Reduced Lymphatic Reserve in Heart Failure With Preserved Ejection Fraction

Abstract

Background: Microvascular dysfunction plays an important role in the pathogenesis of heart failure with preserved ejection fraction (HFpEF). However, no mechanistic link between systemic microvasculature and congestion, a central feature of the syndrome, has yet been investigated.

Objectives: This study aimed to investigate capillary-interstitium fluid exchange in HFpEF, including lymphatic drainage and the potential osmotic forces exerted by any hypertonic tissue Na⁺ excess.

Methods: Patients with HFpEF and healthy control subjects of similar age and sex distributions (n = 16 per group) underwent: 1) a skin biopsy for vascular immunohistochemistry, gene expression, and chemical (water, Na⁺, and K⁺) analyses; and 2) venous occlusion plethysmography to assess peripheral microvascular filtration coefficient (measuring capillary fluid extravasation) and isovolumetric pressure (above which lymphatic drainage cannot compensate for fluid extravasation).

Results: Skin biopsies in patients with HFpEF showed rarefaction of small blood and lymphatic vessels (p = 0.003 and p = 0.012, respectively); residual skin lymphatics showed a larger diameter (p = 0.007) and lower expression of lymphatic differentiation and function markers (LYVE-1 [lymphatic vessel endothelial hyaluronan receptor 1]: p < 0.05; PROX-1 [prospero homeobox protein 1]: p < 0.001) compared with control subjects. In patients with HFpEF, microvascular filtration coefficient was lower (calf: 3.30 [interquartile range (IQR): 2.33 to 3.88] l × 100 ml of tissue^-1 × min^-1 × mm Hg^-1 vs. 4.66 [IQR: 3.70 to 6.15] μl × 100 ml of tissue^-1 × min^-1 × mm Hg^-1; p < 0.01; forearm: 5.16 [IQR: 3.86 to 5.43] l × 100 ml of tissue^-1 × min^-1 × mm Hg^-1 vs. 5.66 [IQR: 4.69 to 8.38] μl × 100 ml of tissue^-1 × min^-1 × mm Hg^-1; p > 0.05), in keeping with blood vascular rarefaction and the lack of any observed hypertonic skin Na⁺ excess, but the lymphatic drainage was impaired (isovolumetric pressure in patients with HFpEF vs. control subjects: calf 16 ± 4 mm Hg vs. 22 ± 4 mm Hg; p < 0.005; forearm 17 ± 4 mm Hg vs. 25 ± 5 mm Hg; p < 0.001).

Conclusions: Peripheral lymphatic vessels in patients with HFpEF exhibit structural and molecular alterations and cannot effectively compensate for fluid extravasation and interstitial accumulation by commensurate drainage. Reduced lymphatic reserve may represent a novel therapeutic target.

Keywords: edema; heart failure; interstitium; lymphatic; microcirculation; preserved ejection fraction; vascular rarefaction.

Graphical abstract

Figure 1

Chemical Analysis of the Skin (A, B) Water content (% of wet weight [WW]), Na⁺ content (mmol/g of tissue dry weight [DW]), Na⁺ concentration, and [Na⁺ + K⁺] concentration (mmol/l of tissue water) in the outer (epidermis and superficial dermis [ESD]) and the inner (deep dermis [DD]) layers of the skin biopsies. (C) Representative skin sections from a healthy control (HC) subject and a patient with heart failure with preserved ejection fraction (HFpEF) (picrosirius red staining). The dashed line identifies the cutting plane for separation of ESD and DD in all study samples, scale bars are at the bottom, and black-circled areas represent dermal fat, quantified in the sections as % of biopsy area and were more abundant in patients with HFpEF than in HC subjects. (B) Excess dermal fat parallels the reduced volume of distribution of water and Na⁺ and, accordingly, their reduced content observed in the DD of patients with HFpEF. All data in panels A–C are presented as mean ± SD; ∗ p < 0.05, ∗∗p < 0.01. (D) Association between Na and water content, including both ESD and DD values; the slope of the regression line was the same between study groups.

Figure 2

Skin Microvascular Anatomy and Gene Expression Skin microvascular anatomy assessed as density (number of vessels/mm² tissue), average cross-sectional area of the vessels (average size; μm²), and total stain⁺ area (expressed as percentage of the dermal area) of the vessels identified in the section. (A) Blood vessels (BVs) showed a reduction in number and total area⁺; (B) lymphatic vessels (LVs) were reduced in number but, on average, were larger in size in patients with HFpEF compared with HC subjects. (C) Representative sections, showing blood vessels (lectin⁺[red]) and lymphatic vessels (LYVE-1 [lymphatic vessel endothelial hyaluronan receptor 1] [green]); arrowheads indicated small terminal lymphatic vessels, reduced in HFpEF compared with HC subjects, and larger vessels appeared unaffected. Scale bar at bottom right = 200 μm. (D) Gene expressions, presented as ΔCt (the higher the value, the lower the gene expression; β-actin as housekeeping gene); colored bars on top to indicate blood (red), lymphatic (green), or mixed vascular specificity; x = automatically detected outlier (ROUT = 1%) (GraphPad Prism). All data are presented as mean ± SD; ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001. PROX-1 = prospero homeobox protein 1; VEGF = vascular endothelial growth factor; VEGFR = vascular endothelial growth factor receptor; other abbreviations as in Figure 1.

Figure 3

Microvascular Fluid Dynamics Net accumulation of interstitial fluid at different hydraulic pressures, at which cuffs were inflated to oppose venous drainage in the forearm (left) and the calf (right). Each regression line corresponds to a participant. The slope of the lines is the microvascular coefficient of filtration (see Table 2). Red lines (HFpEF) intersect the x-axis at lower pressures compared with blue lines (HC subjects; scale magnification below): the intersect indicates the threshold above which interstitial fluid accumulation starts to develop (isovolumetric pressure [P_i]). Data presented as mean ± SD; ∗∗p < 0.01, ∗∗∗p < 0.001. P_cuff = pressure applied to the cuff; other abbreviations as in Figure 1.

Central Illustration

Microvascular Fluid Dynamics and Reduced Lymphatic Reserve in HFpEF In healthy subjects, the fluid filtering out of the capillary bed of the blood vasculature (BV) is evenly balanced by commensurate fluid drainage by lymphatic vasculature (LV); as a result, the physiological amount of interstitial fluid is homeostatically preserved. In heart failure with preserved ejection fraction (HFpEF), the net fluid extravasation tends to be lower because of the reduced vascular surface available for fluid exchange (i.e., capillary rarefaction) and the possible diversion of blood flow toward arteriovenous shunts; however, drainage by the impaired lymphatic system is inadequate to meet demands and facilitates accumulation of interstitial fluid at lower venous pressures. Both anatomical and functional defects (light green) could explain the reduced lymphatic reserve.