Endothelial FoxM1 reactivates aging-impaired endothelial regeneration for vascular repair and resolution of inflammatory lung injury.

Aging is a major risk factor of high incidence and increased mortality of acute respiratory distress syndrome (ARDS). Here, we demonstrated that persistent lung injury and high mortality in aged mice after sepsis challenge were attributable to impaired endothelial regeneration and vascular repair. Genetic lineage tracing study showed that endothelial regeneration after sepsis-induced vascular injury was mediated by lung resident endothelial proliferation in young adult mice, whereas this intrinsic regenerative program was impaired in aged mice. Expression of forkhead box M1 (FoxM1), an important mediator of endothelial regeneration in young mice, was not induced in lungs of aged mice. Transgenic FOXM1 expression or in vivo endothelium-targeted nanoparticle delivery of the FOXM1 gene driven by an endothelial cell (EC)-specific promoter reactivated endothelial regeneration, normalized vascular repair and resolution of inflammation, and promoted survival in aged mice after sepsis challenge. In addition, treatment with the FDA-approved DNA demethylating agent decitabine was sufficient to reactivate FoxM1-dependent endothelial regeneration in aged mice, reverse aging-impaired resolution of inflammatory injury, and promote survival. Mechanistically, aging-induced Foxm1 promoter hypermethylation in mice, which could be inhibited by decitabine treatment, inhibited Foxm1 induction after sepsis challenge. In COVID-19 lung autopsy samples, FOXM1 was not induced in vascular ECs of elderly patients in their 80s, in contrast with middle-aged patients (aged 50 to 60 years). Thus, reactivation of FoxM1-mediated endothelial regeneration and vascular repair may represent a potential therapy for elderly patients with ARDS.

The Unexpected Protective Role of Thrombosis in Sepsis-Induced Inflammatory Lung Injury Via Endothelial Alox15.

Background: Patients with sepsis-induced acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) commonly suffer from severe pulmonary thrombosis, but clinical trials of anti-coagulant therapies in sepsis and ARDS patients have failed. ARDS patients with thrombocytopenia also exhibit increased mortality, and widespread pulmonary thrombosis is often seen in coronavirus disease 2019 (COVID-19) ARDS patients. Methods: Employing different amounts of microbeads to induce various levels of pulmonary thrombosis. Acute lung injury was induced by either lipopolysaccharide i.p. or cecal ligation and puncture. Endothelial cell (EC)-targeted nanoparticle coupled with CDH5 promoter was employed to delivery plasmid DNA expressing the CRISPR/Cas9 system for EC-specific gene knockout or expressing Alox15 for EC-specific overexpression. Additionally, thrombocytopenia was induced by genetic depletion of platelets using DTR Pf4Cre mice by breeding Pf4 Cre mice into the genetic background of DTR mice. Results: We show that while severe pulmonary thrombosis or thrombocytopenia augments sepsis-induced ALI, the induction of mild pulmonary thrombosis conversely reduces endothelial cell (EC) apoptosis, ALI, and mortality via sustained expression of endothelial arachidonate 15-lipoxygenase (Alox15). Endothelial Alox15 knockout via EC-targeted nanoparticle delivery of CRISPR/Cas9 plasmid DNA in adult mice abolished the protective impact of mild lung thrombosis. Conversely, overexpression of endothelial Alox15 inhibited the increases in ALI caused by severe pulmonary thrombosis. The clinical relevance of the findings was validated by the observation of reduced ALOX15-expressing ECs in lung autopsy samples of ARDS patients. Additionally, restoration of pulmonary thrombosis in thrombocytopenic mice also normalized endotoxemia-induced ALI. Conclusion: We have demonstrated that moderate levels of thrombosis protect against sepsis-induced inflammatory lung injury via endothelial Alox15. Overexpression of Alox5 inhibits severe pulmonary thrombosis-induced increase of ALI. Thus, activation of ALOX15 signaling represents a promising therapeutic strategy for treatment of ARDS, especially in sub-populations of patients with thrombocytopenia and/or severe pulmonary thrombosis.

Pulmonary Thrombosis Promotes Tumorigenesis via Myeloid Hypoxia-Inducible Factors.

Cancer patients have a greater risk of thrombosis than individuals without cancer. Conversely, thrombosis is a diagnostic predictor of cancer, but the mechanisms by which thrombosis promotes tumor propagation are incompletely understood. Our previous studies showed that hypoxia-inducible factors (HIF) 1α and HIF2α are stabilized in myeloid cells of murine thrombi. We also previously showed that pulmonary thrombosis increases the levels of HIF1α and HIF2α in murine lungs, enhances the levels of tumorigenic factors in the circulation, and promotes pulmonary tumorigenesis. In this study, we aimed to investigate the regulation of thrombosis-induced tumorigenesis by myeloid cell-specific HIFs (i.e., HIF1 and HIF2 in neutrophils and macrophages). Our in vitro studies showed that multiple tumorigenic factors are upregulated in the secretome of hypoxic versus normoxic neutrophils and macrophages, which promotes lung cancer cell proliferation and migration in a myeloid-HIF-dependent manner. Next, we used a mouse model of pulmonary microvascular occlusion to study the impact of pulmonary thrombosis and myeloid HIFs on lung tumorigenesis. Experiments on mice lacking either HIF1α or HIF2α in myeloid cells demonstrated that loss of either factor eliminates the advantage given to pulmonary tumor formation by thrombotic insult. The myeloid HIF-dependent and tumorigenic impact of pulmonary thrombosis on tumor burden may be partly driven by paracrine thymidine phosphorylase (TP), given that TP levels were increased by hypoxia in neutrophil and macrophage supernates, and that plasma TP levels were positively correlated with multiple measures of tumor progression in wild type mice but not myeloid cell-specific HIF1α or HIF2α knockout mice. These data together demonstrate the importance of thrombotic insult in a model of pulmonary tumorigenesis and the essential role of myeloid HIFs in mediating tumorigenic success.

Rabeprazole Promotes Vascular Repair and Resolution of Sepsis-Induced Inflammatory Lung Injury through HIF-1α.

There are currently no effective treatments for sepsis and acute respiratory distress syndrome (ARDS). The repositioning of existing drugs is one possible effective strategy for the treatment of sepsis and ARDS. We previously showed that vascular repair and the resolution of sepsis-induced inflammatory lung injury is dependent upon endothelial HIF-1α/FoxM1 signaling. The aim of this study was to identify a candidate inducer of HIF-1α/FoxM1 signaling for the treatment of sepsis and ARDS. Employing high throughput screening of a library of 1200 FDA-approved drugs by using hypoxia response element (HRE)-driven luciferase reporter assays, we identified Rabeprazole (also known as Aciphex) as a top HIF-α activator. In cultured human lung microvascular endothelial cells, Rabeprazole induced HIF1A mRNA expression in a dose-dependent manner. A dose-response study of Rabeprazole in a mouse model of endotoxemia-induced inflammatory lung injury identified a dose that was well tolerated and enhanced vascular repair and the resolution of inflammatory lung injury. Rabeprazole treatment resulted in reductions in lung vascular leakage, edema, and neutrophil sequestration and proinflammatory cytokine expression during the repair phrase. We next used Hif1a/Tie2Cre knockout mice and Foxm1/Tie2Cre knockout mice to show that Rabeprazole promoted vascular repair through HIF-1α/FoxM1 signaling. In conclusion, Rabeprazole is a potent inducer of HIF-1α that promotes vascular repair and the resolution of sepsis-induced inflammatory lung injury via endothelial HIF-1α/FoxM1 signaling. This drug therefore represents a promising candidate for repurposing to effectively treat severe sepsis and ARDS.

Silent Persistent Left Superior Vena Cava Right-to-Left Shunt as a Unique Cause of Recurrent Brain Abscesses.

We present a novel case of recurrent brain abscesses found to be the result of a silent congenital right-to-left extracardiac shunt, a persistent left superior vena cava draining into the left atrium. The patient's brain abscess was evacuated surgically and treated with antibiotics, and his shunt was subsequently repaired. The case suggests that attention should be paid to evaluation for shunt physiology allowing for bypass of the pulmonary circulation in those with recurrent brain abscesses.

Hypoxia-Inducible Factor Signaling in Inflammatory Lung Injury and Repair.

Inflammatory lung injury is characterized by lung endothelial cell (LEC) death, alveolar epithelial cell (AEC) death, LEC-LEC junction weakening, and leukocyte infiltration, which together disrupt nutrient and oxygen transport. Subsequently, lung vascular repair is characterized by LEC and AEC regeneration and LEC-LEC junction re-annealing, which restores nutrient and oxygen delivery to the injured tissue. Pulmonary hypoxia is a characteristic feature of several inflammatory lung conditions, including acute lung injury (ALI), acute respiratory distress syndrome (ARDS), and severe coronavirus disease 2019 (COVID-19). The vascular response to hypoxia is controlled primarily by the hypoxia-inducible transcription factors (HIFs) 1 and 2. These transcription factors control the expression of a wide variety of target genes, which in turn mediate key pathophysiological processes including cell survival, differentiation, migration, and proliferation. HIF signaling in pulmonary cell types such as LECs and AECs, as well as infiltrating leukocytes, tightly regulates inflammatory lung injury and repair, in a manner that is dependent upon HIF isoform, cell type, and injury stimulus. The aim of this review is to describe the HIF-dependent regulation of inflammatory lung injury and vascular repair. The review will also discuss potential areas for future study and highlight putative targets for inflammatory lung conditions such as ALI/ARDS and severe COVID-19. In the development of HIF-targeted therapies to reduce inflammatory lung injury and/or enhance pulmonary vascular repair, it will be vital to consider HIF isoform- and cell-specificity, off-target side-effects, and the timing and delivery strategy of the therapeutic intervention.

Emerging and Established Histological Techniques for the Analysis of Thrombosis in COVID-19 Lungs.

Coronavirus disease 2019 (COVID-19) is the potentially lethal disease that is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Patients with COVID-19 have an increased risk of thrombosis, but the role of thrombosis in the pathogenesis and progression of severe COVID-19 remains unclear. A better understanding of the contribution of thrombosis to the development and progression of COVID-19 could lead to the development of novel COVID-19 treatments. For this reason, established and emerging histological techniques have recently been used to analyze COVID-19 lungs quantitatively and visually and in two and three dimensions. The gold standard and novel state-of the-art histological techniques that have been used to assess thrombosis in COVID-19 lungs are described in this Mini Review.

Native valve endocarditis and pacemaker infection with Mycobacterium fortuitum.

Endocarditis and cardiac device infection due to Mycobacterium fortuitum is a rare entity in the hospital settings. We report a case of pacemaker infection and native valve endocarditis due to Mycobacterium fortuitum, which was associated with tricuspid valve vegetation. two days after admission with fever, chills, body aches and swelling around her pacemaker, the patient's pacing system was surgically removed. The patient was then discharged at day 16 after surgery and treated with a multidrug regimen of azithromycin, levofloxacin, imipenem/cilastatin, and amikacin for six weeks followed by trimethoprim/sulfamethoxazole plus doxycycline for a further three months.

Oridonin-Loaded Nanoparticles Inhibit Breast Cancer Progression Through Regulation of ROS-Related Nrf2 Signaling Pathway.

Oridonin (ORI) has been shown to inhibit tumor cell growth and proliferation in vitro, while its optimum anti-tumor activity in vivo is limited due to the poor aqueous solubility and bioavailability. In this study, to improve the bioavailability, we developed a nanoparticle-based drug delivery system to facilitate delivery of ORI to breast tumor. ORI was encapsulated in biodegradable nanoparticles (NPs) based on poly-lactic-co-glycolic acid (PLGA) and polyethylene glycol (PEG) to form ORI NPs (ORI-NPs). The resulting ORI-NPs exhibited a mean particle diameter of 100 nm and displayed an efficient cellular uptake by human breast cancer MCF-7 cells. Compared to free ORI that showed no effects on tumor cell proliferation, the ORI-NPs showed significant cytotoxicity and delayed endothelial cell migration, tube formation and angiogenesis. Pharmacokinetics studies showed that ORI-NPs significantly increased the half-life of ORI in the blood circulation. In the nude mouse xenograft model, ORI-NPs markedly inhibited tumor growth and angiogenesis, while ORI did not show any inhibitory effects on the growth of tumor xenografts. The mechanism experiments showed that the antitumor activity of ORI-NPs against breast cancer might be through ROS related Nrf2/HO-1 signaling pathway. Together, these results demonstrated that ORI-loaded PEG-PLGA NPs enhanced bioactivity and bioavailability in vivo over ORI, indicating that ORI-NPs may represent a promisingly effective candidate against breast cancer.

Endothelial cells in the pathogenesis of pulmonary arterial hypertension.

Pulmonary arterial hypertension (PAH) is a devastating disease that involves pulmonary vasoconstriction, small vessel obliteration, large vessel thickening and obstruction, and development of plexiform lesions. PAH vasculopathy leads to progressive increases in pulmonary vascular resistance, right heart failure and, ultimately, premature death. Besides other cell types that are known to be involved in PAH pathogenesis (e.g. smooth muscle cells, fibroblasts and leukocytes), recent studies have demonstrated that endothelial cells (ECs) have a crucial role in the initiation and progression of PAH. The EC-specific role in PAH is multi-faceted and affects numerous pathophysiological processes, including vasoconstriction, inflammation, coagulation, metabolism and oxidative/nitrative stress, as well as cell viability, growth and differentiation. In this review, we describe how EC dysfunction and cell signalling regulate the pathogenesis of PAH. We also highlight areas of research that warrant attention in future studies, and discuss potential molecular signalling pathways in ECs that could be targeted therapeutically in the prevention and treatment of PAH.

Cobalt oxide nanoparticle-synergized protein degradation and phototherapy for enhanced anticancer therapeutics.

How to enable protein degradation pathways including the autophagy-lysosome pathway (ALP) and the ubiquitin-proteasome system (UPS) to enhance the efficacy of anticancer treatments remains a substantial challenge. Cobalt oxide nanoparticles (Co3O4 NPs) have attracted interest in recent years for their potential use as a synergistic anticancer treatment, although their therapeutic mechanisms of action are still poorly understood. Here, we describe the synergistic use of Co3O4 NPs as an autophagy inhibitor, chemosensitizer and photosensitizer, which manipulate protein degradation pathways (ALP and UPS) and photothermal therapy for enhanced anticancer treatments both in vitro and in vivo. We show that Co3O4 NPs can induce autolysosome accumulation and lysosomal functions damage by inhibiting lysosomal proteolytic activity and reducing intracellular ATP levels. Notably, Co3O4 NPs can be combined with the proteasome inhibitor, Carfilzomib (Cfz), to promote the accumulation of autophagic substrates, protein ubiquitination, and endoplasmic reticulum stress, and in doing so, inhibit cancer progression. By taking advantage of their photothermal conversion efficiency, Co3O4 NPs can also serve as photothermal sensitizer, which synergistically enhances the anticancer efficacy of Cfz both in vitro and in vivo. In summary, we provide evidence of a nanomaterial-synergized, photothermal anticancer strategy that synergistically targets cancer cell survival pathways and may eventually serve to enhance the anticancer efficacy of established cancer therapeutics.

Mechanisms of Endothelial Regeneration and Vascular Repair and Their Application to Regenerative Medicine.

Endothelial barrier integrity is required for maintaining vascular homeostasis and fluid balance between the circulation and surrounding tissues and for preventing the development of vascular disease. Despite comprehensive understanding of the molecular mechanisms and signaling pathways that mediate endothelial injury, the regulatory mechanisms responsible for endothelial regeneration and vascular repair are incompletely understood and constitute an emerging area of research. Endogenous and exogenous reparative mechanisms serve to reverse vascular damage and restore endothelial barrier function through regeneration of a functional endothelium and re-engagement of endothelial junctions. In this review, mechanisms that contribute to endothelial regeneration and vascular repair are described. Targeting these mechanisms has the potential to improve outcome in diseases that are characterized by vascular injury, such as atherosclerosis, restenosis, peripheral vascular disease, sepsis, and acute respiratory distress syndrome. Future studies to further improve current understanding of the mechanisms that control endothelial regeneration and vascular repair are also highlighted.

Surviving and thriving in thrombosis research during a global pandemic: Experiences of a vascular scientist diagnosed with COVID-19.

The 2019-2020 COVID-19 outbreak resulted in widespread suffering along with major changes in the ways that researchers carry out their work. This article profiles the experiences of an early-career investigator in thrombosis research who worked through the COVID-19 pandemic and a COVID-19 diagnosis. The aims of this article are to normalize concern regarding COVID-19 in the research community, to provide a perspective on maintaining productivity during stay-at-home periods, and to discuss how the COVID-19 pandemic might alter common research practices in the future. While the COVID-19 outbreak was clearly disruptive and debilitating on a global level, some research practices that were heavily employed during the pandemic may continue to be utilized in scientific research for many years to come.

Cetuximab-Coated Thermo-Sensitive Liposomes Loaded with Magnetic Nanoparticles and Doxorubicin for Targeted EGFR-Expressing Breast Cancer Combined Therapy.

BACKGROUND: One major limitation of cancer chemotherapy is a failure to specifically target a tumor, potentially leading to side effects such as systemic cytotoxicity. In this case, we have generated a cancer cell-targeting nanoparticle-liposome drug delivery system that can be activated by near-infrared laser light to enable local photo-thermal therapy and the release of chemotherapeutic agents, which could achieve combined therapeutic efficiency. METHODS: To exploit the magnetic potential of iron oxide, we prepared and characterized citric acid-coated iron oxide magnetic nanoparticles (CMNPs) and encapsulated them into thermo-sensitive liposomes (TSLs). The chemotherapeutic drug, doxorubicin (DOX), was then loaded into the CMNP-TSLs, which were coated with an antibody against the epidermal growth factor receptor (EGFR), cetuximab (CET), to target EGFR-expressing breast cancer cells in vitro and in vivo studies in mouse model. RESULTS: The resulting CET-DOX-CMNP-TSLs were stable with an average diameter of approximately 120 nm. First, the uptake of TSLs into breast cancer cells increased by the addition of the CET coating. Next, the viability of breast cancer cells treated with CET-CMNP-TSLs and CET-DOX-CMNP-TSLs was reduced by the addition of photo-thermal therapy using near-infrared (NIR) laser irradiation. What is more, the viability of breast cancer cells treated with CMNP-TSLs plus NIR was reduced by the addition of DOX to the CMNP-TSLs. Finally, photo-thermal therapy studies on tumor-bearing mice subjected to NIR laser irradiation showed that treatment with CMNP-TSLs or CET-CMNP-TSLs led to an increase in tumor surface temperature to 44.7°C and 48.7°C, respectively, compared with saline-treated mice body temperature ie, 35.2°C. Further, the hemolysis study shows that these nanocarriers are safe for systemic delivery. CONCLUSION: Our studies revealed that a combined therapy of photo-thermal therapy and targeted chemotherapy in thermo-sensitive nano-carriers represents a promising therapeutic strategy against breast cancer.

Advances in Anti-Tumor Treatments Targeting the CD47/SIRPα Axis.

CD47 is an immunoglobulin that is overexpressed on the surface of many types of cancer cells. CD47 forms a signaling complex with signal-regulatory protein α (SIRPα), enabling the escape of these cancer cells from macrophage-mediated phagocytosis. In recent years, CD47 has been shown to be highly expressed by various types of solid tumors and to be associated with poor patient prognosis in various types of cancer. A growing number of studies have since demonstrated that inhibiting the CD47-SIRPα signaling pathway promotes the adaptive immune response and enhances the phagocytosis of tumor cells by macrophages. Improved understanding in this field of research could lead to the development of novel and effective anti-tumor treatments that act through the inhibition of CD47 signaling in cancer cells. In this review, we describe the structure and function of CD47, provide an overview of studies that have aimed to inhibit CD47-dependent avoidance of macrophage-mediated phagocytosis by tumor cells, and assess the potential and challenges for targeting the CD47-SIRPα signaling pathway in anti-cancer therapy.

Nasal Delivery of Hesperidin/Chitosan Nanoparticles Suppresses Cytokine Storm Syndrome in a Mouse Model of Acute Lung Injury.

The cytokine storm or cytokine storm syndrome (CSS) is associated with high mortality in patients with acute lung injury (ALI) and acute respiratory distress syndrome (ARDS), for example following sepsis or infectious diseases including COVID-19. However, there are no effective treatments for CSS-associated ALI or ALI/ARDS. Thus, there remains an urgent need to develop effective drugs and therapeutic strategies against CSS and ALI/ARDS. Nasal and inhaled drug delivery methods represent a promising strategy in the treatment of inflammatory lung disease as a result of their ability to improve drug delivery to lungs. Improving the nasal mucosa absorption of poorly water-soluble drugs with poor mucosa bioavailability to a therapeutically effective level is another promising strategy in the fight against ALI/ARDS. Here, chitosan nanoparticles loaded with hesperidin (HPD/NPs) were developed for nasal delivery of the anti-inflammatory HPD compound to inflammatory lungs. In vitro and in vivo, HPD/NPs exhibited enhanced cellular uptake in the inflammatory microenvironment compared with free HPD. In a mouse model of inflammatory lung disease, the HPD/NPs markedly inhibited lung injury as evidenced by reduced inflammatory cytokine levels and suppressed vascular permeability compared with free HPD. Collectively, our study demonstrates that nasal delivery of HPD/NPs suppresses CSS and ALI/ARDS in a murine model of inflammatory lung disease, and that nanoparticle-based treatment strategies with anti-inflammatory effects could be used to reduce CSS and ALI in patients with inflammatory lung injury.

The stimulation of thrombosis by hypoxia.

Thrombus formation is increased under conditions of hypoxia in animal models of thrombosis and in human populations, but current therapies for thrombosis do not directly target hypoxia-responsive signaling pathways. The vascular response to hypoxia is controlled primarily by the hypoxia-inducible transcription factors (HIFs), whose target genes include several factors that regulate thrombus formation. In this article, we review the HIF-dependent and HIF-independent signaling pathways that regulate thrombus formation under hypoxic conditions. A better understanding of hypoxia-induced thrombus formation could lead to the development of novel prophylactic therapies for thrombosis.

Hypoxia and HIF activation as a possible link between sepsis and thrombosis.

Risk factors for thrombosis include hypoxia and sepsis, but the mechanisms that control sepsis-induced thrombus formation are incompletely understood. A recent article published in Thrombosis Journal: (i) reviews the role of endothelial cells in the pathogenesis of sepsis-associated microthrombosis; (ii) describes a novel 'two-path unifying theory' of hemostatic discorders; and (iii) refers to hypoxia as a consequence of microthrombus formation in sepsis patients. The current article adds to this review by describing how sepsis and thrombus formation could be linked through hypoxia and activation of hypoxia-inducible transcription factors (HIFs). In other words, hypoxia and HIF activation may be a cause as well as a consequence of thrombosis in sepsis patients. While microthrombosis reduces microvascular blood flow causing local hypoxia and tissue ischemia, sepsis-induced increases in HIF1 activation could conversely increase the expression of coagulant factors and integrins that promote thrombus formation, and stimulate the formation of pro-thrombotic neutrophil extracellular traps. A better understanding of the role of cell-specific HIFs in thrombus formation could lead to the development of novel prophylactic therapies for individuals at risk of thrombosis.