Liposomes-Encapsulating Double-Stranded Nucleic Acid (Poly I:C) for Head and Neck Cancer Treatment.

Polyriboinosinic acid-polyribocytidylic acid (Poly I:C) serves as a synthetic mimic of viral double-stranded dsRNA, capable of inducing apoptosis in numerous cancer cells. Despite its potential, therapeutic benefits, the application of Poly I:C has been hindered by concerns regarding toxicity, stability, enzymatic degradation, and undue immune stimulation, leading to autoimmune disorders. To address these challenges, encapsulation of antitumor drugs within delivery systems such as cationic liposomes is often employed to enhance their efficacy while minimizing dosages. In this study, we investigated the potential of cationic liposomes to deliver Poly I:C into the Head and Neck 12 (HN12) cell line to induce apoptosis in the carcinoma cells and tumor model. Cationic liposomes made by the hydrodynamic focusing method surpass traditional methods by offering a continuous flow-based approach for encapsulating genes, which is ideal for efficient tumor delivery. DOTAP liposomes efficiently bind Poly I:C, confirmed by transmission electron microscopy images displaying their spherical morphology. Liposomes are easily endocytosed in HN12 cells, suggesting their potential for therapeutic gene and drug delivery in head and neck squamous carcinoma cells. Activation of apoptotic pathways involving MDA5, RIG-I, and TLR3 is evidenced by upregulated caspase-3, caspase-8, and IRF3 genes upon endocytosis of Poly(I:C)-encapsulated liposomes. Therapeutic evaluations revealed significant inhibition of tumor growth with Poly I:C liposomes, indicating the possibility of MDA5, RIG-I, and TLR3-induced apoptosis pathways via Poly I:C liposomes in HN12 xenografts in J:NU mouse models. Comparative histological analysis underscores enhanced cell death with Poly I:C liposomes, warranting further investigation into the precise mechanisms of apoptosis and inflammatory cytokine response in murine models for future research.

PAI-1 Deficiency Drives Pulmonary Vascular Smooth Muscle Remodeling and Pulmonary Hypertension.

Pulmonary arterial hypertension (PAH) is a progressive and potentially a rapidly fatal disease characterized by vasoconstriction and remodeling of small pulmonary arteries (PA) leading to increased pulmonary vascular resistance and right heart failure. Central to the remodeling process is a switch of the smooth muscle cells in small PAs (PASMC) to a proliferative, apoptosis-resistant phenotype. There is reason to suspect that the plasminogen activator system may play an important role in the remodeling program in PAH based on its roles in vascular post-injury restenosis, fibrosis, angiogenesis and tumorigenesis. Plasminogen activator inhibitor-1 (PAI-1) is the primary physiological inhibitor of the plasminogen activators - urokinase-type and tissue-type (uPA and tPA, respectively). Immunohisto- chemical and immunoblot analyses revealed that PAI-1 was deficient in smooth muscle areas of small remodeled PAs and early-passage PASMC from subjects with PAH compared to non-PAH controls. PAI1-/- male and female mice developed spontaneous pulmonary vascular remodeling and pulmonary hypertension (PH) as evidenced by significant increase in PA medial thickness, systolic right ventricular pressure, and right ventricular hypertrophy. Lastly, the uPA inhibitors upamostat (WX-671) and amiloride analog BB2-30F down-regulated mTORC1 and SMAD3, restored PAI-1 levels, reduced proliferation, and induced apoptosis in human PAH PASMC. We examined the effect of inhibition of uPA catalytic activity by BB2-30F on the development of SU5416/Hypoxia (SuHx)-induced PH in mice. Vehicletreated SuHx-exposed mice had up-regulated mTORC1 in small PAs, developed pulmonary vascular remodeling and PH, as evidenced by significant increase of PA MT, sRVP, RV hypertrophy, and a significant decrease in the pulmonary artery acceleration time/pulmonary ejection time (PAAT/PET) ratio compared to age- and sex-matched normoxia controls, whereas BB2-30F-treated group was protected from all these pathological changes. Taken together, our data strongly suggest that PAI-1 down- regulation in PASMC from human PAH lungs promotes PASMC hyper-proliferation, remodeling, and spontaneous PH due to unopposed uPA activation. Further studies are needed to determine the potential benefits of targeting the PAI-1/uPA imbalance to attenuate the progression and/or reverse pulmonary vascular remodeling and PH.

Persistent hypoxia promotes myofibroblast differentiation via GPR-81 and differential regulation of LDH isoenzymes in normal and idiopathic pulmonary fibrosis fibroblasts.

Hypoxia, a state of insufficient oxygen availability, promotes cellular lactate production. Lactate levels are increased in lungs from patients with idiopathic pulmonary fibrosis (IPF), a disease characterized by excessive scar formation, and lactate is implicated in the pathobiology of lung fibrosis. However, the mechanisms underlying the effects of hypoxia and lactate on fibroblast phenotype are poorly understood. We exposed normal and IPF lung fibroblasts to persistent hypoxia and found that increased lactate generation by IPF fibroblasts was driven by the FoxM1-dependent increase of lactate dehydrogenase A (LDHA) coupled with decreased LDHB that was not observed in normal lung fibroblasts. Importantly, hypoxia reduced α-smooth muscle actin (α-SMA) expression in normal fibroblasts but had no significant impact on this marker of differentiation in IPF fibroblasts. Treatment of control and IPF fibroblasts with TGF-ß under hypoxic conditions did not significantly change LDHA or LDHB expression. Surprisingly, lactate directly induced the differentiation of normal, but not IPF fibroblasts under hypoxic conditions. Moreover, while expression of GPR-81, a G-protein-coupled receptor that binds extracellular lactate, was increased by hypoxia in both normal and IPF fibroblasts, its inhibition or silencing only suppressed lactate-mediated differentiation in normal fibroblasts. These studies show that hypoxia differentially affects normal and fibrotic fibroblasts, promoting increased lactate generation by IPF fibroblasts through regulation of the LDHA/LDHB ratio and promoting normal lung fibroblast responsiveness to lactate through GPR-81. This supports a novel paradigm in which lactate may serve as a paracrine intercellular signal in oxygen-deficient microenvironments.

Macrophage-Targeted Lipid Nanoparticle Delivery of microRNA-146a to Mitigate Hemorrhagic Shock-Induced Acute Respiratory Distress Syndrome.

The pro-inflammatory response of alveolar macrophages to injurious physical forces during mechanical ventilation is regulated by the anti-inflammatory microRNA, miR-146a. Increasing miR-146a expression to supraphysiologic levels using untargeted lipid nanoparticles reduces ventilator-induced lung injury but requires a high initial dose of miR-146a making it less clinically applicable. In this study, we developed mannosylated lipid nanoparticles that can effectively mitigate lung injury at the initiation of mechanical ventilation with lower doses of miR-146a. We used a physiologically relevant humanized in vitro coculture system to evaluate the cell-specific targeting efficiency of the mannosylated lipid nanoparticle. We discovered that mannosylated lipid nanoparticles preferentially deliver miR-146a to alveolar macrophages and reduce force-induced inflammation in vitro. Our in vivo study using a clinically relevant mouse model of hemorrhagic shock-induced acute respiratory distress syndrome demonstrated that delivery of a low dose of miR-146a (0.1 nmol) using mannosylated lipid nanoparticles dramatically increases miR-146a levels in mouse alveolar macrophages and decreases lung inflammation. These data suggest that mannosylated lipid nanoparticles may have the therapeutic potential to mitigate lung injury during mechanical ventilation.

A Fucosylated Lactose-Presenting Tetravalent Glycocluster Acting as a Mutual Ligand of Pseudomonas aeruginosa Lectins A (PA-IL) and B (PA-IIL)-Synthesis and Interaction Studies.

The Gram-negative bacterium Pseudomonas aeruginosa is an important opportunistic human pathogen associated with cystic fibrosis. P. aeruginosa produces two soluble lectins, the d-galactose-specific lectin PA-IL (LecA) and the l-fucose-specific lectin PA-IIL (LecB), among other virulence factors. These lectins play an important role in the adhesion to host cells and biofilm formation. Moreover, PA-IL is cytotoxic to respiratory cells in the primary culture. Therefore, these lectins are promising therapeutic targets. Specifically, carbohydrate-based compounds could inhibit their activity. In the present work, a 3-O-fucosyl lactose-containing tetravalent glycocluster was synthesized and utilized as a mutual ligand of galactophilic and fucophilic lectins. Pentaerythritol equipped with azido ethylene glycol-linkers was chosen as a multivalent scaffold and the glycocluster was constructed by coupling the scaffold with propargyl 3-O-fucosyl lactoside using an azide-alkyne 1,3-dipolar cycloaddition reaction. The interactions between the glycocluster and PA-IL or PA-IIL were investigated by isothermal titration microcalorimetry and saturation transfer difference NMR spectroscopy. These results may assist in the development of efficient anti-adhesion therapy for the treatment of a P. aeruginosa infection.

Arbuscular Mycorrhizal Fungus Rhizophagus irregularis Influences Artemisia annua Plant Parameters and Artemisinin Content under Different Soil Types and Cultivation Methods.

Artemisinin extracted from Artemisia annua has been used efficiently in malaria treatment since 2005. In this study, the variations in plant parameters (plant biomass, glandular trichome density, essential oil total chemical content, artemisinin production, and polyphenol oxidase (PPO) activity) were tested under different soil types (Luvisol, Gleysol, Anthrosol and sterile peat) and cultivation conditions (potted plants in semi-open field, and open field experiments) for plants inoculated with arbuscular mycorrhizal fungus (AMF) Rizophagus irregularis. Under semi-open field conditions, the AMF colonization of A. annua plant roots varied, and presented the highest percentage in Luvisol and sterile peat. The increase in the root colonization rate positively influenced some plant parameters (biomass, glandular trichome density, artemisinin concentration, essential oil quantity and composition), but no effects on PPO enzyme activity were detected. AMF fungus R. irregularis significantly increased the artemisinin content and essential oil yield of plants cultivated in Luvisol, Gleysol, Anthrosol and in peat. These soil types can offer appropriate conditions for A. annua cultivation and artemisinin production even on a smaller scale. Under open field conditions, low (about 5%) AMF colonization was observed. No differences in artemisin contents were detected, but essential oil yield significantly increased compared to control plants. AMF treatment increased beta-farnesene and germacrene D concentrations in Artemisia plants in the open field experiment.

Primary peritonitis in an adolescent boy / Primer peritonitis egészséges serdülo fiúban

Primary peritonitis is very rare in healthy children without predisposing factors. In the absence of unique factors and signs, the clinical picture does not differ from secondary peritonitis. Therefore, the diagnosis is almost always an intraoperative diagnosis. Case report: We admitted a previously healthy 15-year-old boy with symptoms of acute enteritis. Within 24 hours, he developed acute abdomen and signs of septic shock. Computer tomography of the abdomen revealed air bubbles in the middle of the abdomen and near the terminal ileum. Suspecting perforation, we performed an emergency laparotomy. However, there was no perforation to be found in the background of the purulent peritonitis. We initiated empirical broad-spectrum antimicrobial therapy which we later adjusted. Septic shock and complications were treated successfully. We could not find the source of the primary peritonitis. Since hospital discharge, the child has been asymptomatic. In primary peritonitis, due to the nonspecific, rapidly progressing symptoms, an emergency surgery can not be avoided. With proper antibiotics and supportive therapy, the prognosis is favourable. Orv Hetil. 2020; 161(23): 977-979.

Clonally selected primitive endothelial cells promote occlusive pulmonary arteriopathy and severe pulmonary hypertension in rats exposed to chronic hypoxia.

One current concept suggests that unchecked proliferation of clonally selected precursors of endothelial cells (ECs) contribute to severe pulmonary arterial hypertension (PAH). We hypothesized that clonally selected ECs expressing the progenitor marker CD117 promote severe occlusive pulmonary hypertension (PH). The remodelled pulmonary arteries of PAH patients harboured CD117+ ECs. Rat lung CD117+ ECs underwent four generations of clonal expansion to enrich hyperproliferative ECs. The resulting clonally enriched ECs behaved like ECs, as measured by in vitro and in vivo angiogenesis assays. The same primitive ECs showed a limited ability for mesenchymal lineage differentiation. Endothelial differentiation and function were enhanced by blocking TGF-ß signalling, promoting bone morphogenic protein (BMP) signalling. The transplantation of the EC clones caused arterio-occlusive PH in rats exposed to chronic hypoxia. These EC clones engrafted in the pulmonary arteries. Yet cessation of chronic hypoxia promoted lung cell apoptosis and resolution of vascular lesions. In conclusion, this is to the best of our knowledge, the first report that clonally enriched primitive ECs promote occlusive pulmonary arteriopathy and severe PH. These primitive EC clones further give rise to cells of endothelial and mesenchymal lineage as directed by BMP and TGF-ß signaling.

Investigation of In Vitro Antioxidant and Antibacterial Potential of Silver Nanoparticles Obtained by Biosynthesis Using Beech Bark Extract.

Green synthesis is one of the rapid and best ways for silver nanoparticles (AgNP) synthesis. In the present study, synthesis and bioactivity of AgNPs has been demonstrated using water beech (Fagus sylvatica L.) bark extract. The physical and chemical factors such as time, metal ion solution, and pH, which play a vital role in the AgNPs synthesis, were assessed. The AgNPs were characterized by ultraviolet-visible (UV-Vis) spectrometry, Fourier transform infrared spectroscopy (FT-IR), and transmission electron microscopy (TEM). Antioxidant and antimicrobial activity of the obtained AgNPs was evaluated. AgNPs were characterized by color change pattern, and the broad peak obtained at 420-475 nm with UV-Vis confirmed the synthesis of AgNPs. FT-IR results confirmed that phenols and proteins of beech bark extract are mainly responsible for capping and stabilization of synthesized AgNPs. TEM micrographs showed spherical or rarely polygonal and triangular particles with an average size of 32 nm at pH = 9, and 62 nm at pH = 4. Furthermore, synthesized AgNPs were found to exhibit antioxidant activity and have antibacterial effect against Staphylococcus aureus, methicillin-resistant Staphylococcus aureus (MRSA), Escherichia coli, and Pseudomonas aeruginosa. These results indicate that bark extract of F. sylvatica L. is suitable for synthesizing stable AgNPs, which act as an excellent antimicrobial agent.

An Ab Initio Investigation of the 4,4'-Methlylene Diphenyl Diamine (4,4'-MDA) Formation from the Reaction of Aniline with Formaldehyde.

The most commonly applied industrial synthesis of 4,4'-methylene diphenyl diamine (4,4'-MDA), an important polyurethane intermediate, is the reaction of aniline and formaldehyde. Molecular understanding of the 4,4'-MDA formation can provide strategy to prevent from side reactions. In this work, a molecular mechanism consisted of eight consecutive, elementary reaction steps from anilines and formaldehyde to the formation of 4,4'-MDA in acidic media is proposed using accurate G3MP2B3 composite quantum chemical method. Then G3MP2B3-SMD results in aqueous and aniline solutions were compared to the gas phase mechanism. Based on the gas phase calculations standard enthalpy of formation, entropy and heat capacity values were evaluated using G3MP2B3 results for intermediates The proposed mechanism was critically evaluated and important side reactions are considered: the competition of formation of protonated p-aminobenzylaniline (PABAH⁺), protonated aminal (AMH⁺) and o-aminobenzylaniline (OABAH⁺). Competing reactions of the 4,4'-MDA formation is also thermodynamically analyzed such as the formation of 2,4-MDAH⁺, 3,4-MDAH⁺. AMH⁺ can be formed through loose transition state, but it becomes kinetic dead-end, while formation of significant amount of 2,4-MDA is plausible through low-lying transition state. The acid strength of the key intermediates such as N-methylenebenzeneanilium, PABAH⁺, 4-methylidenecyclohexa-2,5-diene-1-iminium, and AMH⁺ was estimated by relative pKa calculation.

Endothelial cells are a source of Nestin expression in Pulmonary Arterial Hypertension.

Uncontrolled proliferation of endothelial cells is essential to the pathogenesis of pulmonary arterial hypertension (PAH). Both proliferation and cytoskeleton reorganization are associated with upregulation of the intermediate filament protein Nestin. Recently, accumulation of Nestin-expressing cells was found in pulmonary vascular lesions of PAH patients. The goal of this study is to determine if Nestin expression contributes to endothelial proliferation in pulmonary arterial hypertension, using both lung tissues and endothelial cells. Here we found that endothelial cells from complex and plexiform lesions of PAH patients expressed Nestin. These Nestin+ cells further stained positive for the angiogenic factors CXC chemokine ligand 12 and Wnt1. Likewise, in the chronic hypoxia/SU5416 animal model of pulmonary hypertension, Nestin+ endothelial cells were found in occlusive pulmonary vascular lesions. In vitro, both growing rat and human lung endothelial cells expressed Nestin protein. When Nestin was overexpressed in endothelial cells (both rat and human), Nestin overexpression promoted proliferation and expression of CXC chemokine ligand 12. Nestin overexpression further increased angiogenic tube formation in vitro. Conclusions: We found increased Nestin expression from endothelial cells of occlusive lung vascular lesions in severe pulmonary hypertension. Elevated Nestin expression likely contributes to unchecked pulmonary vascular proliferation and angiogenesis, possibly via induction of CXC chemokine ligand 12. Additional studies are required to determine whether targeting Nestin would be beneficial to treat PAH.

From Here to There, Progenitor Cells and Stem Cells Are Everywhere in Lung Vascular Remodeling.

The field of stem cell biology, cell therapy, and regenerative medicine has expanded almost exponentially, in the last decade. Clinical trials are evaluating the potential therapeutic use of stem cells in many adult and pediatric lung diseases with vascular component, such as bronchopulmonary dysplasia (BPD), chronic obstructive pulmonary disease (COPD), idiopathic pulmonary fibrosis (IPF), or pulmonary arterial hypertension (PAH). Extensive research activity is exploring the lung resident and circulating progenitor cells and their contribution to vascular complications of chronic lung diseases, and researchers hope to use resident or circulating stem/progenitor cells to treat chronic lung diseases and their vascular complications. It is becoming more and more clear that progress in mechanobiology will help to understand the various influences of physical forces and extracellular matrix composition on the phenotype and features of the progenitor cells and stem cells. The current review provides an overview of current concepts in the field.

Impact of parenchymal loss on renal function after laparoscopic partial nephrectomy under warm ischemia.

PURPOSE: To elucidate the impact of renal parenchymal loss and the ischemic reperfusion injury (RI) on the renal function after laparoscopic partial nephrectomy (LPN) under warm ischemia (WI). METHODS: Thirty-five patients with a single polar renal mass ≤4 cm and normal contralateral kidney underwent LPN. Transperitoneal LPN with WI using en bloc hilar occlusion was performed. The total differential renal function (T-DRF) using 99mTc-dimercaptosuccinic acid was evaluated preoperatively and postoperatively over a period of 1 year. A special region of interest (ROI) was selected on the non-tumorous pole of the involved kidney, and was compared with the same ROI in the contralateral kidney. The latter comparison was defined as partial differential renal function (P-DRF). Any postoperative decline in the P-DRF of the operated kidney was attributed to the RI. Subtraction of the P-DRF decline from the T-DRF decline was attributed to the parenchymal loss caused by the resection of the tumor and suturing of the normal parenchyma. RESULTS: The mean WI time was 22 min, and the mean weight of resected specimen was 18 g. The mean postoperative eGFR declined to 87 ml/min/1.73 m2 from its baseline mean value of 97 ml/min/1.73 m2 (p value = 0.075). Mean postoperative T-DRF and P-DRF of the operated kidney declined by 7 and 3 %, respectively. CONCLUSIONS: After LPN of small renal mass, decline in renal function is primarily attributed to parenchymal loss caused by tumor resection and suturing of the normal parenchyma rather than the RI.

Iloprost reverses established fibrosis in experimental right ventricular failure.

Prostacyclin and its analogues improve cardiac output and functional capacity in patients with pulmonary arterial hypertension (PAH); however, the underlying mechanism is not fully understood. We hypothesised that prostanoids have load-independent beneficial effects on the right ventricle (RV). Angio-obliterative PAH and RV failure were induced in rats with a single injection of SU5416 followed by 4 weeks of exposure to hypoxia. Upon confirmation of RV dysfunction and PAH, rats were randomised to 0.1 µg·kg(-1) nebulised iloprost or drug-free vehicle, three times daily for 2 weeks. RV function and treadmill running time were evaluated pre- and post-iloprost/vehicle treatment. Pulmonary artery banded rats were treated 8 weeks after surgery to allow for significant RV hypertrophy. Inhaled iloprost significantly improved tricuspid annulus plane systolic excursion and increased exercise capacity, while mean pulmonary artery pressure and the percentage of occluded pulmonary vessels remained unchanged. Rats treated with iloprost had a striking reduction in RV collagen deposition, procollagen mRNA levels and connective tissue growth factor expression in both SU5416/hypoxia and pulmonary artery banded rats. In vitro, cardiac fibroblasts treated with iloprost showed a reduction in transforming growth factor (TGF)-ß1-induced connective tissue growth factor expression, in a protein kinase A-dependent manner. Iloprost decreased TGF-ß1-induced procollagen mRNA expression as well as cardiac fibroblast activation and migration. Iloprost significantly induced metalloproteinase-9 gene expression and activity and increased the expression of autophagy genes associated with collagen degradation. Inhaled iloprost improves RV function and reverses established RV fibrosis partially by preventing collagen synthesis and by increasing collagen turnover.

[New methods in the treatment of localized prostate cancer: use of dynamic arc therapy and kV cone-beam CT positioning]. / Új módszerek a prosztatadaganatok sugárkezelésében: dinamikus ívbesugárzás és kilovoltos "cone-beam" komputertomográfiás ellenorzés.

INTRODUCTION: Prostate cancer is a common disease among elderly male patients in developed countries. In addition to prostatectomy, definitive irradiation plays an increasing role in the treatment of localized disease. AIM: The authors wanted to share their experience obtained with the use of the Novalis TX linear accelerator for the application of dose-escalation, dynamic, intensity modulated arc therapy with the routine usage of cone-beam computer tomography based or image guided radiotherapy in patients with prostate cancer. METHOD: Between 2011, December and 2013, February the authors performed 102 treatments. In 10 low risk and 10 high risk prostate cancer patients (median age: 72.5 years) three-dimensional conformal plans with the same target volume coverage were created and tolerance doses of organs at risk (OAR) were compared. RESULTS: Compared to three-dimensional conformal techniques, intensity modulated arc therapy treatments produced a significantly lower dose at organ at risk that led to a more favorable early toxicity rate. CONCLUSIONS: The intensity modulated arc therapy with image guided radiotherapy proved to be a safe standard treatment mode in the daily routine in the institute of the authors. Late toxicity and local control rates need to be further examined.

Nuclear factor κB inhibition reduces lung vascular lumen obliteration in severe pulmonary hypertension in rats.

NF-κB and IL-6, a NF-κB downstream mediator, play a central role in the inflammatory response of tissues. We aimed to determine the role of the classical NF-κB pathway in severe pulmonary arterial hypertension (PAH) induced by SU5416 and chronic hypoxia (SuHx) in rats. Tissue samples from patients with idiopathic PAH (iPAH) and control subjects were investigated. SuHx rats were treated from Days 1 to 3, 1 to 21, and 29 to 42 with the NF-κB inhibitor pyrrolidine dithiocarbamate (PDTC) and/or from Days 1 to 21 with anti-IL-6 antibody. Nuclear staining for NF-κB, an indicator of the activation of the classical NF-κB pathway, was detected in pulmonary arterial lesions of patients with iPAH and SuHx rats. NF-κB inhibition with PDTC prevented and reduced pulmonary arterial obliteration without reducing muscularization. However, the elevated lung levels of IL-6 were not reduced in PDTC-treated SuHx animals. PDTC treatment prevented or reduced apoptosis of pulmonary artery wall cells and pulmonary arterial obliteration. IL-6 inhibition had only a partial effect on apoptosis and obliteration. Pulmonary arterial media wall thickness was not affected by any of these treatments. Preventive and therapeutic PDTC treatment promoted immune regulation by increasing the number of perivascular CD4(+) T cells, in particular regulatory T cells (early treatment), and by reducing the number of perivascular CD8(+) T lymphocytes and CD45RA(+) B lymphocytes. Therapeutic PDTC treatment further preserved right ventricular function in SuHx animals. Inhibition of NF-κB may represent a therapeutic option for pulmonary arterial obliteration via reduced vessel wall cell apoptosis and improved regulation of the immune system.

CXCR4 inhibition ameliorates severe obliterative pulmonary hypertension and accumulation of C-kit⁺ cells in rats.

Successful curative treatment of severe pulmonary arterial hypertension with luminal obliteration will require a thorough understanding of the mechanism underlying the development and progression of pulmonary vascular lesions. But the cells that obliterate the pulmonary arterial lumen in severe pulmonary arterial hypertension are incompletely characterized. The goal of our study was to evaluate whether inhibition of CXC chemokine receptor 4 will prevent the accumulation of c-kit⁺ cells and severe pulmonary arterial hypertension. We detected c-kit⁺⁻ cells expressing endothelial (von Willebrand Factor) or smooth muscle cell/myofibroblast (α-smooth muscle actin) markers in pulmonary arterial lesions of SU5416/chronic hypoxia rats. We found increased expression of CXC chemokine ligand 12 in the lung tissue of SU5416/chronic hypoxia rats. In our prevention study, AMD3100, an inhibitor of the CXC chemokine ligand 12 receptor, CXC chemokine receptor 4, only moderately decreased pulmonary arterial obliteration and pulmonary hypertension in SU5416/chronic hypoxia animals. AMD3100 treatment reduced the number of proliferating c-kit⁺ α-smooth muscle actin⁺ cells and pulmonary arterial muscularization and did not affect c-kit⁺ von Willebrand Factor⁺ cell numbers. Both c-kit⁺ cell types expressed CXC chemokine receptor 4. In conclusion, our data demonstrate that in the SU5416/chronic hypoxia model of severe pulmonary hypertension, the CXC chemokine receptor 4-expressing c-kit⁺ α-smooth muscle actin⁺ cells contribute to pulmonary arterial muscularization. In contrast, vascular lumen obliteration by c-kit⁺ von Willebrand Factor⁺ cells is largely independent of CXC chemokine receptor 4.