OBJECTIVETo investigate the feasibility of transferring p21 gene into lung tissue with recombinant adenovirus with full-length cDNA of p21 inserted in the Wistar rat model of pulmonary hypertension (PAH) induced by left-to-right shunt, study the expression of the desired gene in vivo, find if overexpression of desired gene can inhibit pulmonary hypertension.

METHODSWith full-length cDNA of p21 transfected HEK293 cells with clonfectin, and was packed, amplified in order to obtain the high-titer recombinant adenovirus (AdCMV-p21). The infection titer was determined by TCID50 method and was diluted into 1.67 x 10(8) pfu/L. Wistar rats were randomly allocated to control group (n = 10), model group (n = 15), test group (n = 10) and test control group (n = 10). In model group and test group left-to-right shunt pulmonary hypertension was developed by using cuff technique. AdCMV-p21 was transfected into test group and test control group using tracheal inhalation. The mPAP, mRVP and RVHI were measured and compared between every two groups. The left lung was immunohistochemically stained to observe the result of transfection. The right lung was HE stained to observe morphological changes in arteria pulmonalis and calculate WT%.

RESULTSThe mRVP, mPAP and WT% in model group and test group were significantly higher than those in control (P < 0.05), which suggested that the rat model of PAH was established successfully. Brown spots in the nucleus of VSMCs of pulmonary artery were seen in test group and test control group, which indicated that AdCMV-p21 was transfected successfully. The rate of transfected cells in test group was (42.8 +/- 11.6)%, which was equal to that of test control group (P > 0.05). In test group, the mPAP was (20.06 +/- 3.40) mm Hg (1 mm Hg = 0.133 kPa), mRVP was (22.53 +/- 2.53) mm Hg, WT% was (30.8 +/- 3.5)%, which were significantly lower than those in model group (P < 0.05), but higher than those in control group and test control group (P < 0.05).

CONCLUSIONThe recombinant adenovirus could successfully carry p21 and transfect the lung tissue of PAH rat model, and full expression of p21. p21 gene could inhibit the development of PAH.

Adenoviridae ; genetics ; Animals ; Hypertension, Pulmonary ; genetics ; therapy ; Male ; Muscle, Smooth, Vascular ; Oncogene Protein p21(ras) ; genetics ; Rats ; Rats, Wistar ; Transfection

OBJECTIVE: To explore the diagnosis, treatment and genetic characteristics of a neonate with severe pulmonary hypertension and respiratory failure. METHODS: Perinatal history, clinical manifestations, laboratory finding and diagnosis and treatment data of the child were collected. Whole exome sequencing was carried out for the child, and Sanger sequencing was used to verify the candidate variants. RESULTS: The female neonate has developed progressive respiratory failure and refractory pulmonary hypertension shortly after birth. Conventional treatment such as mechanical ventilation, vasoactive drugs, and inhaled nitric oxide were ineffective. She has developed sustained pulmonary hypertension after weaning from extracorporeal membrane oxygenation therapy, and had died after the treatment had ceased. Whole exome sequencing revealed that she has harbored a heterozygous de novo variant of c.682_683insGCGGCGGC (p.G234Rfs*148) of the FOXF1 gene, which was predicted as pathogenic based on guidelines from the American College of Medical Genetics and Genomics (ACMG), with evidence items of PVS1_Strong+PM2_Supporting+PS2. Based on her clinical manifestations and result of genetic testing, the child was diagnosed with alveolar capillary dysplasia with misalignment of the pulmonary veins (ACD/MPV). CONCLUSION Discovery of the c.682_683insGCGGCGGC (p.G234 Rfs*148) variant of the FOXF1 gene has expanded the mutational spectrum of the FOXF1 gene, which has facilitated implementation of specific treatment and provided a basis for clinical diagnosis and genetic counseling.
Female ; Humans ; Child ; Infant, Newborn ; Pregnancy ; Persistent Fetal Circulation Syndrome/therapy* ; Hypertension, Pulmonary ; Pulmonary Veins ; Forkhead Transcription Factors/genetics*

Pulmonary arterial hypertension (PAH) is a rare but progressive and currently incurable disease, which is characterized by vascular remodeling in association with muscularization of the arterioles, medial thickening and plexiform lesion formation. Despite our advanced understanding of the pathogenesis of PAH and the recent therapeutic advances, PAH still remains a fatal disease. In addition, the susceptibility to PAH has not yet been adequately explained. Much evidence points to the involvement of epigenetic changes in the pathogenesis of a number of human diseases including cancer, peripheral hypertension and asthma. The knowledge gained from the epigenetic study of various human diseases can also be applied to PAH. Thus, the pursuit of novel therapeutic targets via understanding the epigenetic alterations involved in the pathogenesis of PAH, such as DNA methylation, histone modification and microRNA, might be an attractive therapeutic avenue for the development of a novel and more effective treatment. This review provides a general overview of the current advances in epigenetics associated with PAH, and discusses the potential for improved treatment through understanding the role of epigenetics in the development of PAH.
Animals ; DNA Methylation/drug effects ; Drug Discovery/methods ; *Epigenesis, Genetic/drug effects ; Genetic Therapy/methods ; Humans ; Hypertension, Pulmonary/*genetics/therapy ; MicroRNAs/*genetics

BACKGROUNDHepatocyte growth factor (HGF) inhibits the development of pulmonary artery hypertension (PAH) by reducing pulmonary artery pressure and right ventricle (RV) hypertrophy. However, whether HGF can prevent RV remodeling via inhibiting apoptosis in RV cardiomyocytes and decreasing neurohormonal activation remains unknown.

METHODSThe PAH and subsequent RV remodeling in rats were induced by subcutaneous injection of monocrotaline (MCT). The PAH rats were transfected with adenovirus carrying HGF (Ad-HGF) via intratracheal instillation. Three weeks after transfection, the hemodynamics indexes were measured, serum levels for angiotonin II (ANG II) and brain natriuretic peptide (BNP) were determined by ELISA. Histological analysis was used to assess the RV hypertrophy and fibrosis. The cardiomyocyte apoptosis in RV was assayed by TUNEL staining. The mRNA expression of BNP, angiotensin-converting enzyme (ACE), Bax and Bcl-2 in RV was determined by reverse transcriptase polymerase chain reaction (RT-PCR), the protein expression of transforming growth factor (TGF)-β1 and tumor necrosis factor (TNF)-α in RV was determined by Western blotting.

RESULTSHGF treatment significantly decreased the mean PAH, RV systolic pressure, serum ANG II and BNP levels. HGF treatment also significantly decreased the RV hypertrophy, collagen deposition, and the number of apoptotic cardiomyocytes. Moreover, HGF treatmemt significantly decreased the expression of BNP, ACE, Bax, TGF-β1, and TNF-α, while it significantly increased the expression of Bcl-2.

CONCLUSIONSGene transfer of HGF decreases MCT-induced PAH and improves RV remodeling. This effect is mediated not only by improving the hemodynamics but also by decreasing neurohormonal activation and inhibiting cardiomyocytes apoptosis. HGF gene treatment may be an effective strategy for improving RV remodeling in MCT-induced PAH.

Animals ; Apoptosis ; genetics ; physiology ; Hepatocyte Growth Factor ; genetics ; physiology ; therapeutic use ; Humans ; Hypertension, Pulmonary ; metabolism ; therapy ; Male ; Rats ; Rats, Sprague-Dawley ; Ventricular Remodeling ; genetics ; physiology

OBJECTIVETo investigate the effect of vascular endothelial growth factor (VEGF) gene transfer on the bleomycin(BLM)-induced pulmonary hypertension in immature rabbits.

METHODSImmature rabbits were divided into 4 groups; control, BLM, liposome and trans-gene groups. The systolic, diastolic and mean pulmonary artery pressure (PASP, PADP, MPAP) were measured by micro-catheter, the pathological changes and the expression of VEGFmRNA and eNOSmRNA of endothelial cells in pulmonary arteries were evaluated by HE stain and in situ hybridization.

RESULT(1) The PAP of BLM and liposome groups was higher than that of control and trans-gene groups. The PASP was 16.5+/-2.9, 25.2+/-7.0, 24.4+/-6.0 and 18.3+/-2.7 mmHg; the PADP was 8.8+/-4.2, 13.1+/-3.8, 13.7+/-4.6 and 10.2+/-2.6 mmHg; the MPAP was 12.1+/-4.0, 18.4+/-4.7, 18.4+/-5.1 and 14.1+/-2.5 mmHg in control, BLM, liposome and trans-gene groups respectively. (2) The thickness of wall increased and the cavity became narrow, and the thickness index (TI) and area index (AI) increased in middle and small pulmonary arteries of BLM and liposome groups. The TI was 0.52+/-0.16, 0.65+/-0.16, 0.63+/-0.11 and 0.55+/-0.13; and the AI was 0.74+/-0.17, 0.84+/-0.14, 0.85+/-0.08 and 0.79+/-0.12 in control, BLM, liposome and trans-gene groups,respectively. (3) The level of VEGFmRNA and eNOSmRNA expression in pulmonary arterial endothelial cells decreased in BLM and liposome groups. The level of VEGFmRNA and eNOSmRNA expression in trans-gene group was higher than that in BLM and liposome groups, but lower than that in control group. VEGFmRNA was 0.83+/-0.09, 0.45+/-0.11, 0.45+/-0.13 and 0.65+/-0.18; eNOSmRNA was 0.79+/-0.12, 0.45+/-0.12, 0.50+/-0.14 and 0.56+/-0.08 in control, BLM, liposome and trans-gene groups, respectively.

CONCLUSIONVEGF gene transfer in immature rabbits with BLM-induced pulmonary hypertension could attenuate the increasing of PAP and wall thickness in middle and small pulmonary arteries, and increase the level of VEGFmRNA and eNOSmRNA expression in pulmonary arterial endothelial cells.

Animals ; Animals, Newborn ; Bleomycin ; Endothelium ; metabolism ; Gene Transfer Techniques ; Genetic Therapy ; Hypertension, Pulmonary ; chemically induced ; metabolism ; therapy ; Nitric Oxide Synthase Type III ; biosynthesis ; genetics ; Pulmonary Artery ; metabolism ; RNA, Messenger ; biosynthesis ; genetics ; Rabbits ; Vascular Endothelial Growth Factor A ; biosynthesis ; genetics

OBJECTIVETo investigate the effects of rapamycin (RAP) on pulmonary hypertension (PH) in rats, and to provide new insights into medication selection for the clinical treatment of PH.

METHODSFifty male Sprague-Dawley rats were randomly divided into blank control, PH model, solvent control, RAP 1, and RAP 2 groups. A rat model of PH was induced by left pneumonectomy (PE) and monocrotaline (MCT). At 5 days after PH model establishment, the solvent control group and the RAP 1 group received an intramuscular injection of solvent and RAP, respectively. At 35 days after PH model establishment, the RAP 2 group received an intramuscular injection of RAP. The mean pulmonary artery pressure (mPAP) and the right ventricle/left ventricle plus septum weight ratio (RV/LV+S) were measured in each group. Histopathological changes in the right lung were evaluated by hematoxylin-eosin (HE) staining. The relative expression of alpha-smooth muscle actin (α-SMA) and smooth muscle protein 22-alpha (SM22α) in each group was determined using real-time PCR.

RESULTSAt 35 days after surgery, the PH model and the solvent control groups had significantly higher mPAP and RV/LV+S than the blank control group, while the RAP 1 and the RAP 2 groups had significantly lower mPAP than the solvent control group (P<0.05). The RV/LV+S in the RAP 1 group was significantly lower than that in the solvent control group (P<0.05); however, there was no significant difference in RV/LV+S between the RAP 2 and the solvent control groups (P>0.05). HE staining in the right lung showed the substantially thickened pulmonary artery wall and narrowed arterial lumen in the PH model and the solvent control groups compared with the blank control group. Different degrees of reversal of the pulmonary artery wall thickening were observed after RAP administration. The results of real-time PCR revealed that the relative expression of α-SMA and SM22α in the PH model and the solvent control groups was significantly lower than in the blank control group, while the relative expression of α-SMA and SM22α in the RAP 1 and the RAP 2 groups was significantly higher than in the solvent control group (P<0.05).

CONCLUSIONSRAP can reverse the increase in pulmonary artery pressure and the right ventricular hypertrophy probably by regulation of the phenotypic conversion of vascular smooth muscle cells.

Actins ; genetics ; Animals ; Hemodynamics ; Hypertension, Pulmonary ; drug therapy ; physiopathology ; Hypertrophy, Right Ventricular ; etiology ; Male ; Microfilament Proteins ; genetics ; Muscle Proteins ; genetics ; Pulmonary Artery ; pathology ; RNA, Messenger ; analysis ; Rats ; Rats, Sprague-Dawley ; Sirolimus ; therapeutic use

In this study, we investigated the effects of Panax notoginseng saponins (PNS) on pulmonary vascular remodeling and ADAM10/Notch3 pathway in pulmonary arterial hypertension (PAH). PAH rat model was established, and male Sprague Dawley (SD) rats were randomly divided into control group, monocrotaline (MCT) group and MCT+PNS group, with 10 rats in each group. Rats in the control group were intraperitoneally injected with equal volume of normal saline. Rats in the MCT group was injected intraperitoneally with 60 mg/kg MCT on the first day, and then with the same volume of normal saline every day. Rats in the MCT+PNS group was injected intraperitoneally with 60 mg/kg MCT on the first day, and then with 50 mg/kg PNS every day. The modeling time of each group lasted for 21 days. After the model was established, the mean pulmonary artery pressure (mPAP) was measured by right heart catheterization technique, the right ventricular hypertrophy index (RVHI) was calculated, the microscopic morphology and changes of pulmonary vascular wall were observed by HE and Masson staining, and the expressions of ADAM10, Notch3, Hes-1, P27, PCNA, Caspase-3 proteins and mRNA in pulmonary vascular tissue of rats were detected by Western blot and qPCR. The expression and localization of Notch3 and α-SMA were detected by immunofluorescence staining. The protein expression of ADAM10 was detected by immunohistochemical staining. The results showed that compared with the control group, mPAP, RVHI, pulmonary vessels and collagen fibers in the MCT group were significantly increased, the expressions of ADAM10, Notch3, Hes-1, and PCNA protein and mRNA were significantly increased, while the expressions of P27 and Caspase-3 protein and mRNA were decreased significantly. Compared with the MCT group, mPAP and RVHI were significantly decreased, pulmonary vessels were significantly improved and collagen fibers were significantly reduced, the expressions of protein and mRNA of ADAM10, Notch3, Hes-1, and PCNA were decreased in MCT+PNS group, but the expressions of protein and mRNA of P27 and Caspase-3 were increased slightly. The results of immunofluorescence showed that Notch3 and α-SMA staining could overlap, which proved that Notch3 was expressed in smooth muscle cells. The expression of Notch3 in the MCT group was increased significantly compared with that in the control group, while PNS intervention decreased the expression of Notch3. Immunohistochemical staining showed that compared with the control group, the amount of ADAM10 in the MCT group was increased significantly, and the expression of ADAM10 in the MCT+PNS group was decreased compared with the MCT group. These results indicate that PNS can improve the PAH induced by MCT in rats by inhibiting ADAM10/Notch3 signaling pathway.
Animals ; Male ; Rats ; Caspase 3/metabolism* ; Collagen ; Disease Models, Animal ; Hypertension, Pulmonary/drug therapy* ; Monocrotaline/adverse effects* ; Panax notoginseng/chemistry* ; Proliferating Cell Nuclear Antigen/pharmacology* ; Pulmonary Arterial Hypertension ; Pulmonary Artery/metabolism* ; Rats, Sprague-Dawley ; Receptor, Notch3/genetics* ; RNA, Messenger ; Saline Solution ; Signal Transduction ; Saponins/pharmacology*

OBJECTIVEIt has been shown that triptolide can attenuate pulmonary arterial hypertension in rats. This study was designed to investigate the therapeutic effect of triptolide on pulmonary hypertension in rats and possible mechanisms.

METHODSSixty Sprague-Dawley (SD) rats were randomly divided into 6 groups: normal control, model, continuous triptolide-treated, delayed triptolide-treated and two placebo groups for continuous and delayed fashions (n=10 each). The rats from the last 5 groups were injected with monocrotaline (MCT, 60 mg/kg) on day 7 after left pneumonectomy. The rats in the continuous triptolide-treated group received therapy from day 5 to 35 with triptolide (0.25 mg/kg intraperitoneally, every other day) and those in the delayed triptolide-treated received therapy with triptolide (0.20 mg/kg intraperitoneally, daily) from day 21 to 35 after operation. The hemodynamic parameters were detected by catheterization and the pathologic changes of small pulmonary arteries were evaluated by light microscopy 5 weeks post-operation. The expression of matrix metalloproteinases (MMPs) was assessed by immunohistochemistry and quantitative fluorescence PCR of relevant (MMP2 and MMP9) mRNAs.

RESULTSBy day 35 after operation, the mean pulmonary arterial pressure (mPAP, 38.10+/-1.20 vs 16.70+/-1.16 mmHg)the ratio of right ventricle/left ventricle plus septum [RV/(LV+S), 62.45+/-5.28% vs 22.76 +/-3.01%] and the vessel obstructive scores (VOS, 1.736 +/-0.080 vs 0.000 +/-0.000) increased significantly in the Model group compared with those of the normal control group (P < 0.01). The expression of MMP2 and MMP9 and their mRNA expression in lung tissues obviously also elevated in the Model group (P < 0.05). The continuous and the delayed triptolide-treated groups had significantly lower mPAP (20.80+/-1.03 and 26.20+/-1.03 mmHg, respectively) and less right ventricular hypertrophy and pulmonary arterial neointimal formation compared with the model and the placebo groups. The two treated groups also demonstrated decreased expression of MMP2 and MMP9 and their mRNA expression in lung tissues. There were significant differences in mPAP, RV/(LV+S) and VOS between the two triptolide-treated groups.

CONCLUSIONSTriptolide attenuates the development of pulmonary hypertention and right ventricular hypertrophy and promotes regression of pulmonary arterial neointimal formation in pneumonectomized rats that received MCT, possibly through an inhibition of MMPs activity.

Animals ; Diterpenes ; pharmacology ; therapeutic use ; Epoxy Compounds ; pharmacology ; therapeutic use ; Hypertension, Pulmonary ; drug therapy ; enzymology ; Immunohistochemistry ; Lung ; enzymology ; Male ; Matrix Metalloproteinase 2 ; analysis ; genetics ; Matrix Metalloproteinase 9 ; analysis ; genetics ; Phenanthrenes ; pharmacology ; therapeutic use ; RNA, Messenger ; analysis ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo observe the effect of sesamin (Ses) on pulmonary vascular remodeling in rats with monocrotaline ( MCT)-induced pulmonary hypertension (PH).

METHODTotally 48 male Sprague-Dawley (SD) rats were fed adaptively for one week and then divided into the normal control group, the MCT group, the MCT +Ses (50 mg x kg(-1)) group and the MCT + Ses (100 mg x kg(-1)) group, with 12 rats in each group. The PH rat model was induced through the subcutaneous injection with MCT(60 mg x kg(-1)). After the administration for four weeks, efforts were made to measure the right ventricular systolic pressure( RVSP) and mean pulmonary artery pressure (mPAP) through right jugular vein catheterization, and isolate right ventricle( RV) and left ventricle( LV) +septum (S) and measure their length to calculate RV/ ( LV + S) and ratio of RV to tibial length. Pathologic changes in arterioles were observed by HE staining. Masson's trichrome stain was used to demonstrate changes in collagen deposition of arterioles. The alpha-smooth muscle actin (alpha-SMA) expression in pulmonary arteries was measured by immunohistochemisty. The total antioxidative capacity (T-AOC) and malondialdehyde (MDA) content in pulmonary arteries were determined by the colorimetric method. The protein expressions of collagen I, NOX2 and NOX4 were analyzed by Real-time PCR and Western blot.

RESULTAfter the administration for 4 weeks, Ses could attenuate RVSP and mPAP induced by MCT, RV/ (LV + S) and ratio of RV to Tibial length, alpha-SMA and collagen I expressions and remodeling of pulmonary vessels and right ventricle. Meanwhile, Ses could obviously inhibit the expressions of NOX2, NOX4 and MDA content and increase T-AOC.

CONCLUSIONSesamin could ameliorate pulmonary vascular remodeling induced by monocrotaline in PH rats. Its mechanism may be related to expressions of NOX2 and NOX4 expression and reduction in oxidative stress injury.

Animals ; Dioxoles ; administration & dosage ; Disease Models, Animal ; Drugs, Chinese Herbal ; administration & dosage ; Humans ; Hypertension, Pulmonary ; drug therapy ; enzymology ; genetics ; physiopathology ; Lignans ; administration & dosage ; Lung ; blood supply ; enzymology ; metabolism ; Male ; Membrane Glycoproteins ; genetics ; metabolism ; Monocrotaline ; adverse effects ; NADPH Oxidase 2 ; NADPH Oxidase 4 ; NADPH Oxidases ; genetics ; metabolism ; Pulmonary Artery ; drug effects ; metabolism ; physiopathology ; Rats ; Rats, Sprague-Dawley ; Vascular Remodeling ; drug effects

OBJECTIVEDuring the development of hypoxic pulmonary hypertension, the quantity of the protein and mRNA of alpha1-adrenergic receptor (alpha1-adrenergic receptor,alpha1-AR) in the lung tissue increased, while no particular reports were found about the change of the alpha1-AR subtypes during that course. The study aimed to understand the quantity and location of alpha1-AR subtypes mRNA expression in control and hypoxia rats,and the effect of phentolamine in hypoxic pulmonary hypertension.

METHODSFifty-five male Wistar rats were divided randomly into 5 groups: control group (n = 10), hypoxia 2 weeks (n = 13), hypoxia 4 weeks (n = 10), saline control group (n = 10) and hypoxia 4 weeks plus phentolamine group (n = 12). Semi-quantitative reverse transcription and polymerase chain reaction (RT-PCR) were used to examine the mRNA expression of alpha1-AR subtypes in each group. In situ hybridization was also used to detect the location of the alpha1-AR in lungs of normal rats.

RESULTS(1) The pulmonary artery pressure increased with the extension of hypoxia. (2) The results of RT-PCR showed that the mRNA expression of alpha1A-AR was the most,alpha1B-AR the second and alphaZD-AR was the least in all of the groups. (3) The expression of alpha1A-AR and alpha1B-AR mRNA increased with the extension of hypoxia, and the expressions among groups showed difference. (4) The expression of alpha1D-AR also increased with the extension of hypoxia but no difference was found among groups. (5) No difference was found in mRNA quantity of all three subtypes between phentolamine group and hypoxia saline group. (6) In situ hybridization showed that mRNA of the three alpha1-AR subtypes located mainly in the artery and venous smooth muscle cells and endothelial cells.

CONCLUSIONSThis study suggested that alpha1-AR subtypes worked in the development of hypoxia pulmonary hypertension. More research on alpha1-AR subtypes may help the clinical treatment of pulmonary hypertension.

Adrenergic alpha-Antagonists ; pharmacology ; Animals ; Disease Models, Animal ; Hypertension, Pulmonary ; drug therapy ; etiology ; genetics ; Hypoxia ; complications ; In Situ Hybridization ; Lung ; metabolism ; pathology ; Male ; Phentolamine ; pharmacology ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Wistar ; Receptors, Adrenergic, alpha-1 ; classification ; genetics ; Reverse Transcriptase Polymerase Chain Reaction