Experimental Rat and Mouse Carotid Artery Surgery: Injury and Remodeling Studies

In cardiovascular research, translation of benchtop findings to the whole body environment is often critical in order to gain a more thorough and comprehensive clinical evaluation of the data with direct extrapolation to the human condition. In particular, developmental and/or pathophysiologic vascular growth studies often employ in vitro approaches such as cultured cells or tissue explant models in order to analyze specific cellular, molecular, genetic, and/or biochemical signaling factors under pristine controlled conditions. However, validation of in vitro data in a whole body setting complete with neural, endocrine, and other systemic contributions provides an essential proof of concept from a clinical perspective. Several well-characterized experimental in vivo models exist that provide excellent proof-of-concept tools to examine vascular growth and remodeling in the whole body. This paper will examine the rat carotid artery balloon injury model, the mouse carotid artery wire denudation injury model, and rat and mouse carotid artery ligation models with particular emphasis on minimally invasive surgical access to the site of intervention. Discussion will include key scientific and technical details as well as caveats, limitations, and considerations for the practical use of each of these valuable experimental models.

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The Potential of sGC Modulators for the Treatment of Age-Related Fibrosis: A Mini-Review

Gerontology ◽

10.1159/000450946 ◽

2016 ◽

Vol 63 (3) ◽

pp. 216-227 ◽

Cited By ~ 13

Author(s):

Peter Sandner ◽

Peter Berger ◽

Christoph Zenzmaier

Keyword(s):

Transforming Growth Factor ◽

Whole Body ◽

In Vivo Models ◽

Age Related ◽

Cgmp Signaling ◽

Fibrotic Diseases ◽

Sgc Activators ◽

Sgc Stimulators

Fibrotic diseases cause high rates of morbidity and mortality, and their incidence increases with age. Despite intense research and development efforts, effective and well-tolerated antifibrotic treatments are scarce. Transforming growth factor-β signaling, which is widely considered the most important profibrotic factor, causes a pro-oxidant shift in redox homeostasis and a concomitant decrease in nitric oxide (NO) signaling. The NO/cyclic guanosine monophosphate (cGMP) signaling cascade plays a pivotal role in the regulation of cell and organ function in whole-body hemostasis. Increases in NO/cGMP can lead to relaxation of smooth muscle cells triggering vasorelaxation. In addition, there is consistent evidence from preclinical in vitro and in vivo models that increased cGMP also exerts antifibrotic effects. However, most of these findings are descriptive and the molecular pathways are still being investigated. Furthermore, in a variety of fibrotic diseases and also during the natural course of aging, NO/cGMP production is low, and current treatment approaches to increase cGMP levels might not be sufficient. The introduction of compounds that specifically target and stimulate soluble guanylate cyclase (sGC), the so called sGC stimulators and sGC activators, might be able to overcome these limitations and could be ideal tools for investigating antifibrotic mechanisms in vitro and in vivo as they may provide effective treatment strategies for fibrotic diseases. These drugs increase cGMP independently from NO via direct modulation of sGC activity, and have synergistic and additive effects to endogenous NO. This review article describes the NO/cGMP signaling pathway and its involvement in fibrotic remodeling. The classes of sGC modulator drugs and their mode of action are described. Finally, the preclinical in vitro and in vivo findings and antifibrotic effects of cGMP elevation via sGC modulation are reviewed. sGC stimulators and activators significantly attenuate tissue fibrosis in a variety of internal organs and in the skin. Moreover, these compounds seem to have multiple intervention sites and may reduce extracellular matrix formation, fibroblast proliferation, and myofibroblast activation. Thus, sGC stimulators and sGC activators may offer an efficacious and tolerable therapy for fibrotic diseases, and clinical trials are currently underway to assess the potential benefit for patients with systemic sclerosis.

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Molecular Targets of Androgen Signaling that Characterize Skeletal Muscle Recovery and Regeneration

Nuclear Receptor Signaling ◽

10.1621/nrs.13005 ◽

2015 ◽

Vol 13 (1) ◽

pp. nrs.13005 ◽

Cited By ~ 11

Author(s):

James G. MacKrell ◽

Benjamin C. Yaden ◽

Heather Bullock ◽

Keyue Chen ◽

Pamela Shetler ◽

...

Keyword(s):

Skeletal Muscle ◽

Molecular Targets ◽

Gene Signature ◽

Muscle Recovery ◽

In Vivo Models ◽

Adult Skeletal Muscle ◽

Injury Model ◽

Androgen Regulation

The high regenerative capacity of adult skeletal muscle relies on a self-renewing depot of adult stem cells, termed muscle satellite cells (MSCs). Androgens, known mediators of overall body composition and specifically skeletal muscle mass, have been shown to regulate MSCs. The possible overlapping function of androgen regulation of muscle growth and MSC activation has not been carefully investigated with regards to muscle regeneration. Therefore, the aim of this study was to examine coinciding androgen-mediated genetic changes in an in vitro MSC model and clinically relevant in vivo models. A gene signature was established via microarray analysis for androgen-mediated MSC engagement and highlighted several markers including follistatin (FST), IGF-1, C-X-C chemokine receptor 4 (CXCR4), hepatocyte growth factor (HGF) and glucocorticoid receptor (GR/Nr3c1). In an in vivo muscle atrophy model, androgen re-supplementation significantly increased muscle size and expression of IGF-1, FST, and HGF, while significantly decreasing expression of GR. Biphasic gene expression profiles over the 7-day re-supplementation period identifed temporal androgen regulation of molecular targets involved in satellite cell engagement into myogenesis. In a muscle injury model, removal of androgens resulted in delayed muscle recovery and regeneration. Modifications in the androgen signaling gene signature, along with reduced Pax7 and MyoD expression, suggested that limited MSC activation and increased inflammation contributed to the delayed regeneration. However, enhanced MSC activation in the androgen-deplete mouse injury model was driven by an androgen receptor (AR) agonist. These results provide novel in vitro and in vivo evidence describing molecular targets of androgen signaling, while also increasing support for translational use of AR agonists in skeletal muscle recovery and regeneration.

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Cellular Origin and Regulation of D-and L-Serine in in Vitro and in Vivo Models of Cerebral Ischemia

Journal of Cerebral Blood Flow & Metabolism ◽

10.1038/jcbfm.2014.164 ◽

2014 ◽

Vol 34 (12) ◽

pp. 1928-1935 ◽

Cited By ~ 12

Author(s):

Takato Abe ◽

Masataka Suzuki ◽

Jumpei Sasabe ◽

Shinichi Takahashi ◽

Miyuki Unekawa ◽

...

Keyword(s):

Cerebral Ischemia ◽

Nmda Receptor ◽

Cultured Cells ◽

Long Term Potentiation ◽

Artery Occlusion ◽

In Vitro Experiments ◽

In Vivo Models ◽

Cellular Origin

D-Serine is known to be essential for the activation of the N-methyl-D-aspartate (NMDA) receptor in the excitation of glutamatergic neurons, which have critical roles in long-term potentiation and memory formation. D-Serine is also thought to be involved in NMDA receptor-mediated neurotoxicity. The deletion of serine racemase (SRR), which synthesizes D-Serine from L-Serine, was recently reported to improve ischemic damage in mouse middle cerebral artery occlusion model. However, the cell type in which this phenomenon originates and the regulatory mechanism for D-/L-Serine remain elusive. The D-/L-Serine content in ischemic brain increased until 20 hours after recanalization and then leveled off gradually. The results of in vitro experiments using cultured cells suggested that D-Serine is derived from neurons, while L-Serine seems to be released from astroglia. Immunohistochemistry studies of brain tissue after cerebral ischemia showed that SRR is expressed in neurons, and 3-phosphoglycerate dehydrogenase (3-PGDH), which synthesizes L-Serine from 3-phosphoglycerate, is located in astrocytes, supporting the results of the in vitro experiments. A western blot analysis showed that neither SRR nor 3-PGDH was upregulated after cerebral ischemia. Therefore, the increase in D-/L-Serine was not related to an increase in SRR or 3-PGDH, but to an increase in the substrates of SRR and 3-PGDH.

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BH3-only proteins Puma and Bim are rate-limiting for γ-radiation– and glucocorticoid-induced apoptosis of lymphoid cells in vivo

Blood ◽

10.1182/blood-2005-04-1595 ◽

2005 ◽

Vol 106 (13) ◽

pp. 4131-4138 ◽

Cited By ~ 186

Author(s):

Miriam Erlacher ◽

Ewa M. Michalak ◽

Priscilla N. Kelly ◽

Verena Labi ◽

Harald Niederegger ◽

...

Keyword(s):

Dna Damage ◽

Target Genes ◽

Cultured Cells ◽

Lymphoid Cells ◽

Whole Body ◽

Γ Radiation ◽

B Cell Leukemia ◽

Induced Apoptosis

Numerous p53 target genes have been implicated in DNA damage–induced apoptosis signaling, but proapoptotic Bcl-2 (B-cell leukemia 2) family members of the BH3 (Bcl-2 homolog region [BH] 3)–only subgroup appear to play the critical initiating role. In various types of cultured cells, 3 BH3-only proteins, namely Puma (p53 up-regulated modulator of apoptosis), Noxa, and Bim (Bcl-2 interacting mediator of cell death), have been shown to initiate p53-dependent as well as p53-independent apoptosis in response to DNA damage and treatment with anticancer drugs or glucocorticoids. In particular, the absence of Puma or Bim renders thymocytes and mature lymphocytes refractory to varying degrees to death induced in vitro by growth factor withdrawal, DNA damage, or glucocorticoids. To assess the in vivo relevance of these findings, we subjected mice lacking Puma, Noxa, or Bim to whole-body γ-radiation or the glucocorticoid dexamethasone and compared lymphocyte survival with that in wild-type and BCL2–transgenic mice. Absence of Puma or Bcl-2 overexpression efficiently protected diverse types of lymphocytes from the effects of γ-radiation in vivo, and loss of Bim provided lower but significant protection in most lymphocytes, whereas Noxa deficiency had no impact. Furthermore, both Puma and Bim were found to contribute significantly to glucocorticoid-induced killing. Our results thus establish that Puma and Bim are key initiators of γ-radiation– and glucocorticoid-induced apoptosis in lymphoid cells in vivo.

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Effects of Controlled Dual Growth Factor Delivery on Bone Regeneration Following Composite Bone-Muscle Injury

10.1101/2020.03.25.008813 ◽

2020 ◽

Author(s):

Ramesh Subbiah ◽

Albert Cheng ◽

Marissa A. Ruehle ◽

Marian H. Hettiaratchi ◽

Luiz E. Bertassoni ◽

...

Keyword(s):

Growth Factors ◽

Growth Factor ◽

Bone Tissue ◽

Treatment Strategy ◽

Growth Factor Delivery ◽

Treatment Groups ◽

Vascular Growth ◽

Injury Model ◽

Dual Growth Factor Delivery

AbstractThe objective of this study was to investigate the controlled release of two growth factors (BMP-2 and VEGF) as a treatment strategy for clinically challenging composite injuries, consisting of a segmental bone defect and volumetric muscle loss. This is the first investigation of dual growth factor delivery in a composite injury model using an injectable smart delivery system consisting of heparin microparticles and alginate gel. The loading efficiency of growth factors into these biomaterials was found to be >90%, revealing a strong affinity of VEGF and BMP-2 to heparin and alginate. The system could achieve simultaneous or sequential release of VEGF and BMP-2 by varying the loading strategy. Single growth factor delivery (VEGF or BMP-2 alone) significantly enhanced vascular growth in vitro. However, no synergistic effect was observed for dual growth factor (BMP-2 + VEGF) delivery. Effective bone healing was achieved in all treatment groups (BMP-2, simultaneous or sequential delivery of BMP-2 and VEGF) in the composite injury model. The mechanics of the regenerated bone reached a maximum strength of ∼52% of intact bone with sequential delivery of VEGF and BMP-2. Overall, simultaneous or sequential co-delivery of low-dose BMP-2 and VEGF failed to fully restore the mechanics of bone in this injury model. Given the severity of the composite injury, VEGF alone may not be sufficient to establish mature and stable blood vessels when compared with previous studies co-delivering BMP-2+VEGF enhanced bone tissue regeneration. Hence, future studies are warranted to develop an alternative treatment strategy focusing on better control over growth factor dose, spatiotemporal delivery, and additional growth factors to regenerate fully functional bone tissue.HighlightsWe developed a smart growth factor delivery system using heparin microparticles and alginate that facilitates tunable delivery of VEGF and BMP-2 in a simultaneous or sequential manner by merely varying the loading strategy.In vitro, both VEGF and BMP-2 alone promoted vascular growth; however, VEGF was significantly more potent, and there was no detectable benefit of co-delivery.In vivo, both BMP-2 alone and co-delivery of VEGF and BMP-2 promoted bone formation in the challenging bone/muscle polytrauma model; however, none of the treatment groups restored biomechanical properties to that of uninjured bone.

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Testing the Efficacy and Toxicity of Adenylyl Cyclase Inhibitors against Enteric Pathogens Using In Vitro and In Vivo Models of Infection

Infection and Immunity ◽

10.1128/iai.01114-09 ◽

2010 ◽

Vol 78 (4) ◽

pp. 1740-1749 ◽

Cited By ~ 8

Author(s):

Scott T. Moen ◽

Carla A. Blumentritt ◽

Terry M. Slater ◽

Shilpa D. Patel ◽

Christopher B. Tutt ◽

...

Keyword(s):

Epithelial Cells ◽

Adenylyl Cyclase ◽

Cultured Cells ◽

Potent Inhibitor ◽

Suppressive Effect ◽

Intestinal Epithelial ◽

In Vivo Models ◽

Adp Ribosyltransferase

ABSTRACT Enterotoxigenic Escherichia coli (ETEC) produces the ADP-ribosyltransferase toxin known as heat-labile enterotoxin (LT). In addition to the toxic effect of LT resulting in increases of cyclic AMP (cAMP) and disturbance of cellular metabolic processes, this toxin promotes bacterial adherence to intestinal epithelial cells (A. M. Johnson, R. S. Kaushik, D. H. Francis, J. M. Fleckenstein, and P. R. Hardwidge, J. Bacteriol. 191:178-186, 2009). Therefore, we hypothesized that the identification of a compound that inhibits the activity of the toxin would have a suppressive effect on the ETEC colonization capabilities. Using in vivo and in vitro approaches, we present evidence demonstrating that a fluorenone-based compound, DC5, which inhibits the accumulation of cAMP in intoxicated cultured cells, significantly decreases the colonization abilities of adenylyl cyclase toxin-producing bacteria, such as ETEC. These findings established that DC5 is a potent inhibitor both of toxin-induced cAMP accumulation and of ETEC adherence to epithelial cells. Thus, DC5 may be a promising compound for treatment of diarrhea caused by ETEC and other adenylyl cyclase toxin-producing bacteria.

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New Alcohol and Onyx Mixture for Embolization: Feasibility and Proof of Concept in Both In Vitro and In Vivo Models

CardioVascular and Interventional Radiology ◽

10.1007/s00270-016-1559-z ◽

2017 ◽

Vol 40 (5) ◽

pp. 735-743

Author(s):

Mohammad Saeed Kilani ◽

Fatemeh Zehtabi ◽

Sophie Lerouge ◽

Gilles Soulez ◽

Jean Michel Bartoli ◽

...

Keyword(s):

Proof Of Concept ◽

In Vivo Models

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Über die Antigenität eines chemisch induzierten neurogenen Sarkoms der Ratte [Antigenicity of a chemically induced neurogenic rat sarcoma—author's trans]

Veterinary Pathology ◽

10.1177/030098588001700409 ◽

1980 ◽

Vol 17 (4) ◽

pp. 477-489

Author(s):

H. Elling ◽

D. Stavrou

Keyword(s):

Tumor Cells ◽

Cultured Cells ◽

Fluorescent Antibody ◽

Whole Body ◽

Plexus Brachialis ◽

Target Cells ◽

Chemically Induced ◽

S 100 ◽

Antibody Titers

Antigenicity of a chemically induced neurogenic rat sarcoma A neurogenic sarcoma was induced in the plexus brachialis of a male Long-Evans rat by administration of N-methyl-N-nitrosourea in the drinking water. The tumor was established in vitro and designated 76LE-NS-369. Cells from tissue culture grew as tumors when isografted in young rats. 76LE-NS-369 cells did not react with antiserum directed against gliaspecific S-100 protein. We used the cultured cells as target cells, and found specific antibodies in the sera of tumor-bearing and immunized rats by indirect fluorescent antibody stain and a complement-dependent antibody-mediated microcytotoxicity assay. In immunization experiments, incubation of tumor cells with Vibrio cholerae neuraminidase yielded higher antibody titers than an antigen preparation with untreated cells. Incubation with neuraminidase enhanced the sensitivity of tumor cells to antibody and complement in vitro, whereas trypsinized cells showed complete loss of reactivity with autologous antisera. The specificity of antisera was tested by absorption with tumor, lyophilized rat whole body and rat nerve tissues.

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Assessment of R18, COG1410, and APP96-110 in excitotoxicity and traumatic brain injury

Translational Neuroscience ◽

10.1515/tnsci-2017-0021 ◽

2017 ◽

Vol 8 (1) ◽

Cited By ~ 14

Author(s):

Li Shan Chiu ◽

Ryan S. Anderton ◽

Jane L. Cross ◽

Vince W. Clark ◽

Adam B. Edwards ◽

...

Keyword(s):

Traumatic Brain Injury ◽

Brain Injury ◽

Treatment Time ◽

Calcium Influx ◽

Axonal Injury ◽

Closed Head Injury ◽

Adhesive Tape ◽

In Vivo Models ◽

Injury Model

AbstractCationic arginine-rich and poly-arginine peptides (referred to as CARPs) have potent neuroprotective properties in in vitro excitotoxicity and in vivo models of stroke. Traumatic brain injury (TBI) shares many pathophysiological processes as stroke, including excitotoxicity. Therefore, we evaluated our lead peptide, poly-arginine R18, with the COG1410 and APP96-110 peptides, which have neuroprotective actions following TBI. In an in vitro cortical neuronal glutamic acid excitotoxicity injury model, R18 was highly neuroprotective and reduced neuronal calcium influx, while COG1410 and APP96-110 displayed modest neuroprotection and were less effective at reducing calcium influx. In an impact-acceleration closed-head injury model (Marmarou model), R18, COG1410, and APP96-110 were administered intravenously (300 nmol/kg) at 30 minutes after injury in male Sprague-Dawley rats. When compared to vehicle, no peptide significantly improved functional outcomes, however the R18 and COG1410 treatment groups displayed positive trends in the adhesive tape test and rotarod assessments. Similarly, no peptide had a significant effect on hippocampal neuronal loss, however a significant reduction in axonal injury was observed for R18 and COG1410. In conclusion, this study has demonstrated that R18 is significantly more effective than COG1410 and APP96-110 at reducing neuronal injury and calcium influx following excitotoxicity, and that both R18 and COG1410 reduce axonal injury following TBI. Additional dose response and treatment time course studies are required to further assess the efficacy of R18 in TBI.

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Advanced late-onset retinitis pigmentosa with dominant-acting D477G RPE65 mutation is responsive to oral synthetic retinoid therapy

BMJ Open Ophthalmology ◽

10.1136/bmjophth-2020-000462 ◽

2020 ◽

Vol 5 (1) ◽

pp. e000462 ◽

Cited By ~ 2

Author(s):

Paul F Kenna ◽

Marian M Humphries ◽

Anna-Sophia Kiang ◽

Philippe Brabet ◽

Laurent Guillou ◽

...

Keyword(s):

Retinitis Pigmentosa ◽

Cultured Cells ◽

Advanced Disease ◽

Late Onset ◽

Visual Fields ◽

Therapeutic Interventions ◽

Proof Of Concept ◽

Retinyl Acetate ◽

Synthetic Retinoid

ObjectivesNo therapeutic interventions are currently available for autosomal dominant retinitis pigmentosa (adRP). An RPE65 Asp477Gly transition associates with late-onset adRP, reduced RPE65 enzymatic activity being one feature associated with this dominant variant. Our objective: to assess whether in a proof-of-concept study, oral synthetic 9 cis-retinyl acetate therapy improves vision in such advanced disease.Methods and analysisA phase 1b proof-of-concept clinical trial was conducted involving five patients with advanced disease, aged 41–68 years. Goldmann visual fields (GVF) and visual acuities (VA) were assessed for 6–12 months after 7-day treatment, patients receiving consecutive oral doses (40 mg/m2) of 9-cis-retinyl acetate, a synthetic retinoid replacement.ResultsPathological effects of D477G variant were preliminarily assessed by electroretinography in mice expressing AAV-delivered D477G RPE65, by MTS [3-(4,5-dimethylthiazol-2-yl)−5-(3-carboxyme- thoxyphenyl)−2-(4-sulfophenyl)−2H-tetrazolium] assays on RPE viability and enzyme activity in cultured cells. In addition to a mild dominant effect reflected in reduced electroretinographics in mice, and reduced cellular function in vitro, D477G exhibited reduced enzymatic RPE65 activity in vitro. In patients, significant improvements were observed in GVF from baseline ranging from 70% to 200% in three of five subjects aged 67–68 years, with largest improvements at 7–10 months. Of two GVF non-responders, one had significant visual acuity improvement (5–15 letters) from baseline after 6 months.ConclusionFamilies with D477G variant have been identified in Ireland, the UK, France, the USA and Canada. Effects of single 7-day oral retinoid supplementation lasted at least 6 months, possibly giving visual benefit throughout remaining life in patients with advanced disease, where gene therapy is unlikely to prove beneficial.

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