S100A8 Mouse

What is S100A8 and what are its primary structural characteristics in mice?

S100A8, also known as CP-10, Calgranulin A, or MRP8, is a 10 kDa member of the S100 family of calcium-binding proteins in mice. The mouse S100A8 protein consists of 89 amino acids with distinct structural modules including:

• An N-terminal alpha-helix

• A calcium-binding EF-hand segment

• A short central linker region

• A second EF-hand segment

• A C-terminal alpha-helix

Mouse S100A8 protein is encoded by the gene located on chromosome 1. The amino acid sequence of mouse S100A8 shares 80% homology with rat S100A8 and 57% with human S100A8 .

How does S100A8 typically interact with its binding partners in mouse models?

S100A8 in mice primarily functions through:

• Homodimerization (S100A8/S100A8)

• Heterodimerization with S100A9 (S100A8/S100A9)

• In the presence of Ca²⁺, these heterodimers can form heterotetramers

The S100A8/A9 heterodimer is more stable than S100A8 homodimers, which explains why isolated S100A8 has greater turnover in the absence of S100A9. This was demonstrated in knockdown experiments where S100A9 depletion also resulted in loss of S100A8 protein .

The dimeric complexes are found both intracellularly and extracellularly, where they bind to heparan sulfate and function as chemoattractants for polymorphonuclear neutrophils (PMNs) and macrophages .

What are the standard methods for detecting S100A8 in mouse experimental samples?

What are the mechanisms underlying S100A8's protective effects in mouse models of sepsis?

S100A8 demonstrates significant protective effects in mouse sepsis models through several mechanisms:

• Attenuation of inflammatory cytokine production:

  • S100A8 decreases LPS-induced expression of proinflammatory cytokines (IL-6, TNF-α) in peritoneal macrophages

  • This occurs through inhibition of TLR4-mediated signaling in an autocrine manner

• Signaling pathway modulation:

  • S100A8 downregulates phosphorylated p38 mitogen-activated protein kinases (MAPKs)

  • This reduces inflammatory responses to lipopolysaccharide (LPS) and bacteria

• Cytokine binding and sequestration:

  • S100A8/A9 displays high-affinity noncovalent binding with proinflammatory IL-1β, IL-6, and TNF-α

  • This suggests a potential cytokine capture mechanism

• Promotion of anti-inflammatory factors:

  • S100A8 promotes anti-inflammatory IL-10 expression in certain cell types

  • This causes impaired LPS-induced neutrophil infiltration and reduces inflammatory cytokine induction

• Reduction of leukocyte adhesion and migration:

  • S100A8 negatively regulates these processes by reducing p38 MAPK phosphorylation

These protective mechanisms were demonstrated in multiple mouse sepsis models including lethal endotoxemia, Escherichia coli injection, and cecal ligation and puncture .

How does obesity and diabetes affect S100A8 expression and function in mouse sepsis models?

Obesity and diabetes significantly alter S100A8 dynamics during sepsis in mice:

• Altered S100A8 regulation:

  • db/db, ob/ob, and western diet-fed mice show reduced upregulation of S100A8 induced by LPS treatment

  • This reduction is observed in both serum and peritoneal cells

• Survival implications:

  • These obese/diabetic mouse models demonstrate shorter survival after LPS injection

  • S100A8 supplementation significantly prolonged survival in these models

• Cellular mechanisms:

  • Macrophages from diabetic mice express and secrete higher levels of S100A9 compared to non-diabetic mice

  • Elevated glucose levels result in increased expression of S100A8 in isolated macrophages

  • S100A9 immunoreactivity is increased in macrophage-rich lesions in diabetic mice and in diabetic Apoe^(-/-)^ mice

• Adaptive response hypothesis:

  • The time-dependent increase in circulating S100A8 levels after LPS injection suggests S100A8 upregulation represents an adaptive response triggered by endotoxemia

  • This adaptive response appears impaired in obesity/diabetes models

These findings indicate S100A8 supplementation may have therapeutic potential in sepsis, particularly in patients with obesity and diabetes comorbidities .

What is the relationship between S100A8 and TLR4 signaling in mouse models of cerebral ischemia?

The interaction between S100A8 and TLR4 plays a critical role in cerebral ischemia reperfusion (I/R) injury:

• Expression correlation:

  • S100A8 expression increases sharply in mouse brains after I/R injury

  • TLR4-deficient mice (C3H/HeJ) show significantly lower expression of I/R-induced S100A8 than wild-type (C3H/HeN) mice

• Functional outcomes:

  • TLR4-deficient mice have lower infarct volumes and better neurological outcomes after cerebral I/R

  • These mice also display less severe nerve cell swelling, reduced vacuolization, and less variation in cell nucleus shape

• Cellular localization:

  • S100A8-positive cells are almost exclusively observed in the ischemic hemisphere in model groups

  • More S100A8-positive cells are found in the ischemic brain of wild-type mice compared to TLR4-deficient mice

• Mechanistic relationship:

  • S100A8 has been identified as an endogenous ligand of TLR4

  • The S100A8-activated TLR4 signal pathway appears to lead to chained amplification of inflammatory reactions in early stages of cerebral I/R injury

These findings suggest that S100A8 interaction with TLR4 is involved in brain damage and inflammation triggered by I/R injury, indicating potential therapeutic approaches targeting this pathway .

How does S100A8/A9 contribute to cardiac hypertrophy in mouse models?

S100A8/A9 plays a significant role in promoting cardiac hypertrophy through specific mechanisms:

• Knockdown effects:

  • AAV9-mediated knockdown of S100A9 (which also depletes S100A8) significantly attenuates pressure overload-induced cardiac hypertrophy

  • This intervention prevents the increase in heart weight/body weight ratio, heart weight/tibia length ratio, cardiomyocyte cross-sectional area, and fibrotic area

• Functional improvements:

  • S100A9 deficiency decreases left ventricular internal diameter at end-diastole and interventricular septal thickness

  • It increases ejection fraction in mice with hypertrophy caused by pressure overload

• Molecular markers:

  • S100A9 deficiency reduces expression of hypertrophic markers:

    • ANP (atrial natriuretic peptide)

    • BNP (brain natriuretic peptide)

    • β-MHC (β-myosin heavy chain)

    • α-MHC (alpha myosin heavy chain)

  • Fibrotic markers are also reduced:

    • TGF-β (transforming growth factor-β)

    • Col I (collagen type I)

    • Col III (collagen type III)

    • α-SMA (α-smooth muscle actin)

These findings indicate that targeting S100A8/A9 may be a promising therapeutic approach for treating cardiac hypertrophy .

What are the mechanisms by which S100A8/A9 mediates mitochondrial dysfunction in sepsis-induced muscle atrophy?

S100A8/A9 drives mitochondrial dysfunction in sepsis-induced muscle atrophy through a specific signaling cascade:

• Clinical correlation:

  • Skeletal muscle atrophy increases disease severity and 60-day mortality in septic patients

  • Elevation of skeletal muscle index (ΔSMI) is an independent risk factor for mortality

• S100A8/A9 upregulation:

  • Mouse models of sepsis show skeletal muscle atrophy associated with upregulation of S100A8/A9

• Mitochondrial mechanism:

  • S100A8/A9 binding to RAGE (Receptor for Advanced Glycation End Products) induces:

    • Drp1 phosphorylation

    • Mitochondrial fragmentation

    • Energy exhaustion

    • Myocyte atrophy

• Intervention effects:

  • Inhibition of S100A8/A9 significantly improves mitochondrial function and alleviates muscle atrophy

  • RAGE ablation or administration of Drp1 inhibitor reduces Drp1-mediated mitochondrial fission

  • These interventions improve mitochondrial morphology and function

• Exacerbation effects:

  • Administration of recombinant S100A8/A9 protein worsens mitochondrial energy exhaustion and myocyte atrophy

This mechanistic pathway suggests targeting S100A8/A9-RAGE-initiated mitochondrial fission could offer a promising therapeutic intervention against septic muscle atrophy .

What experimental approaches should researchers use to investigate S100A8/A9's role in atherosclerosis in mouse models?

Why is S100A8 essential for embryonic development in mice, and what techniques are used to study this phenotype?

S100A8 plays a critical and non-redundant role in mouse embryonic development:

• Lethal phenotype:

  • Targeted disruption of the S100A8 gene causes rapid and synchronous embryo resorption by day 9.5 of development

  • This occurs in 100% of homozygous null embryos

  • Until this point, there is no evidence of developmental delay in S100A8^(-/-)^ embryos and decidualization is normal

• Temporal expression pattern:

  • S100A8 mRNA is expressed without S100A9 mRNA between 6.5 and 8.5 days postcoitum

  • Expression occurs within fetal cells infiltrating the deciduum near the ectoplacental cone

• Maternal-fetal interaction:

  • PCR genotyping at 7.5-8.5 days postcoitum suggests null embryos are infiltrated with maternal cells before overt signs of resorption

  • This implies a role for S100A8 in preventing maternal rejection of the implanting embryo

• Research significance:

  • This was the first evidence for non-redundant function of a member of the S100 gene family

  • The S100A8 null mouse provides a model for studying fetal-maternal interactions during implantation

Research techniques to study this phenotype include targeted gene disruption, temporal PCR genotyping, and analysis of maternal cell infiltration during early development .

What are the controversies regarding S100A8's role in inflammation - protective or harmful?

The role of S100A8 in inflammation presents several apparent contradictions that researchers must consider:

As noted in the literature: "It remains controversial whether S100A8 has a harmful or protective effect on survival in sepsis" . This controversy suggests that S100A8's function is highly context-dependent, influenced by:

• Concentration

• Timing of expression

• Disease model

• Cell type

• Presence of binding partners (especially S100A9)

• Route of administration (endogenous vs. exogenous)

Researchers should carefully consider these factors when designing experiments to study S100A8's role in inflammatory conditions .

What are the critical considerations when using recombinant S100A8 protein in mouse experiments?

When using recombinant mouse S100A8 protein in experiments, researchers should consider:

• Source and purity:

  • E. coli-derived recombinant mouse S100A8 protein (Met1-Glu89) is commonly used

  • Carrier-free preparations are available for applications where BSA might interfere

• Reconstitution and storage:

  • Typical reconstitution at 250 μg/mL in water

  • For stability, use a manual defrost freezer and avoid repeated freeze-thaw cycles

  • Store at -20 to -70°C for long-term storage (up to 6 months)

• Biological activity verification:

  • Validate activity by testing induction of CXCL1/KC secretion in C3H10T1/2 mouse embryonic fibroblast cells

  • The ED50 for this effect is typically 1.5-9 μg/mL

• Quality control:

  • Verify using SDS-PAGE under reducing and non-reducing conditions

  • Pure S100A8 should show bands at approximately 7 kDa

• Experimental design considerations:

  • Dose-response relationships are critical as different concentrations may have opposing effects

  • Consider timing of administration relative to inflammatory stimulus

  • Observe for both local and systemic effects

These considerations ensure experimental reproducibility and valid interpretation of results when using recombinant S100A8 in mouse models .

How should researchers design experiments to study S100A8 in mouse models of sepsis?