Integrating myocardial metabolic imaging and stress myocardial contrast echocardiography to improve the diagnosis of coronary microvascular diseases in rabbits.

Background: Persistent challenges associated with misdiagnosis and underdiagnosis of coronary microvascular disease (CMVD) necessitate the exploration of noninvasive imaging techniques to enhance diagnostic accuracy. Therefore, we aimed to integrate multimodal imaging approaches to achieve a higher diagnostic rate for CMVD using high-quality myocardial metabolism imaging (MMI) and myocardial contrast echocardiography (MCE). This combination diagnostic strategy may help address the urgent need for improved CMVD diagnosis. Methods: In this study, we established five distinct pretreatment groups, each consisting of nine male rabbit: a fasted group, a nonfasted group, a sugar load group, an acipimox group, and a combination group of nonfasted rabbits administered insulin. Moreover, positron emission tomography-computed tomography (PET/CT) scan windows were established at 30-, 60-, and 90-minute intervals. We developed 10 CMVD models and conducted a diagnosis of CMVD through an integrated analysis of MMI and MCE, including image acquisition and processing. For each heart segment, we calculated the standardized uptake value (SUV) based on body weight (SUVbw), as well as certain ratios of SUV including SUV of the heart (SUVheart) to that of the liver (SUVliver) and SUVheart to SUV of the lung (SUVlung). Additionally, we obtained three coronary SUVbw uptake values. To clarify the relationship between SUVbw uptake values and echocardiographic parameters of the myocardial contrast agent more thoroughly, we conducted a comprehensive analysis across different pretreatment protocols. Receiver operating characteristic (ROC) curve analysis was employed to evaluate the diagnostic accuracy of each parameter in the context of CMVD. Results: In the context of MMI, the nonfasted-plus-insulin group, as observed during the 60-minute examination, exhibited a noteworthy total 18F-fluorodeoxyglucose (18F-FDG) uptake of 47.44±6.53 g/mL, which was found to be statistically different from the other groups. To ascertain the reliability of the results, two double-blind investigators independently assessed the data and achieved a good level of agreement, according to the intraclass correlation coefficient (ICC) (0.957). The SUVbw of the nonfasted-plus-insulin group exhibited a moderate correlation with the microvascular blood flow reserve (MBFR) parameters derived from the MCE examination, as evidenced by a r value of 0.686. For the diagnosis of CMVD disease, the diagnostic accuracy of the combined diagnostic method [area under the curve (AUC) =0.789; 95% confidence interval (CI): 0.705-0.873] was significantly higher than that of the MBFR (AUC =0.697; 95% CI: 0.597-0.797) and SUVbw (AUC =0.715; 95% CI: 0.622-0.807) methods (P<0.05). Conclusions: Our study demonstrated the feasibility of a simple premedication approach involving free feeding and intravenous insulin in producing high-quality gated heart 18F-FDG PET/CT images in adult male New Zealand white rabbits. This technique holds considerable potential for ischemic heart disease research in rabbits and can enhance CMVD diagnosis via the comprehensive assessment of myocardial metabolism and perfusion.

Reference values of carotid intima-media thickness and arterial stiffness in Chinese adults based on ultrasound radio frequency signal: A nationwide, multicenter study.

BACKGROUND: Carotid intima-media thickness (IMT) and diameter, stiffness, and wave reflections, are independent and important clinical biomarkers and risk predictors for cardiovascular diseases. The purpose of the present study was to establish nationwide reference values of carotid properties for healthy Chinese adults and to explore potential clinical determinants. METHODS: A total of 3053 healthy Han Chinese adults (1922 women) aged 18-79 years were enrolled at 28 collaborating tertiary centers throughout China between April 2021 and July 2022. The real-time tracking of common carotid artery walls was achieved by the radio frequency (RF) ultrasound system. The IMT, diameter, compliance coefficient, ß stiffness, local pulse wave velocity (PWV), local systolic blood pressure, augmented pressure (AP), and augmentation index (AIx) were then automatically measured and reported. Data were stratified by age groups and sex. The relationships between age and carotid property parameters were analyzed by Jonckheere-Terpstra test and simple linear regressions. The major clinical determinants of carotid properties were identified by Pearson's correlation, multiple linear regression, and analyses of covariance. RESULTS: All the parameters of carotid properties demonstrated significantly age-related trajectories. Women showed thinner IMT, smaller carotid diameter, larger AP, and AIx than men. The ß stiffness and PWV were significantly higher in men than women before forties, but the differences reversed after that. The increase rate of carotid IMT (5.5 µm/year in women and 5.8 µm/year in men) and diameter (0.03 mm/year in both men and women) were similar between men and women. For the stiffness and wave reflections, women showed significantly larger age-related variations than men as demonstrated by steeper regression slopes (all P for age by sex interaction <0.05). The blood pressures, body mass index (BMI), and triglyceride levels were identified as major clinical determinants of carotid properties with adjustment of age and sex. CONCLUSIONS: The age- and sex-specific reference values of carotid properties measured by RF ultrasound for healthy Chinese adults were established. The blood pressures, BMI, and triglyceride levels should be considered for clinical application of corresponding reference values.

Establishment of nonobstructive coronary microcirculatory disorders in rabbits using three established methods and a comparative study.

To compare the merits and drawbacks of three approaches for establishing a rabbit model of nonobstructive coronary microcirculatory disease, namely, open thoracic subtotal ligation of coronary arteries, ultrasound-guided cardiac microsphere injection, and sodium laurate injection. New Zealand rabbits were allocated to four groups: a normal group (Blank group), an Open-chest group (Open-chest), a microsphere group (Echo-M), and a sodium laurate group (Echo-SL), each comprising 10 rabbits. The rabbits were sacrificed 24 h after the procedures, and their echocardiography, stress myocardial contrast echocardiography, pathology, and surgical times were compared. The results demonstrated varying degrees of reduced cardiac function in all three experimental groups, the Open-chest group exhibiting the most significant decline. The myocardial filling in the affected areas was visually analyzed by myocardial contrast echocardiography, revealing sparse filling at rest but more after stress. Quantitative analysis of perfusion parameters (ß, A, MBF) in the affected myocardium showed reduced values, the Open-chest group having the most severe reductions. No differences were observed in stress myocardial acoustic imaging parameters between the Echo-M and Echo-SL groups. Among the pathological presentations, the Open-chest model predominantly exhibited localized ischemia, while the Echo-M model was characterized by mechanical physical embolism, and the Echo-SL model displayed in situ thrombosis as the primary pathological feature. Inflammatory responses and collagen deposition were observed in all groups, with the severity ranking of Open-chest > Echo-SL > Echo-M. The ultrasound-guided intracardiac injection method used in this experiment outperformed open-chest surgery in terms of procedural efficiency, invasiveness, and maneuverability. This study not only optimizes established cardiac injection techniques but also offers valuable evidence to support clinical investigations through a comparison of various modeling methods.

Functional evaluation of constructed pseudo-endogenous microRNA-targeted myocardial ultrasound nanobubble.

Background: Stem cell transplantation is one of the treatment methods for acute myocardial infarction (AMI). MicroRNA-1 contributes to the study of the essential mechanisms of stem cell transplantation for treating AMI by targeted regulating the myocardial microenvironment after stem cell transplantation at the post-transcriptional level. Thus, microRNA-1 participates in regulating the myocardial microenvironment after stem cell transplantation, a promising strategy for the Stem cell transplantation treatment of AMI. However, the naked microRNA-1 synthesized is extremely unstable and non-targeting, which can be rapidly degraded by circulating RNase. Herein, to safely and effectively targeted transport the naked microRNA-1 synthesized into myocardial tissue, we will construct pseudo-endogenous microRNA-targeted myocardial ultrasound nanobubble pAd-AAV-9/miRNA-1 NB and evaluate its characteristics, targeting, and function. Methods: The pAd-AAV-9/miRNA-1 gene complex was linked to nanobubble NBs by the "avidin-biotin bridging" method to prepare cardiomyocyte-targeted nanobubble pAd-AAV-9/miRNA-1 NB. The shape, particle size, dispersion, and stability of nanobubbles and the connection of pAd-AAV-9/miRNA-1 gene complex to nanobubble NB were observed. The virus loading efficiency was determined, and the myocardium-targeting imaging ability was evaluated using contrast-enhanced ultrasound imaging in vivo. The miRNA-1 expression level in myocardial tissue and other vital organs ex vivo of SD rats was considered by Q-PCR. Also, the cytotoxic effects were assessed. Results: The particle size of NBs was 504.02 ± 36.94 nm, and that of pAd-AAV-9/miRNA-1 NB was 568.00 ± 37.39 nm. The particle size and concentration of pAd-AAV-9/miRNA-1 NBs did not change significantly within 1 h at room temperature (p > 0.05). pAd-AAV-9/miRNA-1 NB had the highest viral load rate of 86.3 ± 2.2% (p < 0.05), and the optimum viral load was 5 µL (p < 0.05). pAd-AAV-9/miRNA-1 NB had good contrast-enhanced ultrasound imaging in vivo. Quantitative analysis of miRNA-1 expression levels in vital organs ex vivo of SD rats by Q-PCR showed that pAd-AAV-9/miRNA-1 NB targeted the myocardial tissue. Q-PCR indicated that the expression level of miRNA-1 in the myocardium of the pAd-AAV-9/miRNA-1 NB + UTMD group was significantly higher than that of the pAd-AAV-9/miRNA-1 NB group (p < 0.05). pAd-AAV-9/miRNA-1 NB had no cytotoxic effect on cardiomyocytes (p > 0.05). Conclusion: The pAd-AAV-9/miRNA-1 NB constructed in this study could carry naked miRNA-1 synthesized in vitro for targeted transport into myocardial tissue successfully and had sound contrast-enhanced imaging effects in vivo.

Optimization of the cotransfection of SERCA2a and Cx43 genes for myocardial infarction complications.

As our previous study showed, the therapeutic effect of two genes (SERCA2a and Cx43) on heart failure after myocardial infarction (MI) was greater than that of single gene (SERCA2a or Cx43) therapy for bone marrow stem cell (BMSC) transplantation. Based on previous research, the aim of this study was to investigate the optimal ratio of codelivery of SERCA2a and Cx43 genes for MI therapy after biotinylated microbubble (BMB) transplantation via ultrasonic-targeted microbubble destruction (UTMD). Forty rats underwent left anterior descending (LAD) ligation and BMSC injection into the infarct and border zones. Four weeks later, the genes SERCA2a and Cx43 were codelivered at different ratios (1:1, 1:2 and 2:1) into the infarcted heart via UTMD. Cardiac mechanoelectrical function was determined at 4 wks after gene delivery, and the hearts of the rats were harvested for measurement of MI size and detection of SERCA2a and Cx43 expression. Q-PCR analysis of the expression of Nkx2.5 and GATA4 in the myocardial infarct zone and measurement of neovascularization in infarcted hearts. After comparing the therapeutic effects of different cogene ratios, the SERCA2a/Cx43-1:2 group showed remarkable cardiac electrical stability and strengthened the role of anti-arrhythmia. In conclusion, the optimum ratio of the SERCA2a/Cx43 gene is 1:2, which is advantageous for maintaining cardiac electrophysiological stability.

Screening of the best time window for MSC transplantation to treat acute myocardial infarction with SDF-1α antibody-loaded targeted ultrasonic microbubbles: An in vivo study in miniswine.

The present study aimed to screen the best time window for the transplantation of bone marrow mesenchymal stem cells (MSCs) after acute myocardial infarction (MI) through targeted ultrasound microbubbles loaded with SDF-1α antibody. Thirty-six MI miniswine were randomly divided into six experimental groups according to the duration after infarction (1 day, 3 days, 1 week, 2 weeks, 3 weeks, and 4 weeks after infarction). MSCs were labeled with BrdU and then injected through the coronary artery in the stem cell transplantation group to detect the number of transplanted MSCs at different time points after MI. Three miniswine were randomly selected as the control group (sham operation: open chest without ligation of the coronary artery). All SDF-1α groups and control groups were injected with a targeted microbubble ultrasound contrast agent. The values of the myocardial perfusion parameters (A, ß, and A × ß) were determined. A T, ß T, and (A × ß)T varied with time and peaked 1 week after MI (P < 0.05). The number of transplanted stem cells in the myocardium through coronary injection of MSCs at 1 week was the greatest and consistent with the changing tendency of A T, ß T, and (A × ß)T (r = 0.658, 0.778, 0.777, P < 0.05). ß T(X), (A × ß)T(X), and the number of transplanted stem cells was used to establish the regression equation as follows: Y = 36.11 + 17.601X; Y = 50.023 + 3.348X (R 2 = 0.605, 0.604, P < 0.05). The best time window for transplanting stem cells was 1 week after MI. The myocardial perfusion parameters of the SDF-1α targeted contrast agent can be used to predict the number of transplanted stem cells in the myocardial tissue.

Contrast-Enhanced Ultrasound Combined With 2D Strain Imaging and Histopathological Multimodal Assessment of Carotid Plaque Vulnerability.

OBJECTIVE: The aim of this study was to explore the value of contrast-enhanced ultrasound (CEUS) combined with 2-D strain imaging in evaluating carotid plaque vulnerability and the correlations among CEUS perfusion parameters, strain parameters and histopathological findings in different plaque segments. METHODS: Patients with carotid artery stenosis who underwent carotid endarterectomy (CEA) at the First Affiliated Hospital of Xinjiang Medical University from September 2020 to June 2021 underwent preoperative carotid artery 2-D ultrasonography and CEUS. The plaques were divided into three segments: the proximal end of the shoulder, central cap and distal end of the shoulder. The peak intensity (PI) value and strain rate parameters of the regions of interest were analyzed. Plaques were divided into a stable group (8 cases) and an unstable group (19 cases). The microvascular density (MVD) and vascular endothelial growth factor (VEGF) expression of each patch in the unstable group were analyzed. RESULTS: The peak strain during the systolic period in each plaque segment in both groups showed the following pattern: proximal end shoulder > distal end shoulder > top (p < 0.05). The PI value for CEUS is also represented. In the unstable group, the PI values of each segment of the plaque were positively correlated with the MVD, near-center PI value and VEGF average optical density value. The average optical density of each segment was positively correlated with the MVD (p < 0.05). There were positive correlations between the PI values of the proximal and distal shoulder and the strain values (p < 0.05), and the MVD value of each segment, VEGF value and strain value were positively correlated (p < 0.05). CONCLUSION: PI and the pathological tissue components represented by CEUS were positively correlated with the mechanical parameters of the plaque along the long axis. There may be overlap between the high shear stress area of the plaque and the neovascular aggregation area, and the combination of the two has certain significance for assessing the vulnerability of the plaque.

Dual-modal molecular imaging and therapeutic evaluation of coronary microvascular dysfunction using indocyanine green-doped targeted microbubbles.

Coronary microvascular dysfunction (CMD), which causes a series of cardiovascular diseases, seriously endangers human health. However, precision diagnosis of CMD is still challenging due to the lack of sensitive probes and complementary imaging technologies. Herein, we demonstrate indocyanine green-doped targeted microbubbles (named T-MBs-ICG) as dual-modal probes for highly sensitive near-infrared (NIR) fluorescence imaging and high-resolution ultrasound imaging of CMD in mouse models. In vitro results show that T-MBs-ICG can specifically target fibrin, a specific CMD biomarker, via the cysteine-arginine-glutamate-lysine-alanine (CREKA) peptide modified on the surface of microbubbles. We further employ T-MBs-ICG to achieve NIR fluorescence imaging of injured myocardial tissue in a CMD mouse model, leading to a signal-to-background ratio (SBR) of up to 50, which is 20 fold higher than that of the non-targeted group. Furthermore, ultrasound molecular imaging of T-MBs-ICG is obtained within 60 s after intravenous injection, providing molecular information on ventricular and myocardial structures and fibrin with a resolution of 1.033 mm × 0.466 mm. More importantly, we utilize comprehensive dual-modal imaging of T-MBs-ICG to evaluate the therapeutic efficacy of rosuvastatin, a cardiovascular drug for the clinical treatment of CMD. Overall, the developed T-MBs-ICG probes with good biocompatibility exhibit great potential in the clinical diagnosis of CMD.

Pre-transplantation of Bone Marrow Mesenchymal Stem Cells Amplifies the Therapeutic Effect of Ultrasound-Targeted Microbubble Destruction-Mediated Localized Combined Gene Therapy in Post-Myocardial Infarction Heart Failure Rats.

Although stem cell transplantation and single-gene therapy have been intensively discussed separately as treatments for myocardial infarction (MI) hearts and have exhibited ideal therapeutic efficiency in animal models, clinical trials turned out to be disappointing. Here, we deliver sarcoplasmic reticulum Ca2+-ATPase 2a (SERCA2a) and connexin 43 (Cx43) genes simultaneously via an ultrasound-targeted microbubble destruction (UTMD) approach to chronic MI hearts that have been pre-treated with bone marrow mesenchymal stem cells (BMSCs) to amplify cardiac repair. First, biotinylated microbubbles (BMBs) were fabricated, and biotinylated recombinant adenoviruses carrying the SERCA2a or Cx43 gene were conjugated to the surface of self-assembled BMBs to form SERCA2a-BMBs, Cx43-BMBs or dual gene-loaded BMBs. Then, the general characteristics of these bubbles, including particle size, concentration, contrast signal and gene loading capacity, were examined. Second, a rat myocardial infarction model was created by ligating the left anterior descending coronary artery and injecting BMSCs into the infarct and border zones. Four weeks later, co-delivery of SERCA2a and Cx43 genes to the infarcted heart were delivered together to the infarcted heart using the UTMD approach. Cardiac mechano-electrical function was determined 4 wk after gene transfection, and the infarcted hearts were collected for myocardial infarct size measurement and detection of expression of SERCA2a, Cx43 and cardiac-specific markers. Finally, to validate the role of BMSC transplantation, MI rats transplanted or not with BMSCs were transfected with SERCA2a and Cx43, and the cardiac mechano-electrical function of these two groups of rats was recorded and compared. General characteristics of the self-assembled gene-loaded BMBs were qualified, and the gene loading rate was satisfactory. The self-assembled gene-loaded BMBs were in microscale and exhibit satisfactory dual-gene loading capacity. High transfection efficiency was achieved under ultrasound irradiation in vitro. In addition, rats in which SERCA2a and Cx43 were overexpressed simultaneously had the best contractile function and electrical stability among all experimental groups. Immunofluorescence assay revealed that the levels of SERCA2a and/or Cx43 proteins were significantly elevated, especially in the border zone. Moreover, compared with rats that did not receive BMSCs, rats pre-treated with BMSCs have better mechano-electrical function after transfection with SERCA2a and Cx43. Collectively, we report a promising cardiac repair strategy for post-MI hearts that exploits the providential advantages of stem cell therapy and UTMD-mediated localized co-delivery of specific genes.

Heat shock protein 70 attenuates hypoxia­induced apoptosis of pulmonary microvascular endothelial cells isolated from neonatal rats.

Pulmonary microvascular endothelial cell (PMVEC) apoptosis is the initial stage of adult pulmonary hypertension (PH), which involves high pulmonary arterial pressure and pulmonary vascular remodeling. However, the mechanism regulating PMVEC apoptosis and its involvement in the early stages of neonatal hypoxic PH (HPH) pathogenesis are currently unclear. The present study aimed to investigate the effects of heat shock protein 70 (HSP70) on hypoxia­induced apoptosis in PMVECs. PMVECs isolated from neonatal Sprague­Dawley rats were transfected with lentivirus with or without HSP70, or treated with the synthetic HSP70 inhibitor N­formyl­3,4­methylenedioxy­benzylidene-g-butyrolactam under hypoxic conditions (5% O2) for 24, 48 or 72 h. PMVEC apoptosis was evaluated by performing flow cytometry and mitochondrial membrane potential (MMP) assays. The expression levels of HSP70, hypoxia­inducible factor­1α (HIF­1α) and apoptosis­associated proteins were determined by conducting reverse transcription­quantitative PCR and western blotting. Following 24, 48 or 72 h of hypoxia, the apoptotic rates of PMVECs were significantly elevated compared with cells under normoxic conditions. The MMP was significantly reduced, whereas the mRNA and protein expression levels of HIF­1α, cytochrome c (cyt C), caspase­3 and HSP70 were enhanced by hypoxia compared with those under normoxic conditions. Additionally, the mRNA and protein expression levels of B­cell lymphoma 2 (Bcl­2) were significantly downregulated in the hypoxia group compared with those in the normoxia group. In hypoxic PMVECs, HSP70 overexpression decreased the apoptotic rate and the expression levels of cyt C, downregulated the expression levels of caspase­3 and HIF­1α, and increased the MMP and the expression levels of Bcl­2. HSP70 inhibition resulted in the opposite outcomes compared with those of HSP70 overexpression. Therefore, the results of the present study suggested that HSP70 may inhibit mitochondrial pathway­mediated apoptosis in isolated neonatal rat PMVECs in early­stage hypoxia, which may be associated with HSP70­mediated HIF­1α downregulation. Overall, HSP70 may be protective against neonatal HPH through the HSP70/HIF­1α pathway.

General Characteristics of Microbubble-Adenovirus Vectors Carrying Genes.

INTRODUCTION: Transferring genes safely, targeting cells and achieving efficient transfection are urgent problems in gene therapy that need to be solved. Combining microbubbles (MBs) and viruses to construct double vectors has become a promising approach for gene delivery. Understanding the characteristic performance of MBs that carry genes is key to promoting effective gene transfer. Therefore, in this study, we constructed MB-adenovirus vectors and discussed their general characteristics. METHODS: We constructed MB-adenovirus vectors carrying the chemokine (C-X-C motif) ligand 12 (Cxcl12) and bone morphogenetic protein-2 (Bmp2) genes (pAd-Cxcl12 and pAd-Bmp2, respectively) to explore the general characteristics of double vectors carrying genes. RESULTS: The MB-adenovirus vectors had stable physical properties, and no significant differences in diameter, concentration, or pH were noted compared with naked MBs (p > 0.05). Flow cytometry and RT-PCR were used to detect the gene-loading capacity of MBs. The gene-loading efficiency of MBs increased with increasing virus amounts and was highest (91%) when 10.0 µL of virus was added. Beyond 10.0 µL of added virus, the gene-loading efficiency of MBs decreased with the continuous addition of virus. The maximum amounts of pAd-Cxcl12 and pAd-Bmp2 in 100 µL of MBs were approximately 14 and 10 µL, respectively. CONCLUSIONS: This study indicates that addition of an inappropriate viral load will result in low MB loading efficiency, and the maximum amount of genes loaded by MBs may differ based on the genes carried by the virus.

Can Combined Screening of Ultrasound and Elastography Improve Breast Cancer Identification Compared with MRI in Women with Dense Breasts-a Multicenter Prospective Study.

Objectives: To assess the performance of elastography (ES) and ultrasound (US) in predicting the malignancy of breast lesions and to compare their combined diagnostic value with that of magnetic resonance imaging (MRI). Materials and Methods: The study prospectively enrolled 242 female patients with dense breasts treated in 35 heath care facilities in China between November 2018 and October 2019. Based on conventional US and elastography, radiologists classified the degree of suspicion of breast lesions according to the US Breast Imaging Reporting and Data System (BI-RADS) criteria. The diagnostic value was compared between US BI-RADS and MRI BI-RADS, with pathological results used as the reference standard. Results: The results demonstrated that irregular tumor shape, a nonparallel growth orientation, indistinct margins, angular contours, microcalcifications, color Doppler flow and ES score on US imaging were significantly related to breast cancer in dense breasts (P=0.001; P=0.001; P=0.008; P<0.001; P=0.019; P=0.008; P=0.002, respectively). The sensitivity, specificity, PPV, NPV, accuracy and AUC of US BI-RADS category were 94.7%, 90.7%, 95.8%, 88.0%, 93.4% and 0.93 (95%CI, 0.88-0.97), respectively, while those of MRI BI-RADS category were 98.2%, 57.5%, 84.3%, 83.3%, 86.0% and 0.78 (95%CI, 0.71-0.85), respectively. MRI BI-RADS showed a significantly higher sensitivity than US BI-RADS (98.2% vs 94.7%, P=0.043), whereas US BI-RADS showed significantly higher specificity (90.7% vs 57.5%, P<0.001). US BI-RADS showed better diagnostic efficiency in differentiating nodules in dense breasts than MRI BI-RADS (AUC 0.93 vs 0.78, P<0.001). Conclusion: By combining the use of ES and conventional US, US BI-RADS had better diagnostic efficiency in differentiating nodules in dense breasts than MRI. For the diagnosis of malignant tumors in patients with dense breasts, MRI and US BI-RADS can be used as supplemental diagnostic tools to detect lesions, with US BI-RADS considered the preferred adjunctive resource.

In Vivo Ultrasound Molecular Imaging of SDF-1 Expression in a Swine Model of Acute Myocardial Infarction.

Background: Stem cell therapy of acute myocardial infarction (AMI) is proving to be a promising approach to repair the injured myocardia. The time window for stem cell transplantation is crucial yet difficult to determine since it produces different therapeutic effects at different times after myocardial infarction. Stromal cell-derived factor-1 (SDF- 1) plays a pivotal role in the mobilization, homing, proliferation, and differentiation of transplanted stem cells. Here, by using ultrasound molecular imaging via targeted microbubbles, we determined the dynamic expression of SDF-1 in a swine model of AMI in vivo. Methods: Twenty-four miniswine were randomly selected for the control group and the AMI model group, which underwent ligation of the left anterior descending coronary artery (LAD). The AMI animals were randomly divided into six experimental groups according to the duration of the myocardial infarction. All animals were subjected to ultrasound molecular imaging through injections with targeted microbubbles (T + T group) or nontargeted control microbubbles (T + C group). The values of the myocardial perfusion parameters (A, ß, and A × ß) were determined using Q-Lab (Philips ultrasound, version 9.0), and the expression level of SDF-1 was analyzed by real-time polymerase chain reaction (RT-PCR). Results: Our results showed that the expression of SDF-1 gradually increased and peaked at 1 week after AMI. The trend is well reflected by ultrasound molecular imaging in the myocardial perfusion parameters. The A, ß, and A × ß values correlated with SDF-1 in the T + T group (r = 0.887, 0.892, and 0.942; P < 0.05). Regression equations were established for the relationships of the A, ß, and A × ß values (X) with SDF-1 (Y): Y = 0.699X - 0.6048, Y = 0.4698X + 0.3282, and Y = 0.0945X + 0.6685, respectively (R 2 = 0.772, 0.7957, and 0.8871; P < 0.05). Conclusions: Our finding demonstrated that ultrasound molecular imaging could be used to evaluate the expression dynamics of SDF-1 after AMI.

Biosynthetic nanobubbles for targeted gene delivery by focused ultrasound.

Ultrasound-targeted microbubble destruction (UTMD) has recently drawn considerable attention in biomedicine applications due to its great potential to locally enhance gene delivery. However, conventional microbubbles have a microscale particle size and polydisperse particle size distribution, which makes it difficult for them to directly come into contact with tumor cells and to efficiently deliver therapeutic genes via ultrasound cavitation effects. In the current study, we developed a kind of novel cationic biosynthetic nanobubble (CBNB) as an ultrasonic gene delivery carrier through coating PEI on the surface of these biosynthetic nanobubbles (BNBs). The BNBs, produced from an extremely halophilic archaeon (Halobacterium NRC-1), possess a nanoscale size and can produce stable contrast signals both in vitro and in vivo. Surface modification with PEI polymer greatly increased the DNA loading capability of BNBs, leading to significantly improved gene transfection efficiency when combining with ultrasound. To our knowledge, this is the first report to apply biosynthetic bubbles as non-viral gene carriers which can effectively deliver genes into tumor cells with the aid of ultrasound cavitation. Our study provides a powerful tool for image-guided and efficient gene delivery using biosynthetic nanoscale contrast agents.

Apical hypertrophic cardiomyopathy with apical endomyocardial fibrosis and calcification: Two case reports.

RATIONALE: Apical hypertrophic cardiomyopathy (AHCM) is a rare form of hypertrophic cardiomyopathy which affects predominantly the apex of the left ventricle. Generally, left ventricular enlargement is not present in AHCM; additionally, endomyocardial fibrosis, and calcification are also rare. PATIENT CONCERNS: A 61-year-old female (Case 1) and a 60-year-old female (Case 2) both presented with the symptoms of atypical chest pain, dyspnoea, exercise intolerance, palpitations. DIAGNOSIS: Magnetic resonance and single-photon emission computed tomography (SPECT) revealed apical hypertrophic cardiomyopathy. Furthermore, 2D-transthoracic echocardiogram showed left atrium and ventricular enlargement, as well as endomyocardial fibrosis and calcification. Based on these findings, the patients were diagnosed with AHCM. INTERVENTIONS: Both the patients were treated with ACEI, metoprolol, and aspirin. Additionally, both these patient underwent genetic test. OUTCOMES: The results of the genetic test of the 2 cases for hypertrophic cardiomyopathy (HCM) were negative. However, the gene mutation for dilated cardiomyopathy (TMPO) was detected in one of the cases. No change in condition during follow-up. LESSONS: In past reports, Apical hypertrophic cardiomyopathy has been shown to have a benign prognosis. But in this case report, the imaging studies of the 2 patients suggest a poor prognosis. Furthermore, diagnosing cardiomyopathy should require multimodality imaging examinations to rule out differential diagnoses.

Ultrasound-Targeted Microbubble Destruction-Mediated Co-Delivery of Cxcl12 (Sdf-1alpha) and Bmp2 Genes for Myocardial Repair.

Acute myocardial infarction (AMI) is a serious threat to human health. Stem cells can serve as ideal seed cells for myocardial repair and regeneration; however, insufficient homing to the infarcted areas and poor myocardial differentiation limit their further clinical application. Chemokine (C-X-C motif) ligand 12 (CXCL12) is an important stem cell homing factor, and Bone morphogenetic protein-2 (BMP2) is a differentiation-promoting factor. In this study, we designed microbubbleadenovirus complexes and investigated the functional benefit of co-delivery of Cxcl12 and Bmp2 genes in adenoviral vectors by ultrasound-targeted microbubble destruction (UTMD) for myocardial repair in AMI rats. By transfection of rat bone marrow mesenchymal stem cells (BMSCs), high transfection efficiency was achieved under ultrasound irradiation, and successful myocardial differentiation of the transfected BMSCs was induced after ultrasound-mediated-transfection of Bmp2 in vitro. In the AMI rat model, co-delivery of Cxcl12 and Bmp2 at a ratio of 2:1 may result in significantly higher myocardial repair efficacy compared to transfection of Cxcl12 or Bmp2 only. Higher myocardial repairing efficacy was achieved with co-delivery of Cxcl12 and Bmp2 at a ratio of 2:1 than using a ratio of 1:1 or 1:2, as evidenced by smaller infarction size, higher micro-vessel density, and better recovery of cardiac function at 28 days after treatment. Hence, UTMD-mediated co-transfection of Cxcl12 and Bmp2 significantly promoted the repair/regeneration of the infarcted myocardium, thereby providing a promising approach for the repair of cardiac injury.

Distribution Pattern of Atherosclerotic Stenosis in Chinese Patients with Stroke: A Multicenter Registry Study.

The aim of this multicenter study was to demonstrate the distribution pattern of atherosclerotic stenosis and its trend with aging between extracranial and intracranial arteries and its distribution between the anterior and posterior circulations in Chinese patients hospitalized with ischemic stroke. In addition, the risk factors for the distribution pattern were illustrated. From June 2015 to May 2016, 9,346 patients with ischemic stroke from 20 hospitals were enrolled. Carotid artery ultrasonography and transcranial color-coded sonography/transcranial Doppler were used to evaluate the extracranial and intracranial arteries. The distribution pattern of atherosclerotic stenosis and its trend with aging were analyzed. Logistic regression was used to analyze the risk factors for the distribution pattern. Among the 9,346 patients, 2,882 patients (30.8%) had at least one artery with a degree of stenosis ≥50%. Among patients with arterial stenosis, the proportion of patients with intracranial artery stenosis was higher than those with extracranial artery stenosis (52.6% vs. 27.6%), and the proportion of anterior circulation artery stenosis was higher than that in the posterior circulation (52.2% vs.26.2%). With aging, the proportion of intracranial artery stenosis alone decreased; at the same time, the proportion of extracranial artery stenosis and extracranial plus intracranial artery stenosis increased (trend χ2=6.698, P=0.001). Hypertension (OR 1.416, P=0.008) and family history of stroke (OR 1.479, P=0.014) were risk factors for intracranial artery stenosis. Male, aging, and smoking were factors more related to extracranial artery stenosis. Aging (OR 1.022, P<0.001) and hypertension (OR 1.392, P=0.019) were related to posterior circulation artery stenosis. Intracranial arteries and anterior circulation arteries were susceptible to stenosis in Chinese patients with ischemic stroke. However, the distribution pattern of atherosclerotic stenosis was dynamic and varied with aging. Aging and different risk factors contribute to this distribution pattern.

Antithrombotic effects and related mechanisms of Salvia deserta Schang root EtOAc extracts.

Salvia deserta Schang (SDS) belongs to the same family as Salvia miltiorrhiza bunge, one of the antithrombotic Chinese herbal medicines. In our study, EtOAc root extracts were analyzed for their effects on adenosine diphosphate (ADP)-induced platelet aggregation in rabbits and FeCl3-induced rat common carotid artery thrombosis as well as on rat blood plasma concentrations of thromboxane B2 (TXB2), 6-keto-prostaglandin F1 alpha (6-keto-PGF1α), antithrombin-III (AT-III), protein C (PC), plasminogen (PLG), plasminogen activator inhibitor (PAI-1), von Willebrand factor (vWF) and tissue-type plasminogen activator (t-PA). EtOAc extracts from SDS roots had significant inhibitory effects on ADP-induced maximum platelet aggregation rate (10.2 ± 2.6 vs control 35.7 ± 5.2; P < 0.05), reduced the FeCl3-induced rat common carotid artery thrombus weight and thrombus area ratio (P < 0.05), significantly decreased plasma TXB2, vWF and PAI-1 levels and increased 6-keto-PGF1α and t-PA levels in a dose dependent manner (all P < 0.05). Thus, the ratio of TXB2/6-keto-PGF1α was significantly decreased (P < 0.05), while the ratio of t-PA/PAI-1 was significantly increased (P < 0.05). In addition, enhanced AT-III and PC activities indicated coagulation inactivation effects of EtOAc SDS root extracts. EtOAc extraction from SDS showed antithrombotic effects, which are likely due to platelet adhesion and aggregation inhibition as well as anticoagulant activities.

Homing of endogenous bone marrow mesenchymal stem cells to rat infarcted myocardium via ultrasound-mediated recombinant SDF-1α adenovirus in microbubbles.

Stem cells can promote myocardial regeneration and accelerate the formation of new blood vessels. As such, transplanted stem cells represent a promising treatment modality for acute myocardial infarction (AMI). Stem cells spontaneously home to the infarcted myocardium using chemotaxis, in which the stromal cell-derived factor (SDF-1α) has been shown to be one of the most important chemokines. However, spontaneously secreted SDF-1α is short-lived, and therefore does not meet the needs of tissue repair. In this study, adenoviruses carrying SDF-1α genes were loaded on microbubble carriers and the adenoviruses were released into AMI rats by ultrasound targeted microbubble destruction. The possibility of in vivo self-transplantation of bone marrow mesenchymal stem cells (BMSCs) induced by overexpression of SDF-1α in the infarcted myocardium was explored by detecting the number of BMSCs homing from the peripheral blood to the myocardial infarcts. The concentration of SDF-1α in peripheral blood was significantly higher after transfection, and the number of BMSCs was significantly higher in the peripheral blood and infarcted area. Further analyses indicated that the number of homing BMSCs increased with increased SDF-1α expression. In conclusion, our results suggest that ultrasound mediated transduction of exogenous SDF-1α genes into myocardial infarcted AMI rats can effectively promote the homing of endogenous BMSCs into the heart. Moreover, the number of homing stem cells was controlled by the level of SDF-1α expression.