In vivo experimental intracranial aneurysm models: a systematic review.

Animal models are necessary to develop and test innovations in aneurysm therapy before clinical introduction. This review aims at identifying the most likely candidates for standardizing preclinical testing of aneurysm devices. We systematically searched electronic databases for publications on animal aneurysm models from 1961-2008 to assess the methodologic quality of the studies and collect data on the patency and angiographic and pathologic outcomes of treatments. There has been a steady increase in the annual number of publications with time. Species that were most frequently used were dogs, rabbits, and rodents, followed by swine. Most publications are single-laboratory studies with variables and poorly validated outcome measures, a small number of subjects, and limited standardization of techniques. The most appropriate models to test for recurrences after endovascular occlusion were the surgical bifurcation model in dogs, and the elastase-induced aneurysm model in rabbits. A standardized multicenter study is needed to improve the preclinical evaluation of endovascular devices in aneurysm therapy.

Mechanisms of occlusion and recanalization in canine carotid bifurcation aneurysms embolized with platinum coils: an alternative concept.

BACKGROUND AND PURPOSE: Endovascular treatment of aneurysms may result in complete or incomplete occlusions or may be followed by recurrences. The goal of the present study was to better define pathologic features associated with so-called healing or recurrences after coiling and to propose an alternative concept to the currently accepted view. MATERIALS AND METHODS: Experimental canine venous pouch aneurysms were created by using a T-type (group A, N = 29) or a Y-type constructed bifurcation (group B, N = 37) between the carotid arteries. Coil embolization was performed 2 weeks later; and angiography, immediately after and at 12 weeks. Angiographic results, neointima formation at the neck, endothelialization, and organization of thrombus were compared between groups by using qualitative scores and immunohistochemistry. RESULTS: Angiographic results at 3 months were significantly better in group A than in group B (P = .001). Macroscopic neointimal scores were also better (P = .012). Only 10/32 aneurysms with satisfactory results at angiography were completely sealed by neointima formation. Animals with residual or recurrent aneurysms had significantly worse neointimal scores than those with completely occluded ones (P = .0003). On histologic sections, the neointima was constantly present in "healed" and in recurrent aneurysms. This neointima was a multicellular layer of alpha-actin+ cells in a collagenous matrix, covered with a single layer of nitric oxide synthetase (NOS+) endothelial cells, whether it completely occluded the neck of the aneurysm or dived into the recurring or residual space between the aneurysm wall and the coil mass embedded in organizing thrombus. CONCLUSION: Complete angiographic occlusions at 3 months can be associated with incomplete neointimal closure of the neck at pathology. Thrombus organization, endothelialization, and neointima formation can occur concurrently with recurrences.

Effects of stenting the parent artery on aneurysm filling and gene expression of various potential factors involved in healing of experimental aneurysms.

SUMMARY: Intracranial stents are increasingly used in the endovascular treatment of aneurysms, but very little is known regarding their effect on the cellular and molecular evolution of aneurysms. Bilateral venous pouch lateral wall carotid aneurysms were created in 20 dogs. All dogs then underwent angiography and balloon-expandable stenting of one aneurysm four to six weeks later. Fifteen dogs underwent aneurysm harvesting at one day (n=3), four days (n=4), seven days (n=3), and 14 days (n=5) for mRNA expression analysis, using axial sections taken from the aneurysm neck and fundus for RTPCR amplification of four cytokines or growth factors: TNF-a, TGF-b1, MCP-1, and PDGFBB; two adhesion molecules: VCAM-1 and PECAM-1; five matrix modifying agents; MMP- 2, 9, TIMPs 1, 3, 4, and two cellular markers: CD34 and a-SMA. Five other dogs, sacrificed at 12 weeks, were examined for extent of filling of the aneurysm neck with organized tissue and for neointima formation at the aneurysm ostium. Angiography was performed prior to sacrifice in all animals, and compared with initial studies. Eleven out of 20 stented aneurysms showed a favorable angiographic evolution, while none of the 20 nonstented aneurysms improved (p=0.001). Pathology showed partially occluded aneurysms, with neointima formation around the stent struts.Observed trends in mRNA expression, that stenting increased expression of genes involved in organization and neointima formation, agreed with experimental hypotheses, but differences between stented and non-stented aneurysms did not reach statistical significance. Parent vessel stenting was associated with angiographic improvement of aneurysm appearance. Modifications in mRNA expression patterns following stenting deserve further study to better establish potential molecular targets to promote aneurysm healing.

Alpha8beta1 integrin is upregulated in myofibroblasts of fibrotic and scarring myocardium.

Integrins mediate cell attachment to the extracellular matrix (ECM) regulating migration, proliferation, and differentiation. We previously reported the presence of alpha8beta1 integrin on cultured cardiac fibroblasts. Extending this information, we localized alpha8beta1 integrin in normal rat myocardial tissue, and investigated its expression pattern in rats chronically infused with angiotensin II (Ang II, 500 ng/kg/min), a well-recognized profibrotic factor. Alpha8beta1-integrin expression was analyzed by binding assay, western blotting, and immunohistochemistry. In normal myocardium, immunohistochemical staining for alpha8 was found in fibroblasts, as well as in the epicardium, endocardium, and valves. Vascular smooth muscle cells (VSMCs) of the media of cardiac arteries also stained positively. After 14-d-Ang II infusion, staining for fibronectin, as well as collagen staining by Sirius red, revealed extensive interstitial and perivascular fibrosis. Increased expression of alpha8 integrin in ventricular smooth muscle (SM) alpha-actin-positive fibroblasts (myofibroblasts) was also recorded. The upregulation of alpha8beta1 integrin was confirmed by binding assay and by western blotting. Microscopic scars, a characteristic of reparative fibrosis, were invaded by matrix proteins and by strongly alpha8- and SM alpha-actin-positive myofibroblasts. The results indicate that, in rat adult myocardium, alpha8beta1 integrin is expressed in fibroblasts and VSMC. In Ang II-infused animals, alpha8beta1-integrin expression was enhanced in the left ventricle and arteries. The coordinate regulation of alpha8beta1 integrin on fibroblasts and ECM proteins raises the possibility that this integrin is implicated in the deposition of matrix components leading to fibrosis.

Upregulation of alpha(8)beta(1)-integrin in cardiac fibroblast by angiotensin II and transforming growth factor-beta1.

Using a novel pharmacological tool with (125)I-echistatin to detect integrins on the cell, we have observed that cardiac fibroblasts harbor five different RGD-binding integrins: alpha(8)beta(1), alpha(3)beta(1), alpha(5)beta(1), alpha(v)beta(1), and alpha(v)beta(3). Stimulation of cardiac fibroblasts by angiotensin II (ANG II) or transforming growth factor-beta1 (TGF-beta1) resulted in an increase of protein and heightening by 50% of the receptor density of alpha(8)beta(1)-integrin. The effect of ANG II was blocked by an AT(1), but not an AT(2), receptor antagonist, or by an anti-TGF-beta1 antibody. ANG II and TGF-beta1 increased fibronectin secretion, smooth muscle alpha-actin synthesis, and formation of actin stress fibers and enhanced attachment of fibroblasts to a fibronectin matrix. The alpha(8)- and beta(1)-subunits were colocalized by immunocytochemistry with vinculin or beta(3)-integrin at focal adhesion sites. These results indicate that alpha(8)beta(1)-integrin is an abundant integrin on rat cardiac fibroblasts. Its positive modulation by ANG II and TGF-beta1 in a myofibroblast-like phenotype suggests the involvement of alpha(8)beta(1)-integrin in extracellular matrix protein deposition and cardiac fibroblast adhesion.

Biochemical characteristics of beta-adrenoceptors in rats after an 18-day spaceflight (LMS-STS78).

BACKGROUND: To ascertain whether there was autonomic adaptation with the development of adrenoceptor hypersensitivity under microgravity, the biochemical properties of the beta-adrenoceptors were determined using (125I)iodocyanopindolol (ICYP) binding in rats flown for 18 d onboard the space shuttle. METHODS: This study was performed on heart and kidneys of 3 groups of 12 animals: the flight and 2 ground control (vivarium and AEM) groups. To distinguish the possible role of the corticosteroids, half of each animal group was bilaterally adrenalectomized (ADX rats) with an aldosterone and corticosterone supplementation while the other half was SHAM operated. RESULTS: The Scatchard analysis of the ICYP-binding in both organs revealed no significant alterations in the dissociation constant (Kd) and in the maximal binding capacity (Bmax) between SHAM flight and control groups. The Kd of the beta-adrenoceptors in the cardiac atria of the SHAM flight rats (74 +/- 5 pm) was significantly higher (p < 0.05) than in those of the ADX flight rats (60 +/- 3 pm) while the Bmax was nonsignificantly higher (1925 +/- 370 in SHAM flight rats vs. 1482 +/- 283 fmol x mg(-1) protein in ADX flight rats). No significant change was determined for the Bmax and Kd values in the kidneys of the ADX and SHAM flight rats. CONCLUSIONS: This work performed on animals did not show any obvious effect of microgravity on the beta-adrenergic function in the heart and kidneys. Inflight rodent sacrifice protocols should definitely ensure assessment of the influence of microgravity on the animals.

Effect of a 14-day hindlimb suspension on beta-adrenoreceptors in rats.

Exposure to long-term simulated microgravity exhibits reduced sympathetic nervous system activity. This study tested the hypothesis that the hypersensitivity of adrenoreceptors would explain partly many other features of the hemodynamic consequences of return from space. The biochemical properties of the beta adrenoreceptors (betaAR) were determined using 125I-cyanopindolol (125I-CYP) binding in three rat groups: (1) The first experimental group consisted of 24 h-restrained orthostatic rats in the horizontal position, to test the early effect of the attachment to the suspension device; (2) the second experimental group consisted of 24 h-restrained antiorthostatic rats, to test the early effect of the suspension; (3) the third experimental group consisted of 14 day-restrained antiorthostatic rats, to test the long term effect of the suspension. The study was performed in two organs involved in blood pressure regulation, i.e. the heart (atria and ventricles were separated) and kidneys. The Scatchard analysis of 125I-cyanopindolol binding in both organs indicated no significant alterations in the dissociation constant (Kd) and the maximum binding capacity (Bmax) in the three experimental groups. These results do not allow the conclusion about the SNS adaptation pattern to simulated microgravity. Thus, the hypothesis that betaAR are involved in the cardiovascular adaptation to simulated microgravity is not verified in this model where, as a matter of fact, cardiovascular deconditioning is not verified even if this model is widely used.

Restraint vs. hindlimb suspension on fluid and electrolyte balance in rats.

To determine the effect of hindlimb suspension on body fluid volume, salt and water balance, and relevant hormones, two series of experiments were performed in an experimental protocol including periods of isolation (7 days), horizontal attachment (7 days), and suspension (14 days). 1) During the first experiment, water and electrolyte balance, arginine vasopressin (AVP), and guanosine 3',5'- cyclic monophosphate (cGMP) were determined in urine, atrial natriuretic peptide in plasma and atria, and renin concentration and AVP in plasma in 30 rats. 2) During the second experiment, blood volume and extracellular fluid volume were measured by a dilution technique (Evans blue and sodium thiocyanate) in another 30 rats. We observed a pronounced and early effect of horizontal attachment on the renal variables. After 48 h, diuresis (49%), natriuresis (44%), kaliuresis (36%), osmotic load (39%), creatinine (28%), and AVP excretion (155%) were significantly increased in attached rats (P < 0.05). There was no short-term (24-h) effect of suspension on urine flow and Na+, K+, creatinine, and AVP excretion, but the urine cGMP decreased significantly (45%; P < 0.05). Significant decreases in natriuresis, kaliuresis, urine creatinine, and osmotic load occurred in the suspension group 7 days after suspension. After the 14-day tail suspension, plasma volume and extracellular fluid volume measured in suspended rats were not different from isolated rat values, whereas plasma volume increased by 15% (P < 0.05) in the attached rats. Plasma immunoreactive plasma atrial natriuretic levels of suspended rats were significantly reduced by 35% vs. isolated rats (P < 0.001) and by 18% vs. attached rats (P < 0.05). By using this experimental protocol, the physiological alterations revealed that suspension produced some acute and long-term effects, but the fixation to the suspension device, restraint, and confinement have their own influence on fluid distribution and renal function.

Central and peripheral sympathetic activities in rats during recovery from simulated weightlessness.

Rats were tail suspended, keeping their forelimbs weight bearing for 14 days, and then allowed to recover for a short (6-h) or a long (24-h) period to assess the behavior of the sympathetic nervous system after weightless simulation. Sympathetic activity was determined by measuring norepinephrine (NE) turnover in the brain stem cell groups involved in central blood pressure control and in organs playing a key role in the cardiovascular regulation (heart and kidneys). The NE turnover was greatly reduced in the rostral (-56%; P < 0.001) and caudal (-73%; P < 0.001) A2 nucleus of suspended rats but was unchanged in the A1, A5, and A6 cell groups compared with attached rats. The NE turnover in the cardiac atria (-34%; P < 0.001) and ventricles (-35%; P < 0.001) and kidneys (-31%; P < 0.001) was decreased after suspension. The central and peripheral sympathetic activities returned to normal within 24 h of release from suspension, but there was hyperactivity after 6 h of recovery. This raises the problem of interpreting the results obtained in animals killed a few hours after return from spaceflight.

A day/night rhythm of vasopressin and oxytocin in rat retina, pineal and harderian gland.

Arginine vasopressin (AVP), oxytocin (OT) and neurophysins (Np) have been found in the pineal gland and the retina of the rat. Because the retina, pineal gland and Harderian gland (HG) serve analogous functions, we undertook a study to determine the presence of these peptides in these three organs of rats. They were detected by two specific methods: HPLC and specific radioimmunoassays. For Np, total neurophysins (NpT) were measured. To determine a 24 hr rhythm, the animals were maintained under a light/dark cycle of 12 hr/12 hr for 3 weeks. The pineal glands, retinae and HG were collected. Day/night rhythms of AVP, OT and NpT were demonstrated in the retina and HG; but the pineal gland had only AVP rhythm. A significant decrease in the rhythms at 4 a.m. was demonstrated in the retina and HG. The 24 hr variation of AVP in the retina seemed parallel to that of the HG.

Effects of propranolol and swim-training on blood pressure, plasma electrolytes, and vasopressin in spontaneously hypertensive and normotensive rats.

The aim of this work was to study the influence of beta-adrenoreceptor blockade on the adaptation to exercise of one of the hormonal systems (arginine vasopressin) involved in the regulation of blood volume and pressure in spontaneously hypertensive rats (SHR). Systolic blood pressure (SBP) was measured in SHR and WKY rats during 11 wk of swim training. At the end of the training program we determined post-exercise values of plasma arginine-vasopressin (pAVP), osmolality (pOsm), K+ (pK+), Na+ (pNa+), hemoglobin (Hgb), and hematocrit (Hct) in SHR and WKY rats. The following groups were studied: control (C), propranolol treated (PC), swim trained (S), and propranolol-treated and swim-treated (PS). SBP was significantly reduced by swim training or propranolol, bu these beneficial effects on SBP were attenuated when propranolol and swim training were combined. pNa+ and pOsm were significantly reduced by training alone in SHR. This reduction of pNa+ and, consequently, of pOsmol without any modification of other parameters could suggest an Na+ loss. In contrast, the SHR group treated with propranolol alone showed a significant reduction in Hct, suggesting an increased plasma volume without Na+ loss. PS SHR showed a significant reduction of Hgb, Hct, proteins, pNa+, and pOsmol, probably as a consequence of the additive effects of swimming- and propranolol-induced hypervolemia with Na+ loss. The slight and nonsignificant reduction in pAVP observed with either training or propranolol treatment alone became much more pronounced and statistically significant when the 2 treatments were combined. WKY rats showed a much smaller response to exercise and beta-adrenoreceptor blockade than SHR. We conclude that the hypervolemia suggested in PS SHR could be a possible cause of attenuation of the beneficial effects of either swimming or propranolol on SBP.