scholarly journals A Simple and Effective Flow Cytometry-Based Method for Identification and Quantification of Tissue Infiltrated Leukocyte Subpopulations in a Mouse Model of Peripheral Arterial Disease

2020 ◽  
Vol 21 (10) ◽  
pp. 3593 ◽  
Author(s):  
Konda Kumaraswami ◽  
Natallia Salei ◽  
Sebastian Beck ◽  
Stephan Rambichler ◽  
Anna-Kristina Kluever ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Arterial Disease ◽  
Tissue Samples ◽  
Collateral Arteries ◽  
Optimized Protocol ◽  
Adductor Muscles ◽  
Protocol Flow ◽  
Peripheral Arterial ◽  
Perivascular Infiltration ◽  
Detailed Protocol

Arteriogenesis, the growth of a natural bypass from pre-existing arteriolar collaterals, is an endogenous mechanism to compensate for the loss of an artery. Mechanistically, this process relies on a locally and temporally restricted perivascular infiltration of leukocyte subpopulations, which mediate arteriogenesis by supplying growth factors and cytokines. Currently, the state-of-the-art method to identify and quantify these leukocyte subpopulations in mouse models is immunohistology. However, this is a time consuming procedure. Here, we aimed to develop an optimized protocol to identify and quantify leukocyte subpopulations by means of flow cytometry in adductor muscles containing growing collateral arteries. For that purpose, adductor muscles of murine hindlimbs were isolated at day one and three after induction of arteriogenesis, enzymatically digested, and infiltrated leukocyte subpopulations were identified and quantified by flow cytometry, as exemplary shown for neutrophils and macrophages (defined as CD45+/CD11b+/Ly6G+ and CD45+/CD11b+/F4/80+ cells, respectively). In summary, we show that flow cytometry is a suitable method to identify and quantify leukocyte subpopulations in muscle tissue, and provide a detailed protocol. Flow cytometry constitutes a timesaving tool compared to histology, which might be used in addition for precise localization of leukocytes in tissue samples.

scholarly journals Angiogenic response to passive movement and active exercise in individuals with peripheral arterial disease

2013 ◽  
Vol 115 (12) ◽  
pp. 1777-1787 ◽  
Author(s):  
B. Hoier ◽  
M. Walker ◽  
M. Passos ◽  
P. J. Walker ◽  
A. Green ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Peripheral Arterial Disease ◽  
Arterial Disease ◽  
Passive Movement ◽  
Vegf Receptor ◽  
Tissue Samples ◽  
Angiogenic Response ◽  
Control Subjects ◽  
Thrombospondin 1 ◽  
Peripheral Arterial

Peripheral arterial disease (PAD) is caused by atherosclerosis and is associated with microcirculatory impairments in skeletal muscle. The present study evaluated the angiogenic response to exercise and passive movement in skeletal muscle of PAD patients compared with healthy control subjects. Twenty-one PAD patients and 17 aged control subjects were randomly assigned to either a passive movement or an active exercise study. Interstitial fluid microdialysate and tissue samples were obtained from the thigh skeletal muscle. Muscle dialysate vascular endothelial growth factor (VEGF) levels were modestly increased in response to either passive movement or active exercise in both subject groups. The basal muscle dialysate level of the angiostatic factor thrombospondin-1 protein was markedly higher ( P < 0.05) in PAD patients compared with the control subjects, whereas soluble VEGF receptor-1 dialysate levels were similar in the two groups. The basal VEGF protein content in the muscle tissue samples was ∼27% lower ( P < 0.05) in the PAD patients compared with the control subjects. Analysis of mRNA expression for a range of angiogenic and angiostatic factors revealed a modest change with active exercise and passive movement in both groups, except for an increase ( P < 0.05) in the ratio of angiopoietin-2 to angiopoietin-1 mRNA in the PAD group with both interventions. PAD patients and aged individuals showed a similar limited angiogenic response to active exercise and passive movement. The limited increase in muscle extracellular VEGF combined with an elevated basal level of thrombospondin-1 in muscle extracellular fluid of PAD patients may restrict capillary growth in these patients.

scholarly journals Optimized Protocol for Characterization of Mouse Gut Innate Lymphoid Cells

2020 ◽  
Vol 11 ◽  
Author(s):  
Ana Valle-Noguera ◽  
María José Gómez-Sánchez ◽  
Mathilde J. H. Girard-Madoux ◽  
Aranzazu Cruz-Adalia
Keyword(s):  
Flow Cytometry ◽  
Innate Lymphoid Cells ◽  
Tissue Homeostasis ◽  
Innate Immune ◽  
Lymphoid Cells ◽  
Cell Marker ◽  
Single Cell Suspension ◽  
Optimized Protocol ◽  
Detailed Protocol

Since their discovery, innate lymphoid cells (ILCs) have gradually been gaining greater relevance in the field of immunology due to their multiple functions in the innate immune response. They can mainly be found in mucosal and barrier organs like skin, gut, and lungs, and have been classified into five main types (NKs, ILC1s, ILC2s, ILC3s, and Lti cells) according to their function and development. They all play major roles in functions such as tissue homeostasis, early pathogen defense, regulation of inflammation, or tissue remodeling. ILCs are mostly tissue-resident cells tightly bound to the tissue structure, a fact that requires long and complex protocols that do not always provide sufficient yield for analysis. This suggests the need for optimized approaches aimed at ensuring that enriched and viable ILC samples are obtained, in order to furnish quality results. Herein a detailed protocol is established for obtaining a single-cell suspension highly enriched in lymphoid cells from mouse gut in order to identify the different subsets of ILCs by means of flow cytometry. The cell marker panel and flow cytometry gating strategies for identification and quantification of all the different ILC populations are provided for simultaneous analysis. Moreover, the protocol described includes a procedure for studying the different cytokines produced by ILC3s involved in maintaining the integrity of the gut barrier and defending against extracellular pathogens. As a result, herein an efficient method is presented for studying mouse ILCs within the lamina propria of the small intestine and colon; this can constitute a useful tool for future investigations in the field.

scholarly journals Endothelial Msx1 transduces hemodynamic changes into an arteriogenic remodeling response

2015 ◽  
Vol 210 (7) ◽  
pp. 1239-1256 ◽  
Author(s):  
Ine Vandersmissen ◽  
Sander Craps ◽  
Maarten Depypere ◽  
Giulia Coppiello ◽  
Nick van Gastel ◽  
...  
Keyword(s):  
Shear Stress ◽  
Adhesion Molecule ◽  
Knockout Mice ◽  
Fluid Shear Stress ◽  
Arterial Disease ◽  
Intercellular Adhesion Molecule ◽  
Intercellular Adhesion Molecule 1 ◽  
Collateral Arteries ◽  
Peripheral Arterial

Collateral remodeling is critical for blood flow restoration in peripheral arterial disease and is triggered by increasing fluid shear stress in preexisting collateral arteries. So far, no arterial-specific mediators of this mechanotransduction response have been identified. We show that muscle segment homeobox 1 (MSX1) acts exclusively in collateral arterial endothelium to transduce the extrinsic shear stimulus into an arteriogenic remodeling response. MSX1 was specifically up-regulated in remodeling collateral arteries. MSX1 induction in collateral endothelial cells (ECs) was shear stress driven and downstream of canonical bone morphogenetic protein–SMAD signaling. Flow recovery and collateral remodeling were significantly blunted in EC-specific Msx1/2 knockout mice. Mechanistically, MSX1 linked the arterial shear stimulus to arteriogenic remodeling by activating the endothelial but not medial layer to a proinflammatory state because EC but not smooth muscle cellMsx1/2 knockout mice had reduced leukocyte recruitment to remodeling collateral arteries. This reduced leukocyte infiltration in EC Msx1/2 knockout mice originated from decreased levels of intercellular adhesion molecule 1 (ICAM1)/vascular cell adhesion molecule 1 (VCAM1), whose expression was also in vitro driven by promoter binding of MSX1.

Management of Peripheral Arterial Disease

2006 ◽  
Vol 39 (3) ◽  
pp. 44
Author(s):  
WILLIAM E. GOLDEN ◽  
ROBERT H. HOPKINS
Keyword(s):  
Peripheral Arterial Disease ◽  
Arterial Disease ◽  
Peripheral Arterial

High tolerance of progressive elastic compression in peripheral arterial disease

VASA ◽  
2019 ◽  
Vol 48 (5) ◽  
pp. 413-417 ◽  
Author(s):  
Serge Couzan ◽  
Jean-François Pouget ◽  
Claire Le Hello ◽  
Céline Chapelle ◽  
Silvy Laporte ◽  
...  
Keyword(s):  
Venous Insufficiency ◽  
Ankle Brachial Index ◽  
Arterial Disease ◽  
Return Flow ◽  
High Tolerance ◽  
Compression Stockings ◽  
Short Term ◽  
Poor Tolerance ◽  
The Mean ◽  
Peripheral Arterial

Summary. Background: Theoretically progressive compression stockings, which produce a higher compression at the calf than at the ankle level, improve venous return flow without exacerbating peripheral arterial insufficiency (PAD). We aimed to evaluate the short-term tolerance of elastic progressive compression stockings on peripheral arterial vascularisation in patients with symptomatic PAD and associated mild venous insufficiency. Patients and methods: Monocentric, prospective, open pilot study of 18 patients (acceptability study, 6 x 6 plan) evaluating the short-term tolerance of progressive compression stockings (18 ± 2 mmHg at calf and 8 ± 2 mmHg at ankle level) in patients with PAD (ankle brachial index ABI > 0.60 < 0.75) and chronic venous insufficiency (C1s–C4 stages of the CEAP classification). Day 15 tolerance was evaluated by a composite primary criteria comprising: no decrease > 15 % of ABI on each side, no decrease > 15 % of toe brachial index (TBI) on each side and no decrease > 25 % of the number of active plantar flexions performed while standing. Results: The proportion of men was 77.8 %, mean age was 77.3 ± 7.5 years and no patient were diabetic. At inclusion, the mean low ABI was 0.60 ± 0.04 and the mean high ABI was 0.77 ± 0.18. The mean low TBI was 0.32 ± 0.09 and the mean high TBI 0.46 ± 0.15. The mean number of active standing plantar flexions was 33.0 ± 5.0. The majority of the patients were classified in CEAP C2s and C3 classes (class 2: 16.7 %, class C2s: 27.8 %, class C3: 44.4 %, class C4: 5.6 % and class C4s: 5.6 %). Poor tolerance occurred in no patient. By day 30, no patient had worsening of their arterial and venous symptoms. No adverse events occurred during the study. Conclusions: These results suggest a high tolerance of progressive elastic stockings (18 ± 2 mmHg at calf and 8 ± 2 mmHg at ankle level) in symptomatic PAD.

Surgical and endovascular hybrid approach in peripheral arterial disease of the lower limbs

VASA ◽  
2016 ◽  
Vol 45 (5) ◽  
pp. 417-422 ◽  
Author(s):  
Anouk Grandjean ◽  
Katia Iglesias ◽  
Céline Dubuis ◽  
Sébastien Déglise ◽  
Jean-Marc Corpataux ◽  
...  
Keyword(s):  
Mortality Rate ◽  
Peripheral Arterial Disease ◽  
Limb Salvage ◽  
Hybrid Approach ◽  
Arterial Disease ◽  
Early Mortality ◽  
Secondary Patency ◽  
Limb Salvage Rate ◽  
Peripheral Arterial ◽  
Salvage Rate

Abstract. Background: Multilevel peripheral arterial disease is frequently observed in patients with intermittent claudication or critical limb ischemia. This report evaluates the efficacy of one-stage hybrid revascularization in patients with multilevel arterial peripheral disease. Patients and methods: A retrospective analysis of a prospective database included all consecutive patients treated by a hybrid approach for a multilevel arterial peripheral disease. The primary outcome was the patency rate at 6 months and 1 year. Secondary outcomes were early and midterm complication rate, limb salvage and mortality rate. Statistical analysis, including a Kaplan-Meier estimate and univariate and multivariate Cox regression analyses were carried out with the primary, primary assisted and secondary patency, comparing the impact of various risk factors in pre- and post-operative treatments. Results: 64 patients were included in the study, with a mean follow-up time of 428 days (range: 4 − 1140). The technical success rate was 100 %. The primary, primary assisted and secondary patency rates at 1 year were 39 %, 66 % and 81 %, respectively. The limb-salvage rate was 94 %. The early mortality rate was 3.1 %. Early and midterm complication rates were 15.4 % and 6.4 %, respectively. The early mortality rate was 3.1 %. Conclusions: The hybrid approach is a major alternative in the treatment of peripheral arterial disease in multilevel disease and comorbid patients, with low complication and mortality rates and a high limb-salvage rate.

Markers of arterial stiffness in peripheral arterial disease

VASA ◽  
2015 ◽  
Vol 44 (5) ◽  
pp. 341-348 ◽  
Author(s):  
Marc Husmann ◽  
Vincenzo Jacomella ◽  
Christoph Thalhammer ◽  
Beatrice R. Amann-Vesti
Keyword(s):  
Arterial Stiffness ◽  
Peripheral Arterial Disease ◽  
Pulse Wave ◽  
Augmentation Index ◽  
Arterial Disease ◽  
Care Physician ◽  
Additional Information ◽  
Non Invasive ◽  
Assessment Techniques ◽  
Peripheral Arterial

Abstract. Increased arterial stiffness results from reduced elasticity of the arterial wall and is an independent predictor for cardiovascular risk. The gold standard for assessment of arterial stiffness is the carotid-femoral pulse wave velocity. Other parameters such as central aortic pulse pressure and aortic augmentation index are indirect, surrogate markers of arterial stiffness, but provide additional information on the characteristics of wave reflection. Peripheral arterial disease (PAD) is characterised by its association with systolic hypertension, increased arterial stiffness, disturbed wave reflexion and prognosis depending on ankle-brachial pressure index. This review summarises the physiology of pulse wave propagation and reflection and its changes due to aging and atherosclerosis. We discuss different non-invasive assessment techniques and highlight the importance of the understanding of arterial pulse wave analysis for each vascular specialist and primary care physician alike in the context of PAD.

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