Rantes Mouse

What is RANTES/CCL5 and what are its primary functions in mice?

RANTES is an 8kD member of the C-C family of chemokines that functions as a proinflammatory chemoattractant for several cell types including CD4+ T cells, monocytes, and eosinophils . In mice, RANTES cDNA encodes a 91 amino acid protein . Unlike other members of the chemokine superfamily, RANTES has been observed to be downregulated upon activation in T cells .

RANTES serves multiple biological functions in mice, including:

• Recruitment of immune cells to inflammatory sites

• Modulation of glutamate transmission in neuronal tissues

• Promotion of angiogenesis and vascular network formation

• Involvement in atherosclerotic plaque formation

Which receptors does mouse RANTES interact with and how do they affect signaling?

Mouse RANTES primarily signals through three G protein-coupled receptors:

• CCR1

• CCR3

• CCR5

These receptors have differential impacts on RANTES-mediated functions. In mouse spinal cord synaptosomes, antibodies against CCR1 and CCR5 prevented RANTES-induced facilitation of glutamate exocytosis, while anti-CCR3 antibody was ineffective. Similarly, CCR1 antagonist BX513 and CCR5 antagonist D-Ala-peptide T-amide (DAPTA) blocked RANTES-induced effects, whereas the CCR3 antagonist SB 328437 showed no activity in this system .

How should RANTES be handled in laboratory settings?

For optimal handling of recombinant mouse RANTES:

• Centrifuge the vial before opening

• Reconstitute by gently pipetting recommended solution down the sides of the vial

• Do not vortex

• Allow several minutes for complete reconstitution

• For prolonged storage, dilute to working aliquots in a 0.1% BSA solution

• Store at -80°C and avoid repeated freeze-thaw cycles

What methods are available for detecting RANTES in mouse samples?

For protein detection, the Quantikine ELISA Kit represents a standard method for quantifying mouse RANTES in:

• Cell culture supernatants

• Serum

• Platelet-poor plasma

For immunoblotting applications, antibodies such as the R6G9 monoclonal antibody can be used at concentrations ≤2 μg/mL on samples like lysates of LPS-activated Raw 264.7 cells .

What are the performance characteristics of ELISA assays for mouse RANTES?

The mouse/rat CCL5/RANTES Quantikine ELISA kit demonstrates high precision and recovery rates:

Intra-Assay Precision:

Inter-Assay Precision:

Recovery of RANTES in different mouse sample types:

How does RANTES modulate glutamate transmission in mouse neural tissues?

RANTES (0.01-1 nM) facilitates the 15 mM K+-evoked overflow of [³H]D-aspartate ([³H]D-ASP) from mouse spinal cord synaptosomes, although it does not affect spontaneous release. This modulation involves:

• Signaling through CCR1 and CCR5 receptors (but not CCR3)

• PLC-dependent mobilization of Ca²⁺ from IP₃-sensitive stores

• No involvement of adenylyl cyclase

Interestingly, RANTES effects differ between brain regions. In mouse cortex, CCR1 and CCR5 antagonists prevented RANTES-mediated [³H]D-ASP release, while RANTES-induced inhibition of K⁺-evoked [³H]D-ASP exocytosis was antagonized by all three receptor antagonists including the CCR3 antagonist .

What role does RANTES play in mouse models of vascular injury and repair?

RANTES demonstrates significant pro-angiogenic effects in mouse hindlimb ischemia models. RANTES-loaded polysaccharide-based microparticles have been shown to:

• Improve clinical scores in mice with hindlimb ischemia

• Induce revascularization of ischemic tissues

• Promote muscle regeneration in injured mouse limbs

The mechanisms underlying these effects involve interactions with endothelial progenitor cells (EPCs) through:

• Increased spreading and migration of EPCs

• Formation of vascular networks

• Engagement with CCR5, syndecan-4, and CD44 receptors on EPCs

What structural elements of RANTES are critical for its function in mice?

Two critical structural aspects of RANTES are essential for its pro-angiogenic effects:

• Chemokine oligomerization: Studies using [E66A]-RANTES mutant with impaired oligomerization ability demonstrate that this property is essential for RANTES-induced angiogenesis in vitro.

• Glycosaminoglycan binding: Research with [⁴⁴AANA⁴⁷]-RANTES mutant (mutated in the main glycosaminoglycan binding site) shows that this interaction is also necessary for angiogenic activity .

These findings indicate that both oligomerization and GAG binding are required for RANTES to effectively promote angiogenesis in mouse models.

How does RANTES influence atherosclerosis progression in mice?

In hypercholesterolemic mouse models, blocking the RANTES pathway using the CC chemokine antagonist Met-RANTES reduces atherosclerosis progression by:

• Diminishing expression of major chemokines and chemokine receptors

• Decreasing leukocyte infiltration into vascular lesions

• Increasing collagen-rich atheroma formation, a feature associated with stable atheroma

The treatment was well-tolerated and did not affect serum lipid profiles, suggesting a direct effect on the inflammatory component of atherosclerosis rather than on lipid metabolism .

What is the relationship between RANTES and eosinophil recruitment in mouse allergic inflammation?

In OVA-sensitized mice, injection of 500 ng of RANTES into the peritoneal cavity did not induce significant leukocyte infiltration compared to vehicle controls. The total number of cells, eosinophils, neutrophils, and mononuclear cells remained similar between RANTES-injected and vehicle-injected mice .

This suggests that while RANTES is important for eosinophil recruitment in allergic contexts, additional factors or specific inflammatory environments may be required for its full activity.

How do RANTES receptor antagonists affect inflammatory responses in mouse models?

RANTES receptor antagonists demonstrate significant anti-inflammatory effects:

• Met-RANTES (a CC chemokine antagonist) reduces atherosclerosis progression in hypercholesterolemic mouse models

• In neuronal tissues, specific receptor antagonists show differential effects:

  • BX513 (CCR1 antagonist) prevents RANTES-induced facilitation of glutamate exocytosis

  • DAPTA (CCR5 antagonist) blocks RANTES-induced effects on glutamate release

  • SB 328437 (CCR3 antagonist) shows context-dependent activity

These findings suggest that targeted antagonism of specific RANTES receptors might allow for selective modulation of RANTES functions in inflammatory conditions.

What are the key experimental controls for RANTES studies in mice?

When designing mouse experiments to study RANTES function, researchers should include:

• Appropriate vehicle controls for RANTES administration

• Receptor antagonist controls to confirm receptor specificity

• Antibody specificity controls when using anti-RANTES or anti-receptor antibodies

• Concentration-response analyses to determine optimal RANTES dosing

• Time-course studies to capture the dynamic nature of RANTES responses

• Cross-validation using multiple detection methods (ELISA, Western blot, qPCR)

How should researchers design delivery systems for RANTES in mouse models?

RANTES delivery methods significantly impact experimental outcomes. Options include:

• Direct administration of recombinant RANTES protein

• Sustained-release formulations such as RANTES-loaded polysaccharide-based microparticles, which have demonstrated improved efficacy in hindlimb ischemia models

• RANTES mutants to study structural requirements for activity

• Local versus systemic delivery depending on the target tissue and research question

How can researchers effectively study RANTES-receptor interactions in mouse models?

To investigate RANTES-receptor interactions:

• Use receptor-specific antagonists (BX513 for CCR1, DAPTA for CCR5, SB 328437 for CCR3)

• Apply receptor-specific antibodies to block individual receptors

• Consider using receptor knockout mice when available

• Employ multiple approaches to confirm receptor involvement, as RANTES can signal through multiple receptors simultaneously

• Investigate tissue-specific receptor expression patterns to explain differential responses

How do researchers explain different RANTES effects in various mouse tissues?

Differences in RANTES effects across mouse tissues may be attributed to:

• Differential receptor expression: The relative expression of CCR1, CCR3, and CCR5 varies between tissues

• Tissue-specific signaling pathways: RANTES activates PLC-dependent pathways in spinal cord but may engage different signaling in other tissues

• Context-dependent factors: Local inflammatory environments may modify RANTES activity

• Concentration-dependent effects: RANTES demonstrates different activities at varying concentrations

Why might exogenous RANTES administration produce variable results in different mouse models?

Variability in responses to exogenous RANTES administration may result from:

• Pre-existing inflammatory state: RANTES effects depend on the baseline activation status of target tissues

• Delivery method: Direct protein administration versus sustained-release formulations

• RANTES oligomerization status: Native versus mutant forms with altered oligomerization capacity

• Presence of co-factors: Other cytokines or chemokines may synergize with or antagonize RANTES

• Target cell availability: The abundance and activation state of RANTES-responsive cells varies between models