Characterization of the cellular components of mouse collecting lymphatic vessels reveals that lymphatic muscle cells are the innate pacemaker cells regulating lymphatic contractions

Zawieja, S.D.; Pea, G.A.; Broyhill, S.E.; Patro, A.; Bromert, K.H.; Norton, C. E.; Kim, H. J.; Sivasankaran, S. K.; Li, M.; Castorena-Gonzalez, J.A.; Drumm, B.T.; Davis, M.J.

doi:10.1101/2023.08.24.554619

Cited by 3 publications

(1 citation statement)

References 130 publications

(176 reference statements)

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“…Next, we assessed the effects of GSK101 on the contractile function of lymphatic vessels from Myh11 -CreER T2 ; Ip3r1 fx/fx ; Ip3r2 fx/fx ; Ip3r3 fx/fx mice, lacking all three isoforms of Ip3Rs in smooth muscle cells. Lymphatic vessels from Myh11 -CreER T2 ; Ip3r1 fx/fx ; Ip3r2 fx/fx ; Ip3r3 fx/fx mice displayed a significantly lower contractile frequency than those from WT controls, this is in association with the critical role that Ip3r1 plays in the regulation of pressure-dependent chronotropy and contractility, as recently shown by our groups 57,62,63 . Upon stimulation with GSK101, lymphatic vessels lacking Ip3Rs in LMCs did not display robust and sustained constriction (Figure 12C).…”

Section: Resultssupporting

confidence: 75%

TRPV4-Expressing Tissue-Resident Macrophages Regulate the Function of Collecting Lymphatic Vessels via Thromboxane A2 Receptors in Lymphatic Muscle Cells

Schulz,

Akerstrom,

Song

et al. 2024

Preprint

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Rationale: TRPV4 channels are critical regulators of blood vascular function and have been shown to be dysregulated in many disease conditions in association with inflammation and tissue fibrosis. These are key features in the pathophysiology of lymphatic system diseases, including lymphedema and lipedema; however, the role of TRPV4 channels in the lymphatic system remains largely unexplored. TRPV4 channels are calcium permeable, non-selective cation channels that are activated by diverse stimuli, including shear stress, stretch, temperature, and cell metabolites, which may regulate lymphatic contractile function. Objective: To characterize the expression of TRPV4 channels in collecting lymphatic vessels and to determine the extent to which these channels regulate the contractile function of lymphatics. Methods and Results: Pressure myography on intact, isolated, and cannulated lymphatic vessels showed that pharmacological activation of TRPV4 channels with GSK1016790A (GSK101) led to contractile dysregulation. The response to GSK101 was multiphasic and included, 1) initial robust constriction that was sustained for ≥1 minute and in some instances remained for ≥4 minutes; and 2) subsequent vasodilation and partial or complete inhibition of lymphatic contractions associated with release of nitric oxide. The functional response to activation of TRPV4 channels displayed differences across lymphatics from four anatomical regions, but these differences were consistent across different species (mouse, rat, and non-human primate). Importantly, similar responses were observed following activation of TRPV4 channels in arterioles. The initial and sustained constriction was prevented with the COX inhibitor, indomethacin. We generated a controlled and spatially defined single-cell RNA sequencing (scRNAseq) dataset from intact and microdissected collecting lymphatic vessels. Our data uncovered a subset of macrophages displaying the highest expression of Trpv4 compared to other cell types within and surrounding the lymphatic vessel wall. These macrophages displayed a transcriptomic profile consistent with that of tissue-resident macrophages (TRMs), including differential expression of Lyve1, Cd163, Folr2, Mrc1, Ccl8, Apoe, Cd209f, Cd209d, and Cd209g; and at least half of these macrophages also expressed Timd4. This subset of macrophages also highly expressed Txa2s, which encodes the thromboxane A2 (TXA2) synthase. Inhibition of TXA2 receptors (TXA2Rs) prevented TRPV4-mediated contractile dysregulation. TXA2R activation on LMCs caused an increase in mobilization of calcium from intracellular stores through Ip3 receptors which promoted store operated calcium entry and vasoconstriction. Conclusions: Clinical studies have linked cancer-related lymphedema with an increased infiltration of macrophages. While these macrophages have known anti-inflammatory and pro-lymphangiogenic roles, as well as promote tissue repair, our results point to detrimental effects to the pumping capacity of collecting lymphatic vessels mediated by activation of TRPV4 channels in macrophages. Pharmacological targeting of TRPV4 channels in LYVE1-expressing macrophages or pharmacological targeting of TXA2Rs may offer novel therapeutic strategies to improve lymphatic pumping function and lymph transport in lymphedema.

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Section: Resultssupporting

confidence: 75%

TRPV4-Expressing Tissue-Resident Macrophages Regulate the Function of Collecting Lymphatic Vessels via Thromboxane A2 Receptors in Lymphatic Muscle Cells

Schulz,

Akerstrom,

Song

et al. 2024

Preprint

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Transcriptional, developmental, and functional parallels of lymphatic and venous smooth muscle

Arroyo-Ataz,

Yagüe,

Breda

et al. 2024

Preprint

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Lymphatic muscle cells (LMCs) are indispensable for lymphatic vessel contraction and their aberrant recruitment or absence is associated with both primary and secondary lymphedema. Despite their critical role in lymphatic vessel function, the transcriptomic and developmental basis that confer the unique contractile properties to LMCs are largely undefined. In this study, we employed single-cell RNA sequencing (scRNAseq), lineage tracing and in vivo imaging to investigate the basis for the hybrid cardiomyocyte and blood vascular smooth muscle cell (SMC) characteristics that have been described for LMCs. Using scRNAseq, the transcriptomes of LMC and venous SMCs from the murine hindlimb exhibited more similarities than differences, although both were markedly distinct from that of arteriole SMCs in the same tissue. Functionally, both lymphatic vessels and blood vessels in the murine hindlimb displayed pulsatile contractility. However, despite expressing genes that overlap with the venous SMC transcriptome, through lineage tracing we show that LMCs do not originate from Myh11+ SMC progenitors. Previous studies have shown that LMCs express cardiac-related genes, whereas in our study we found that arteriole SMCs, but not LMCs, expressed cardiac-related genes. Through lineage tracing, we demonstrate that a subpopulation of LMCs and SMCs originate from WT1+ mesodermal progenitors, which are known to give rise to SMCs. LMCs, however, do not derive from Nkx2.5+ cardiomyocyte progenitors. Overall, our findings suggest that venous SMCs and LMCs and may derive from a related mesodermal progenitor and adopt a similar gene expression program that enable their contractile properties.

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Cells and ionic conductances contributing to spontaneous activity in bladder and urethral smooth muscle

Drumm,

Gupta,

Mircea

et al. 2024

The Journal of Physiology

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Smooth muscle organs of the lower urinary tract comprise the bladder detrusor and urethral wall, which have a reciprocal contractile relationship during urine storage and micturition. As the bladder fills with urine, detrusor smooth muscle cells (DSMCs) remain relaxed to accommodate increases in intravesical pressure while urethral smooth muscle cells (USMCs) sustain tone to occlude the urethral orifice, preventing leakage. While neither organ displays coordinated regular contractions as occurs in small intestine, lymphatics or renal pelvis, they do exhibit patterns of rhythmicity at cellular and tissue levels. In rabbit and guinea‐pig urethra, electrical slow waves are recorded from USMCs. This activity is linked to cells expressing vimentin, c‐kit and Ca2+‐activated Cl− channels, like interstitial cells of Cajal in the gastrointestinal tract. In mouse, USMCs are rhythmically active (firing propagating Ca2+ waves linked to contraction), and this cellular rhythmicity is asynchronous across tissues and summates to form tone. Experiments in mice have failed to demonstrate a voltage‐dependent mechanism for regulating this rhythmicity or contractions in vitro, suggesting that urethral tone results from an intrinsic ability of USMCs to ‘pace’ their own Ca2+ mobilization pathways required for contraction. DSMCs exhibit spontaneous transient contractions, increases in intracellular Ca2+ and action potentials. Consistent across numerous species, including humans, this activity relies on voltage‐dependent Ca2+ influx in DSMCs. While interstitial cells are present in the bladder, they do not ‘pace’ the organ in an excitatory manner. Instead, specialized cells (PDGFRα+ interstitial cells) may ‘negatively pace’ DSMCs to prevent bladder overexcitability. image

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Characterization of the cellular components of mouse collecting lymphatic vessels reveals that lymphatic muscle cells are the innate pacemaker cells regulating lymphatic contractions

Cited by 3 publications

References 130 publications

TRPV4-Expressing Tissue-Resident Macrophages Regulate the Function of Collecting Lymphatic Vessels via Thromboxane A2 Receptors in Lymphatic Muscle Cells

TRPV4-Expressing Tissue-Resident Macrophages Regulate the Function of Collecting Lymphatic Vessels via Thromboxane A2 Receptors in Lymphatic Muscle Cells

Transcriptional, developmental, and functional parallels of lymphatic and venous smooth muscle

Cells and ionic conductances contributing to spontaneous activity in bladder and urethral smooth muscle

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