Light sheet microscopy-based 3-dimensional histopathology of the lymphatic vasculature in Emberger syndrome

Abstract

Introduction Lymphovascular diseases represent a heterogenous group of inherited and sporadic disorders and refer to a range of possible underlying pathologies and pathogenesis.Emberger Syndrome, an inherited form of lymphedema, is characterized by bilateral lower limb lymphedema, however, upper limbs do not show any signs of swelling.To identify disease-associated histopathological alterations in patients with Emberger Syndrome and to elucidate potential histological differences between the lymphatic vasculature of upper and lower limbs, a detailed knowledge on the 3-dimensional tissue and vessel architecture is essential. However, the current gold standard in 2-dimensional histology provides only very limited spatial information. Material and methods To elucidate the underlying vascular pathology in Emberger Syndrome on the cellular level, we applied the 3-dimensional visualization and analysis approach VIPAR (volume information-based histopathological analysis by 3D reconstruction and data extraction) to entire wholemount immunofluorescence-stained human tissue samples. VIPAR is a light sheet microscopy-based imaging technique, which allows 3-dimensional reconstruction of entire tissue biopsies followed by automated and semi-automated analysis of vascular parameters in 3-dimensional space. Results Using VIPAR we could show that in Emberger Syndrome the dermal lymphatic vasculature is intact and non-disrupted.However, lower limbs showed an hypoplastic lymphatic vasculature with absence of lymphatic valves in pre-collecting and collecting vessels. In contrast to the lower limbs, the lymphatic vasculature of the upper limbs showed no morphological alterations of lymphatic vessels and lymphatic valves compared to healthy controls. Discussion Based on the 3-dimensional histopathological analysis we were able to perform a detailed phenotyping of lymphatic vessels in the upper and lower limb in Emberger Syndrome and to identify the underlying vascular pathology. In addition, we could show vascular alteration between the upper and lower limbs indicating a vascular heterogeneity of dermal lymph vessels causing the lower limb lymphedema.

Pathogenic variants in MDFIC cause recessive central conducting lymphatic anomaly with lymphedema

“…The failure of lymphatic vessel valves to form or function in mice deficient in genes including Foxc2 and Gata2 , or harboring a pathogenic variant in Kras , results in lymphatic phenotypes including edema, chylous ascites, and chylothorax ( 14 , 23 , 26 , 28 ). Patients with primary lymphedema syndromes or lymphatic malformations caused by pathogenic variants in FOXC2 , GATA2 , or KRAS also exhibit lymphatic vessel valve defects and, in some cases, venous valve defects, underscoring the impact of valve defects on lymphatic function in human lymphatic disease ( 14 , 23 , 29 , 30 ).…”