inner diameters. MLVs responded consistently to NE (1 nM-10 µM) but the responsiveness of MLVs and TD to SP (0.1 nM-10 µM) was poor: TD reacted only with 10 µM SP, whereas MLVs were sensitive to nanomolar concentrations and the contractile response declined with higher concentrations. CONCLUSIONS: Under in vitro isometric conditions, human MLVs generate maximum tension when stretched to a passive level corresponding to 22 mmHg, and the majority of MLVs are responsive when normalized to this pressure. MLVs respond to NE and SP though NE produces a more consistent response in the concentration range tested.

Human lymphatic vessel contractile activity is inhibited in vitro but not in vivo by the calcium channel blocker nifedipine.

Calcium channel blockers (CCB) are widely prescribed anti-hypertensive agents. The commonest side-effect, peripheral oedema, is attributed to a larger arterial than venous dilatation causing increased fluid filtration. Whether CCB treatment is detrimental to human lymphatic vessel function and thereby exacerbates oedema formation is unknown. We observed that spontaneous lymphatic contractions in isolated human vessels (thoracic duct and mesenteric lymphatics) maintained under isometric conditions were inhibited by therapeutic concentrations (nanomolar) of the CCB nifedipine while higher than therapeutic concentrations of verapamil (micromolar) were necessary to inhibit activity. Nifedipine also inhibited spontaneous action potentials measured by sharp microelectrodes. Furthermore, noradrenaline did not elicit normal increases in lymphatic vessel tone when maximal constriction was reduced to 29.4 ± 4.9% of control in the presence of 20 nmol l(-1) nifedipine. Transcripts for the L-type calcium channel gene CACNA1C were consistently detected from human thoracic duct samples examined and the CaV1.2 protein was localized by immunoreactivity to lymphatic smooth muscle cells. While human lymphatics ex vivo were highly sensitive to nifedipine, this was not apparent in vivo when nifedipine was compared to placebo in a randomized, double-blinded clinical trial: conversely, lymphatic vessel contraction frequency was increased and refill time was faster despite all subjects achieving target nifedipine plasma concentrations. We conclude that human lymphatic vessels are highly sensitive to nifedipine in vitro but that care must be taken when extrapolating in vitro observations of lymphatic vessel function to the clinical situation, as similar changes in lymphatic function were not evident in our clinical trial comparing nifedipine treatment to placebo.