Xbp1s-Ddit3 promotes MCT-induced pulmonary hypertension

Pulmonary hypertension (PH) is a life-threatening disease characterized by vascular remodeling. Exploring new therapy target is urgent. The purpose of this study is to investigate whether and how spliced x-box binding protein 1 (xbp1s), a key component of endoplasmic reticulum stress (ERS), contributes to the pathogenesis of PH. Forty male SD rats were randomly assigned to four groups: Control, Monocrotalin (MCT), MCT+AAV-CTL (control), and MCT+AAV-xbp1s. The xbp1s protein levels were found to be elevated in lung tissues of the MCT group. Intratracheal injection of adeno-associated virus serotype 1 carrying xbp1s shRNA (AAV-xbp1s) to knock down the expression of xbp1s effectively ameliorated the MCT-induced elevation of right ventricular systolic pressure (RVSP), total pulmonary resistance (TPR), right ventricular hypertrophy and medial wall thickness of muscularized distal pulmonary arterioles. The abnormally increased positive staining rates of PCNA and Ki67 and decreased positive staining rates of TUNEL in pulmonary arterioles were also reversed in the MCT+AAV-xbp1s group. For mechanistic exploration, bioinformatic prediction of the protein network was performed on the STRING database, and further verification was performed by qRT-PCR, western blots and coimmunoprecipitation. Ddit3 (DNA damage-inducible transcript 3) was identified as a downstream protein that interacted with xbp1s. Overexpression of Ddit3 restored the decreased proliferation, migration, and cell viability caused by silencing of xbp1s. The protein level of Ddit3 was also highly consistent with xbp1s in the animal model. Taken together, our study demonstrated that xbp1s-Ddit3 may be a potential target to interfere with vascular remodeling in PH.

Correlation of postoperative magnetic resonance image measurements with persisting acetabular dysplasia in open reduction of developmental hip dysplasia

Objectives: The aim of this study was to evaluate correlation of post-reduction magnetic resonance imaging (MRI)-based parameters with residual acetabular dysplasia in developmental dysplasia of the hip (DDH) patients who underwent open reduction. Patients and methods: A total of 62 hips of 54 children (5 males, 57 females; mean age: 8.5±3.5 months; range, 0 to 24 months) with a diagnosis of DDH who underwent open reduction between January 2012 and January 2017 were retrospectively analyzed. The acetabular head index (AHI), head coverage index (HCI), sphericity, bony acetabular index (BAI), cartilage acetabular index (CAI), anterior acetabular index (AAI), posterior acetabular index (PAI), abduction angle (AA), and acetabular medial wall thickness were measured by MRI. The correlation between MRI measurements and residual acetabular dysplasia was evaluated. Results: The mean follow-up was 23.7±10.1 (range, 12 to 56) months. The mean age at the final examination was 47.6±10.4 months. The age at the time of operation (r=0.250, p=0.049), medial wall thickness (r=0.304, p=0.016), AAI (r=0.729, p<0.001), PAI (r=0.590, p<0.001), and early postoperative AI (r=0.900, p<0.001) at the third postoperative month were positively correlated with the last follow-up AI. The AHI (r=-0.512, p<0.001), sphericity (r=-0,661, p<0.001), and HCI (r=-0.554, p< 0.001) were negatively correlated with the last follow-up AI. Conclusion: Post-reduction MRI parameters can be used to evaluate correlation with persistent acetabular dysplasia in DDH patients.

Aortic Thickness: A Forgotten Paradigm in Risk Stratification of Aortic Disease

Background This study aimed at risk-stratifying aortic dilatation using aortic wall thickness (AWT) and comparing methods of AWT assessment. Methods Demographic, epidemiological, and perioperative data on 72 consecutive aortic surgeries (age = 62 years[standard deviation (SD) = 12] years) performed by a single surgeon were collected from hospital database. Aortic thickness was measured on computed tomography scans, as well as intraoperatively in four quadrants, at the level of aortic sinuses, as well as midascending aorta, using calipers. Aortic wall stress was calculated using standard mathematical formulae. Results The ascending aorta was 48.2 (SD = 8) mm and the mean thickness at ascending aorta level was 1.9 (SD = 0.3) mm. There was congruence between imaging and intraoperative measurements of thickness, as well as between the radiologist and surgeon. Preoperatively, 16 patients had multiple imaging studies showing an average rate of growth of 1.2 mm per year without significant difference in thickness. The wider the aorta, the thinner was the lateral or convex wall. Aortic stenosis (p = 0.01), lateral to medial wall thickness ratio (p = 0.04), and history of hypertension (p = 0.00), all had protective effect on aortic root stress. The ascending aortic stress was directly affected by age (p = 0.03) and inversely related to lateral to medial wall thickness ratio (p = 0.03). Conclusion Aortic thickness can be measured preoperatively and easily confirmed intraoperatively. Risk stratification based on both aortic thickness and diameter (stress calculations) would better predict acute aortic events in dilated aortas and define aortic resection criteria more objectively.

Effect of antenatal tetramethylpyrazine on lung development and YAP expression in a rat model of experimental congenital diaphragmatic hernia

Tetramethylpyrazine (TMP) is a chemical compound found in extracts derived from the Chinese medicinal plant. Due to its remarkable therapeutic effects, availability, and low cost and toxicity, TMP has been used to treat cardiovascular diseases and pulmonary hypertension in China. The aim of this study was to investigate the therapeutic effects and underlying mechanism of TMP on lung development using a rat model of nitrofen-induced congenital diaphragmatic hernia (CDH). Pregnant rats were divided into three groups: control, CDH, and CDH+TMP. Nitrofen was used to induce CDH. In the CDH and CDH+TMP, Fetuses only with left diaphragmatic hernias were chosen for analysis. Lung and body weight were recorded and lung histologic evaluations, image analysis, and western blot analysis of YAP, p-YAP and LATS1 were performed after lung processing. A marked abnormal structure was observed, as evidenced by pulmonary hypoplasia and vascular remodeling, in the CDH. These abnormalities were improved in the CDH+TMP. There were significant differences between the CDH and CDH+TMP in percentage of medial wall thickness, arteriole muscularization, radial alveolar counts, AA%, and alveolar septal thickness. YAP expression was markedly increased in the CDH compared to the control, which was not affected by antenatal TMP administration. However, prenatal TMP intervention significantly increased expression of LATS1 and phosphorylation of YAP in the CDH fetuses. Our results demonstrate that antenatal TMP administration improved vascular remodeling and promoted lung development in a rat model of CDH, potentially through increasing expression of LATS1 and phosphorylation of YAP.

P6377Cardiopulmonary effects of nitric oxide supplementation in a model of chronic hypoxia pulmonary hypertension

Objective We previously demonstrated that the inhibition of phosphodiesterase type-5, an enzyme that degrades cGMP into inactive 5'-GMP, attenuates pulmonary remodeling and right ventricular (RV) hypertrophy (RVH) during exposure to chronic hypoxia (CH). The nitric oxide (NO) pathway is thought to play a major role in these changes. In this study, we investigate whether L-Arginine (L-ARG), a substrate of endothelial NO synthase (eNOS) and molsidomine (MOL), a NO donor, might alleviate the cardiovascular and pulmonary dysfunction led by CH. Methods Male rats (n=80; 10/group) were maintained in normoxic (21% O2) or hypoxic chambers (10% O2) for 14 days. Hypoxic rats were subdivided in four groups: untreated control, treated with L-ARG (45 mg/kg), eNOS inhibitor N-nitro-L-arginine methyl ester (L-NAME, 45 mg/kg) or MOL (15 mg/kg). Drugs were given daily in the drinking water. After sacrifice, we measured RV systolic pressure (RVSP), RV contractility (dP/dt), RVH, the lung/body weight ratio, the pulmonary vessels medial wall thickness, and the cardiac and pulmonary eNOS phosphorylation. Results Although CH increased RVSP, RV contractility and RVH, these increases were attenuated by L-ARG and MOL. Whereas L-ARG attenuated the RVH increase, MOL and L-NAME were ineffective. No treatment prevented the increase in lung/body weight ratio. Under all conditions, the lung tissue water content was unchanged, indicating no edema development. By contrast, CH rats developed a marked increase in medial wall thickness of small (0–100 mm) pulmonary arteries, while larger arteries were not affected. This increase was attenuated by L-ARG or MOL. Although CH decreased cardiac and pulmonary phosphorylated eNOS, L-ARG and MOL restored the normoxic level. Conclusions NO supplementation during CH attenuates RVSP, RVH and pulmonary remodeling, probably due to increased phosphorylation of eNOS. Despite normalizing RVSP, MOL does not influence RVH development.

Abstract 13158: Impact of Enhancing Fetal Lung Vascular and Alveolar Development by Slow-release Novel Synthetic Prostacyclin Agonist for Treatment of Fetal Lung Hypoplasia

Background: Lung hypoplasia and persistent pulmonary hypertension associated with critical pathologies, such as congenital diaphragmatic hernia (CDH), is the major cause of mortality in the neonates. Although prostaglandins pathway is known to play a pivotal role in the lung development, effectiveness of postnatal treatment by prostaglandin agonists was reported to be suboptimal. We hypothesized that prenatal treatment by ONO-1301SR, slow-release form of a novel synthetic prostacyclin agonist with thromboxane inhibitory activity, might promote development of lungs having hypoplasia in the fetal period. Methods: On embryonic day (E) 9.5, nitrofen was given to pregnant Sprague-Dawley rats to produce a CDH-relating lung hypoplasia model, while normal rats (n=4) received vehicle only. At the same day, either ONO-1301SR or placebo was randomly given to the nitrofen-treated rats (placebo; n=7, ONO; n=5). On E21.5, the normal fetuses, and the fetuses having CDH were analyzed. Results: Prenatal ONO-1301SR administration had no influence on incidence of nitrofen-induced CDH. One-seventh of maternal blood concentration of ONO-1301 was transferred to fetal blood at E21.5, warranting efficient placental permeability of this molecule. Prostacyclin receptor was expressed in pulmonary arterial smooth muscle cells in the fetus. Lung-to-body weight ratio (%) in the ONO group (1.88±0.07) was greater than that in the placebo group (1.60±0.04, p<0.01). Histologically, medial wall thickness in the ONO group was two-third thinner than that in the placebo group (p<0.01). In addition, the number of Ttf-1-positive cells and capillary density were 1.5 times or more greater in the ONO group than that in the placebo group (p<0.05), associated with a greater expression of VEGF and SDF-1 in the ONO group, suggesting enhanced development of the alveolar and capillary network. Conclusions: Prenatal ONO-1301SR administration promoted efficient development of hypoplastic lungs associated with CDH in the nitrofen-induced rat model, indicating potential of this treatment for pathologies having lung hypoplasia.

Spaceflight on the Bion-M1 biosatellite alters cerebral artery vasomotor and mechanical properties in mice

Conditions during spaceflight, such as the loss of the head-to-foot gravity vector, are thought to potentially alter cerebral blood flow and vascular resistance. The purpose of the present study was to determine the effects of long-term spaceflight on the functional, mechanical, and structural properties of cerebral arteries. Male C57BL/6N mice were flown 30 days in a Bion-M1 biosatellite. Basilar arteries isolated from spaceflight (SF) ( n = 6), habitat control (HC) ( n = 6), and vivarium control (VC) ( n = 16) mice were used for in vitro functional and mechanical testing and histological structural analysis. The results demonstrate that vasoconstriction elicited through a voltage-gated Ca2+ mechanism (30–80 mM KCl) and thromboxane A2 receptors (10−8 − 3 × 10−5 M U46619) are lower in cerebral arteries from SF mice. Inhibition of Rho-kinase activity (1 μM Y27632) abolished group differences in U46619-evoked contractions. Endothelium-dependent vasodilation elicited by acetylcholine (10 μM, 2 μM U46619 preconstriction) was virtually absent in cerebral arteries from SF mice. The pressure-diameter relation was lower in arteries from SF mice relative to that in HC mice, which was not related to differences in the extracellular matrix protein elastin or collagen content or the elastin/collagen ratio in the basilar arteries. Diameter, medial wall thickness, and medial cross-sectional area of unpressurized basilar arteries were not different among groups. These results suggest that the microgravity-induced attenuation of both vasoconstrictor and vasodilator properties may limit the range of vascular control of cerebral perfusion or impair the distribution of brain blood flow during periods of stress.

Macitentan reverses early obstructive pulmonary vasculopathy in rats: early intervention in overcoming the survivin-mediated resistance to apoptosis

It remains unknown whether current disease-targeting therapy can histologically reverse obstructive pulmonary vasculopathy and how the timing of the therapy influences the antiremodeling effects of the compound. We test the hypothesis that a novel endothelin receptor antagonist macitentan reverses the early and/or late stages of occlusive pulmonary vascular disease (PVD) in rats. Rats with pulmonary arterial hypertension (PAH), which were produced by combined exposure to a vascular endothelial growth factor receptor inhibitor Sugen 5416 and hypobaric hypoxia for 3 wk, were assigned to receive macitentan or vehicle during 3–5 wk (early study) or during 5–8 wk (late study) after Sugen injection. Compared with vehicle-treated PAH rats and PAH rats evaluated before treatment initiation, the macitentan-treated rats showed decreases in the proportion of occlusive lesions in the early study, a finding consistent with the reversal of right ventricular systolic pressure and indexes of right ventricular hypertrophy and medial wall thickness. Macitentan ameliorated but did not reverse the proportion of occlusive lesions in the late study. Although macitentan decreased the proportion of Ki67+ lesions in both studies, macitentan increased the proportion of cleaved caspase 3+ lesions and suppressed an antiapoptotic molecule survivin expression in the early study but not in the late study. In conclusion, macitentan reversed early but not late obstructive PVD in rats. This reversal was associated with the suppression of survivin-related resistance to apoptosis and proliferation of cells in PVD.

Antenatal imatinib treatment reduces pulmonary vascular remodeling in a rat model of congenital diaphragmatic hernia

The pathophysiology of congenital diaphragmatic hernia (CDH) is constituted by pulmonary hypoplasia and pulmonary hypertension (PH). We previously reported successful treatment with imatinib of a patient with CDH. This study examines the effect of antenatal imatinib administration on the pulmonary vasculature in a rat model of CDH. Pregnant rats were given nitrofen to induce CDH. Controls were given olive oil. Half of the CDH fetuses and half of the controls were treated with imatinib antenatally E17-E21, rendering four groups: Control, Control+Imatinib, CDH, and CDH+Imatinib. Lung sections were obtained for morphometry and immunohistochemistry, and protein was purified for Western blot. Effects of nitrofen and imatinib on Ki-67, caspase-3, PDGF-B, and PDGF receptors were analyzed. Imatinib significantly reduced medial wall thickness in pulmonary arteries of rats with CDH. It also normalized lumen area and reduced the proportion of fully muscularized arteries. Imatinib also caused medial thinning in the control group. Cell proliferation was increased in CDH, and this proliferation was significantly reduced by imatinib. PDGF-B and PDGFR-β were upregulated in CDH, and imatinib treatment resulted in a downregulation. PDGFR-α remained unchanged in CDH but was significantly downregulated by imatinib. Antenatal imatinib treatment reduces development of medial wall thickness and restores lumen area in pulmonary arteries in nitrofen-induced CDH. The mechanism is reduced cell proliferation. Imatinib is an interesting candidate for antenatal therapy for PH in CDH, but potential side effects need to be investigated and more specific targeting of PDGF signaling is needed.

Computed Tomographic Measurement of Cervical Pedicles for Transpedicular Fixation in a Malay Population

Purpose. To measure the cervical pedicles and assess the feasibility of transpedicular fixation in a Malay population. Methods. 960 computed tomography (CT) scans of bilateral C2 to C7 pedicles of 80 Malays were compared. 22 men and 24 women aged <60 (mean, 37.3; range, 18–56) years were defined as young patients, whereas 18 men and 16 women aged ≥60 (mean, 63.9; range, 60–76) years as elderly patients. An inner diameter of <3.0 mm (85% of a 3.5-mm screw) was defined as ‘unfeasible’ and a medial or lateral wall thickness of <1.0 mm as ‘unsafe’ for cervical pedicle screw fixation. Results. In the respective young versus elderly groups, the inner diameters ranged from 1.94 to 2.80 mm versus 2.51 to 3.37 mm in men, and from 1.52 to 2.31 mm versus 1.64 to 2.46 mm in women. Medial wall thickness ranged from 1.25 to 1.46 mm versus 1.13 to 1.48 mm in men, and from 1.28 to 1.72 mm versus 1.10 to 1.24 mm in women. Lateral wall thickness ranged from 0.80 to 0.90 mm versus 0.66 to 0.88 mm in men, and from 0.85 to 0.99 mm versus 0.59 to 0.86 mm in women. Conclusion. The cervical spine of Malays may not be adequate to accommodate a 3.5-mm pedicle screw for transpedicular fixation, as this procedure may risk adjacent vital structures.