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Introduction: Hyperbaric oxygen (HBO) has been used for the treatment of cerebral palsy for more than 20 years, but its efficacy and safety are still controversial. In this systematic review and meta-analysis, we evaluated the currently promulgated data related to the efficacy of HBO for patients with cerebral palsy. Methods: We searched the PubMed/Medline, Embase, Web of Science, Cochrane Library, China National Knowledge Infrastructure, and Wanfang databases (from their inception to April 2020) for randomized controlled trials published in English or Chinese. Two researchers used the Cochrane Collaboration tool for data extraction and an independent quality assessment. The extracted data were analyzed by Review Manager 5.3 software. Results: A total of 25 studies consistent with the inclusion criteria were included, with a total of 2,146 people, which included 1,185 participants in the HBO group and 961 in the control group. This meta-analysis showed that when compared with the controls, HBO therapy can improve the gross motor functions evaluated by the Gross Motor Function Measure (n = 696, SMD 0.29, 95% CI [0.07–0.51], Z = 2.62, p = 0.009) and Gross Motor Function Classification System (n = 248, MD –0.40, 95% CI [–0.52 to –0.27], Z = 6.28, p < 0.00001), global developmental level evaluated by Gesell (n = 560, RR 1.30, 95% CI [1.19–1.42], Z = 6.03, p < 0.00001) and developmental quotient (n = 374, MD 8.25, 95% CI [6.48–10.01], Z = 9.15, p < 0.00001) and language expression (n = 270, MD 4.34, 95% CI [2.30–6.38], Z = 4.17, p < 0.00001) and comprehension (n = 270, MD 4.87, 95% CI [2.87–6.88], Z = 4.76, p < 0.00001). HBO therapy only caused mild ear pain. However, the quality of the data for all outcomes evaluated by the Grading of Recommendations Assessment, Development, and Evaluation analysis was very low. Conclusions: HBO therapy may produce a much more efficient clinical experiment result than the control group with cerebral palsy patients, and HBO therapy is well tolerated and relatively safe for the included participants.

Introduction The ubiquitously expressed nonhistone nuclear protein high‐mobility group box protein 1 (HMGB1) has different functions related to posttranslational modifications and cellular localization. In the nucleus, HMGB1 modulates gene transcription, replication and DNA repair as well as determines chromosomal architecture. When the post‐transcriptional modified HMGB1 is released into the extracellular space, it triggers several physiological and pathological responses and initiates innate immunity through interacting with its reciprocal receptors (i.e., TLR4/2 and RAGE). The effect of HMGB1‐mediated inflammatory activation on different systems has received increasing attention. HMGB1 is now considered to be an alarmin and participates in multiple inflammation‐related diseases. In addition, HMGB1 also affects the occurrence and progression of tumors. However, most studies involving HMGB1 have been focused on adults or mature animals. Due to differences in disease characteristics between children and adults, it is necessary to clarify the role of HMGB1 in pediatric diseases. Methods and Results Through systematic database retrieval, this review aimed to first elaborate the characteristics of HMGB1 under physiological and pathological conditions and then discuss the clinical significance of HMGB1 in the pediatric diseases according to different systems. Conclusions HMGB1 plays an important role in a variety of pediatric diseases and may be used as a diagnostic biomarker and therapeutic target for new strategies for the prevention and treatment of pediatric diseases.

Purpose Histological analysis of artery tissue samples is a widely used method for diagnosis and quantification of cardiovascular diseases. However, the variable and labor-intensive tissue staining procedures hinder efficient and informative histological image analysis. Procedures In this study, we developed a deep learning-based method to transfer bright-field microscopic images of unlabeled tissue sections into equivalent bright-field images of histologically stained versions of the same samples. We trained a convolutional neural network to build maps between the unstained images and histologically stained images using a conditional generative adversarial network model. Results The results of a blind evaluation by board-certified pathologists illustrate that the virtual staining and standard histological staining images of rat carotid artery tissue sections and those involving different types of stains showed no major differences. Quantification of virtual and histological H&E staining in carotid artery tissue sections showed that the relative errors of intima thickness, intima area, and media area were lower than 1.6 %, 5.6 %, and 12.7 %, respectively. The training time of deep learning network was 12.857 h with 1800 training patches and 200 epoches. Conclusions This virtual staining method significantly mitigates the typically laborious and time-consuming histological staining procedures and could be augmented with other label-free microscopic imaging modalities.

Background and aims Type 2 diabetes mellitus (T2DM) is a well-recognized independent risk factor for ASCVD, the aim of this study was to investigate the effects of a dipeptidyl peptidase-4 inhibitor, sitagliptin, on prevention of progression of coronary atherosclerosis assessed by three-dimensional quantitative coronary angiography (3D-QCA) in T2DM patients with coronary artery disease (CAD). Methods This was a prospective, randomized, double-center, open-label, blinded end point, controlled 18-month study in patients with CAD and T2DM. A total of 149 patients, who had at least 1 atherosclerotic plaque with 20%–80% luminal narrowing in a coronary artery, and had not undergone intervention during a clinically indicated coronary angiography or percutaneous coronary intervention, were randomized to sitagliptin group (n = 74) or control group (n = 75). Atherosclerosis progression was measured by repeat 3D-QCA examination in 88 patients at study completion. The primary outcome was changes in percent atheroma volume (PAV) from baseline to study completion measured by 3D-QCA. Secondary outcomes included change in 3D-QCA-derived total atheroma volume (TAV) and late lumen loss (LLL). Results The primary outcome of PAV increased of 1.69% (95%CL, −0.8%–4.2%) with sitagliptin and 5.12% (95%CL, 3.49%–6.74%) with the conventional treatment (p = 0.023). The secondary outcome of change in TAV in patients treated with sitagliptin increased of 6.45 mm3 (95%CL,-2.46 to 6.36 mm3) and 9.45 mm3 (95%CL,-4.52 to 10.14 mm3) with conventional treatment (p = 0.023), however, no significant difference between groups was observed (p = 0.175). Patients treated with sitagliptin had similar LLL as compared with conventional antidiabetics (−0.06, 95%CL, −0.22 to 0.03 vs. −0.08, −0.23 to −0.03 mm, p = 0.689). Conclusions In patients with type 2 diabetes and coronary artery disease, treatment with sitagliptin resulted in a significantly lower rate of progression of coronary atherosclerosis compared with conventional treatment.

Previous studies have demonstrated that a non-invasive light-flickering regime and auditory tone stimulation could affect Aβ and tau metabolism in the brain. As a non-invasive technique, repetitive transcranial magnetic stimulation (rTMS) has been applied for the treatment of neurodegenerative disorders. This study explored the effects of rTMS on Aβ and tau levels in rhesus monkey cerebrospinal fluid (CSF). This is a single-blind, self-controlled study. Three different frequencies (low frequency, 1 Hz; high frequencies, 20 Hz and 40 Hz) of rTMS were used to stimulate the bilateral-dorsolateral prefrontal cortex (DLPFC) of the rhesus monkey. A catheterization method was used to collect CSF. All samples were subjected to liquid chip detection to analyze CSF biomarkers (Aβ42, Aβ42/Aβ40, tTau, pTau). CSF biomarker levels changed with time after stimulation by rTMS. After stimulation, the Aβ42 level in CSF showed an upward trend at all frequencies (1 Hz, 20 Hz, and 40 Hz), with more significant differences for the high-frequencies (p < 0.05) than for the low frequency. After high-frequency rTMS, the total Tau (tTau) level of CSF immediately increased at the post-rTMS timepoint (p < 0.05) and gradually decreased by 24 h. Moreover, the results showed that the level of phosphorylated Tau (pTau) increased immediately after 40 Hz rTMS (p < 0.05). The ratio of Aβ42/Aβ40 showed an upward trend at 1 Hz and 20 Hz (p < 0.05). There was no significant difference in the tau levels with low-frequency (1 Hz) stimulation. Thus, high-frequencies (20 Hz and 40 Hz) of rTMS may have positive effects on Aβ and tau levels in rhesus monkey CSF, while low-frequency (1 Hz) rTMS can only affect Aβ levels.

BACKGROUND: Antithrombotic therapy was commonly applied in treating Kawasaki disease (KD) children; however, the effects and mechanisms of different plans were not fully elucidated. In this study, we aimed to evaluate different antithrombotic drugs. METHODS: Eighty-two children diagnosed with KD in Hebei Children’s Hospital from January 2017 to January 2020 were recruited. For cohort study, KD children were divided into a series of groups according to whether they were complicated with coronary artery lesions (CAL), drug therapy plan, and the presence of liver damage. The thromboelastogram (TEG) indexes [clotting time (R), clot formation time (K), clot formation angle (α), maximum amplitude of the clot (MA), arachidonic acid (AA), and adenosine diphosphate (ADP)] were employed to evaluate the relationship between disease state and drug treatment efficacy. Meanwhile, children were divided into different therapy groups according to their degree of CAL, the treatment efficiency was evaluated by TEG indexes, and the bleeding ratio was recorded. In addition, the warfarin metabolic gene was detected to explain the changes of coagulation parameters in children treated with warfarin. RESULTS: The R value and coagulation index (CoI) were significantly lower (P

The adenylate cyclase activating polypeptide 1 (pituitary) receptor (ADCYAP1R1) gene is associated with the hypothalamic-pituitary-adrenal (HPA) axis, which controls stress responses. The single-nucleotide polymorphism of ADCYAP1R1, rs2267735, has been investigated in many studies to test its association with posttraumatic stress disorder (PTSD), but the results have not been consistent. It is worth systematically exploring the role of rs2267735 in PTSD development. In this study, we analyzed rs2267735 in 1,132 trauma-exposed Chinese individuals (772 females and 360 males). We utilized the PTSD checklist for DSM-5 (PCL-5) to measure the PTSD symptoms. Then, we analyzed the main, G × E (rs2267735 × trauma exposure), and G × G (with other HPA axis gene polymorphisms) effects of rs2267735 on PTSD severity (total symptoms). There were no significant main or G × E effects (P > 0.05). The G × G ADCYAP1R1-FKBP5 interaction (rs2267735 × rs1360780) was associated with PTSD severity (beta = −1.31 and P = 0.049) based on all subjects, and the G × G ADCYAP1R1-CRHR1 interaction (rs2267735 × rs242924) was correlated with PTSD severity in men (beta = −4.72 and P = 0.023). Our study indicated that the ADCYAP1R1 polymorphism rs2267735 may affect PTSD development through diverse gene-gene interactions.

In this paper, a novel image encryption algorithm based on the Once Forward Long Short Term Memory Structure (OF-LSTMS) and the Two-Dimensional Coupled Map Lattice (2DCML) fractional-order chaotic system is proposed. The original image is divided into several image blocks, each of which is input into the OF-LSTMS as a pixel sub-sequence. According to the chaotic sequences generated by the 2DCML fractional-order chaotic system, the parameters of the input gate, output gate and memory unit of the OF-LSTMS are initialized, and the pixel positions are changed at the same time of changing the pixel values, achieving the synchronization of permutation and diffusion operations, which greatly improves the efficiency of image encryption and reduces the time consumption. In addition the 2DCML fractional-order chaotic system has better chaotic ergodicity and the values of chaotic sequences are larger than the traditional chaotic system. Therefore, it is very suitable to image encryption. Many simulation results show that the proposed scheme has higher security and efficiency comparing with previous schemes.

Background Reendothelialisation is the natural pathway that inhibits neointimal hyperplasia and in-stent restenosis. Circulating endothelial progenitor cells (EPCs) derived from bone marrow (BM) might contribute to endothelial repair. However, the temporal and spatial distributions of reendothelialisation and neointimal hyperplasia after EPC transplantation in injured arteries are currently unclear. Methods A carotid balloon injury (BI) model was established in Sprague-Dawley rats, and PKH26-labelled BM-derived EPCs were transplanted after BI. The carotid arteries were harvested on the first, fourth, seventh, and 14th day post-injury and analysed via light-sheet fluorescence microscopy and pathological staining (n = 3). EPC and human umbilical vein endothelial cell culture supernatants were collected, and blood samples were collected before and after transplantation. The paracrine effects of VEGF, IGF-1, and TGF-β1 in cell culture supernatants and serum were analysed by enzyme-linked immunosorbent assay (n = 4). Results Transplanted EPCs labelled with PKH26 were attached to the injured luminal surface the first day after BI. In the sham operation group, the transplanted EPCs did not adhere to the luminal surface. From the fourth day after BI, the mean fluorescence intensity of PKH26 decreased significantly. However, reendothelialisation and inhibition of neointimal hyperplasia were significantly promoted by transplanted EPCs. The degree of reendothelialisation of the EPC7d and EPC14d groups was higher than that of the BI7d and BI14d groups, and the difference in neointimal hyperplasia was observed between the EPC14d and BI14d groups. The number of endothelial cells on the luminal surface of the EPC14d group was higher than that of the BI14d group. The number of infiltrated macrophages in the injured artery decreased in the EPC transplanted groups. Conclusions Transplanted EPCs had chemotactic enrichment and attached to the injured arterial luminal surface. Although decreasing significantly after the fourth day at the site of injury after transplantation, transplanted EPCs could still promote reendothelialisation and inhibit neointimal hyperplasia. The underlying mechanism is through paracrine cytokines and not differentiation into mature endothelial cells.

ObjectiveTo investigate the correlation between clinical characteristics and cardiac injury of COVID-2019 pneumonia.MethodsIn this retrospective, single-center study, 41 consecutive corona virus disease 2019 (COVID-2019) patients (including 2 deaths) of COVID-2019 in Beijing Youan Hospital, China Jan 21 to Feb 03, 2020, were involved in this study. The high risk factors of cardiac injury in different COVID-2019 patients were analyzed. Computed tomographic (CT) imaging of epicardial adipose tissue (EAT) has been used to demonstrate the cardiac inflammation of COVID-2019.ResultsOf the 41 COVID-2019 patients, 2 (4.88%), 32 (78.05%), 4 (9.75%) and 3 (7.32%) patients were clinically diagnosed as light, mild, severe and critical cases, according to the 6th guidance issued by the National Health Commission of China. 10 (24.4%) patients had underlying complications, such as hypertension, CAD, type 2 diabetes mellites and tumor. The peak value of TnI in critical patients is 40-fold more than normal value. 2 patients in the critical group had the onset of atrial fibrillation, and the peak heart rates reached up to 160 bpm. CT scan showed low EAT density in severe and critical patients.ConclusionOur results indicated that cardiac injury of COVID-2019 was rare in light and mild patients, while common in severe and critical patients. Therefore, the monitoring of the heart functions of COVID-2019 patients and applying potential interventions for those with abnormal cardiac injury related characteristics, is vital to prevent the fatality.

Blood-brain barrier (BBB) contributes to maintenance of brain homeostasis. Gut microbiome composition affected BBB development and expression of tight junction proteins in rodents. However, we still do not know if there’s any direct effect of gut microbiol composition on BBB permeability and function in normal adult animals. In current study, we determined temporal and spatial change of BBB permeability in rhesus monkeys receiving either oral or intravenous amoxicillin-clavulanic acid (AC), by monitoring CSF/serum albumin ratio (AR) and the volume transfer constant (Ktrans). We showed that oral, but not intravenous AC led to a significant alteration in gut microbiol composition and increase of BBB permeability in all monkeys, especially in thalamus area. Acetic acid and propionic acid might play a pivotal role in this newly found communication between gut and central nervous system. Antibiotics-induced gut microbiol composition change, especially the decreasing of acetic and propionic acid producing phyla and genera, leads to increase of BBB permeability, which may contribute to a variety of neurological and psychological diseases.

The blood-brain barrier (BBB) contributes to the maintenance of brain homeostasis. Gut microbiome composition affects BBB development and expression of tight junction proteins in rodents. However, we still do not know if there is any direct effect of gut microbial composition on BBB permeability and function in normal adult animals. In the current study, we determined temporal and spatial changes in BBB permeability of rhesus monkeys receiving amoxicillin-clavulanic acid (AC) by monitoring the cerebrospinal fluid/serum albumin ratio and the volume transfer constant (Ktrans ). We showed that oral, but not intravenous, AC was associated with subsequent significant alteration in gut microbial composition and an increase in BBB permeability in all monkeys, especially in the thalamus area. Acetic and propionic acids might play a pivotal role in this newly found communication between the gut and the central nervous system. Antibiotic-induced gut microbial composition change, especially the decrease in acetic acid- and propionic acid-producing phyla and genera, might have a potential effect on the increase in BBB permeability, which may contribute to a variety of neurological and psychological diseases.

Background Relatively little is known about the role of phosphatidylserine (PS) in procoagulant activity (PCA) in patients with non-ST-elevated myocardial infarction (NSTEMI) after stent implantation. This study was designed to evaluate whether exposed PS on microparticles (MPs) and blood cells were involved in the hypercoagulable state in NSTEMI patients with stent implantation. Methods NSTEMI patients (n=90) and healthy controls (n=20) were included in our study. PS exposure on MPs and blood cells was analyzed with flow cytometer and confocal microscope. PCA was evaluated by clotting time, purified coagulation complex assays and fibrin production assays. Results Baseline levels of MPs and PS + blood cells were significantly higher (P + blood cells. Specifically, PS + MPs, PS + platelets and erythrocytes peaked at 18h following stent implantation, while PS + leukocytes peaked on day 2. In addition, circulating MPs (mostly derived from platelets, leukocytes, erythrocytes and endothelial cells) cooperating with PS + blood cells, contributed to markedly shortened coagulation time and markedly increased FXa/thrombin/fibrin (all P Conclusions Our results suggest that PS + MPs and blood cells play a procoagulant role in NSTEMI patients following stent implantation. Blockade of PS could become a novel therapeutic modality for the prevention of thrombosis in these patients.

Purpose/Aim of the study: Cerebrovascular reactivity (CVR) is an important marker for assessing cerebrovascular disease. This study assessed the CVR by perfusion computed tomography (CT) and CO2 inhalation tests in patients with unilateral middle cerebral artery (MCA) stenosis disease. Materials and Methods: Thirty-one patients with unilateral MCA stenosis disease diagnosed by digital subtraction angiography were studied. Patients were divided into two groups according to the degree of stenosis: severe and moderate. The regional cerebral blood flow (CBF) before and after CO2 inhalation was determined by perfusion CT. Regional CVR values were obtained by the following formula: increase (%) = (post-CBF) − (pre-CBF)/(pre-CBF) × 100%. Results: No significant differences in the mean CBF in the MCA stenosis region were found between the affected and contralateral sides before the CO2 inhalation test; after the test, CBF was more significantly decreased on the affected side than on the contralateral side. The ch...

Yinbing Zhang,1,2 Haifeng Liu,3 Yingqian Zhang,4 Qiong Wu,1 Yanyan Zhang,1,2 Jie Zhang,1,2 Xiangshan Zhou,3 He Jiao,5 Feng Fan,6 Qi Xue,7 Xin Wang,1,2 Zhihui Zhong1,2 1Laboratory of Nonhuman Primate Disease Modeling Research, State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China; 2Sichuan Kangcheng Biomed Co., Ltd., Chengdu, China; 3Angde Biotech Pharmaceutical Co., Ltd., Liaocheng, China; 4Department of Physiology, Southwest Medical University, Luzhou, China; 5Department of Interventional therapy, West China Hospital, Sichuan University, Chengdu, China; 6Department of Neurointervention, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, China; 7Food and Drug Administration of Shibei District Government, Qingdao, China Purpose: In order to evaluate the thrombolytic effects of reteplase in pulmonary thromboembolism (PTE), we developed a novel canine PTE model. The efficacy of reteplase against PTE in comparison to alteplase was clarified for the first time, and this PTE model could be further applied to studies of novel thrombolytic therapies. Patients and methods: Twenty-four dogs were divided into four groups: sham operation, vehicle, alteplase, and reteplase. Autologous thrombi/saline were injected into the pulmonary artery, and thrombolytic agents were administrated. Thrombus formation and dissolution were monitored by real-time digital subtraction angiography (DSA), and pulmonary pressures were measured simultaneously. Blood coagulation, blood gas, hematology, and histopathologic examinations were used as subsidiary methods. Results: The canine PTE model was established with a significant decrease of blood flow and ~75% blocking area. Administration of reteplase (0.6 mg/kg) resulted in effective thrombus dissolution with a recovery of over 80% blood flow, as effective as alteplase (1.6 mg/kg). Correspondingly, the elevated pulmonary systolic, diastolic, and mean arterial pressures declined to the normal level. Blood coagulation was changed by reteplase, with a dramatic elongation of prothrombin time, activated partial thromboplastin time, and thrombin time, even longer than alteplase. In contrast to the vehicle group, no obvious pathological changes were found in the two thrombolytic groups. Hematological, blood biochemical, and blood gas results also indicated that reteplase had no adverse reactions in this PTE model. Conclusion: Reteplase proved to be an effective and safe therapy for PTE for the first time, and a small dosage of reteplase exerted an efficacy comparable to the routine dosage of alteplase. Our findings indicated the potential of reteplase as clinical treatment against PTE. This technically innovative, stability- and validity-proved canine PTE model developed by minimally invasive surgery and DSA resembled major clinical features. This may further facilitate our understanding of thrombotic disorders and development of prophylactic and therapeutic approaches. Keywords: reteplase, canine pulmonary thromboembolism model, thrombolytic therapy

BACKGROUND The aim of this study was to evaluate the effects of carotid artery angioplasty and carotid artery stenting (CAS) on cerebral blood flow (CBF) and cerebrovascular reactivity (CVR) in patients with preoperative cerebrovascular hemodynamic impairment. MATERIAL AND METHODS Seventeen patients with unilateral severe internal carotid artery (ICA) stenosis and ipsilateral CVR impairment underwent CAS. CBF and CVR were measured by single photon emission computed tomography (SPECT) with inhalation of carbon dioxide (CO2) one week before and three months after CAS. Sixty-eight ROIs in the middle cerebral artery (MCA) territory were analyzed in 17 patients. RESULTS Before CAS, CVR was impaired in all ROIs. CBF was impaired in 16 ROIs (23.5%). The percentage of ROIs with impaired CBF was significantly increased in patients with ≥90% carotid artery stenosis (p=0.047) without collateral flow through the circle of Willis (p=0.005). CAS significantly increased CVR in ROIs with a normal preoperative CBF and impaired CVR, indicating mild hemodynamic impairment (0.9±6.7% vs. 4.9±8.6%) (p=0.014). CAS significantly increased CBF in ROIs with preoperative impaired CBF and impaired CVR, indicating severe hemodynamic impairment (79.1±7.5% vs. 86.7±10.0%) (p

Purpose/aim of the study: Cerebrovascular reactivity (CVR) reflects the vasodilatory reserve of cerebral resistance vessels, which is an important marker for assessing cerebrovascular disease. The present study is to investigate whether CVR impairment increases adverse long-term outcome risk of patients with ≥ 50% symptomatic unilateral middle cerebral artery (MCA) stenosis (ischemic stroke (IS) or transient ischemic attack (TIA)).Digital subtraction angiography (DSA) was used to assess the degree of stenosis, and perfusion CT and 5% COSeventy-three patients with ≥ 50% symptomatic unilateral MCA stenosis, involving 31 non-CVR impairment cases and 42 CVR impairment cases, were included in the present study. Finally, IS occurred in six CVR impairment patients, and no endpoint happened in the non-CVR impairment group. Therefore, the annual rate of IS was 14.29% in the CVR impairment group and 0% in the non-CVR impairment group (P = 0.035). Besides, further Kaplan-Meier analysis found CVR impairment was closely associated with the IS risk (Kaplan-Meier Log-rank 4.719, P = 0.030).Our results showed that for patients with ≥ 50% symptomatic unilateral MCA stenosis, there was significant difference between non-CVR impairment cases and CVR impairment cases in the annual rate of IS. It suggests that CVR impairment increases the risk of adverse long-term outcomes.

Hypoxia/reoxygenation (H/R) is one of the cellular stresses in pathological conditions, such as myocardial infarction, stroke and organ transplantation. Oxidative stress caused by reactive oxygen species (ROS) is a crucial element of H/R injury in vascular endothelial cells (ECs). Xanthine oxidase (XO) has been recognized to contribute to H/R injury. Of note, xanthine oxidoreductase is synthesized as xanthine dehydrogenase (XDH) and needs to be converted to XO to become a source of superoxide. Hepatocyte growth factor (HGF) has been found to protect ECs against H/R injury. The relation, however, between HGF and XO in ECs under H/R conditions remains to be determined. Primary cultured rat cardiac microvascular endothelial cells (CMECs) were exposed to 4 h of hypoxia and followed by 1 h of reoxygenation. Generation of ROS and cytosolic Ca2+ concentration was measured by flow cytometry qualification of DCFHDA and fluo-3 AM staining cells, respectively. XDH mRNA was qualified by qRT-PCR analysis. XO activity was determined by colorimetric assay and XO protein levels were determined by Western blot. Cell apoptosis was assessed by caspase-3 activity and Annexin V/PI staining. After H/R, cellular ROS production significantly increased. Both XO activity and XO protein increased after H/R. Cellular ROS elevation was inhibited by allopurinol (a potent XO inhibitor), indicting XO accounting for the generation of ROS after H/R. In addition, XDH mRNA increased after H/R, indicating a de novo XDH synthesis, which needs to be converted to XO to become a source of superoxide. Pretreatment of HGF inhibited the elevation of XO activity and XO protein level after H/R; however, HGF has no effect on the increase of XDH mRNA. We also find an increase of the cytosolic Ca2+ in CMECs after H/R. BAPTA-AM, a cell-permeable Ca2+ chelator, prevented the increase of XO activity and XO protein levels, implicating the elevated cytosolic Ca2+ concentration involvement in XO conversion and XO activation. HGF inhibited the elevation of cytosolic Ca2+ concentration in CMECs after H/R. Furthermore, HGF ameliorated H/R-induced CMECs apoptosis. These findings suggest a novel mechanism whereby HGF inhibited XO-generated ROS production after H/R treatment. H/R induces a de novo synthesis of XDH, the XO precursor. In addition, H/R increases cytosolic Ca2+ concentration and promotes a Ca2+ -involved XO conversion and XO activation. HGF has no effect on the increase of XDH mRNA; however, HGF inhibited the elevation of XO protein level and XO activity after H/R in the post-transcriptional level primarily by inhibiting the increase of cytosolic Ca2+ concentration. HGF protects CMECs from H/R-induced apoptosis by inhibiting the elevation of XO protein level and XO activity.

Microvascular endothelial cells (CMECs) oxidative damage resulting from hypoxia/reoxygenation (H/R) injury is responsible for microcirculation perfusion disturbances and the progression of cardiac dysfunction. However, few strategies are available to reverse such pathologies. Here, we studied the effects and mechanisms of liraglutide on CEMCs oxidative damage, focusing in particular on calcium overload-triggered free radical injury signals and the GLP-1R/PI3K/Akt/survivin survival pathways. The results indicate that H/R increased IP3R expression but reduced SERCA2a expression, which rapidly raised intracellular Ca(2+) levels, subsequently leading to Ca(2+)-dependent xanthine oxidase (XO) activation, reactive oxygen species (ROS) production and the cellular apoptosis of CMECs. However, liraglutide pretreatment abrogated Ca(2+)-mediated oxidative apoptosis. Furthermore, liraglutide regulated the rate of IP3R/SERCA2a gene transcription and conserved SERCA2a-ATPase activity via the maintenance of ATP production under H/R, which drove excessive Ca(2+) reflux to the sarcoplasmic reticulum (SR) and inhibited Ca(2+) release from the SR, ultimately restoring Ca(2+) homeostasis. Furthermore, the regulatory role of liraglutide on Ca(2+) balance in conjunction with its up-regulation of superoxide dismutase, glutathione and glutathione peroxidase collectively scavenged the excess ROS under H/R. Moreover, we showed that liraglutide strengthened Akt phosphorylation and subsequently survivin expression. In addition, both the blockade of the GLP-1R/PI3K/Akt pathways and the siRNA-mediated knockdown of survivin abolished the protective effects of liraglutide on SR-Ca(2+) function and CMECs oxidative apoptosis. In summary, this study confirmed that H/R induced CMECs oxidative damage through the SR-Ca(2+)-XO-ROS injury signals and that liraglutide pretreatment may suppress such CMECs damage through the PI3K/Akt/survivin pathways.

Blood–brain barrier (BBB) disruption plays an important role in pathophysiological progress of ischemic stroke. However, our knowledge of the dynamic change of BBB permeability and its mechanism remains limited. In the current study, we used a non-human primate (NHP) MCAO model and a serial CSF sampling method that allowed us to determine the dynamic change of BBB permeability by calculating the CSF/serum albumin ratio (AR). We showed that AR increased rapidly and significantly after ischemia, and the fold increase of AR is highly correlated with the infarction size during the subacute phase. Moreover, we determined the temporal change of MMP-1, MMP-2, MMP-3, MMP-9, MMP-10, MMP-13, TIMP-1, and TIMP-2 in CSF and serum. Each MMP and TIMP showed different change patterns when comparing their values in CSF and serum. Based on the longitudinal dataset, we showed that the fold increase of MMP-9 in serum and CSF are both correlated to infarction size. Among the measured MMPs and TIMPs, only MMP-2, MMP-13, and TIMP-2 in CSF correlated with AR to some extent. Our data suggest there is no single MMP or TIMP fully responsible for BBB breakdown, which is regulated by a much more complicated signal network and further investigations of the mechanisms are needed.

Diabetic patients suffer from undesired intimal hyperplasia after angioplasty. Nicorandil has a trend to reduce the rate of target lesion revascularization. However, whether nicorandil inhibits intimal hyperplasia and the possible mechanisms underlying it remain to be determined. We aimed at

We build the hypothesis that nicorandil may promote reendothelialization impaired by rapamycin through inhibiting XO-generated ROS formation and promoting eNOS expression. In vivo carotid balloon injury model and in vitro CMECs were used to indentify this hypothesis. Sprague-Dawley (SD) rats (male

A whole genome scale proteome array consisting of 908 open reading frames encoded in Chlamydia trachomatis genome and plasmid was used to profile anti-chlamydial Ab responses. A total of 719 chlamydial proteins was recognized by one or more antisera from 99 women urogenitally infected with C. trachomatis. Revealing such a large C. trachomatis ANTIGENome in humans might partially be attributed to the significantly improved detection sensitivity of the whole genome scale proteome array assay because both linear and conformation-dependent Abs were detected by the array assay. Twenty-seven of the 719 Ags were recognized by ≥50% antisera, thus designated as immunodominant Ags. Comparison of Ag profiles recognized by live chlamydial organism-infected versus dead organism-immunized hosts led to the identification of infection-dependent or in vivo expressed Ags. The infection-dependent Ags induced Abs only in live organism-infected, but not in dead organism-immunized hosts. Many of these Ags were highly expressed during replication, but only minimally packaged into the infectious elementary bodies. Because inactivated whole chlamydial organism-based vaccines failed to induce protection in humans, identification of the infection-dependent or in vivo expressed immunodominant Ags in humans should greatly facilitate the selection of promising chlamydial subunit vaccine candidates for further evaluation. This approach may also be applicable to other pathogens.

A study was conducted to evaluate the inhibitory effects of vascular endothelial growth factor receptor 2 (VEGFR2) monoclonal antibody, BC001, against laser-induced choroidal neovascularization (CNV).We induced the experimental CNV in rhesus monkey eyes using laser photocoagulation. Monkeys were randomly assigned to 4 groups that received a single intravitreal administration of BC001 at 0 (vehicle-treated group), 0.05, 0.2, and 0.5 mg/eye. Fundus fluorescein angiography, optical coherence tomography, and histological studies were used for evaluations. The ocular recovery was determined by comparing changes of fluorescein leaking area and thickness of disrupted retina around the laser burn spot before and after drug administration. Choroidal blood vessels were stained and quantified by lectin staining. Hematoxylin and eosin staining was performed to determine the general histological complications.An intravitreal injection of BC001 at 0.05, 0.2, and 0.5 mg per eye at 20 days after laser burn significantly reduced the CNV-induced fluorescein leakage, retina pathology, and aberrant choroidal vessel growth and did not change intraocular pressure or induce any immune response.BC001 confers significant inhibitory effects against laser-induced CNV in rhesus monkeys, thereby suggesting that prevention of VEGFR2 activation may be promising as an alternative therapeutic target for exudative age-related macular degeneration.

Although the presence of C-type natriuretic peptide (CNP) in gastrointestinal tract has been demonstrated, the effect of CNP on interstitial cells of Cajal (ICC), pacemaker cells in gastrointestinal tract, is still unclear. This study was designed to investigate the effect of CNP on pacemaker currents of ICC and possible mechanisms. We used immunocytochemistry techniques to exhibit natriuretic peptide receptors (NPR) and recorded membrane currents by using whole-cell patch clamp technique on cultured ICC. Our experiment showed that NPR-A and NPR-B were expressed in ICC from murine small intestine. Whole cell recordings further showed that the amplitude of pacemaker currents in intestinal small networks of ICC was 322+/-22pA and the frequency was 16.25+/-0.95Hz. CNP significantly reduced the amplitude of pacemaker currents in small networks of ICC in a dose-dependent manner, and the amplitude was inhibited by 23.95%, 61.76% and 81.67%, the amplitude values in 329+/-28.0pA, 311.2+/-14.8pA and 295+/-26.5pA before treatment with CNP and 237.9+/-27.5pA, 119.6+/-18.5pA and 57.2+/-13.5pA after treatment with 0.01 micromolxL(-1), 0.1 micromolxL(-1) and 1pmolxL(-1) CNP, respectively. The frequencies of pacemaker currents were also significantly reduced from 16.25+/-0.95Hz of control to 13+/-0.9Hz, 12+/-0.8Hz and 3+/-0.2Hz by 0.01micromolxL 1, 0.1micromolxL(-1) and 1 micromol x L(-1) CNP, respectively. CNP also inhibited the amplitude of pacemaker currents in single ICC. The inhibitory effect of CNP was mimicked by 8-Br-cGMP, a membrane permeable cGMP analogue, which suggests that CNP could inhibit pacemaker currents via NPR-B-particulate guanylate cyclase (pGC)-cGMP signal pathway.

Objective The objective of the study was to assess antibodies against Chlamydia trachomatis heat shock proteins (HSP) in patients with tubal factor infertility (TFI), infertility controls (IFC), and fertile controls (FC). HSPs assist organisms in surviving caustic environments such as heat. Study design Twenty-one TFI, 15 IFC, and 29 FC patients were enrolled after laparoscopic tubal assessment. The titers of antibodies against C trachomatis organisms and 14 chlamydial HSPs were compared among the 3 groups. Results TFI patients developed significantly higher levels of antibodies against C trachomatis and specifically recognizing chlamydial HSP60 and caseinolytic protease (Clp) P, a subunit of the ATP-dependent Clp protease complex involved in the degradation of abnormal proteins. Conclusion In addition to confirming high titers of antibodies against C trachomatis organisms and HSP60 in TFI patients, we identified a novel link of TFI with anti-ClpP antibodies. These findings may provide useful information for developing a noninvasive screening test for TFI and constructing subunit anti- C trachomatis vaccines.

We have previously shown that individuals infected with Chlamydia trachomatis can develop a robust antibody response to a Chlamydia type III secretion effector protein called Tarp and that immunization with Tarp induces protection against challenge infection in mice. The current study aimed to map the immunodominant regions of the Tarp protein by expressing 11 fragments of Tarp as glutathione S -transferase (GST) fusion proteins and detecting the reactivity of these fusion proteins with antisera from patients infected with C. trachomatis in the urogenital tract or in the ocular tissue and from rabbits immunized with C. trachomatis organisms. A major immunodominant region was strongly recognized by all antibodies. This region covers amino acids 152 to 302, consisting of three repeats (amino acids 152 to 201, 202 to 251, and 252 to 302). Each of the repeats contains multiple tyrosine residues that are phosphorylated by host cell kinases when Tarp is injected into host cells. Several other minor immunodominant regions were also identified, including those comprising amino acids 1 to 156, 310 to 431, and 582 to 682 (recognized by antisera from both humans and rabbits), that comprising amino acids 425 to 581 (recognized only by human antisera), and that comprising amino acids 683 to 847 (preferentially recognized by rabbit antisera). This immunodominance was also confirmed by the observations that six out of the nine monoclonal antibodies (MAbs) bound to the major immunodominant region and that the other three each bound to one of the minor fragments, comprising amino acids 1 to 119, 120 to 151, and 310 to 431. The antigenicity analyses have provided important information for further understanding the structure and function of Tarp.

Chlamydia trachomatis genome is predicted to encode a type III secretion system consisting of more than forty open reading frames (ORFs). To test whether these ORFs are expressed and immunogenic during chlamydial infection in humans, we expressed 55 chlamydial ORFs covering all putative type III secretion components plus control molecules as fusion proteins and measured the reactivity of these fusion proteins with antibodies from patients infected with C. trachomatis in the urogenital tract (24 antisera) or in the ocular tissue (8 antisera). Forty-five of the 55 proteins were recognized by at least one of the 32 human antisera, suggesting that these proteins are both expressed and immunogenic during chlamydial infection in humans. Tarp, a putative type III secretion effector protein, was identified as a novel immunodominant antigen due to its reactivity with the human antisera at high frequency and titer. The expression and immunogenicity of Tarp were confirmed in cell culture and mouse systems. Tarp was mainly associated with the infectious form of chlamydial organisms and became undetectable between 13 and 24 hours during the infection cycle in cell culture. Mice intravaginally infected with C. muridarum developed Tarp-specific humoral and cellular immune responses. More importantly, immunization of mice with Tarp induced Th1-dominant immunity that significantly reduced the shedding of live organisms from the lower genital tract and attenuated inflammatory pathologies in the fallopian tube tissues. These observations have demonstrated that Tarp, an immunodominant antigen identified by human antisera, can induce protective immunity against chlamydial infection and pathology in mice.