Hongshan Zhong

Performance of a modified rabbit model of abdominal aortic aneurysm induced by topical application of porcine elastase: 5-month follow-up study.

OBJECTIVES To modify the method for creating an abdominal aortic aneurysm in rabbits, and to study its performance. MATERIALS AND METHODS A total of 24 New Zealand white rabbits were induced topically with 10 μl of porcine elastase (0, 0.1, 5 and 10 units μl(-1)) to define the optimal concentration (groups A-D). Twelve aneurysms were induced with 10 units μl(-1) of 10 μl elastase to serve as a follow-up group (group E) to serve as a follow-up. A 1.5-cm aortic segment was isolated and induced with elastase solution for 30 min. RESULTS All animals in groups D and E developed AAA by day 5. Aneurysms in Group E were stable over 100 days. Partial destruction to disappearance of elastic lamellae and smooth muscle cells (SMCs) was seen in elastase-treated animals by day 5. Regenerated elastin and proliferated SMCs were present in group E. Matrix metalloproteinases 2 and 9 and RAM11 showed strong expression in group D, but expression decreased in group E after day 15. CONCLUSIONS The rabbit AAA model induced via topical application of porcine elastase at 10 units μl(-1) for 30 min appears easy and simple, with shorter induction and more rapid aortic dilation. The model is stable over 100 days and is useful to study the formation and progress of AAAs.

A Novel In Vivo Rabbit Model of Abdominal Aortic Aneurysm Induced by Periarterial Incubation of Papain

PURPOSE The objective of this study was to determine the possibility of creating a novel animal model of abdominal aortic aneurysm (AAA) in rabbits by the periarterial application of papain. MATERIALS AND METHODS Twelve New Zealand white rabbits were randomized into two groups: (1) the papain group, which received 2mg of papain (n = 8) and (2) the control group, which received physiologic saline solution (n = 4). A 1-cm aortic segment proximal to the bifurcation was isolated, and its adventitia was incubated with papain for 20 minutes. The rabbits underwent intravenous digital subtraction angiography (IVDSA) 5 and 21 days after the operation. The animals were then humanely killed for histomorphometric and immunohistochemical studies. RESULTS All animals in the papain group developed AAA, with an average aneurysm diameter of 4.0 ± 0.6 and 4.1 ± 0.4 mm on days 5 and 21, respectively. No aneurysms were seen in the control group. On day 5, the papain-incubated aortas exhibited thinned and disorganized aortic walls, with decreased smooth muscle cells (SMCs) and fragmented and almost nonexistent elastic lamella. Media thickening, intimal hyperplasia, and smooth muscle cell regeneration were obvious on day 21. Immunostaining of matrix metalloproteinase (MMP)-9 and RAM11 showed strong expression in the papain group. On the contrary, the control group did not present histologic alterations and showed almost no expression of MMP-9 and RAM11. CONCLUSIONS A novel in vivo rabbit model of AAA can be induced through periarterial application of papain for 20 minutes. This model is similar to an elastase-induced aneurysm model and could be useful to clarify AAA pathogenesis and endovascular treatment intervention.

Low intensity ultrasound promotes the sensitivity of rat brain glioma to Doxorubicin by down-regulating the expressions of p-glucoprotein and multidrug resistance protein 1 in vitro and in vivo.

The overall prognosis for malignant glioma is extremely poor, and treatment options are limited in part because of multidrug resistant proteins. Our previous findings suggest low intensity ultrasound (LIUS) can induce apoptosis of glioma cells. Given this finding, we were interested in determining if LIUS could help treat glioma by inhibiting multidrug resistant proteins, and if so, which pathways are involved. In this study, the toxicity sensitivity and multidrug resistance proteins of glioma induced by LIUS were investigated using CCK-8, immunohistochemistry, immunofluorency, and RT-PCR in tissue samples and cultured cells. LIUS inhibited increase of C6 cells in an intensity- and time-dependent manner. The toxicity sensitivity of C6 cells increased significantly after LIUS sonication (intensity of 142.0 mW/cm2) or Doxorubicin (DOX) at different concentration, particularly by the combination of LIUS sonication and DOX. The expressions of P-gp and MRP1 decreased significantly post-sonication at intensity of 142.0 mW/cm2 both in vitro and in vivo. The expressions of p110 delta (PI3K), NF-κB-p65, Akt/PKB, and p-Akt/PKB were downregulated by LIUS sonication and DOX treatment separately or in combination at the same parameters in rat glioma. These results indicate that LIUS could increase the toxicity sensitivity of glioma by down-regulating the expressions of P-gp and MRP1, which might be mediated by the PI3K/Akt/NF-κB pathway.

Low frequency and intensity ultrasound induces apoptosis of brain glioma in rats mediated by caspase-3, Bcl-2, and survivin

Low frequency and intensity ultrasound (LFU) sonication can selectively induce brain tumor cell apoptosis without damaging neural cells, while also enhancing drug delivery to brain tumors. To explore the underlying mechanisms of related pathways in LFU-induced apoptosis, we investigated the expression of proteins associated with LFU-induced apoptosis. C6 cells were used for in vitro experiments and C6 tumor-bearing rats were used during in vivo experiments. 3-[4.5-Dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide; thiazolyl blue (MTT) assay was used to detect C6 cell viability in vitro. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) analysis was used to check the apoptotic cells, and they were counted and analyzed both in vitro and in vivo. Transmission electron microscopy (TEM) was used to illustrate the ultrastructure of apoptotic nuclei of cancer cells in vivo. The expressions of caspase-3, Bcl-2, and survivin proteins were assessed by immunofluorescence, immunohistochemistry and Western blot analysis in vivo. C6 cell viability decrease was statistically significant; the numbers of apoptotic C6 cells in the LFU sonication groups were higher than those in the control group both in vitro and in vivo. The expression of caspase-3 increased, yet the expressions of Bcl-2 and survivin decreased significantly 6h after LFU sonication, compared with the control group in vivo. This study suggests that LFU can induce apoptosis in vitro and in vivo, and that three signaling proteins, caspase-3, Bcl-2, and survivin, might be involved in LFU-induced apoptosis.

Different long-term outcomes of abdominal aortic aneurysm and intracranial aneurysm models: hemodynamic change may also play an essential role in the initiation and progression of abdominal aortic aneurysm in rabbits.

Self-healing phenomenon was found in the periarterial elastase-induced abdominal aortic aneurysm (AAA) in rabbit. This kind of aneurysm model does not progress and heals spontaneously in the long term, which is quite different from the performance of AAA disease in human. In order to better mimic human AAA and overcome this shortcoming of traditional AAA model in rabbit, we studied the pathogenesis of cerebral aneurysm (CA) model in small animal, which shows an excellent long-term patency and progressive enlargement. We found that hemodynamic conditions, such as turbulence flow, high blood flow, and shear stress, play an important role in the formation and progression of CA. So, we hypothesize that hemodynamic change may also play an essential role in the initiation and progression of rabbit AAA, and self-healing will be overcome if hemodynamic condition changes by coarctation of infra-renal aorta after elastase incubation.

(2-Hydroxypropyl)-β-Cyclodextrin Is a New Angiogenic Molecule for Therapeutic Angiogenesis

Background Peripheral artery disease (PAD), which is caused by atherosclerosis, results in progressive narrowing and occlusion of the peripheral arteries and inhibits blood flow to the lower extremities. Therapeutic angiogenesis is a promising strategy for treating ischemia caused by PAD. Nitric oxide (NO) has been shown to be a key mediator of angiogenesis. It has been demonstrated that β-cyclodextrincan stimulate vessel growth in rabbit corneas. In this study, we assessed the mechanism of action and therapeutic potential of a new angiogenic molecule, (2-hydroxypropyl)-β-cyclodextrin (2HP-β-CD). Methods and Results 2HP-β-CD significantly increased vascular endothelial growth factor A (VEGF-A) and platelet-derived growth factor BB (PDGF-BB) peptides in human umbilical vein endothelial cells (HUVECs) and also increased basic fibroblast growth factor (bFGF) peptide in human aortic smooth muscle cells (HASMCs). 2HP-β-CD stimulated both proliferation and migration of HUVECs in an endothelial nitric oxide synthase (eNOS)/NO-dependent manner, whereas NO was found to be involved in proliferation, but not migration, of HASMCs. In a unilateral hindlimb ischemia model in mice, 2HP-β-CD injections not only promoted blood flow recovery and increased microvessel densities in ischemic muscle, but also promoted coverage of the vessels with smooth muscle cells, thus stabilizing the vessels. Administration of 2HP-β-CD increased the expression of several angiogenic factors, including VEGF-A, PDGF-BB and transforming growth factor beta-1 (TGF-β1) in ischemic muscle. Injections of 2HP-β-CD also stimulated protein kinase B and extracellular regulated protein kinases (ERK), leading to an increase in phosphorylation of eNOS in ischemic muscle. Treatment with the NOS inhibitor, Nω-nitro-L-arginine methyl ester (L-NAME), showed that stimulation of blood flow induced by 2HP-β-CD was partially dependent on NO. Conclusions Therapeutic angiogenesis by 2HP-β-CD may be beneficial to patients with PAD.

A New Unibody Branched Stent-graft for Reconstruction of the Canine Aortic Arch

OBJECTIVES To evaluate the feasibility and safety of a new unibody branched stent-graft for reconstruction of the canine aortic arch. MATERIALS AND METHODS Twenty adult hybrid dogs were used for the experiments. Ten dogs were implanted with single-branched stent-grafts; the other ten dogs were implanted with double-branched stent-grafts. The stent-grafts were implanted transluminally via the abdominal aorta. The branched limbs were caught and pulled into supra-aortic vessels using gooseneck snare wires introduced via the axillary arteries. The animals were euthanized 4 months after implantation. RESULTS One of the ten dogs implanted with a single-branched stent-graft died from failure of the implantation procedure, and two of the ten dogs implanted with double-branched stent-grafts died from failure of the procedure and excessive blood loss. After month 4, the remaining unibody branched stent-grafts were patent and did not migrate. CONCLUSIONS This new unibody branched stent-graft could be used to reconstruct the aortic arch. This is a total endovascular technique, and compared to other branched stent-grafts appears to be safer and easier to implant.

Transauricular intra-arterial and intravenous digital subtraction angiography for abdominal aortic aneurysm imaging in a rabbit model

Aim The aim of this study was to evaluate transauricular digital subtraction angiography (DSA) as an alternative to conventional intra-arterial DSA for rabbit abdominal aortic aneurysm (AAA). Materials and methods AAA models were created in 8 New Zealand white rabbits by sewing vein patch. The diameters of aortic arteries were measured by DSA via ear vein and ear central artery. The common carotid artery (CCA) was exposed and cannulated for DSA as conventional angiography. Diameter size was measured and compared. Results Aortic diameters, tested by DSA via ear vein, ear central artery and CCA were 7.9 ± 1.2 mm, 7.8 ± 1.0 mm and 7.9 ± 1.1 mm respectively, with no significant differences. Angiography via CCA as standard procedure, correlation in aneurysm neck diameter was r = 0.93 for IVDSA and r = 0.96 for angiography via central artery (P < 0.01); Correlation in AAA diameter was r = 0.99 for IVDSA and r = 0.99 for angiography via central artery (P < 0.0001). Conclusions Transauricular DSA shows good correlation to conventional DSA, can be used repeatedly with less invasiveness, and suitable for rabbit AAA follow-up study.

Combination of Periaortic Elastase Incubation and Cholesterol-Rich Diet: A Novel Model of Atherosclerosis in Rabbit Abdominal Aorta

Atherosclerosis, the leading cause of most cardiovascular disease, is a progressive multifaceted inflammatory disease characterized by extracellular matrix degradation and extensive remodeling of artery wall. However, its mechanism has not been completely understood, and animal models are useful to study its pathogenetic process. An analysis of literature on the nature of atherosclerosis indicates that focal accumulation of smooth muscle cells (SMCs) into the intima by plasma factors is fundamental to the entire process of plaque growth. In our previous study, vascular SMCs proliferation was obvious in elastase-induced aorta by day 15, which led to intimal hyperplasia and regression of rabbit aneurysm. Model induced by combination of balloon injury and an atherogenic diet in rabbits is the conventional, but most largely used experimental model of atherosclerosis. Since proliferation and accumulation of intimal SMCs are found in elastase-induced aorta, and hypercholesterolemia is usually induced by cholesterol-rich diets in rabbits, a novel atherosclerosis model may be induced by combination periaortic elastase incubation and cholesterol-rich diet.

A Feasibility Study of a New Unibody Branched Stent Graft Applied to Reconstruct the Canine Aortic Arch

OBJECTIVES The aim was to evaluate the feasibility and safety of a new unibody branched stent graft for the reconstruction of the canine aortic arch. METHODS The unibody branched stent grafts included single branched stent grafts and double branched stent grafts. The main stent graft and branched limbs were sutured together. The branched stent grafts were folded into the introducer system, which consisted of a double channel catheter, a detachable sleeve, and an introducer sheath. The branched stent grafts were introduced and deployed into the aortic arch by the delivery system. Twenty adult mongrel dogs were used for the experiments. Ten dogs were implanted with single branched stent grafts; the other 10 were implanted with double branched stent grafts. The surviving animals were followed up for 3 months. Computed tomography angiography (CTA) was performed to observe the status of the branched stent grafts. RESULTS All the unibody branched stent grafts were successfully implanted into the canine aortic arches. The technical success rate was 100%. There was no cerebral infarction, paraplegia or incision infection. CTA showed that all the branched stent grafts were patent; there was no endoleak or stent migration. CONCLUSIONS The unibody branched stent graft system could be used to reconstruct the aortic arch. The animal experimental procedures demonstrated the safety and feasibility of the unibody branched stent graft system.