Fropofol prevents disease progression in mice with hypertrophic cardiomyopathy.

AIMS: Increased myofilament contractility is recognized as a crucial factor in the pathogenesis of hypertrophic cardiomyopathy (HCM). Direct myofilament desensitization might be beneficial in preventing HCM disease progression. Here, we tested whether the small molecule fropofol prevents HCM phenotype expression and disease progression by directly depressing myofilament force development. METHODS AND RESULTS: Force, intracellular Ca2+, and steady-state activation were determined in isolated trabecular muscles from wild-type (WT) and transgenic HCM mice with heterozygous human α-myosin heavy chain R403Q mutation (αMHC 403/+). αMHC 403/+ HCM mice were treated continuously with fropofol by intraperitoneal infusion for 12 weeks. Heart tissue was analysed with histology and real-time PCR of prohypertrophic and profibrotic genes. Fropofol decreased force in a concentration-dependent manner without significantly altering [Ca2+]i in isolated muscles from both WT and αMHC 403/+ HCM mouse hearts. Fropofol also depressed maximal Ca2+-activated force and increased the [Ca2+]i required for 50% activation during steady-state activation. In whole-animal studies, chronic intra-abdominal administration of fropofol prevented hypertrophy development and diastolic dysfunction. Chronic fropofol treatment also led to attenuation of prohypertrophic and profibrotic gene expression, reductions in cell size, and decreases in tissue fibrosis. CONCLUSIONS: Direct inhibition of myofilament contraction by fropofol prevents HCM disease phenotypic expression and progression, suggesting that increased myofilament contractile force is the primary trigger for hypertrophy development and HCM disease progression.

Influence of Age on Clinical Presentation, Therapeutic Options, and Outcome in Chinese Patients with Acute Aortic Dissection.

It has been shown in previous studies that Chinese patients with acute aortic dissection (AD) were approximately 10 years younger than patients from western countries. However, there is a lack of studies concerning the age-related differences in clinical characteristics and outcomes in Chinese patients with acute AD. A total of 1,061 patients with AD (570 type A and 491 type B AD) were enrolled between 2006 and 2008. The clinical characteristics were compared between the patients in our study and those in the International Registry of Acute Aortic Dissection (IRAD). Compared with patients in the IRAD, those in our study were relatively younger, comprised more males, and had a higher proportion of Marfan syndrome but received fewer surgical interventions. When stratified by 10-year age, younger patients were more likely to have type A AD, familial AD, and Marfan syndrome, whereas older patients tended to comprise more females and type B AD. As age increased, the proportion of surgical intervention gradually decreased regardless of the type of AD. During a median follow-up of 2.2 years, 147 patients died, of whom 94 (63.9%) had type A AD and 53 (36.1%) had type B AD. Long-term mortality increased with increasing age, especially in patients above 70 years old. Furthermore, the recurrence rate of AD was higher in both the young and the older patients. In conclusion, compared with western patients with AD, Chinese patients have distinct characteristics and more attention should be paid to the young and older patients because of their high long-term mortality and recurrence rate.

Long non-coding RNA CASC2 suppresses pulmonary artery smooth muscle cell proliferation and phenotypic switch in hypoxia-induced pulmonary hypertension.

BACKGROUND: In this study, we aimed to investigate whether and how lncRNA CASC2 was involved in hypoxia-induced pulmonary hypertension (PH)-related vascular remodeling. METHODS: The expression of lncRNAs or mRNAs was detected by qRT-PCR, and western blot analysis or immunochemistry was employed for detecting the protein expression. Cell number assay and EdU (5-ethynyl-2'-deoxyuridine) staining were performed to assess cell proliferation. Besides, flow cytometry and wound healing assay were employed for assessments of cell apoptosis and cell migration, respectively. Rat model of hypoxic PH was established and the hemodynamic measurements were performed. Hematoxylin and eosin (HE) and Masson's trichrome staining were carried out for pulmonary artery morphometric analysis. RESULTS: The expression of lncRNA CASC2 was decreased in hypoxia-induced rat pulmonary arterial tissues and pulmonary artery smooth muscle cells (PASMCs). Up-regulation of lncRNA CASC2 inhibited cell proliferation, migration yet enhanced apoptosis in vitro and in vivo in hypoxia-induced PH. Western blot analysis and immunochemistry showed that up-regulation of lncRNA CASC2 greatly decreased the expression of phenotype switch-related marker α-SMA in hypoxia-induced PH. Furthermore, it was indicated by the pulmonary artery morphometric analysis that lncRNA CASC2 suppressed vascular remodeling of hypoxia-induced rat pulmonary arterial tissues. CONCLUSION: LncRNA CASC2 inhibited cell proliferation, migration and phenotypic switch of PASMCs to inhibit the vascular remodeling in hypoxia-induced PH.

Fropofol decreases force development in cardiac muscle.

Supranormal contractile properties are frequently associated with cardiac diseases. Anesthetic agents, including propofol, can depress myocardial contraction. We tested the hypothesis that fropofol, a propofol derivative, reduces force development in cardiac muscles via inhibition of cross-bridge cycling and may therefore have therapeutic potential. Force and intracellular Ca2+ concentration ([Ca2+]i) transients of rat trabecular muscles were determined. Myofilament ATPase, actin-activated myosin ATPase, and velocity of actin filaments propelled by myosin were also measured. Fropofol dose dependently decreased force without altering [Ca2+]i in normal and pressure-induced hypertrophied-hypercontractile muscles. Similarly, fropofol depressed maximum Ca2+-activated force ( Fmax) and increased the [Ca2+]i required for 50% of Fmax (Ca50) at steady state without affecting the Hill coefficient in both intact and skinned cardiac fibers. The drug also depressed cardiac myofibrillar and actin-activated myosin ATPase activity. In vitro actin sliding velocity was significantly reduced when fropofol was introduced during rigor binding of cross-bridges. The data suggest that the depressing effects of fropofol on cardiac contractility are likely to be related to direct targeting of actomyosin interactions. From a clinical standpoint, these findings are particularly significant, given that fropofol is a nonanesthetic small molecule that decreases myocardial contractility specifically and thus may be useful in the treatment of hypercontractile cardiac disorders.-Ren, X., Schmidt, W., Huang, Y., Lu, H., Liu, W., Bu, W., Eckenhoff, R., Cammarato, A., Gao, W. D. Fropofol decreases force development in cardiac muscle.

The effect of admission serum potassium levels on in-hospital and long-term mortality in type A acute aortic dissection.

BACKGROUND: Mild fluctuations in serum potassium (K+) levels are related to the prognosis of cardiovascular disease. This study aimed to determine the effect of admission serum potassium levels on in-hospital and long-term mortality in patients with Stanford type A acute aortic dissection (AAD). MATERIALS AND METHODS: A total of 588 consecutive patients with type A AAD were enrolled, and they were grouped according to admission serum potassium level: <3.5, 3.5 to <4.0, 4.0 to <4.5, 4.5 to <5.0, and ≥5.0mmol/L. Clinical outcomes were in-hospital death and long-term all-cause mortality. RESULTS: The in-hospital and long-term all-cause mortality rates were 10.7% and 16.3%, respectively. A U-shaped relationship was observed between admission serum potassium levels and both in-hospital death and long-term mortality. Univariate Cox regression identified potassium levels outside the interval of <3.5 to 4.5mmol/L to be a risk factor for both in-hospital and long-term death. After adjusting for age, gender, surgery and other risk factors, potassium levels outside the interval of <3.5 to 4.5mmol/L still had a significant association with long-term death [hazard ratio (HR)=1.72, 95% confidence interval (95% CI): 1.07-2.74, P=0.024]. Surgical intervention was the main protective factor associated with both in-hospital (HR=0.01, 95% CI 0.01-0.06, P<0.001) and long-term survival (HR=0.06, 95% CI 0.03-0.12, P<0.001). CONCLUSIONS: Among patients with Stanford type A AAD, admission serum potassium levels other than 3.5 to 4.5mmol/L might be associated with an increased risk of in-hospital death and long-term mortality.

Onset seasons and clinical outcomes in patients with Stanford type A acute aortic dissection: an observational retrospective study.

OBJECTIVES: To evaluate the association of onset season with clinical outcome in type A acute aortic dissection (AAD). DESIGN: A single-centre, observational retrospective study. SETTING: The study was conducted in Fuwai Hospital, the National Centre for Cardiovascular Disease, Beijing, China. PARTICIPANTS: From 2008 to 2010, a set of consecutive patients with type A AAD, confirmed by CT scanning, were enrolled and divided into four groups according to onset season: winter (December, January and February), spring (March, April and May), summer (June, July and August) and autumn (September, October and November). The primary end points were in-hospital death and all-cause mortality during follow-up. RESULTS: Of the 492 cases in this study, 129 occurred in winter (26.2%), 147 in spring (29.9%), 92 in summer (18.7%), and 124 in autumn (25.2%). After a median follow-up of 20.4 months (IQR 9.7-38.9), the in-hospital mortality in cases occurring in autumn was higher than in the other three seasons (23.4% vs 8.4%, p<0.01). Long-term mortality was comparable among the four seasonal groups (p=0.63). After adjustment for age, gender and other risk factors, onset in autumn was still an independent factor associated with increased risk of in-hospital mortality (HR 2.05; 95% CI 1.15 to 3.64, p=0.02) in addition to surgical treatment. Further analysis showed that the seasonal effect on in-hospital mortality (autumn vs other seasons: 57.4% vs 27.3%, p<0.01) was only significant in patients who did not receive surgical treatment. No seasonal effect on long-term clinical outcomes was found in this cohort. CONCLUSIONS: Onset in autumn may be a factor that increases the risk of in-hospital death from type A AAD, especially in patients who receive conservative treatment. Immediate surgery improves the short-term and long-term outcomes regardless of onset season.

Admission white blood cell count predicts short-term clinical outcomes in patients with uncomplicated Stanford type B acute aortic dissection.

OBJECTIVES: Inflammation has been shown to be related with acute aortic dissection (AAD). The present study aimed to evaluate the association of white blood cell counts (WBCc) on admission with both in-hospital and long-term all-cause mortality in patients with uncomplicated Stanford type B AAD. METHODS: From 2008 to 2010, a total of 377 consecutive patients with uncomplicated type B AAD were enrolled and then followed up. Clinical data and WBCc on admission were collected. The primary end points were in-hospital death and long-term all-cause death. RESULTS: The in-hospital death rate was 4.2%, and the long-term all-cause mortality rate was 6.9% during a median follow-up of 18.9 months. WBCc on admission was identified as a risk factor for in-hospital death by univariate Cox regression analysis as both a continuous variable and a categorical variable using a cut off of 11.0 × 109 cell/L (all P < 0.05). After adjusting for age, sex and other risk factors, elevated admission WBCc was still a significant predictor for in-hospital death as both a continuous variable [hazard ratio (HR): 1.052, 95% CI: 1.024-1.336, P = 0.002] and a categorical variable using a cut off of 11.0 × 109 cell/L (HR: 2.056, 95% CI: 1.673-5.253, P = 0.034). No relationship was observed between WBCc on admission and long-term all-cause death. CONCLUSIONS: Our results indicate that elevated WBCc upon admission might be used as a predictor for increased risk of in-hospital death in uncomplicated type B AAD. There might be no predictive value of WBCc for the long-term survival of type B AAD.

Impact of Admission White Blood Cell Count on Short- and Long-term Mortality in Patients With Type A Acute Aortic Dissection: An Observational Study.

Studies have shown inflammation is involved in the development of acute aortic dissection (AAD). The hypothesis that white blood cell count (WBCc) on admission may have an impact on the short- and long-term outcomes of type A AAD was tested in a large-scale, prospective observational cohort study.From 2008 to 2010, a total of 570 consecutive patients with type A AAD in Fuwai hospital were enrolled and were followed up. Baseline characteristics and WBCc on admission were collected. The primary outcomes were 30-day and long-term all-cause mortality.During a median of 1.89 years of follow-up, the 30-day and long-term all-cause mortality were 10.7% and 6.5%, respectively. Univariate Cox regression analysis identified admission WBCc as an independent predictor of 30-day mortality when considered as a continuous variable or as a categorical variable using the cutoff of 11.0 × 10 cells/L (all P < 0.05). After adjustment for age, sex, C-reactive protein, D-dimer, and surgical intervention, elevated admission WBCc (>11.0 × 10 cells/L) remained an independent predictor of 30-day mortality of AAD (hazard ratio = 3.31, 95% confidence interval 1.38-7.93, P = 0.007). No impact of admission WBCc was observed on the long-term all-cause mortality.In conclusion, elevated admission WBCc may be valuable as a predictor of 30-day mortality, and may be useful in the risk stratification of type A AAD during hospitalization.

Negative feedback-defective PRPS1 mutants drive thiopurine resistance in relapsed childhood ALL.

Relapse is the leading cause of mortality in children with acute lymphoblastic leukemia (ALL). Among chemotherapeutics, thiopurines are key drugs in ALL combination therapy. Using whole-exome sequencing, we identified relapse-specific mutations in the phosphoribosyl pyrophosphate synthetase 1 gene (PRPS1), which encodes a rate-limiting purine biosynthesis enzyme, in 24/358 (6.7%) relapsed childhood B cell ALL (B-ALL) cases. All individuals who harbored PRPS1 mutations relapsed early during treatment, and mutated ALL clones expanded exponentially before clinical relapse. Our functional analyses of PRPS1 mutants uncovered a new chemotherapy-resistance mechanism involving reduced feedback inhibition of de novo purine biosynthesis and competitive inhibition of thiopurine activation. Notably, the de novo purine synthesis inhibitor lometrexol effectively abrogated PRPS1 mutant-driven drug resistance. These results highlight the importance of constitutive activation of the de novo purine synthesis pathway in thiopurine resistance, and they offer therapeutic strategies for the treatment of relapsed and thiopurine-resistant ALL.

Impact of d-Dimer Levels on Admission on Inhospital and Long-Term Outcome in Patients With Type A Acute Aortic Dissection.

Limited studies with relatively small sample sizes have reported that elevated d-dimer levels on admission were associated with increased risk of short-term mortality in patients with type A acute aortic dissection (AAD). However, there were unavailable data regarding the impact of admission d-dimer levels on long-term outcomes. Our present study aimed to evaluate the association of admission d-dimer levels with both inhospital and long-term all-cause mortality in patients with type A AAD. A total of 212 consecutive patients with type A AAD were enrolled. d-Dimer levels were measured on admission, and patients were followed up prospectively. The primary end points were inhospital and long-term all-cause mortality. The median length of follow-up was 18.8 months (interquartile range 6.7 to 24.4 months). The inhospital and long-term all-cause mortality rates were 12.7% and 12.4%, respectively. Compared with the survivors, the nonsurvivors had significantly higher d-dimer levels (p <0.001). When divided into 4 groups according to admission d-dimer quartiles, patients in Q4 (>6.10 µg/ml) had the highest inhospital and long-term mortality among groups. After multivariate adjustment, the d-dimer level in Q4 (>6.10 µg/ml) was an independent risk factor for inhospital mortality (hazard ratio [HR] 6.12, 95% confidence interval 1.35 to 27.89, p = 0.019) in addition to surgical treatment; however, this was not an independent predictor for long-term mortality. In conclusion, our study with a relatively large sample size suggested that elevated admission d-dimer levels (>6.10 µg/ml) might be a predictor for increased risk of inhospital mortality, and urgent-emergent surgery might be needed in patients with elevated d-dimer levels on admission. However, d-dimer levels at admission failed to predict long-term mortality.

Comparison of two tranexamic acid dose regimens in patients undergoing cardiac valve surgery.

OBJECTIVE: Tranexamic acid (TA), a synthetic antifibrinolytic drug, has been shown to reduce postoperative bleeding and the need for allogeneic blood transfusion in cardiac surgery. However, the optimal dose regimen of TA is still under debate. The aim of this study was to evaluate whether a lower-dose TA regimen produced equivalent efficacy to its higher-dose counterpart in reducing postoperative bleeding and transfusion needs. DESIGN: A prospective, randomized, double-blind trial. SETTING: National Center for Cardiovascular Diseases & University Hospital, Beijing, People's Republic of China. PARTICIPANTS: One hundred seventy-five patients undergoing cardiac valve surgery were enrolled in the study. INTERVENTIONS: All patients were divided randomly into 2 groups. The lower-dose TA group received a loading dose of 10 mg/kg, maintenance dose of 2 mg/kg/h, and a cardiopulmonary bypass pump prime dose of 40 mg; the higher-dose TA group received a loading dose of 30 mg/kg, maintenance dose of 16 mg/kg/h, and a pump prime dose of 2 mg/kg. MEASUREMENTS AND MAIN RESULTS: The amount of postoperative bleeding, the amount and frequency of allogeneic transfusion, mortality, and morbidities were recorded. There was no significant difference in the volume of 24-hour postoperative bleeding between the lower-dose group and the higher-dose group. Other measurements also showed no statistical difference between the 2 groups, including the amount and frequency of allogeneic transfusion, mortality, and morbidities. CONCLUSION: Lower-dose TA regimen was as effective as the higher-dose regimen in reducing postoperative bleeding and transfusion needs in patients undergoing cardiac valve surgery.

Expression of feline angiotensin converting enzyme 2 and its interaction with SARS-CoV S1 protein.

Feline angiotensin converting enzyme 2 (fACE2) gene was amplified from domestic cat lung with RT-PCR, cloned and sequenced. The complete coding region is 2418bp in length and is the closest to human ACE2 among known ACE2 homologs of non-primate animals. The N terminal fragment 19- 367 aa was expressed in Escherishia coli. Both Western blotting and ELISA demonstrated that fACE2 could react with SARS-CoV S1 protein as efficiently as ACE2 of Vero E6 cells did.

Modification of SARS-CoV S1 gene render expression in Pichia pastoris.

S1 gene fragment containing receptor-binding region was amplified by several sets of primers using Over-Lap PCR. The native S1 gene was modified at A + T abundant regions; n.t.777-1683, n.t.1041-1050, n.t.1236-1248, n.t.1317-1335, n.t.1590-1605; based on the same amino acid sequences. The modified gene was cloned into a yeast expression vector pPIC9K. The resultant plasmid pPIC9K- S1 was transformed into Pichia pastoris GS 115 and the protein expression was induced with methanol. SDS-PAGE confirmed that the recombinant SI was secreted in the supernatant of induced GS 115. The protein yield reached 69 mg/l. ELISA and Western blot demonstrated that the S1 could react with the convalescent sera of people infected by SARS-CoV. Furthermore, ligand blot assay showed that the recombinant S1 could react with ACE2, the receptor of SARS-CoV. The molecular mass of expressed S1 was about 70 kDa, which was higher than that of the 30 kDa expected. PNGase F deglycosylation resulted in a protein band of 30 kDa. In conclusion, the S1 gene modification rendered the high-level expression of S1 in P. pastoris GS 115 and the protein was secreted as a biologically active form which was hyperglycosylated.

[Study on immunogenicity of the N-terminal polypeptide of RTX toxin I of Actinobacillus pleuropneumoniae].

ApxI is one of the most important virulence factors of Actinobacillus pleuropneumoniae (APP). To study the immunogenicity of the ApxI, the complete coding sequence (3146bp) and its 5'-terminal 1140 bp fragment of the apxIA gene were separately cloned into the prokaryotic expression vector pET-28a, and expressed in the E. coli BL21 (DE3) with induction by IPTG. The expression products, rApxIA and rApxIAN, were present in a form of inclusion bodies and showed the same immunological reactivity as natural ApxI (nApxI) in Western-blot analysis. BALB/c mice were intraperitoneally immunized with the rApxIA, rApxIAN and nApxI respectively. The serum antibody levels of the rApxIAN immunized mice were significantly lower than those immunized with rApxIA or nApxI in an ApxI-specific ELISA, but serum neutralization test demonstrated that immunized mice with rApxIAN, rApxIA and nApxI could generate similar levels of antibodies neutralizing the hemolytic activity of the natural ApxI. The rApxIAN was able to elicite 80% protection rate against APP serovar 1 and 100% against serovar 2 when challenged at a dose of one LD50 after 2 weeks of boost immunization.