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NF-κB–SEAP reporter HEK293 cells expressing human TLR8

Trafficking & signaling in HEK-Blue™ hTLR8 cells
(click to enlarge and see legend)

HEK-Blue™ hTLR8 cells were engineered from the human embryonic kidney HEK293 cell line to study the Toll-like receptor 8 (TLR8)-dependent NF-κB pathway. This important pattern recognition receptor (PRR) recognizes viral single-stranded RNA (ssRNA) structures and subsequently triggers NF-κB and IRF immune responses [1].

More details

Description

HEK-Blue™ hTLR8 cells express the human TLR8 gene as well as an NF-κB/AP-1-inducible SEAP reporter gene. SEAP (secreted embryonic alkaline phosphatase) levels produced upon TLR8 stimulation can be readily determined by performing the assay in HEK-Blue™ Detection, a cell culture medium that allows for real-time detection of SEAP. Alternatively, SEAP activity may be monitored using QUANTI-Blue™, a SEAP detection reagent.

Due to the expression of hTLR8, HEK-Blue™ hTLR8 cells strongly and stably respond to TLR8-specific ligands (see figures). Interestingly, while HEK-Blue™ murine (m)TLR8 cells do not respond to the TLR8 agonist ssRNA40, HEK-Blue™ hTLR8 cells respond to this ligand (see figures). Thus, findings regarding mouse TLR8 are not transposable to its human counterpart [1,2].

HEK-Blue™ hTLR8 cells do not respond to TLR7-specific ligands, since HEK293 cells do not express endogenous TLR7 (see figures). However, as they express endogenous levels of TLR3, TLR5, and NOD1 [in-house data], HEK-Blue™ hTLR8 cells may respond to their cognate ligands such as Poly(I:C), flagellin, or Tri-DAP (see figure).

As TLR8 activates both the NF-κB and IRF signaling pathways, InvivoGen also offers HEK-Dual™ hTLR8 cells; a cell line expressing a dual reporter system comprising an NF-κB-inducible SEAP and an IRF-inducible-Lucia luciferase (learn more here).

Of note, HEK293 cells express endogenous levels of various PRRs, including TLR3 and TLR5. Therefore, they might respond to their cognate ligands (see figures).

Key Features

Verified expression of human (h)TLR8
Strong and stable response to TLR8-specific ligands
Distinct monitoring of NF-κB by assessing the SEAP activities

Applications

Defining the distinct role of TLR8 in the NF-κB-dependent pathway
Comparing human and murine TLR8-responses using the HEK-Blue™ mTLR8 cell line
Screening for novel TLR8-specific agonists or inhibitors in comparison with their parental cell line HEK-Blue™ Null1

References:

1. Heil F. et al., 2004. Species-specific recognition of single-stranded RNA via Toll-like receptor 7 and 8. Science. 303:1526.
2. Eigenbrod T. & Dalpke A.H., 2015. Bacterial RNA: an underestimated stimulus for innate immune responses. J. Immunol. 195:411.
3. Georg P. & Sander L.E., 2019. Innate sensors that regulate vaccine responses. Curr. Op. Immunol. 59:31.

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