Potential long-term effects of SARS-CoV-2 infection on the pulmonary vasculature: Multilayered cross-talks in the setting of coinfections and comorbidities.

The Coronavirus Disease 2019 (COVID-19) caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) and its sublineages pose a new challenge to healthcare systems worldwide due to its ability to efficiently spread in immunized populations and its resistance to currently available therapies. COVID-19, although targeting primarily the respiratory system, is also now well established that later affects every organ in the body. Most importantly, despite the available therapy and vaccine-elicited protection, the long-term consequences of viral infection in breakthrough and asymptomatic individuals are areas of concern. In the past two years, investigators accumulated evidence on how the virus triggers our immune system and the molecular signals involved in the cross-talk between immune cells and structural cells in the pulmonary vasculature to drive pathological lung complications such as endothelial dysfunction and thrombosis. In the review, we emphasize recent updates on the pathophysiological inflammatory and immune responses associated with SARS-CoV-2 infection and their potential long-term consequences that may consequently lead to the development of pulmonary vascular diseases.

Persistent circulation of soluble and extracellular vesicle-linked Spike protein in individuals with postacute sequelae of COVID-19.

SARS-CoV-2, the causative agent of COVID-19 disease, has resulted in the death of millions worldwide since the beginning of the pandemic in December 2019. While much progress has been made to understand acute manifestations of SARS-CoV-2 infection, less is known about post-acute sequelae of COVID-19 (PASC). We investigated the levels of both Spike protein (Spike) and viral RNA circulating in patients hospitalized with acute COVID-19 and in patients with and without PASC. We found that Spike and viral RNA were more likely to be present in patients with PASC. Among these patients, 30% were positive for both Spike and viral RNA; whereas, none of the individuals without PASC were positive for both. The levels of Spike and/or viral RNA in the PASC+ve patients were found to be increased or remained the same as in the acute phase; whereas, in the PASC-ve group, these viral components decreased or were totally absent. Additionally, this is the first report to show that part of the circulating Spike is linked to extracellular vesicles without any presence of viral RNA in these vesicles. In conclusion, our findings suggest that Spike and/or viral RNA fragments persist in the recovered COVID-19 patients with PASC up to 1 year or longer after acute SARS-CoV-2 infection.

Extracellular Vesicle TGF-ß1 Is Linked to Cardiopulmonary Dysfunction in Human Immunodeficiency Virus.

Extracellular vesicles (EVs) have emerged as important mediators in cell-cell communication; however, their relevance in pulmonary hypertension (PH) secondary to human immunodeficiency virus (HIV) infection is yet to be explored. Considering that circulating monocytes are the source of the increased number of perivascular macrophages surrounding the remodeled vessels in PH, this study aimed to identify the role of circulating small EVs and EVs released by HIV-infected human monocyte-derived macrophages in the development of PH. We report significantly higher numbers of plasma-derived EVs carrying higher levels of TGF-ß1 (transforming growth factor-ß1) in HIV-positive individuals with PH compared with individuals without PH. Importantly, levels of these TGF-ß1-loaded, plasma-derived EVs correlated with pulmonary arterial systolic pressures and CD4 counts but did not correlate with the Dl CO or viral load. Correspondingly, enhanced TGF-ß1-dependent pulmonary endothelial injury and smooth muscle hyperplasia were observed. HIV-1 infection of monocyte-derived macrophages in the presence of cocaine resulted in an increased number of TGF-ß1-high EVs, and intravenous injection of these EVs in rats led to increased right ventricle systolic pressure accompanied by myocardial injury and increased levels of serum ET-1 (endothelin-1), TNF-α, and cardiac troponin-I. Conversely, pretreatment of rats with TGF-ß receptor 1 inhibitor prevented these EV-mediated changes. Findings define the ability of macrophage-derived small EVs to cause pulmonary vascular modeling and PH via modulation of TGF-ß signaling and suggest clinical implications of circulating TGF-ß-high EVs as a potential biomarker of HIV-associated PH.

NADPH oxidase-mediated endothelial injury in HIV- and opioid-induced pulmonary arterial hypertension.

We previously demonstrated that the combined exposure of human pulmonary microvascular endothelial cells (HPMECs) to morphine and viral protein(s) results in the oxidative stress-mediated induction of autophagy, leading to shift in the cells from early apoptotic to apoptosis-resistant proliferative status associated with the angioproliferative remodeling observed in pulmonary arterial hypertension (PAH). In this study, we tried to delineate the major source of HIV-1 protein Tat and morphine induced oxidative burst in HPMECs and its consequences on vascular remodeling and PAH in an in vivo model. We observed switch from the initial increased expression of NADPH oxidase (NOX) 2 in response to acute treatment of morphine and HIV-Tat to later increased expression of NOX4 on chronic treatment in the endoplasmic reticulum of HPMECs without any alterations in the mitochondria. Furthermore, NOX-dependent induction of autophagy was observed to play a pivotal role in regulating the endothelial cell survival. Our in vivo findings showed significant increase in pulmonary vascular remodeling, right ventricular systolic pressure, and Fulton index in HIV-transgenic rats on chronic administration of morphine. This was associated with increased oxidative stress in lung tissues and rat pulmonary microvascular endothelial cells. Additionally, endothelial cells from morphine-treated HIV-transgenic rats demonstrated increased expression of NOX2 and NOX4 proteins, inhibition of which ameliorated their increased survival upon serum starvation. In conclusion, this study describes NADPH oxidases as one of the main players in the oxidative stress-mediated endothelial dysfunction on the dual hit of HIV-viral protein(s) and opioids.

Extracellular vesicles: novel communicators in lung diseases.

The lung is the organ with the highest vascular density in the human body. It is therefore perceivable that the endothelium of the lung contributes significantly to the circulation of extracellular vesicles (EVs), which include exosomes, microvesicles, and apoptotic bodies. In addition to the endothelium, EVs may arise from alveolar macrophages, fibroblasts and epithelial cells. Because EVs harbor cargo molecules, such as miRNA, mRNA, and proteins, these intercellular communicators provide important insight into the health and disease condition of donor cells and may serve as useful biomarkers of lung disease processes. This comprehensive review focuses on what is currently known about the role of EVs as markers and mediators of lung pathologies including COPD, pulmonary hypertension, asthma, lung cancer and ALI/ARDS. We also explore the role EVs can potentially serve as therapeutics for these lung diseases when released from healthy progenitor cells, such as mesenchymal stem cells.

HIV-Nef Protein Persists in the Lungs of Aviremic Patients with HIV and Induces Endothelial Cell Death.

It remains a mystery why HIV-associated end-organ pathologies persist in the era of combined antiretroviral therapy (ART). One possible mechanism is the continued production of HIV-encoded proteins in latently HIV-infected T cells and macrophages. The proapoptotic protein HIV-Nef persists in the blood of ART-treated patients within extracellular vesicles (EVs) and peripheral blood mononuclear cells. Here we demonstrate that HIV-Nef is present in cells and EVs isolated from BAL of patients on ART. We hypothesize that HIV-Nef persistence in the lung induces endothelial apoptosis leading to endothelial dysfunction and further pulmonary vascular pathologies. The presence of HIV-Nef in patients with HIV correlates with the surface expression of the proapoptotic endothelial-monocyte-activating polypeptide II (EMAPII), which was implicated in progression of pulmonary emphysema via mechanisms involving endothelial cell death. HIV-Nef protein induces EMAPII surface expression in human embryonic kidney 293T cells, T cells, and human and mouse lung endothelial cells. HIV-Nef packages itself into EVs and increases the amount of EVs secreted from Nef-expressing T cells and Nef-transfected human embryonic kidney 293T cells. EVs from BAL of HIV+ patients and Nef-transfected cells induce apoptosis in lung microvascular endothelial cells by upregulating EMAPII surface expression in a PAK2-dependent fashion. Transgenic expression of HIV-Nef in vascular endothelial-cadherin+ endothelial cells leads to lung rarefaction, characterized by reduced alveoli and overall increase in lung inspiratory capacity. These changes occur concomitantly with lung endothelial cell apoptosis. Together, these data suggest that HIV-Nef induces endothelial cell apoptosis via an EMAPII-dependent mechanism that is sufficient to cause pulmonary vascular pathologies even in the absence of inflammation.

Macrophage-derived extracellular vesicles mediate smooth muscle hyperplasia: role of altered miRNA cargo in response to HIV infection and substance abuse.

Our previous studies consistently demonstrate enhanced pulmonary vascular remodeling in HIV-infected intravenous drug users, and in simian immunodeficiency virus-infected macaques or HIV-transgenic rats exposed to opioids or cocaine. Although we reported an associated increase in perivascular inflammation, the exact role of inflammatory cells in the development of pulmonary vascular remodeling remains unknown. In this study, HIV-infected and cocaine (H+C)-treated human monocyte derived macrophages released a higher number of extracellular vesicles (EVs), compared to HIV-infected or uninfected cocaine-treated macrophages, with a significant increase in the particle size range to 100-150 nm. Treatment of primary human pulmonary arterial smooth muscle cells (HPASMCs) with these EVs resulted in a significant increase in smooth muscle proliferation. We also observed a significant increase in the miRNA-130a level in the EVs derived from H+C-treated macrophages that corresponded with the decrease in the expression of phosphatase and tensin homolog and tuberous sclerosis 1 and 2 and activation of PI3K/protein kinase B signaling in HPASMCs on addition of these EVs. Transfection of HPASMCs with antagomir-130a-ameliorated the EV-induced effect. Thus, we conclude that EVs derived from H+C-treated macrophages promote pulmonary smooth muscle proliferation by delivery of its prosurvival miRNA cargo, which may play a crucial role in the development of PAH.-Sharma, H., Chinnappan, M., Agarwal, S., Dalvi, P., Gunewardena, S., O'Brien-Ladner, A., Dhillon, N. K. Macrophage-derived extracellular vesicles mediate smooth muscle hyperplasia: role of altered miRNA cargo in response to HIV infection and substance abuse.

Drug-induced pulmonary arterial hypertension: a primer for clinicians and scientists.

Drug-induced pulmonary arterial hypertension (D-PAH) is a form of World Health Organization Group 1 pulmonary hypertension (PH) defined by severe small vessel loss and obstructive vasculopathy, which leads to progressive right heart failure and death. To date, 16 different compounds have been associated with D-PAH, including anorexigens, recreational stimulants, and more recently, several Food and Drug Administration-approved medications. Although the clinical manifestation, pathology, and hemodynamic profile of D-PAH are indistinguishable from other forms of pulmonary arterial hypertension, its clinical course can be unpredictable and to some degree dependent on removal of the offending agent. Because only a subset of individuals develop D-PAH, it is probable that genetic susceptibilities play a role in the pathogenesis, but the characterization of the genetic factors responsible for these susceptibilities remains rudimentary. Besides aggressive treatment with PH-specific therapies, the major challenge in the management of D-PAH remains the early identification of compounds capable of injuring the pulmonary circulation in susceptible individuals. The implementation of pharmacovigilance, precision medicine strategies, and global warning systems will help facilitate the identification of high-risk drugs and incentivize regulatory strategies to prevent further outbreaks of D-PAH. The goal for this review is to inform clinicians and scientists of the prevalence of D-PAH and to highlight the growing number of common drugs that have been associated with the disease.

Effect of Cocaine on Pulmonary Vascular Remodeling and Hemodynamics in Human Immunodeficiency Virus-Transgenic Rats.

Human immunodeficiency virus (HIV)-related pulmonary arterial hypertension has been found to be more prevalent in intravenous drug users. Our earlier cell-culture findings reported down-regulation of bone morphogenetic protein receptors (BMPRs) in combination with enhanced proliferation of human pulmonary arterial smooth muscle cells (PASMCs) in the presence of HIV-Trans-activator of transcription (Tat) and cocaine compared with either treatment alone. Here, we report physiologic evidence of significant increases in mean pulmonary arterial pressure in HIV-transgenic (Tg) rats intraperitoneally administered 40 mg/kg body weight cocaine (HIV-cocaine group) once daily for 21 days when compared with HIV-Tg rats given saline (HIV group) or wild-type (WT) Fischer 334 rats treated with (WT-cocaine group) and without cocaine (WT group). In addition, right ventricle systolic pressure was also found to be significantly higher in the HIV-cocaine rats compared with the WT group. Significant down-regulation in protein expression of BMPR-2 and BMPR-1B was observed in total lung extract from HIV-cocaine rats compared with the other three groups. Furthermore, the PASMCs isolated from HIV-cocaine rats demonstrated a higher level of proliferation and lower levels of apoptosis compared with cells isolated from other rat groups. Interestingly, corroborating our earlier cell-culture findings, we observed higher expression of BMPR-2 and BMPR-1B messenger RNA and significantly lower levels of BMPR-2 and BMPR-1B protein in HIV-cocaine PASMCs compared with cells isolated from all other groups. In conclusion, our findings support an additive effect of cocaine and HIV on smooth muscle dysfunction, resulting in enhanced pulmonary vascular remodeling with associated elevation of mean pulmonary arterial pressure and right ventricle systolic pressure in HIV-Tg rats exposed to cocaine.

Ligand-Independent Activation of Platelet-Derived Growth Factor Receptor ß during Human Immunodeficiency Virus-Transactivator of Transcription and Cocaine-Mediated Smooth Muscle Hyperplasia.

Our previous study supports an additive effect of cocaine to human immunodeficiency virus infection in the development of pulmonary arteriopathy through enhancement of proliferation of pulmonary smooth muscle cells (SMCs), while also suggesting involvement of platelet-derived growth factor receptor (PDGFR) activation in the absence of further increase in PDGF-BB ligand. Redox-related signaling pathways have been shown to regulate tyrosine kinase receptors independent of ligand binding, so we hypothesized that simultaneous treatment of SMCs with transactivator of transcription (Tat) and cocaine may be able to indirectly activate PDGFR through modulation of reactive oxygen species (ROS) without the need for PDGF binding. We found that blocking the binding of ligand using suramin or monoclonal IMC-3G3 antibody significantly reduced ligand-induced autophosphorylation of Y1009 without affecting ligand-independent transphosphorylation of Y934 residue on PDGFRß in human pulmonary arterial SMCs treated with both cocaine and Tat. Combined treatment of human pulmonary arterial SMCs with cocaine and Tat resulted in augmented production of superoxide radicals and hydrogen peroxide when compared with either treatment alone. Inhibition of this ROS generation prevented cocaine- and Tat-mediated Src activation and transphosphorylation of PDGFRß at Y934 without any changes in phosphorylation of Y1009, in addition to attenuation of smooth muscle hyperplasia. Furthermore, pretreatment with an Src inhibitor, PP2, also suppressed cocaine- and Tat-mediated enhanced Y934 phosphorylation and smooth muscle proliferation. Finally, we report total abrogation of cocaine- and Tat-mediated synergistic increase in cell proliferation on inhibition of both ligand-dependent and ROS/Src-mediated ligand-independent phosphorylation of PDGFRß.

Downregulation of bone morphogenetic protein receptor axis during HIV-1 and cocaine-mediated pulmonary smooth muscle hyperplasia: implications for HIV-related pulmonary arterial hypertension.

OBJECTIVE: Our previous findings support an additive effect of cocaine to HIV infection in the development of pulmonary arteriopathy through enhanced proliferation of human pulmonary smooth muscle cells. We now examined the role of antiproliferative bone morphogenetic protein receptor (BMPR) axis in HIV protein and cocaine-mediated pulmonary smooth muscle hyperplasia. APPROACH AND RESULTS: Stimulation of BMPR axis resulted in attenuation of synergistic increase in the proliferation of human pulmonary arterial smooth muscle cells in response to cocaine and HIV protein, transactivator of transcription (Tat). Interestingly, an increase in mRNA but decrease in protein levels of BMPR with correlated decrease in the activation of Sma- and MAD-related family protein 1/5/8 and Id1 gene expression was observed on combined treatment with cocaine and Tat when compared with the untreated cells at all time points tested. Although longer exposure to either cocaine or Tat alone also resulted in a significant decrease in the BMPR protein expression, the abrogation on combined treatment was still significantly more when compared with that of the monotreatments. Significant increase in mRNA but downmodulation of BMPR protein expression was also observed in the lung extracts from HIV-infected intravenous drug users (HIV+IVDU) when compared with that from HIV-infected non-IVDUs (HIV) or uninfected IVDUs (IVDU). Furthermore, significant decrease in BMPR protein expression was also observed in HIV or IVDUs when compared with normal controls that correlated with in vitro findings on chronic exposure to cocaine or HIV protein alone. CONCLUSIONS: Simultaneous exposure of pulmonary smooth muscle cells to viral protein(s) and cocaine exacerbates downregulation of BMPR axis that may result in enhanced pulmonary vasculature aberrations in HIV+IVDUs.

Analysis of MicroRNA Cargo in Circulating Extracellular Vesicles from HIV-Infected Individuals with Pulmonary Hypertension.

The risk of developing pulmonary hypertension (PH) in people living with HIV is at least 300-fold higher than in the general population, and illicit drug use further potentiates the development of HIV-associated PH. The relevance of extracellular vesicles (EVs) containing both coding as well as non-coding RNAs in PH secondary to HIV infection and drug abuse is yet to be explored. We here compared the miRNA cargo of plasma-derived EVs from HIV-infected stimulant users with (HIV + Stimulants + PH) and without PH (HIV + Stimulants) using small RNA sequencing. The data were compared with 12 PH datasets available in the GEO database to identify potential candidate gene targets for differentially altered miRNAs using the following functional analysis tools: ingenuity pathway analysis (IPA), over-representation analysis (ORA), and gene set enrichment analysis (GSEA). MiRNAs involved in promoting cell proliferation and inhibition of intrinsic apoptotic signaling pathways were among the top upregulated miRNAs identified in EVs from the HIV + Stimulants + PH group compared to the HIV + Stimulants group. Alternatively, the downregulated miRNAs in the HIV + Stimulants + PH group suggested an association with the negative regulation of smooth muscle cell proliferation, IL-2 mediated signaling, and transmembrane receptor protein tyrosine kinase signaling pathways. The validation of significantly differentially expressed miRNAs in an independent set of HIV-infected (cocaine users and nondrug users) with and without PH confirmed the upregulation of miR-32-5p, 92-b-3p, and 301a-3p positively regulating cellular proliferation and downregulation of miR-5571, -4670 negatively regulating smooth muscle proliferation in EVs from HIV-PH patients. This increase in miR-301a-3p and decrease in miR-4670 were negatively correlated with the CD4 count and FEV1/FVC ratio, and positively correlated with viral load. Collectively, this data suggest the association of alterations in the miRNA cargo of circulating EVs with HIV-PH.

Enhanced pulmonary arteriopathy in simian immunodeficiency virus-infected macaques exposed to morphine.

RATIONALE: HIV-associated pulmonary arterial hypertension (PAH) is likely a more prevalent noninfectious complication of AIDS than previously recognized. Furthermore, the majority of HIV-PAH cases occur in individuals with a history of intravenous drug use. In this study we used a simian immunodeficiency (SIV) macaque model and a primary cell-culture system to investigate the association between drug abuse and HIV infection in HIV-PAH development. METHODS: The archival lung tissues from macaques previously used to study the effect of morphine on SIV infection-associated neuropathogenesis were analyzed for pulmonary vascular changes. The direct effect of HIV proteins and illicit drugs was investigated on oxidative stress, survival, and proliferation of human pulmonary microvascular endothelial cells. MEASUREMENTS AND MAIN RESULTS: SIVmacR71/17E-infected rhesus macaques treated with morphine (VM group) demonstrated significant pulmonary vascular remodeling, including the presence of early and advanced complex (plexiform) lesions, when compared with either the SIV-infected (V group) or morphine-treated uninfected (M group) macaques. However, both the V (two of five) and VM (two of six) groups included some animals with Pneumocystis jirovecii pneumonia. The endothelial cells lining the vessels with medial hypertrophy or initial-stage intimal lesions in lung sections from VM macaques demonstrated an increase in positivity for both terminal dUTP nick-end labeling and Ki67. Oxidative stress-mediated enhanced apoptosis followed by enhanced proliferation of endothelial cells was observed on simultaneous treatment with viral proteins and drugs of abuse compared with either treatment alone. CONCLUSIONS: Our findings suggest that SIV/HIV protein(s) and morphine interact to cause the proliferation of apoptosis-resistant endothelial cells leading to angio-obliteration.

HIV and Drug Use: A Tale of Synergy in Pulmonary Vascular Disease Development.

Over the past two decades, with the advent and adoption of highly active anti-retroviral therapy, HIV-1 infection, a once fatal and acute illness, has transformed into a chronic disease with people living with HIV (PWH) experiencing increased rates of cardio-pulmonary vascular diseases including life-threatening pulmonary hypertension. Moreover, the chronic consequences of tobacco, alcohol, and drug use are increasingly seen in older PWH. Drug use, specifically, can have pathologic effects on the cardiovascular health of these individuals. The "double hit" of drug use and HIV may increase the risk of HIV-associated pulmonary arterial hypertension (HIV-PAH) and potentiate right heart failure in this population. This article explores the epidemiology and pathophysiology of PAH associated with HIV and recreational drug use and describes the proposed mechanisms by which HIV and drug use, together, can cause pulmonary vascular remodeling and cardiopulmonary hemodynamic compromise. In addition to detailing the proposed cellular and signaling pathways involved in the development of PAH, this article proposes areas ripe for future research, including the influence of gut dysbiosis and cellular senescence on the pathobiology of HIV-PAH. © 2023 American Physiological Society. Compr Physiol 13:4659-4683, 2023.

Notch3 deletion regulates HIV-1 gene expression and systemic inflammation to ameliorate chronic kidney disease.

Antiretroviral therapy (ART) has profoundly decreased HIV-1 associated morbidity. However, despite ART, immune cells remain latently infected and slowly release viral proteins, leading to chronic inflammation and HIV associated comorbidities. Thus, new strategies are needed to reduce the inflammatory effects of HIV-1. In previous studies we found that gamma secretase inhibitor (GSIXX) ameliorated renal lesions of HIV-Tg26 mice carrying replication defective HIV-1 PNL4-3 by inhibiting Notch activation. Since gamma secretase inhibition is not a safe strategy in humans, here we examined the specific role of the Notch3 pathway in the pathogenesis of the renal lesions and outcome of HIV-Tg26 mice. We found that Notch3 is activated in podocytes and other renal cells in HIV-Tg26 mice and human biopsies with HIV-1 associated Nephropathy (HIVAN). Knockdown of Notch3 in HIV-Tg26 mice revealed a marked reduction in the mortality rate, improvement in renal injury and function. RNA sequencing and immunolabeling data revealed that Notch3 deletion drastically reduced infiltrating renal macrophages in HIV-Tg-N3KO mice in association with renal reduction of HIV-nef mRNA expression levels. In fact, bone marrow derived macrophages from HIV-Tg26 mice showed a significant activation of Notch3 signaling. Further, systemic levels of TNF-alpha and MCP-1 and other inflammatory chemokines and cytokines were reduced in Tg-N3KO mice as compared to HIV-Tg26 mice and this translated to a marked reduction of HIV-induced skin lesions. Taken together, these studies strongly point to a dual inhibitory/therapeutic effect of Notch3 inhibition on HIV-induced systemic, skin and renal lesions independently of ART.

Exploring extracellular vesicles as mediators of clinical disease and vehicles for viral therapeutics: Insights from the COVID-19 pandemic.

The COVID-19 pandemic has challenged researchers to rapidly understand the capabilities of the SARS-CoV-2 virus and investigate potential therapeutics for SARS-CoV-2 infection. COVID-19 has been associated with devastating lung and cardiac injury, profound inflammation, and a heightened coagulopathic state, which may, in part, be driven by cellular crosstalk facilitated by extracellular vesicles (EVs). In recent years, EVs have emerged as important biomarkers of disease, and while extracellular vesicles may contribute to the spread of COVID-19 infection from one cell to the next, they also may be engineered to play a protective or therapeutic role as decoys or "delivery drivers" for therapeutic agents. This review explores these roles and areas for future study.

Effect of cocaine on human immunodeficiency virus-mediated pulmonary endothelial and smooth muscle dysfunction.

Human immunodeficiency virus (HIV)-associated pulmonary arterial hypertension (PAH) is a devastating, noninfectious complication of acquired immune deficiency syndrome, and the majority of HIV-PAH cases occur in individuals with a history of intravenous drug use (IVDU). However, although HIV-1 and IVDU have been associated with PAH independently or in combination, the pathogenesis of the disproportionate presence of HIV-PAH in association with IVDU has yet to be characterized. The objective of this study was to obtain a better understanding of the interactions between HIV-1 and cocaine to help uncover the mechanism(s) of the development of HIV-PAH. We observed that exposure of HIV-infected macrophages or HIV-Trans-Activator of Transcription (Tat)-treated pulmonary endothelial cells to cocaine enhanced the expression of platelet-derived growth factor (PDGF)-BB. Simultaneous treatment with Tat and cocaine, on the other hand, exacerbated both the disruption of tight junction proteins (TJPs), with enhanced permeability in pulmonary endothelial cells, and the proliferation of pulmonary smooth muscle cells (pSMCs) compared with either treatment alone. Histological examination of HIV plus IVDU human lung sections showed signs of early pulmonary arteriopathy, severe down-modulation of TJPs, and increased expression of PDGF-BB compared with the lung sections from individuals who are infected with HIV and without history of IVDU. Interestingly, blocking of PDGF receptor signaling with the receptor antagonist or small interfering RNA has been shown to inhibit the increase in proliferation of pSMCs on Tat and cocaine exposure. Our results, therefore, support an additive effect of cocaine to HIV infection in the development of pulmonary arteriopathy through enhancement of endothelial dysfunction and proliferation of pSMCs, while also suggesting PDGF-PDGF receptor axis as a potential target for use in clinical intervention.

Hypoxia-inducible factor-1 α/platelet derived growth factor axis in HIV-associated pulmonary vascular remodeling.

BACKGROUND: Human immunodeficiency virus (HIV) infected patients are at increased risk for the development of pulmonary arterial hypertension (PAH). Recent reports have demonstrated that HIV associated viral proteins induce reactive oxygen species (ROS) with resultant endothelial cell dysfunction and related vascular injury. In this study, we explored the impact of HIV protein induced oxidative stress on production of hypoxia inducible factor (HIF)-1α and platelet-derived growth factor (PDGF), critical mediators implicated in the pathogenesis of HIV-PAH. METHODS: The lungs from 4-5 months old HIV-1 transgenic (Tg) rats were assessed for the presence of pulmonary vascular remodeling and HIF-1α/PDGF-BB expression in comparison with wild type controls. Human primary pulmonary arterial endothelial cells (HPAEC) were treated with HIV-associated proteins in the presence or absence of pretreatment with antioxidants, for 24 hrs followed by estimation of ROS levels and western blot analysis of HIF-1α or PDGF-BB. RESULTS: HIV-Tg rats, a model with marked viral protein induced vascular oxidative stress in the absence of active HIV-1 replication demonstrated significant medial thickening of pulmonary vessels and increased right ventricular mass compared to wild-type controls, with increased expression of HIF-1α and PDGF-BB in HIV-Tg rats. The up-regulation of both HIF-1α and PDGF-B chain mRNA in each HIV-Tg rat was directly correlated with an increase in right ventricular/left ventricular+septum ratio. Supporting our in-vivo findings, HPAECs treated with HIV-proteins: Tat and gp120, demonstrated increased ROS and parallel increase of PDGF-BB expression with the maximum induction observed on treatment with R5 type gp-120CM. Pre-treatment of endothelial cells with antioxidants or transfection of cells with HIF-1α small interfering RNA resulted in abrogation of gp-120CM mediated induction of PDGF-BB, therefore, confirming that ROS generation and activation of HIF-1α plays critical role in gp120 mediated up-regulation of PDGF-BB. CONCLUSION: In summary, these findings indicate that viral protein induced oxidative stress results in HIF-1α dependent up-regulation of PDGF-BB and suggests the possible involvement of this pathway in the development of HIV-PAH.

Extracellular vesicle-mediated endothelial apoptosis and EV-associated proteins correlate with COVID-19 disease severity.

Coronavirus disease-2019 (COVID-19), caused by the novel severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), has lead to a global pandemic with a rising toll in infections and deaths. Better understanding of its pathogenesis will greatly improve the outcomes and treatment of affected patients. Here we compared the inflammatory and cardiovascular disease-related protein cargo of circulating large and small extracellular vesicles (EVs) from 84 hospitalized patients infected with SARS-CoV-2 with different stages of disease severity. Our findings reveal significant enrichment of proinflammatory, procoagulation, immunoregulatory and tissue-remodelling protein signatures in EVs, which remarkably distinguished symptomatic COVID-19 patients from uninfected controls with matched comorbidities and delineated those with moderate disease from those who were critically ill. Specifically, EN-RAGE, followed by TF and IL-18R1, showed the strongest correlation with disease severity and length of hospitalization. Importantly, EVs from COVID-19 patients induced apoptosis of pulmonary microvascular endothelial cells in the order of disease severity. In conclusion, our findings support a role for EVs in the pathogenesis of COVID-19 disease and underpin the development of EV-based approaches to predicting disease severity, determining need for patient hospitalization and identifying new therapeutic targets.