Echocardiography, a non‐invasive and cost‐effective method for monitoring cardiac function, is commonly used for evaluation and pre‐clinical diagnostics of pulmonary hypertension (PH). Previous echocardiographic studies in experimental models of PH are fragmentary in terms of the evaluation of right ventricle (RV) function. In this study, three rodent models of PH: a mouse model of hypoxia‐induced PH, a rat model of hypoxia+Sugen induced PH and a rat model of monocrotaline‐induced PH, were employed to measure RV fractional area change (RVFAC), RV free wall thickness (RVFWT), pulmonary acceleration time (PAT), pulmonary ejection time (PET), and tricuspid annular plane systolic excursion (TAPSE). We found that, in these models, RVFWT significantly increased, but RVFAC, PAT, or PAT/PET ratios and TAPSE values significantly decreased. Accurate and complete TAPSE patterns were demonstrated in the three rodent models of PH. The RV echocardiography data matched the corresponding invasive hemodynamic and heart histologic data in each model. This serves as a reference study for real‐time and non‐invasive evaluation of RV function in rodent models of PH using echocardiography.

Hemin, the oxidized prosthetic moiety of hemoglobin, has been implicated in the pathogenesis of acute chest syndrome in patients with sickle cell disease by virtue of its endothelial-activating properties. In this study, we examined whether hemin can cause lung microvascular endothelial barrier dysfunction. By assessing transendothelial resistance using electrical cell impedance sensing, and by directly measuring trans-monolayer fluorescein isothiocyanate-dextran flux, we found that hemin does cause endothelial barrier dysfunction in a concentration-dependent manner. Pretreatment with either a Toll-like receptor 4 inhibitor, TAK-242, or an antioxidant, N-acetylcysteine, abrogated this effect. Increased monolayer permeability was found to be associated with programmed cell death by necroptosis, as evidenced by Trypan blue staining, terminal deoxynucleotidyl transferase dUTP nick-end labeling assay, Western blotting for activated forms of key effectors of cell death pathways, and studies utilizing specific inhibitors of necroptosis and apoptosis. Further studies examining the role of endothelial cell necroptosis in promoting noncardiogenic pulmonary edema during acute chest syndrome are warranted and may open a new avenue of potential treatments for this devastating disease.

Sphingosine kinase 1 (SPHK1) and the sphingosine-1-phosphate (S1P) signaling pathway have been shown to play a role in pulmonary arterial hypertension (PAH). S1P is an important stimulus for pulmonary artery smooth muscle cell (PASMC) proliferation and pulmonary vascular remodeling. We aimed to examine the specific roles of SPHK1 in PASMCs during pulmonary hypertension (PH) progression. We generated smooth muscle cell-specific, Sphk1-deficient (Sphk1^f/f TaglnCre⁺) mice and isolated Sphk1-deficient PASMCs from SPHK1 knockout mice. We demonstrated that Sphk1^f/f TaglnCre⁺ mice are protected from hypoxia or hypoxia/Sugen-mediated PH, and pulmonary vascular remodeling and that Sphk1-deficient PASMCs are less proliferative compared with ones isolated from wild-type (WT) siblings. S1P or hypoxia activated yes-associated protein 1 (YAP1) signaling by enhancing its translocation to the nucleus, which was dependent on SPHK1 enzymatic activity. Further, verteporfin, a pharmacologic YAP1 inhibitor, attenuated the S1P-mediated proliferation of hPASMCs, hypoxia-mediated PH, and pulmonary vascular remodeling in mice and hypoxia/Sugen-mediated severe PH in rats. Smooth muscle cell-specific SPHK1 plays an essential role in PH via YAP1 signaling, and YAP1 inhibition may have therapeutic potential in treating PH.

Community associated methicillin-resistant Staphylococcus aureus (CA-MRSA) is an emerging threat to human health throughout the world. Rodent MRSA pneumonia models mainly focus on the early innate immune responses to MRSA lung infection. However, the molecular pattern and mechanisms of recovery from MRSA lung infection are largely unknown. In this study, a sublethal mouse MRSA pneumonia model was employed to investigate late events during the recovery from MRSA lung infection. We compared lung bacterial clearance, bronchoalveolar lavage fluid (BALF) characterization, lung histology, lung cell proliferation, lung vascular permeability and lung gene expression profiling between days 1 and 3 post MRSA lung infection. Compared to day 1 post infection, bacterial colony counts, BALF total cell number and BALF protein concentration significantly decreased at day 3 post infection. Lung cDNA microarray analysis identified 47 significantly up-regulated and 35 down-regulated genes (p<0.01, 1.5 fold change [up and down]). The pattern of gene expression suggests that lung recovery is characterized by enhanced cell division, vascularization, wound healing and adjustment of host adaptive immune responses. Proliferation assay by PCNA staining further confirmed that at day 3 lungs have significantly higher cell proliferation than at day 1. Furthermore, at day 3 lungs displayed significantly lower levels of vascular permeability to albumin, compared to day 1. Collectively, this data helps us elucidate the molecular mechanisms of the recovery after MRSA lung infection.

Blood paper cards provide an effective DNA storage method. In this study, we used three DNA dissolving reagents (Tris-EDTA [TE] buffer, Tris–HCl buffer, and water) and one common commercially available kit (DN131 from MRC Inc) to elute DNA from 105 human blood paper cards collected up to 10 years ago. These DNA samples were used as templates for amplification of a single nucleotide polymorphism (SNP, C125T) region of human caspase-12 by PCR and a specific Taqman genotyping assay using the same amount of DNA. We show that DNA isolated by Tris–HCl buffer has higher yield and quality in comparison to DN131 solution. PCR success rate to amplify caspase-12 C125T SNP using Tris–HCl is comparable to the method using DN131 (89.5% vs 87.6%). The Taqman genotyping success rate using Tris–HCl is higher than using DN131 (81.9% vs 70.5%). Using TE or water, PCR success rates are lower than using DN131 (73.3% [TE]; 72.4% [H₂O]), but Taqman genotyping success rates are comparable to the method using DN131 (70.5% [TE]; 79.1% [H₂O]). We concluded that using Tris–HCl is a reliable and effective method to elute DNA from old human blood paper cards. The crude DNA isolated by Tris–HCl can be used to study genetic polymorphisms in human populations.