Background: Mutations in PIEZO1 cause human lymphatic malformations. We have previously uncovered an Orai1-mediated mechanotransduction pathway that triggers lymphatic sprouting through Notch downregulation in response to fluid flow. However, the identity of its upstream mechanosensor remains unknown. This study aimed to identify and characterize the molecular sensor that translates the flow-mediated external signal to the Orai1-regulated lymphatic expansion. Methods: Various mutant mouse models, cellular, biochemical, and molecular biology tools, and a mouse tail lymphedema model were employed to elucidate the role of Piezo1 in flow-induced lymphatic growth and regeneration. Results: Piezo1 was found to be abundantly expressed in lymphatic endothelial cells. Piezo1 knockdown in cultured lymphatic endothelial cells inhibited the laminar flow-induced calcium influx and abrogated the flow-mediated regulation of the Orai1 downstream genes, such as Klf2 , Dtx1 , Dtx3L , and Notch1 , which are involved in lymphatic sprouting. Conversely, stimulation of Piezo1 activated the Orai1-regulated mechanotransduction in the absence of fluid flow. Piezo1-mediated mechanotransduction was significantly blocked by Orai1 inhibition, establishing the epistatic relationship between Piezo1 and Orai1. Lymphatic-specific conditional Piezo1 knockout largely phenocopied sprouting defects shown in Orai1- or Klf2- knockout lymphatics during embryo development. Postnatal deletion of Piezo1 induced lymphatic regression in adults. Ectopic Dtx3L expression rescued the lymphatic defects caused by Piezo1 knockout, affirming that the Piezo1 promotes lymphatic sprouting through Notch downregulation. Consistently, transgenic Piezo1 expression or pharmacological Piezo1 activation enhanced lymphatic sprouting. Finally, we assessed a potential therapeutic value of Piezo1 activation in lymphatic regeneration and found that a Piezo1 agonist, Yoda1, effectively suppressed postsurgical lymphedema development. Conclusions: Piezo1 is an upstream mechanosensor for the lymphatic mechanotransduction pathway and regulates lymphatic growth in response to external physical stimuli. Piezo1 activation presents a novel therapeutic opportunity for preventing postsurgical lymphedema. The Piezo1-regulated lymphangiogenesis mechanism offers a molecular basis for Piezo1-associated lymphatic malformation in humans.

BACKGROUND The impairment of cutaneous wound healing results in chronic, non-healing wounds that are caused by altered wound environment oxygenation, tissue injury, and permissive microbial growth. Current modalities for the treatment of these wounds inadequately address the complex changes involved in chronic wound pathogenesis. Consequently, stem cell therapies have emerged as a potential therapeutic modality to promote cutaneous regeneration through trophic and paracrine activity. AIM To investigate current literature regarding use of stem cell therapies for the clinical treatment of chronic, non-healing wounds. METHODS PubMed, EMBASE, Cochrane Library, Web of Science, and Scopus were queried with combinations of the search terms “mesenchymal stem cells,” “adult stem cells,” “embryonic stem cells,” “erythroid precursor cells,” “stem cell therapies,” and “chronic wounds” in order to find relevant articles published between the years of 2000 and 2019 to review a 20-year experience. Reference lists from the articles were reviewed to identify additional pertinent articles. Retrieved manuscripts (reviews, case reports/series, retrospective/prospective studies, and clinical trials) were evaluated by the authors for their depiction of clinical stem cell therapy use. Data were extracted from the articles using a standardized collection tool. RESULTS A total of 43 articles describing the use of stem cell therapies for the treatment of chronic wounds were included in this review. While stem cell therapies have been explored in in vitro and in vivo applications in the past, recent efforts are geared towards assessing their clinical role. A review of the literature revealed that adipose-derived stem cells, bone marrow-derived stem cells, bone marrow-derived mononuclear cells, epidermally-derived mesenchymal stem cells, fibroblast stem cells, keratinocyte stem cells, placental mesenchymal stem cells, and umbilical cord mesenchymal stem cells have all been employed in the treatment of chronic wounds of various etiologies. Most recently, embryonic stem cells have emerged as a novel stem cell therapy with the capacity for multifaceted germ cell layer differentiation. With the capacity for self-renewal and differentiation, stem cells can enrich existing cell populations in chronic wounds in order to overcome barriers impeding the progression of wound healing. Further, stem cell therapies can be utilized to augment cell engraftment, signaling and activity, and resultant patient outcomes. CONCLUSION Assessing observed clinical outcomes, potential for stem cell use, and relevant therapeutic challenges allows wound care stakeholders to make informed decisions regarding optimal treatment approaches for their patients’ chronic wounds.

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Soft tissue sarcomas are a heterogenous group of malignant tumors that represent approximately 1% of adult malignancies. Although these tumors occur throughout the body, the majority involved the lower extremity. Management may involve amputation but more commonly often includes wide local resection by an oncologic surgeon and involvement of a plastic surgeon for reconstruction of larger and more complex defects. Postoperative wound complications are challenging for the surgeon and patient but also impact management of adjuvant chemotherapy and radiation therapy. To explore risk factors for wound complications, we reviewed our single-institution experience of lower-extremity soft tissue sarcomas from April 2009 to September 2016. We identified 127 patients for retrospective review and analysis. The proportion of patients with wound complications in the cohort was 43.3%. Most notably, compared with patients without wound complications, patients with wound complications had a higher proportion of immediate reconstruction (34.5% vs 15.3%; P = 0.05) and a marginally higher proportion who received neoadjuvant radiation (30.9% vs 16.7%; P = 0.06).

Head and neck lymphedema (HNL) is a disfiguring disease affecting over 90% of patients treated for head and neck cancer. Animal models of lymphedema are used to test pharmacologic and microsurgical therapies; however, no animal model for HNL is described in the literature to date. In this study we describe the first reproducible rat model for HNL. Animals were subjected to two surgical protocols: (1) lymphadenectomy plus irradiation; and (2) sham surgery and no irradiation. Head and neck expansion was measured on post-operative days 15, 30 and 60. Magnetic resonance imaging (MRI) was acquired at the same time points. Lymphatic drainage was measured at day 60 via indocyanine green (ICG) lymphography, after which animals were sacrificed for histological analysis. Postsurgical lymphedema was observed 100% of the time. Compared to sham-operated animals, lymphadenectomy animals experienced significantly more head and neck swelling at all timepoints (P < 0.01). Lymphadenectomy animals had significantly slower lymphatic drainage for 6 days post-ICG injection (P < 0.05). Histological analysis of lymphadenectomy animals revealed 83% greater subcutis thickness (P = 0.008), 22% greater collagen deposition (P = 0.001), 110% greater TGFβ1+ cell density (P = 0.04), 1.7-fold increase in TGFβ1 mRNA expression (P = 0.03), and 114% greater T-cell infiltration (P = 0.005) compared to sham-operated animals. In conclusion, animals subjected to complete lymph node dissection and irradiation developed changes consistent with human clinical postsurgical HNL. This was evidenced by significant increase in all head and neck measurements, slower lymphatic drainage, subcutaneous tissue expansion, increased fibrosis, and increased inflammation compared to sham-operated animals.