The Effect of Hemodynamic Remodeling on the Survival of Arterialized Venous Flaps

Objective

To evaluate the effect of hemodynamic remodeling on the survival status of the arterialized venous flaps (AVFs) and investigate the mechanism of this procedure.

Materials and Methods

Two 7 x 9 cm skin flaps in each rabbit (n=36) were designed symmetrically in the abdomen. The thoracoepigastric pedicle and one femoral artery were used as vascular sources. Four groups were included: Composite skin grafts group and arterial perfusion group were designed in one rabbit; AVF group and hemodynamic remodeling group by ligation of the thoracoepigastric vein in the middle were outlined in another rabbit. Flap viability, status of vascular perfusion and microvasculature, levels of epidermal metabolite and water content in each group were assessed.

Results

Highly congested veins and simple trunk veins were found using angiography in the AVF group; while a fairly uniform staining and plenty of small vessels were observed in the hemodynamic remodeling group. The metabolite levels of the remodeling group are comparable with those in the arterial perfusion group. There was no statistically significant difference in the percentage of flap survival between the arterial perfusion group and hemodynamic remodeling group; however, significant difference was seen between the AVF group and the hemodynamic remodeling group.

Conclusions

Under the integrated perfusion mode, the AVFs are in an over-perfusion and non-physiological hemodynamic state, resulting in unreliability and unpredictability in flap survival; under the separated perfusion mode produced by remodeling, a physiological-like circulation will be created and therefore, better flap survival can be expected.     

Improved Visualization of Intracranial Vessels with Intraoperative Coregistration of Rotational Digital Subtraction Angiography and Intraoperative 3D Ultrasound

Introduction

Ultrasound can visualize and update the vessel status in real time during cerebral vascular surgery. We studied the depiction of parent vessels and aneurysms with a high-resolution 3D intraoperative ultrasound imaging system during aneurysm clipping using rotational digital subtraction angiography as a reference.

Methods

We analyzed 3D intraoperative ultrasound in 39 patients with cerebral aneurysms to visualize the aneurysm intraoperatively and the nearby vascular tree before and after clipping. Simultaneous coregistration of preoperative subtraction angiography data with 3D intraoperative ultrasound was performed to verify the anatomical assignment.

Results

Intraoperative ultrasound detected 35 of 43 aneurysms (81%) in 39 patients. Thirty-nine intraoperative ultrasound measurements were matched with rotational digital subtraction angiography and were successfully reconstructed during the procedure. In 7 patients, the aneurysm was partially visualized by 3D-ioUS or was not in field of view. Post-clipping intraoperative ultrasound was obtained in 26 and successfully reconstructed in 18 patients (69%) despite clip related artefacts. The overlap between 3D-ioUS aneurysm volume and preoperative rDSA aneurysm volume resulted in a mean accuracy of 0.71 (Dice coefficient).

Conclusions

Intraoperative coregistration of 3D intraoperative ultrasound data with preoperative rotational digital subtraction angiography is possible with high accuracy. It allows the immediate visualization of vessels beyond the microscopic field, as well as parallel assessment of blood velocity, aneurysm and vascular tree configuration. Although spatial resolution is lower than for standard angiography, the method provides an excellent vascular overview, advantageous interpretation of 3D-ioUS and immediate intraoperative feedback of the vascular status. A prerequisite for understanding vascular intraoperative ultrasound is image quality and a successful match with preoperative rotational digital subtraction angiography.     

Catecholamine and Volume Therapy for Cardiac Surgery in Germany – Results from a Postal Survey

Background

Management of cardiac surgery patients is a very standardized procedure in respective local institutions. Yet only very limited evidence exists concerning optimal indication, safety and efficacy of hemodynamic monitoring catecholamine and fluid therapy.

Methods

Between April and May 2013, all 81 German anaesthesia departments involved in cardiac surgery care were asked to participate in a questionnaire addressing the institutional specific current practice in hemodynamic monitoring, catecholamine and volume therapy.

Results

51 (63%) questionnaires were completed and returned. All participating centers used basic hemodynamic monitoring (i.e. invasive arterial blood pressure and central venous pressure), supplemented by transesophageal echocardiography. Pulmonary arterial catheter and calibrated trend monitoring devices were also routinely available. In contrast, non-calibrated trend monitoring and esophageal doppler ultrasound devices were not commonly in use. Cerebral oximetry is increasingly emerging, but lacks clear indications. The majority of patients undergoing cardiac surgery, especially in university hospitals, required catecholamines during perioperative care, In case of low cardiac output syndrome, dobutamine (32%), epinephrine (30%) or phosphodiesterase inhibitors (8%) were first choice. In case of hypotension following vasoplegia, norepinephrine (96%) represented the most common catecholamine. 88% of the participating centers reported regular use of colloid fluids, with hydroxyethyl starches (HES) being first choice (64%).

Conclusions

Choice of hemodynamic monitoring is homogenous throughout German centers treating cardiac surgery patients. Norepinephrine is the first line catecholamine in cases of decrease in peripheral vascular resistance. However, catecholamine choice for low cardiac output syndrome varies considerably. HES was the primary colloid used for fluid resuscitation. After conduct of this survey, HES use was restricted by European regulatory authorities in critically ill patients and should only be considered as second-line fluid in surgical patients without renal impairment or severe coagulopathy. Large clinical studies addressing catecholamine and fluid therapy in cardiac surgery patients are lacking.     

Diet Induced Mild Hypercholesterolemia in Pigs: Local and Systemic Inflammation,Effects on Vascular Injury – Rescue by High-Dose Statin Treatment

Objective

The aim of the present study was to comprehensively evaluate systemic and local inflammation as well as progression of vascular inflammation in normal and mechanically injured vessels in a large animal model of mild hypercholesterolemia. Our aim was also to test the effect of high-dose statin treatment on these processes.

Methods

Pigs were kept for 120 days on a standard diet (SD, n=7), high-cholesterol diet (HCD, n=7) or high-cholesterol diet with Atorvastatin starting after 50 days (STATIN, n=7). Left carotid artery balloon injury was conducted in all groups after 60 days of diet treatment. Biochemical analysis together with evaluation of blood and tissue markers of vascular injury and inflammation were performed in all groups at the end of experiment.

Results

HCD compared to SD induced systemic inflammation demonstrated by increased number of circulating monocytes and lymphocytes. HCD compared to SD induced also local inflammation demonstrated by adipocyte hypertrophy and infiltration of T-lymphocytes in abdominal white adipose tissue, activation of hepatic stellate cells with infiltration of T- and B-lymphocytes and macrophages in the liver and increased macrophage content in lung parenchyma. These changes were accompanied by increased Intima/Media thickness, stenosis, matrix deposition and activated T-cell infiltrates in injured but not in uninjured contralateral carotid artery as we previously reported. The treatment with high-dose statin attenuated all aspects of systemic and local inflammation as well as pathological changes in injured carotid artery.

Conclusions

Diet related mild hypercholesterolemia induce systemic and local inflammation in the liver, lung and adipose tissue that coincide with enhanced inflammation of injured vessel but is without deleterious effect on uninjured vessels. High dose statin attenuated systemic and local inflammation and protected injured vessels. However, finding exact role of reduced systemic and remote inflammation in vascular protection requires further studies.     

Does Antenatal Maternal Psychological Distress Affect Placental Circulation in the Third Trimester?

Introduction

Some types of antenatal maternal psychological distress may be associated with reduced fetal growth and birthweight. A stress-mediated reduction in placental blood flow has been suggested as a mechanism. Previous studies have examined this using ultrasound-derived arterial resistance measures in the uterine (UtA) and umbilical (UA) arteries, with mixed conclusions. However, a reduction in placental volume blood flow may occur before changes in arterial resistance measures are seen. Fetoplacental volume blood flow can be quantified non-invasively in the umbilical vein (UV). Our objective was to study whether specific types of maternal psychological distress affect the placental circulation, using volume blood flow quantification in addition to arterial resistance measures.

Methods

This was a prospective observational study of 104 non-smoking pregnant women (gestational age 30 weeks) with uncomplicated obstetric histories. Psychological distress was measured by General Health Questionnaire-28 (subscales anxiety and depression) and Impact of Event Scale-22 (subscales intrusion, avoidance and arousal). UtA and UA resistance measures and UV volume blood flow normalized for fetal abdominal circumference, were obtained by Doppler ultrasound.

Results

IES intrusion scores above the mean were associated with a reduction in normalized UV volume blood flow (corresponding to –0.61 SD; P = 0.003). Adjusting for UA resistance increased the strength of this association (difference –0.66 SD; P<0.001). Other distress types were not associated with UV volume blood flow. Maternal distress was not associated with arterial resistance measures, despite adjustment for confounders.

Conclusions

Intrusive thoughts and emotional distress regarding the fetus were associated with reduced fetoplacental volume blood flow in third trimester. Uterine and umbilical artery resistance measures were not associated with maternal distress. Our findings support a decrease in fetoplacental blood flow as a possible pathway between maternal distress and reduced fetal growth.     

Optic Nerve Head Blood Flow Autoregulation during Changes in Arterial Blood Pressure in Healthy Young Subjects

Aim

In the present study the response of optic nerve head blood flow to an increase in ocular perfusion pressure during isometric exercise was studied. Based on our previous studies we hypothesized that subjects with an abnormal blood flow response, defined as a decrease in blood flow of more than 10% during or after isometric exercise, could be identified.

Methods

A total of 40 healthy subjects were included in this study. Three periods of isometric exercise were scheduled, each consisting of 2 minutes of handgripping. Optic nerve head blood flow was measured continuously before, during and after handgripping using laser Doppler flowmetry. Blood pressure was measured non-invasively in one-minute intervals. Intraocular pressure was measured at the beginning and the end of the measurements and ocular perfusion pressure was calculated as 2/3*mean arterial pressure –intraocular pressure.

Results

Isometric exercise was associated with an increase in ocular perfusion pressure during all handgripping periods (p < 0.001). By contrast no change in optic nerve head blood flow was seen. However, in a subgroup of three subjects blood flow showed a consistent decrease of more than 10% during isometric exercise although their blood pressure values increased. In addition, three other subjects showed a consistent decline of blood flow of more than 10% during the recovery periods.

Conclusion

Our data confirm previous results indicating that optic nerve head blood flow is autoregulated during an increase in perfusion pressure. In addition, we observed a subgroup of 6 subjects (15%) that showed an abnormal response, which is in keeping with our previous data. The mechanisms underlying this abnormal response remain to be shown.     

Hemodynamic and Metabolic Correlates of Perinatal White Matter Injury Severity

Background and Purpose

Although the spectrum of perinatal white matter injury (WMI) in preterm infants is shifting from cystic encephalomalacia to milder forms of WMI, the factors that contribute to this changing spectrum are unclear. We hypothesized that the variability in WMI quantified by immunohistochemical markers of inflammation could be correlated with the severity of impaired blood oxygen, glucose and lactate.

Methods

We employed a preterm fetal sheep model of in utero moderate hypoxemia and global severe but not complete cerebral ischemia that reproduces the spectrum of human WMI. Since there is small but measurable residual brain blood flow during occlusion, we sought to determine if the metabolic state of the residual arterial blood was associated with severity of WMI. Near the conclusion of hypoxia-ischemia, we recorded cephalic arterial blood pressure, blood oxygen, glucose and lactate levels. To define the spectrum of WMI, an ordinal WMI rating scale was compared against an unbiased quantitative image analysis protocol that provided continuous histo-pathological outcome measures for astrogliosis and microgliosis derived from the entire white matter.

Results

A spectrum of WMI was observed that ranged from diffuse non-necrotic lesions to more severe injury that comprised discrete foci of microscopic or macroscopic necrosis. Residual arterial pressure, oxygen content and blood glucose displayed a significant inverse association with WMI and lactate concentrations were directly related. Elevated glucose levels were the most significantly associated with less severe WMI.

Conclusions

Our results suggest that under conditions of hypoxemia and severe cephalic hypotension, WMI severity measured using unbiased immunohistochemical measurements correlated with several physiologic parameters, including glucose, which may be a useful marker of fetal response to hypoxia or provide protection against energy failure and more severe WMI.     

Evaluation of the Combined Application of Ultrasound Imaging Techniques for Middle Cerebral Artery Stent Surveillance and Follow-Up Study

Objective

In recent years, cerebral artery stenting has become an effective method for the treatment of cerebral artery stenosis. However, methods for assessing efficacy and techniques for follow-up imaging still need to be developed. This study was designed to evaluate the application of transcranial color-coded sonography (TCCS) in assessing stenting of middle cerebral artery (MCA) stenosis. And, two new imaging techniques (vascular enhancement technology (VET) and 3-dimensional (3D) imaging) were tried out and evaluated.

Method

We enrolled 43 patients with cerebral artery stenosis for vascular stent implantation. All patients were examined by ultrasonography and confirmed through digital subtraction angiography. The stenosis was imaged and blood flow parameters were analyzed before and after the procedure using TCCS. VET and 3D imaging model were used in part of the patients. Important postoperative hemodynamic changes were noted.

Results

1) Adequate stent image was present in 41 out of 43 patients as detected by postoperative 2-dimensional imaging. Images lacking clarity were obtained in 2 patients. 2) The perioperative and postoperative (one week follow-up) instantaneous blood flow velocity at the site of stenosis was significantly decreased (P<0.05) when compared with preoperative levels. Differences between postoperative (one week follow-up) and preoperative blood flow velocity were significant (P<0.05). Differences in blood flow velocity at long-term follow-up (six months and two years) compared to one-week values were not statistically significant (P>0.05). 3) VET imaging visualizes the MCA lumen and stent morphology clearly. 3D ultrasound can be used for imaging of the stent shape as well as its inner surface.

Conclusion

TCCD can be considered a quick and effective clinical detection method to evaluate the intracranial arterial hemodynamics changes before and after stenting treatment for MCA stenosis. New imaging technologies 3D and VET can achieve additional image information.     

Modified Uterine Allotransplantation and Immunosuppression Procedure in the Sheep Model

Objective

To develop an orthotopic, allogeneic, uterine transplantation technique and an effective immunosuppressive protocol in the sheep model.

Methods

In this pilot study, 10 sexually mature ewes were subjected to laparotomy and total abdominal hysterectomy with oophorectomy to procure uterus allografts. The cold ischemic time was 60 min. End-to-end vascular anastomosis was performed using continuous, non-interlocking sutures. Complete tissue reperfusion was achieved in all animals within 30 s after the vascular re-anastomosis, without any evidence of arterial or venous thrombosis. The immunosuppressive protocol consisted of tacrolimus, mycophenolate mofetil and methylprednisolone tablets. Graft viability was assessed by transrectal ultrasonography and second-look laparotomy at 2 and 4 weeks, respectively.

Results

Viable uterine tissue and vascular patency were observed on transrectal ultrasonography and second-look laparotomy. Histological analysis of the graft tissue (performed in one ewe) revealed normal tissue architecture with a very subtle inflammatory reaction but no edema or stasis.

Conclusion

We have developed a modified procedure that allowed us to successfully perform orthotopic, allogeneic, uterine transplantation in sheep, whose uterine and vascular anatomy (apart from the bicornuate uterus) is similar to the human anatomy, making the ovine model excellent for human uterine transplant research.     

Effect of Antiprogesterone RU486 on VEGF Expression and Blood Vessel Remodeling on Ovarian Follicles before Ovulation

Background

The success of ovarian follicle growth and ovulation is strictly related to the development of an adequate blood vessel network required to sustain the proliferative and endocrine functions of the follicular cells. Even if the Vascular Endothelial Growth Factor (VEGF) drives angiogenesis before ovulation, the local role exerted by Progesterone (P₄) remains to be clarified, in particular when its concentration rapidly increases before ovulation.

Aim

This in vivo study was designed to clarify the effect promoted by a P₄ receptor antagonist, RU486, on VEGF expression and follicular angiogenesis before ovulation, in particular, during the transition from pre to periovulatory follicles induced by human Chorionic Gonadotropins (hCG) administration.

Material and Methods

Preovulatory follicle growth and ovulation were pharmacologically induced in prepubertal gilts by combining equine Chorionic Gonadotropins (eCG) and hCG used in the presence or absence of RU486. The effects on VEGF expression were analyzed using biochemical and immunohistochemical studies, either on granulosa or on theca layers of follicles isolated few hours before ovulation. This angiogenic factor was also correlated to follicular morphology and to blood vessels architecture.

Results and Conclusions

VEGF production, blood vessel network and follicle remodeling were impaired by RU486 treatment, even if the cause-effect correlation remains to be clarified. The P₄ antagonist strongly down-regulated theca VEGF expression, thus, preventing most of the angiogenic follicle response induced by hCG. RU486-treated follicles displayed a reduced vascular area, a lower rate of endothelial cell proliferation and a reduced recruitment of perivascular mural cells. These data provide important insights on the biological role of RU486 and, indirectly, on steroid hormones during periovulatory follicular phase. In addition, an in vivo model is proposed to evaluate how periovulatory follicular angiogenesis may affect the functionality of the corpus luteum (CL) and the success of pregnancy.     

Clinical Significance of Ischemia-Modified Albumin in the Diagnosis of Doxorubicin-Induced Myocardial Injury in Breast Cancer Patients

Background

Ischemia-modified albumin is an altered serum albumin that forms under conditions of oxidative stress, a state also associated with doxorubicin-induced myocardial injury.

Objective

The aim of this study was to better assess diagnostic and prognostic significance of ischemia-modified albumin in patients with breast cancer undergoing doxorubicin chemotherapy.

Methods

Blood samples were collected from 152 breast cancer patients before and after each cycle of doxorubicin chemotherapy to measure the serum levels of ischemia-modified albumin, cardiac troponin T and creatine kinase-MB. We also monitored cardiac function during a 12 month follow-up.

Results

There was a significant difference in ischemia-modified albumin levels before and after each cycle of chemotherapy and the ischemia-modified albumin concentration positively correlated with the cumulative dose of doxorubicin (r = 0.212, P < 0.05). The combination of ischemia-modified albumin with cardiac troponin T and creatine kinase-MB increased the sensitivity to 0.920 and the specificity to 0.830 in the diagnosis of doxorubicin-induced myocardial injury. The optimal cutoff for ischemia-modified albumin concentration was 112.09 U/ml. The rate of change for ischemia-modified albumin levels correlated negatively with the rate of change for left ventricular ejection fraction at one year (r = –0.221, P < 0.05).

Conclusion

Ischemia-modified albumin may be a clinically potential new marker for diagnosing doxorubicin-induced myocardial injury, and is helpful to predict long-term impairment of cardiac function.     

Bone Mesenchymal Stem Cells Contributed to the Neointimal Formation after Arterial Injury

Objectives

Recent findings suggest that in response to repair-to-injury bone marrow mesenchymal stem cells (BMSCs) participate in the process of angiogenesis. It is unclear what role BMSCs play in the structure of the vessel wall. In present study, we aimed to determine whether BMSCs had the capacity of endothelial cells (ECs).

Methods

BMSCs were separated and cultured. FACS and RT-PCR analysis confirmed the gene expression phenotype. The capacity of migration and adhesion and the ultrastructure of BMSCs were examined. The effect of BMSCs transplantation on the vascular repair was investigated in a murine carotid artery-injured model.

Results

BMSCs could express some markers and form the tube-like structure. The migration and adhesion capacity of BMSCs increased significantly after stimulated. In addition, BMSCs had the intact cell junction. In vivo the local transfer of BMSCs differentiated into neo-endothelial cells in the injury model for carotid artery and contributed to the vascular remodeling.

Conclusion

These results showed that BMSCs could contribute to neointimal formation for vascular lesion and might be associated with the differentiation into ECs, which indicated the important therapeutic implications for vascular diseases.     

Perivascular Delivery of Notch 1 siRNA Inhibits Injury-Induced Arterial Remodeling

Objectives

To determine the efficacy of perivascular delivery of Notch 1 siRNA in preventing injury-induced arterial remodeling.

Methods and Results

Carotid artery ligation was performed to induce arterial remodeling. After 14 days, morphometric analysis confirmed increased vSMC growth and subsequent media thickening and neointimal formation. Laser capture microdissection, quantitative qRT-PCR and immunoblot analysis of medial tissue revealed a significant increase in Notch1 receptor and notch target gene, Hrt 1 and 2 expression in the injured vessels. Perivascular delivery of Notch 1 siRNA by pluronic gel inhibited the injury-induced increase in Notch 1 receptor and target gene expression when compared to scrambled siRNA controls while concomitantly reducing media thickening and neointimal formation to pre-injury, sham-operated levels. Selective Notch 1 knockdown also reversed the injury-induced inhibition of pro-apoptotic Bax expression while decreasing injury-induced anti-apoptotic Bcl-X_L expression to sham-operated control levels. In parallel experiments, proliferative cyclin levels, as measured by PCNA expression, were reversed to sham-operated control levels following selective Notch 1 knockdown.

Conclusion

These results suggest that injury-induced arterial remodeling can be successfully inhibited by localized perivascular delivery of Notch 1 siRNA.     

Arterial Stiffness and Pulse Wave Reflection Are Increased in Patients Suffering from Severe Periodontitis

Aim

This single blind cross-sectional study compared the vascular health of subjects suffering from severe chronic periodontitis, severe aggressive periodontitis and periodontal healthy controls by evaluating pulse wave velocity (PWV), augmentation index (AIx) and pulse pressure amplification (PPA).

Material and Methods

In a total of 158 subjects, 92 suffering from severe periodontitis and 66 matched periodontal healthy controls, PWV, AIx, central and peripheral blood pressure were recorded using an oscillometric device (Arteriograph).

Results

Subjects suffering from severe chronic or aggressive periodontitis exhibited significantly higher PWV (p = 0.00004), higher AIx (p = 0.0049) and lower PPA (p = 0.028) than matched periodontal healthy controls.

Conclusions

The results of this study confirm the association between periodontal inflammation and increased cardiovascular risk shown by impaired vascular health in case of severe periodontitis. As impaired vascular health is a common finding in patients suffering from severe periodontal disease a concomitant routine cardiovascular evaluation may be advised.     

Relationships between HDL-C,hs-CRP,with Central Arterial Stiffness in Apparently Healthy People Undergoing a General Health Examination

Background

Some cardiovascular risk factors have been confirmed to be positively correlated with arterial stiffness. However, it is unclear whether HDL-C, a well-established anti-risk factor, has an independent association with arterial stiffness. The aim of this study was to evaluate the relationship between HDL-C levels and arterial stiffness and the possible role of high-sensitivity C-reactive protein (hs-CRP) in this potential correlation in apparently healthy adults undergoing a general health examination in China.

Materials and Methods

This was a cross-sectional survey. In total, 15,302 participants (age range, 18–82 years; mean, 43.88±8.44 years) were recruited during routine health status examinations. A questionnaire was used and we measured the body mass index, systolic and diastolic blood pressure, and fasting glucose, and serum lipid, uric acid, hs-CRP, and serum creatinine levels of each participant. Central arterial stiffness was assessed by carotid–femoral pulse wave velocity (cf-PWV).

Results

HDL-C levels decreased as cf-PWV increased. Pearson’s correlation analysis revealed that HDL-C levels were associated with cf-PWV (r=−0.18, P<0.001). hs-CRP levels were positively associated with cf-PWV (r=0.13). After adjustment for all confounders, HDL-C was inversely independently associated with all quartiles of cf-PWV. Furthermore, HDL-C was associated with cf-PWV in different quartiles of hs-CRP, and the correlation coefficients (r) gradually decreased with increasing hs-CRP levels (quartiles 1–4).

Conclusions

HDL-C is inversely independently associated with central arterial stiffness. The anti-inflammatory activity of HDL-C may mediate its relationship with cf-PWV. Further, long-term follow-up studies are needed to evaluate whether high HDL-C levels are protective against central artery stiffening through the anti-inflammatory activity of HDL-C.     

The Blood Neutrophil to Lymphocyte Ratio Correlates with Clinical Status in Children with Cystic Fibrosis: A Retrospective Study

Purpose

The blood neutrophil to lymphocyte ratio (NLR) has been identified as a potentially useful marker of clinical outcome in disease states with an inflammatory component. The objective of this study was to evaluate the relationship between NLR and clinical status in children with cystic fibrosis.

Methods

This was a retrospective chart review. Data collected included NLR, body mass index, and forced expiratory volume in 1 second (FEV1) while asymptomatic, and during hospitalizations for pulmonary exacerbation. An NLR breakpoint of 3 was used for comparisons of body mass index and FEV1.

Results

A total of 159 charts were reviewed. An NLR ≥ 3 was significantly associated with lower body mass index and lower FEV1. NLR during hospitalization was significantly higher than NLR while asymptomatic. NLR measured during the first 3 months of life was negatively correlated with FEV1 at age 12.

Conclusion

NLR correlates with clinical status in children with cystic fibrosis and may be a useful biomarker in this population.     

Behavioral and Cardiorespiratory Responses to Bilateral Microinjections of Oxytocin into the Central Nucleus of Amygdala of Wistar Rats,an Experimental Model of Compulsion

Introduction

The central nucleus of amygdala plays an important role mediating fear and anxiety responses. It is known that oxytocin microinjections into the central nucleus of amygdala induce hypergrooming, an experimental model of compulsive behavior. We evaluated the behavioral and cardiorespiratory responses of conscious rats microinjected with oxytocin into the central nucleus of amygdala.

Methods

Male Wistar rats were implanted with guide cannulae into the central nucleus of amygdala and microinjected with oxytocin (0.5 µg, 1 µg) or saline. After 24 h, rats had a catheter implanted into the femoral artery for pulsatile arterial pressure measurement. The pulsatile arterial pressure was recorded at baseline conditions and data used for cardiovascular variability and baroreflex sensitivity analysis. Respiratory and behavioral parameters were assessed during this data collection session.

Results

Microinjections of oxytocin (0.5 µg) into the central nucleus of amygdala produced hypergrooming behavior but did not change cardiorespiratory parameters. However, hypergrooming evoked by microinjections of oxytocin (1 µg) into the central nucleus of amygdala was accompanied by increase in arterial pressure, heart rate and ventilation and augmented the power of low and high (respiratory-related) frequency bands of the systolic arterial pressure spectrum. No changes were observed in power of the low and high frequency bands of the pulse interval spectrum. Baroreflex sensitivity was found lower after oxytocin microinjections, demonstrating that the oxytocin-induced pressor response may involve an inhibition of baroreflex pathways and a consequent facilitation of sympathetic outflow to the cardiovascular system.

Conclusions

The microinjection of oxytocin (1 µg) into the central nucleus of amygdala not only induces hypergrooming but also changes cardiorespiratory parameters. Moreover, specific oxytocin receptor antagonism attenuated hypergrooming but did not affect pressor, tachycardic and ventilatory responses to oxytocin, suggesting the involvement of distinct neural pathways.     

High resolution imaging of vascular function in zebrafish

Rationale

The role of the endothelium in the pathogenesis of cardiovascular disease is an emerging field of study, necessitating the development of appropriate model systems and methodologies to investigate the multifaceted nature of endothelial dysfunction including disturbed barrier function and impaired vascular reactivity.

Objective

We aimed to develop and test an optimized high-speed imaging platform to obtain quantitative real-time measures of blood flow, vessel diameter and endothelial barrier function in order to assess vascular function in live vertebrate models.

Methods and Results

We used a combination of cutting-edge optical imaging techniques, including high-speed, camera-based imaging (up to 1000 frames/second), and 3D confocal methods to collect real time metrics of vascular performance and assess the dynamic response to the thromboxane A₂ (TXA₂) analogue, U-46619 (1 µM), in transgenic zebrafish larvae. Data obtained in 3 and 5 day post-fertilization larvae show that these methods are capable of imaging blood flow in a large (1 mm) segment of the vessel of interest over many cardiac cycles, with sufficient speed and sensitivity such that the trajectories of individual erythrocytes can be resolved in real time. Further, we are able to map changes in the three dimensional sizes of vessels and assess barrier function by visualizing the continuity of the endothelial layer combined with measurements of extravasation of fluorescent microspheres.

Conclusions

We propose that this system-based microscopic approach can be used to combine measures of physiologic function with molecular behavior in zebrafish models of human vascular disease.     

Angiopoietin-Like 7 Is an Anti-Angiogenic Protein Required to Prevent Vascularization of the Cornea

Purpose

We sought to identify the anti-angiogenic molecule expressed in corneal keratocytes that is responsible for maintaining the avascularity of the cornea.

Methods

Human umbilical vein endothelial cells (HUVECs) were cultured with either human dermal fibroblasts or with human corneal keratocytes under serum-free conditions. The areas that exhibited blood vessel formation were estimated by immunostaining the cultures with an antitibody against CD31, a blood vessel marker. We also performed microarray gene-expression analysis and selected one molecule, angiopoietin-like 7 (ANGPTL7) for further functional studies conducted with the keratocytes and in vivo in mice.

Results

Areas showing blood vessel formation in normal serum-free medium were conditions were markedly smaller when HUVECs were co-cultured with corneal keratocytes than when they were co-cultured with the dermal fibroblasts under the same conditions. Microarray analysis revealed that ANGPTL7 expression was higher in keratocytes than in dermal fibroblasts. In vitro, inhibiting ANGPTL7 expression by using a specific siRNA led to greater tube formation than did the transfection of cells with a control siRNA, and this increase in tube formation was abolished when recombinant ANGPTL7 protein was added to the cultures. In vivo, intrastromal injections of an ANGPTL7 PshRNA into the avascular corneal stroma of mice resulted in the growth of blood vessels.

Conclusions

ANGPTL7, which is abundantly expressed in keratocytes, plays a major role in maintaining corneal avascularity and transparency.