Background & Aims

Interleukin 6 (IL-6), its receptor (IL-6R), and the IL-6 signal transducer gp130 (IL-6ST) are well known regulators of innate immunity but also affect neuron morphology and function. Although important novel knowledge is available indicating the importance of peptidergic neurons in regulating goblet cell functions (1), the pathways and mechanisms responsible for the interactions between the peripheral nervous system and the microbiome are not yet fully understood. Therefore, we aimed to identify the alterations in gut motility, epithelial cell function, and potential signaling alterations in transgenic mice with a conditional depletion of Interleukin-6 signal transducer (IL6ST/gp130) in neurons expressing the nociceptor specific ion channel NaV1.8 (SNS-gp130-/-), which are largely protected from signatures of pathological pain (2, 3).

Methods

To assess gut motility, feces were collected, and the number and weight of fecal boli were recorded. An in vitro colon preparation was performed and colon samples were stimulated with depolarizing concentrations of KCl followed by ELISA quantification of calcitonin gene related peptide (CGRP) (4). Colonic mucus thickness and the number of goblet cells were quantified using histological methods. To assess CGRP effects on epithelial cells we compared stimulated with unstimulated colonic CaCo-2 and HT-29MTX cell lines, measured alterations of transepithelial electrical resistance (TEER) upon stimulation, and performed RNA sequencing to detect differentially expressed genes. Finally, we analyzed blood immune cells using FACS and assessed depressive-like behavior in the spared nerve injury (SNI) model for neuropathic pain. All experiments were performed in male and female SNS-gp130-/- and littermate gp130fl/fl control mice (age 4-6 months).

Results

Our data show that IL6ST depletion did not cause significant changes in gut motility. Although the morphology of the colonic epithelium and more specifically of goblet cells was similar in both mouse strains, colonic mucus thickness was decreased in SNS-gp130-/- mice, accompanied by a decreased stimulated CGRP release in these mice. CGRP did not alter TEER of differentiated CaCo-2 cells but induced transcriptional changes in CaCo-2 and HT-29MTX cells that may be related to the changes observed in mucus thickness. However, no differences in immune cell populations were discovered between mouse strains and no benefit of the conditional IL6ST depletion on the performance in the open field test was observed. Both mouse genotypes developed signs of depressive behavior as indicated by longer duration spent in the corners of the testing arena.

Conclusions

In conclusion, our study proposes that the depletion of IL6ST/gp130 in peripheral sensory neurons affects communication between sensory neurons and epithelial cells in the gut and specifically reduces the thickness of the mucus layer possibly by altering transcriptional programs of relevant cell types such as goblet cells. These changes are unrelated to depressive-like behavior in the neuropathic pain models but could affect other behaviors such as motivation to move.

References

1. Yang D, Jacobson A, Meerschaert KA, Sifakis JJ, Wu M, Chen X, Yang T, Zhou Y, Anekal PV, Rucker RA, Sharma D, Sontheimer-Phelps A, Wu GS, Deng L, Anderson MD, Choi S, Neel D, Lee N, Kasper DL, Jabri B, Huh JR, Johansson M, Thiagarajah JR, Riesenfeld SJ, Chiu IM. Nociceptor neurons direct goblet cells via a CGRP-RAMP1 axis to drive mucus production and gut barrier protection. Cell. 2022 Oct 27;185(22):4190-4205
2. Kalpachidou T, Malsch P, Qi Y, Mair N, Geley S, Quarta S, Kummer KK, Kress M. Genetic and functional evidence for gp130/IL6ST-induced transient receptor potential ankyrin 1 upregulation in uninjured but not injured neurons in a mouse model of neuropathic pain. Pain. 2022 Mar 1;163(3):579-589
3. Andratsch M, Mair N, Constantin CE, Scherbakov N, Benetti C, Quarta S, Vogl C, Sailer CA, Uceyler N, Brockhaus J, Martini R, Sommer C, Zeilhofer HU, Müller W, Kuner R, Davis JB, Rose-John S, Kress M. A key role for gp130 expressed on peripheral sensory nerves in pathological pain. J Neurosci. 2009 Oct 28;29(43):13473-83.
4. Roza C, Reeh PW. Substance P, calcitonin gene related peptide and PGE2 co-released from the mouse colon: a new model to study nociceptive and inflammatory responses in viscera, in vitro. Pain. 2001 Sep;93(3):213-219

Presenting Author

Lydia Riehl

Poster Authors

Lydia Riehl

MSc

Medical University Innsbruck, Institute of Physiology

Lead Author

E-mail

Johannes ao.Univ.-Prof. Mag. Dr.rer.nat. Fürst

Medical University Innsbruck, Institute of Physiology

Lead Author

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Susanne Prof. Dr. Sauer

FAU Erlangen, Institute of Physiology and Pathophysiology

Lead Author

E-mail

Kai Kummer

Mag.rer.nat.PhD

Institute of Physiology, Medical University of Innsbruck, Innsbruck, Austria

Lead Author

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Theodora Kalpachidou

Medical University of Innsbruck

Lead Author

E-mail

Michaela Kress

Institute of Physiology, Medical University of Innsbruck, Innsbruck, Austria

Lead Author

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Topics

  • Models: Chronic Pain - Neuropathic