Comparison of l-Carnitine and l-Carnitine HCL salt for targeted lung treatment of pulmonary hypertension (PH) as inhalation aerosols: Design, comprehensive characterization, in vitro 2D/3D cell cultures, and in vivo MCT-Rat model of PH

New insights into the biomechanical properties of cells are revealing the importance of these properties and how they relate to underlying molecular, architectural, and behavioral changes associated with cell state and disease processes. However, the current understanding of how these in vitro biomechanical properties are associated with in vivo processes has been developed based on the traditional monolayer (two-dimensional [2D]) cell culture, which traditionally has not translated well to the three-dimensional (3D) cell culture and in vivo function. Many gold standard methods and tools used to observe the biomechanical properties of 2D cell cultures cannot be used with 3D cell cultures. Fluorescent molecules can respond to external factors almost instantaneously and require relatively low-cost instrumentation. In this review, we provide the background on fluorescent molecular rotors, which are attractive tools due to the relationship of their emission quantum yield with environmental microviscosity. We make the case for their use in both 2D and 3D cell cultures and speculate on their fundamental and practical applications in cell biology.

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A role for platelet-derived growth factor β-receptor in a newborn rat model of endothelin-mediated pulmonary vascular remodeling

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00180.2004 ◽

2005 ◽

Vol 288 (6) ◽

pp. L1162-L1170 ◽

Cited By ~ 46

Author(s):

Robert P. Jankov ◽

Crystal Kantores ◽

Rosetta Belcastro ◽

Soojin Yi ◽

Ross A. Ridsdale ◽

...

Keyword(s):

Pulmonary Hypertension ◽

Smooth Muscle ◽

Rat Model ◽

Vascular Remodeling ◽

Critical Role ◽

Receptor Expression ◽

Newborn Rat ◽

Arterial Smooth Muscle

Newborn rats exposed to 60% O2 for 14 days develop endothelin (ET)-1-dependent pulmonary hypertension with vascular remodeling, characterized by increased smooth muscle cell (SMC) proliferation and medial thickening of pulmonary resistance arteries. Using immunohistochemistry and Western blot analyses, we examined the effect of exposure to 60% O2 on expression in the lung of receptors for the platelet-derived growth factors (PDGF), which are implicated in the pathogenesis of arterial smooth muscle hyperplasia. We observed a marked O2-induced upregulation of PDGF-α and -β receptors (PDGF-αR and -βR) on arterial smooth muscle. This led us to examine pulmonary vascular PDGF receptor expression in 60% O2-exposed rats given SB-217242, a combined ET receptor antagonist, which we found prevented the O2-induced upregulation of PDGF-βR, but not PDGF-αR, on arterial smooth muscle. PDGF-BB, a major PDGF-βR ligand, was found to be a potent in vitro inducer of hyperplasia and DNA synthesis in cultured pulmonary artery SMC from infant rats. A critical role for PDGF-βR ligands in arterial SMC proliferation was confirmed in vivo using a truncated soluble PDGF-βR intervention, which attenuated SMC proliferation induced by exposure to 60% O2. Collectively, these data are consistent with a major role for PDGF-βR-mediated SMC proliferation, acting downstream of increased ET-1 in a newborn rat model of 60% O2-induced pulmonary hypertension.

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In vivo assessment of a single adenine mutation in 5′UTR of Endothelin-1 gene in paediatric cases with severe pulmonary hypertension: an observational study

BMC Research Notes ◽

10.1186/s13104-021-05609-5 ◽

2021 ◽

Vol 14 (1) ◽

Author(s):

Abhishek Kumar ◽

Minati Choudhury ◽

Sakshi Dhingra Batra ◽

Kriti Sikri ◽

Anushree Gupta

Keyword(s):

Pulmonary Hypertension ◽

Congenital Heart ◽

Small Sample Size ◽

Small Sample ◽

Severe Pulmonary Hypertension ◽

Endothelin 1 ◽

Circulating Levels ◽

In Vivo Assessment

Abstract Objective Endothelin-1 plays an important role in the pathogenesis of severe pulmonary hypertension. The + 139 ‘A’, adenine insertion variant in 5′UTR of edn1 gene has been reported to be associated with increased expression of Endothelin-1 in vitro. The aim of present study was to explore the association of this variant with the circulating levels of Endothelin-1 in vivo using archived DNA and plasma samples from 38 paediatric congenital heart disease (cyanotic and acyanotic) patients with severe pulmonary hypertension. Results The plasma Endothelin-1 levels were highly varied ranging from 1.63 to75.16 pg/ml. The + 139 ‘A’ insertion variant in 5′UTR of edn1 was seen in 8 out of 38 cases with only one acyanotic sample demonstrating homozygosity of inserted ‘A’ allele at + 139 site (4A/4A genotype). The plasma Endothelin-1 levels in children with homozygous variant 3A/3A genotype were comparable in cyanotic and acyanotic groups. Lone 4A/4A acyanotic sample had ET-1 levels similar to the median value of ET-1 associated with 3A/3A genotype and was absent in cyanotic group presumably due to deleterious higher ET-1 levels. The discussed observations, limited by the small sample size, are suggestive of homozygous adenine insertion variant posing a risk in cyanotic babies with Severe Pulmonary Hypertension.

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A Collagen-Based Scaffold for Promoting Neural Plasticity in a Rat Model of Spinal Cord Injury

Polymers ◽

10.3390/polym12102245 ◽

2020 ◽

Vol 12 (10) ◽

pp. 2245

Author(s):

Jue-Zong Yeh ◽

Ding-Han Wang ◽

Juin-Hong Cherng ◽

Yi-Wen Wang ◽

Gang-Yi Fan ◽

...

Keyword(s):

Spinal Cord ◽

Spinal Cord Injury ◽

Rat Model ◽

Neural Plasticity ◽

Collagen Scaffold ◽

Glial Scar ◽

Beneficial Effects ◽

Cord Injury

In spinal cord injury (SCI) therapy, glial scarring formed by activated astrocytes is a primary problem that needs to be solved to enhance axonal regeneration. In this study, we developed and used a collagen scaffold for glial scar replacement to create an appropriate environment in an SCI rat model and determined whether neural plasticity can be manipulated using this approach. We used four experimental groups, as follows: SCI-collagen scaffold, SCI control, normal spinal cord-collagen scaffold, and normal control. The collagen scaffold showed excellent in vitro and in vivo biocompatibility. Immunofluorescence staining revealed increased expression of neurofilament and fibronectin and reduced expression of glial fibrillary acidic protein and anti-chondroitin sulfate in the collagen scaffold-treated SCI rats at 1 and 4 weeks post-implantation compared with that in untreated SCI control. This indicates that the collagen scaffold implantation promoted neuronal survival and axonal growth within the injured site and prevented glial scar formation by controlling astrocyte production for their normal functioning. Our study highlights the feasibility of using the collagen scaffold in SCI repair. The collagen scaffold was found to exert beneficial effects on neuronal activity and may help in manipulating synaptic plasticity, implying its great potential for clinical application in SCI.

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Accelerated Repurposing and Drug Development of Pulmonary Hypertension Therapies for COVID-19 Treatment Using an AI-Integrated Biosimulation Platform

Molecules ◽

10.3390/molecules26071912 ◽

2021 ◽

Vol 26 (7) ◽

pp. 1912

Author(s):

Kaushik Chakravarty ◽

Victor G. Antontsev ◽

Maksim Khotimchenko ◽

Nilesh Gupta ◽

Aditya Jagarapu ◽

...

Keyword(s):

Pulmonary Hypertension ◽

Renin Angiotensin System ◽

Mechanistic Modeling ◽

Channel Blockers ◽

Angiotensin System ◽

In Silico Modeling ◽

Site Of Action ◽

Line Of Therapy

The COVID-19 pandemic has reached over 100 million worldwide. Due to the multi-targeted nature of the virus, it is clear that drugs providing anti-COVID-19 effects need to be developed at an accelerated rate, and a combinatorial approach may stand to be more successful than a single drug therapy. Among several targets and pathways that are under investigation, the renin-angiotensin system (RAS) and specifically angiotensin-converting enzyme (ACE), and Ca2+-mediated SARS-CoV-2 cellular entry and replication are noteworthy. A combination of ACE inhibitors and calcium channel blockers (CCBs), a critical line of therapy for pulmonary hypertension, has shown therapeutic relevance in COVID-19 when investigated independently. To that end, we conducted in silico modeling using BIOiSIM, an AI-integrated mechanistic modeling platform by utilizing known preclinical in vitro and in vivo datasets to accurately simulate systemic therapy disposition and site-of-action penetration of the CCBs and ACEi compounds to tissues implicated in COVID-19 pathogenesis.

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Clindamycin-loaded titanium prevents implant-related infection through blocking biofilm formation

Journal of Biomaterials Applications ◽

10.1177/08853282211051183 ◽

2021 ◽

pp. 088532822110511

Author(s):

Youbin Li ◽

Shaochuan Wang ◽

Shidan Li ◽

Jun Fei

Keyword(s):

Surface Modification ◽

Rat Model ◽

Biofilm Formation ◽

High Performance ◽

Bacterial Colonization ◽

Tissue Homogenate ◽

Tartrate Resistant Acid Phosphatase ◽

Layer By Layer

Implant-related infection is a disastrous complication. Surface modification of titanium is considered as an important strategy to prevent implant-related infection. However, there is no recognized surface modification strategy that can be applied in clinic so far. We explored a new strategy of coating. The clindamycin-loaded titanium was constructed by layer-by-layer self-assembly. The release of clindamycin from titanium was detected through high performance liquid chromatography. Different titanium was co-cultured with Staphylococcus aureus for 24 h in vitro, then the effect of different titanium on bacterial colonization and biofilm formation was determined by spread plate method and scanning electron microscopy. Cytotoxicity and cytocompatibility of clindamycin-loaded titanium on MC3T3-E1 cells were measured by CCK8. The antibacterial ability of clindamycin-loaded titanium in vivo was also evaluated using a rat model of osteomyelitis. The number of osteoclasts in bone defect was observed by tartrate-resistant acid phosphatase staining. Bacterial burden of surrounding tissues around the site of infection was calculated by tissue homogenate and colony count. Clindamycin-loaded titanium could release clindamycin slowly within 160 h. It reduced bacterial colonization by three orders of magnitude compare to control ( p < .05) and inhibits biofilm formation in vitro. Cells proliferation and adhesion were similar on three titanium surfaces ( p > .05). In vivo, clindamycin-loaded titanium improved bone healing, reduced microbial burden, and decreased the number of osteoclasts compared control titanium in the rat model of osteomyelitis. This study demonstrated that clindamycin-loaded titanium exhibited good biocompatibility, and showed antibacterial activity both in vivo and in vitro. It is promising and might have potential for clinical application.

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Virulence and Genetic Characterization of Six Baculovirus Strains Isolated From Different Populations of Spodoptera Frugiperda (Lepidoptera: Noctuidae)

10.21203/rs.3.rs-988050/v1 ◽

2021 ◽

Author(s):

Ingrid Zanella-Saenz ◽

Elisabeth A. Herniou ◽

Jorge E. Ibarra ◽

Ma.Cristina Del Rincón-Castro ◽

Ilse Alejandra Huerta-Arredondo

Keyword(s):

Biological Control ◽

Cell Cultures ◽

Spodoptera Frugiperda ◽

Fall Armyworm ◽

The United States ◽

Infection Mechanisms ◽

Different Populations

Abstract Fall armyworm (FAW), Spodoptera frugiperda (Smith, 1797), is a polyphagous, voracious, and economically important agricultural pest. Biological control of FAW is a strategy that must be further explored. This study evaluated six baculovirus strains isolated from infected FAW larvae from Mexico, Argentina, Honduras, and the United States. Five alphabaculoviruses (SfNPV-An2, SfNPV-Arg, SfNPV-Fx, SfNPV-Ho and SfNPV-Sin) and one betabaculovirus (SfGV-RV), were tested against FAW larvae, showing a wide diversity of virulence levels among strains when their estimated LC50s were compared, being SfNPVArg, SfNPV-Ho and SfNPV-Fx more virulent than SfNPV-An 2 , SfNPV-Sin and SfGV-RV. To determine any virulence difference in vitro studies of these isolates, Sf9 cell cultures were used. Interestingly, only ODVs from four of the test SfNPV strains showed infectivity on Sf9 cell cultures, and some differences in virulence were observed. Genomic restriction analyses and partial sequences of lef-8, lef-9 , and polh/granulin genes showed little variability among alphabaculoviruses, both, among them and with previously reported sequences. However, sequences from SfGV-RV were closer to previously reported sequences from the SfGVVG008 strain than the SfGV-Arg and SfGV-VG014 strains. The great difference in the in vivo virulence was not correlated with great similarity among the isolates. The characterization of these six baculoviruses isolates offers the basis for exploring their potential as biological control agents against S. frugiperda, as well the initial studies on their specific infection mechanisms, evolution, and ecology.

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Peptide Blocking Self-Polymerization of Extracellular Calcium-Sensing Receptor Attenuates Hypoxia-Induced Pulmonary Hypertension

Hypertension ◽

10.1161/hypertensionaha.120.16712 ◽

2021 ◽

Author(s):

Rui Xiao ◽

Shengquan Luo ◽

Ting Zhang ◽

Yankai Lv ◽

Tao Wang ◽

...

Keyword(s):

Pulmonary Hypertension ◽

Disulfide Bonds ◽

Acute Hypoxia ◽

Extracellular Domain ◽

Left Ventricular ◽

Extracellular Calcium ◽

Calcium Sensing Receptor ◽

Calcium Sensing

Activation of the CaSR (extracellular calcium-sensing receptor) has been recognized as a critical mediator of hypoxia-induced pulmonary hypertension. Preventive targeting of the early initiating phase as well as downstream events after CaSR activation remains unexplored. As a representative of the G protein-coupled receptor family, CaSR polymerizes on cell surface upon stimulation. Immunoblotting together with MAL-PEG technique identified a reactive oxygen species-sensitive CaSR polymerization through its extracellular domain in pulmonary artery smooth muscle cells upon exposure to acute hypoxia. Fluorescence resonance energy transfer screening employing blocking peptides determined that cycteine129/131 residues in the extracellular domain of CaSR formed intermolecular disulfide bonds to promote CaSR polymerization. The monitoring of intracellular Ca 2+ signal highlighted the pivotal role of CaSR polymerization in its activation. In contrast, the blockade of disulfide bonds formation using a peptide decreased both CaSR and hypoxia-induced mitogenic factor expression as well as other hypoxic-related genes in vitro and in vivo and attenuated pulmonary hypertension development in rats. The blocking peptide did not affect systemic arterial oxygenation in vivo but inhibited acute hypoxia-induced pulmonary vasoconstriction. Pharmacokinetic analyses revealed a more efficient lung delivery of peptide by inhaled nebulizer compared to intravenous injection. In addition, the blocking peptide did not affect systemic arterial pressure, body weight, left ventricular function, liver, or kidney function or plasma Ca 2+ level. In conclusion, a peptide blocking CaSR polymerization reduces its hypoxia-induced activation and downstream events leading to pulmonary hypertension and represents an attractive inhaled preventive alternative worthy of further development.

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Comparison of the hemodynamic effects of nitric oxide and endothelium-dependent vasodilators in intact lungs

Journal of Applied Physiology ◽

10.1152/jappl.1990.68.2.735 ◽

1990 ◽

Vol 68 (2) ◽

pp. 735-747 ◽

Cited By ~ 51

Author(s):

S. L. Archer ◽

K. Rist ◽

D. P. Nelson ◽

E. G. DeMaster ◽

N. Cowan ◽

...

Keyword(s):

Nitric Oxide ◽

Methylene Blue ◽

Pulmonary Hypertension ◽

Feedback Inhibition ◽

Pressor Response ◽

Pulmonary Vasculature ◽

Hemodynamic Effects ◽

Isolated Lung

The effects of endothelium-dependent vasodilation on pulmonary vascular hemodynamics were evaluated in a variety of in vivo and in vitro models to determine 1) the comparability of the hemodynamic effects of acetylcholine (ACh), bradykinin (BK), nitric oxide (NO), and 8-bromo-guanosine 3′,5′-cyclic monophosphate (cGMP), 2) whether methylene blue is a useful inhibitor of endothelium-dependent relaxing factor (EDRF) activity in vivo, and 3) the effect of monocrotaline-induced pulmonary hypertension on the responsiveness of the pulmonary vasculature to ACh. In isolated rat lungs, which were preconstricted with hypoxia, ACh, BK, NO, and 8-bromo-cGMP caused pulmonary vasodilation, which was not inhibited by maximum tolerable doses of methylene blue. Methylene blue did not inhibit EDRF activity in any model, despite causing increased pulmonary vascular tone and responsiveness to various constrictor agents. There were significant differences in the hemodynamic characteristics of ACh, BK, and NO. In the isolated lung, BK and NO caused transient decreases of hypoxic vasoconstriction, whereas ACh caused more prolonged vasodilation. Pretreatment of these lungs with NO did not significantly inhibit ACh-induced vasodilation but caused BK to produce vasoconstriction. Tachyphylaxis, which was agonist specific, developed with repeated administration of ACh or BK but not NO. Tachyphylaxis probably resulted from inhibition of the endothelium-dependent vasodilation pathway proximal to NO synthesis, because it could be overcome by exogenous NO. Pretreatment with 8-bromo-cGMP decreased hypoxic pulmonary vasoconstriction and, even when the hypoxic pressor response had largely recovered, subsequent doses of ACh and NO failed to cause vasodilation, although BK produced vasoconstriction. These findings are compatible with the existence of feedback inhibition of the endothelium-dependent relaxation by elevation of cGMP levels. Responsiveness to ACh was retained in lungs with severe monocrotaline-induced pulmonary hypertension. Many of these findings would not have been predicted based on in vitro studies and illustrate the importance for expanding studies of EDRF to in vivo and ex vivo models.

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