Interleukin 8 (IL-8) is one of the common indicators used in the study of inflammatory mechanisms, angiogenesis, atherosclerosis, and the pathogenesis of cancer, etc. So why can’t we find IL-8 ELISA kits for mice or rats?

1. Introduction of IL-8

Interleukin-8 (IL-8), otherwise known as IL-8, GCP-1, and NAP-1, is a heparin-binding 8-9 kDa alpha member, or CXC family chemokine.There are at least 15 human CXC family members that all adopt a three beta -sheet/one alpha -helix structure. Most CXC chemokines show an N-terminal Glu-Leu-Arg (ELR) tripeptide motif. IL-8 circulates as a monomer, homodimer, and heterodimer with CXCL4/PF4. The monomer is considered the most bio-active, while the heterodimer can potentiate PF4 activity. IL-8 oligomerization is modulated by its interactions with matrix and cell surface glycosaminoglycans (GAGs).

IL-8 bioactivity is mediated through two G-protein-coupled receptors, termed CXCR1/IL-8 RA and CXCR2/IL-8 RB. CXCR1 is 45-50 kDa in size and is used almost exclusively by IL-8. CXCR2 is 35-40 kDa in size and is used by nearly all CXC chemokines. Both CXCR1 and CXCR2 constitutively associate into functional homodimers. They can also heterodimerize, but these complexes dissociate following IL-8 binding. CXCR2 ligation induces leukocyte adhesion to activated vascular endothelium and migration to sites of inflammation, while CXCR1 ligation primes neutrophil antimicrobial activity. IL-8 can also form a complex with Serpin A1/alpha-1 Antitrypsin, and this prevents IL-8 interaction with CXCR1.

In addition to its pro-inflammatory effects, IL-8 is involved in angiogenesis and the pathogenesis of atherosclerosis and cancer. It induces VEGF expression in vascular endothelial cells and functions as an autocrine factor for EC growth and angiogenesis. It is upregulated in atherosclerotic lesions and is elevated in the serum and cerebrospinal fluid following myocardial infarction. In cancer, IL-8 promotes epithelial-mesenchymal transition as well as tumor cell invasiveness and metastasis.

It is important to emphasize that rodents (Rodent definition: are a group of animal species named after the possession of a pair of specialized incisors with ever-growing roots. They are usually small mammals with plant-eating dietary habits. Rodents include numerous species such as rats, squirrels, hamsters, rabbits, etc.) do not have IL-8 gene counterpart!

2. Commercially available ELISA kits for mouse and rat IL-8

Mouse KC cDNA encodes a 96 amino acid (aa)residue precursor protein from which the amino-terminal 19 aa residues are cleaved to generate the 77 aa residue mature KC.The protein sequence of mouse KC shows approximately 63% identity to that of mouse MIP-2,another mouse alpha chemokine.In addition,the protein sequence of KC is about 60%identical to the human GROs.Like other alpha chemokines,mouse KC is a potent neutrophilattractant and activator.The activities of KC and MIP-2 have been shown to be mediated by the unique mouse IL-8 receptor that shows 71% and 68% aa sequence identity to human IL-8 Rβ and IL-8 Ra,respectively.Since an IL-8 homolog has not been identified in mice,its has-been suggested that MIP-2 and KC are the functional homologs of IL-8 and may function as the major proinflammatory alpha chemokines in mice.Increased KC expression has been tried and found associated with neutrophil influx in various inflammatory conditions.

CXCL15 (alias IL-8, wedge, Scyb15, linguine) causes increased chemokine activity. Acts upstream or within hematopoiesis; inflammatory response; and neutrophil chemotaxis. Situated in the extracellular region. Cast in numerous structures including the central nervous system; dorsal aorta; embryonic mesenchyme; genitourinary system; and umbilical vein.

CXCR1 (also referenced as IL-8RA) is the interleukin-8 receptor, a potent neutrophil chemotactic factor.Binding of IL-8 to the receptor causes neutrophil activation. This response is submitted by G proteins that activate the phosphatidylinositol-calcium second messenger system.

CXCR2 (alias IL-8RB) Interleukin-8 receptor, a potent neutrophil chemotactic factor.Binding of IL-8 to the receptor causes neutrophil activation. This response is submitted by G proteins that activate the phosphatidylinositol-calcium second messenger system. Have a high affinity for IL – 8. Also binds with high affinity to CXCL3, GRO / MGSA and NAP – 2.

To gain insight into this indicator of IL – 8, we briefly analyze both protein sequence similarity and receptor, and refer to the literature for recommendations.

3. Protein Sequence Comparison

We obtained 34 sequences when we used NCBI’s Protein blast function to blast mouse data with the protein sequence of Human IL-8. All 34 sequences were downloaded, and MEGA7 was used for sequence comparison and construction of phylogenetic tree, and redundant sequences were removed to reduce confusion and increase readability to participate in later analysis. The homologous sequence of Human IL-8 in rat was also selected.

Fig 1 Phylogenetic tree of Human IL-8 with homologous sequences in mouse and rat

As can be seen in the above figure, Human IL-8 has no one-to-one homologs in mice and rats. Slightly closer relationship with CXCL1, 2, 3, 4, 5 (CXCL5 also known as AMCF-II, Cxcl6, ENA-78, GCP-2, LIX, Scyb5, Scyb6 in mice and rats), and 7 is not supported by high statistics.

4. Receptor-based analysis

In humans, there are 2 receptors for IL-8, CXCR1 and CXCR2. IL-8 binds highly specifically to CXCR1 and low specifically to CXCR2. However, there are 6 other CXCL ligands that bind to CXCR1 and CXCR2 (Fig 2).

Fig 2 CXCL family receptor and ligand binding diagrams（Wang S, et al. 2021）

The rodents mouse and rat also express CXCR1 and CXCR2. recent studies have shown that mouse CXCR1 binds only to CXCL6 and IL-8 from humans. In contrast, CXCR2 binds to all seven other CXCL proteins (Fig 2, Table 1).

So, neither based on sequence homology nor based on receptor binding properties can the one most closely related to human IL-8 be identified. There is not one, but several genes that are more closely related to human IL-8.
In summary, IL-8 gene does not exist in rodents, and the corresponding detection can only be solved by detecting the receptor or similar functional chemokine family such as CXCL1/KC. We have Mouse CXCL1/KC ELISA products in Mult Sciences.

References

1. Lira SA, Zalamea P, Heinrich JN, et al. Expression of the chemokine N51/KC in the thymus and epidermis of transgenic mice results in marked infiltration of a single class of inflammatory cells. J Exp Med. 1994 Dec 1;180(6):2039-48.
2. Tani M, Fuentes ME, Peterson JW, et al. Neutrophil infiltration, glial reaction, and neurological disease in transgenic mice expressing the chemokine N51/KC in oligodendrocytes. J Clin Invest.
3. Godiska R, Chantry D, Dietsch GN, et al. Chemokine expression in murine experimental allergic encephalomyelitis. J Neuroimmunol. 1995 May;58(2):167-76.
4. Farone A, Huang S, Paulauskis J, et al. Airway neutrophilia and chemokine mRNA expression in sulfur dioxide-induced bronchitis. Am J Respir Cell Mol Biol. 1995 Mar;12(3):345-50.
5. Wang S, Gao S, Li Y, et al. Emerging Importance of Chemokine Receptor CXCR4 and Its Ligand in Liver Disease. Front Cell Dev Biol. 2021 Jul 27;9:716842.
6. Matsushima K, Yang D, et al. Interleukin-8: An evolving chemokine. Cytokine. 2022 May;153:155828.
7. Asfaha S, Dubeykovskiy AN, Tomita H,, et al.. Mice that express human interleukin-8 have increased mobilization of immature myeloid cells, which exacerbates inflammation and accelerates colon carcinogenesis. Gastroenterology. 2013 Jan;144(1):155-66.