Skip to main content
The Journal of Basic and Applied Zoology
Download PDF

IL-33 triggers lung autophagy in anaphylaxis mice models

The Journal of Basic and Applied Zoology volume 85, Article number: 55 (2024) Cite this article

Abstract

Background

The relationship between the alarming cytokine interleukin-33 (IL-33) and lung autophagy in systemic anaphylaxis mouse models is not yet fully elucidated, hence, the current study attempts to explain the regulation of lung autophagy in systemic anaphylactic mouse models. IL-33 plays a critical role in endoplasmic reticulum (ER) stress and autophagy regulation via insulin-like growth factor-binding protein-3 (IGFBP-3).

Results

The results of the present study confirmed the induction of systemic anaphylaxis in mice through the elevated mast cell mediators in the peritoneal lavage. Consequently, lung stress triggered IL-33 secretion that influenced autophagy markers; IGFBP-3, activating transcription factor-6 (ATF-6), autophagy related gene 4B (ATG4B), p62, microtubule-associated protein light chain3-II (LC3-II) as well as DNA damage-regulated autophagy modulator 1 (DRAM1).

Conclusion

This research is a trial to investigate lung autophagy in compound 48/80 or ovalbumin-induced systemic anaphylaxis mouse models and pay a particular attention to the role of IL-33 in lung autophagy in such models.

Graphical abstract

Background

Anaphylaxis is an acute fatal allergic reaction that arises after exposure to a trigger (Cianferoni, 2021). Systemic anaphylaxis mouse models are essential tools for elucidating the mechanisms of anaphylaxis and identifying potential therapies. However, the most common anaphylactic animal models used are compound 48/80 and ovalbumin-induced anaphylaxis mouse models (Waheed et al., 2021). It is well known that anaphylaxis stress is one of the vital types of stimuli that causes the release of several mediators from lung tissue (Piper, 1977). Mast cells (MCs) play a key role in the induction of anaphylaxis. Once activated, they secrete histamine and other inflammatory mediators, such as chemokines, cytokines, proteoglycans, and proteases (Kunimura et al., 2023). IL-33 belonging to the IL-1 family is barely secreted by living cells under steady state circumstances. This cytokine has been proposed to act as the first line of defense secreted in response to tissue injury and necroptosis (Shakerian et al., 2022). IL-33 is produced from barrier surface cells involving airway epithelial cells, endothelial cells, and fibroblasts (Lei et al., 2019). It stimulates and amplifies the production of inflammatory mediators, highlighting its involvement in anaphylactic reactions (Shakerian et al., 2022). Importantly, IL33 is suggested to act as a double-edged sword that functions as an extracellular cytokine and a nuclear factor modulating gene expression (Lei et al., 2019). Recent studies scoped on the therapeutic and diagnostic role of autophagy in different diseases (Khoza et al., 2022; Singh et al., 2021). Autophagy is a pro-survival intrinsic defense process that aids the cell in dealing with stress by clearing damaged proteins (Khandia et al., 2019). The process of protein synthesis and folding is sensitive to ER stress that causes accumulation of unfolded proteins. Consequently, cells have evolved an unfolded protein response (UPR) to prevent further accumulation of unfolded proteins to maintain cell homeostasis (Li et al., 2021). In contrast, sustained endoplasmic reticulum stress triggers autophagy to cause destruction of cell physiological functions, which can also lead to cell death through a pro-death apoptotic mechanism (Dąbrowska-Bouta et al., 2022). Hence, autophagy can have both pro-survival and pro-death functions (Wang et al., 2023). Upon our research review, although there are multiple studies on mast cell autophagy in anaphylaxis models, no studies have examined autophagy induced by anaphylaxis stress in lung tissue in these two anaphylactic mice models. So, this research is a trial to investigate lung autophagy mechanism in compound 48/80 or ovalbumin-induced systemic anaphylaxis mouse models and produce new insights to the role of IL-33 in triggering lung autophagy in such anaphylaxis models.

Methods

Ethics and animals

All animals were authorized by the Ethics Committee of Faculty of Women, Ain Shams University (Code: Sci1332403002), which sticks to the Animal Research: Reporting of In Vivo Experiments (ARRIVE) guidelines. Thirty adult male albino mice (Mus musculus) were bred and kept under standard conditions. Before the commencement of the study, animals had spent a week getting used to the experimental conditions.

Materials

Compound 48/80 is a hygroscopic powder with condensation of N-methyl-P-methoxyphenethylamine and formaldehyde (C11H15NO)n. It was obtained from Sigma Aldrich (St. Louis, MO, USA). Ovalbumin (laboratory grade, purity ≥ 90%) is a creamy white powder (C6H8N2O4) that was purchased from Loba Chemie Pvt. Ltd. (Bombay, India). Aluminium hydroxide was purchased from El-Nasr Pharmaceutical Chemical Company, Cairo, Egypt.

Experimental design

Thirty adult male Swiss Albino mice (Mus musculus), weighing 18–20 g, were randomly divided into three equal groups; the first group served as the normal control group (NC), the second group was the compound 48/80-induced anaphylaxis model (C48/80), and the third group was the ovalbumin-induced anaphylaxis model (OVA). The NC group was further divided into two subgroups: one normal control for the compound 48/80-induced anaphylaxis model (C48/80) and another normal control for the ovalbumin-induced anaphylaxis model (OVA). In C48/80 group, systemic anaphylactic shock was induced by a single intraperitoneal injection of 8 mg/kg body weight of compound 48/80 dissolved in 100 μl of phosphate-buffered saline (Waheed et al., 2021). This group was observed for 1 h after anaphylactic shock induction and then anesthetized and euthanized. In OVA group, mice were first sensitized on day 0 by intraperitoneal injection of 100 μg of ovalbumin adsorbed to 2 mg/ml aluminium hydroxide as adjuvant, followed by a booster on day 7 in the same manner. Mice were then challenged intravenously twice on days 14 and 21 with 2.25 mg ovalbumin in 200 μl saline/animal (Je et al., 2015). This group was anesthetized and euthanized 1 h after the second challenge dose on day 21. All animals in all experimental groups were anesthetized and euthanized by an intraperitoneal injection of 50 mg/kg of pentobarbital (Oh & Narver, 2024). Peritoneal lavage was then collected from all mice as previously described by Waheed et al. (2021). The peritoneal lavage was used to evaluate histamine, tryptase, leukotriene C4 (LTC4), prostaglandin D2 (PGD2), tumor necrosis factor-α (TNF-α), IL-4, IL-12, IL-1β, transforming growth factor-beta (TGF-β), IL-8 and hydrogen peroxide (H2O2) levels. Sedimented peritoneal mast cells were washed and resuspended in ice-cold phosphate-buffered saline and photographed by using the Olympus inverted microscope (Model: CKX41, Olympus Corporation, Tokyo, Japan). Lung tissues were isolated and homogenized in a lysis buffer for determination of IL-33, IGFBP-3, ATF-6, ATG4B, LC3-II, ubiquitin-binding protein p62 and DRAM1.

Determination of the mortality rate

To detect the mortality rate, mice were monitored for 1 h after anaphylaxis induction in the C48/80 model. In the OVA model, mice were observed for 1 h after each challenge dose. The mortality rate was calculated by using the following formula:

$$\text{Mortality }({\%})\text{ in each group }= (\frac{number \,of\, dead\, mice }{total \,number \,of \,expermental \,mice })\times 100$$

Determination of MCs mediators by ELISA technique

Evaluation of the levels of MCs mediators was carried out by mouse ELISA kits for the quantitative determination of their concentrations according to the manufacturer’s instructions as follows: Histamine by MyBioSource ELISA kit (catalogue number: MBS725193, San Diego, CA, USA); Tryptase by MyBioSource ELISA kit (catalogue number: MBS027692, San Diego, CA, USA); LTC4 by LSBio ELISA kit (catalogue number: LS-F28464, Shirley, MA, USA); PGD2 by MyBioSource ELISA kit (catalogue number: MBS703802, San Diego, CA, USA).

Determination of MCs cytokines and H2O2 by ELISA technique

Evaluation of the levels of MCs cytokines was carried out by mouse ELISA kits for the quantitative determination of their concentrations according to the manufacturer’s instructions (Abcam, USA) as follows: TNF-α (catalogue number: ab100747); IL-4 (catalogue number: ab100710); IL-12 p70 (catalogue number: ab119531); IL-1β (catalogue number: ab197742); TGF-β (catalogue number: ab119557); IL-8 by MyBioSource ELISA kit (catalogue number: MBS286946, San Diego, CA, USA); H2O2 (catalogue number: ab102500, Abcam, USA).

Lung tissue analysis

Evaluation of IL-33, IGFBP-3, ATF-6, LC3-II and p62 contents in lung tissue was carried out by mouse ELISA kits for the quantitative determination of their concentrations according to the manufacturer’s instructions as follows: Mouse IL-33 ELISA Kit, Elabscience Biotechnology (catalogue number: E-EL-M2642, USA); Mouse IGFBP-3 ELISA Kit, Thermo Fisher Scientific, Waltham, MA USA (catalogue number: EMIGFBP3); Mouse ATF-6 ELISA kit, MyBioSource (catalogue number: MBS720701, San Diego, CA, USA); Mouse LC3-II ELISA kit, Cell Biolabs, Inc., San Diego, CA, USA (catalogue number: CBA-5116). Mouse p62 ELISA kit, MyBioSource, San Diego, CA, USA (catalogue number: MBS3806181).

Real-time PCR amplification of ATG4B and DRAM1 genes

Total mRNAs were extracted from lung tissues using the RNeasy Mini Kit (Qiagen, Hilden, Germany) following the manufacturer’s instructions. cDNA was synthesized by reverse transcription reaction with the reverse transcription RT Kit (Qiagen, Hilden, Germany). The QuantiTect SYBR Green PCR Kit cat no.: 204141 (Qiagen, Germany) was handled to amplify the genes against the β-actin gene which was used as a housekeeping reference gene. All samples were analyzed using the 5 plex Rotor Gene PCR Analyzer (Qiagen, Germany). Programming of the real-time cycler was done as follows: activation step for 15 min at 95 °C for HotStarTaq DNA Polymerase activation. Three-step cycling: denaturation for 15 s at 94 °C, annealing for 30 s at 55 °C, extension for 30 s at 70 °C, for 40 cycles. The relative expression level for ATG4B and DRAM genes was standardized to an internal control (β-actin) and corresponding to calibrator were estimated using the equation 2−∆∆Ct of fold-change low. A primer pair of mice’s Actb gene was brought from OriGene Technologies, Inc., USA (Cat. no. MP200232). Primer pairs of mice’s ATG4B and DRAM1 genes were designed by Primer-Blast and brought from Invitrogen Co. (USA) (Table 1).

Table 1 mRNA quantification of ATG4B and DRAM1 genes and their corresponding primers
Full size table

Statistical analysis

The present research used the statistical package for social science (SPSS v.19). Mean ± S.E. was calculated for all groups. Calculation of statistical difference was performed by ANOVA and LSD tests. Means were considered significant at P < 0.05. *: P < 0.05 was compared to NC group.

Results

Mortality rate of anaphylactic mice models

The mortality rates reached 80% in C48/80-induced systemic anaphylaxis group and 90% in OVA-induced systemic anaphylaxis group.

Mast cell morphology in peritoneal lavage of different experimental groups

The peritoneal MCs of the NC group showed a normal round shape with intact membrane. However, C48/80 and OVA groups showed wide changes in mast cell morphology represented by an irregular membrane, and appearance of the released granules close to the cell surface, showing MCs degranulation (Fig. 1).

Fig. 1
figure 1

Photographs of mice peritoneal mast cells. A The control group showing the intact round shaped mast cells (arrow), B and C the C48/80 and OVA groups showing extensively degranulated mast cells and their secretory granules that were scattered outside the domain of the cells (arrow). The scale bar equals 100 µm. C48/80: compound 48/80, OVA: ovalbumin

Full size image

MC mediators in peritoneal lavage of experimental groups

In the present study, histamine and tryptase peritoneal lavage levels elucidated a significant increase (P < 0.05) in both C48/80 and OVA models as compared to the NC group. In the C48/80 model, histamine and tryptase showed 23.98 and 5.38-fold increase respectively compared to normal control animals. While in the OVA model, these mediators proved 9.72 and 3.64-fold increase respectively compared to their normal controls. LTC4 and PGD2 showed significant increases (P < 0.05) in the C48/80 group when compared to NC group. This increase was represented by an 8.32 and 4.46-fold increase, respectively. However, LTC4 and PGD2 expressed a 2.93 and 2.11-fold increase, respectively, in the OVA group compared to the NC group (Figs. 2, 3).

Fig. 2
figure 2

Levels of histamine, tryptase, LTC4 and PGD2 in peritoneal lavage of the compound 48/80-induced anaphylaxis model. *: Significant changes (P < 0.05) when compared to the normal control group. NC: normal control group; C48/80: compound 48/80 induced systemic anaphylaxis group; LTC4: Leukotriene C4; PGD2: Prostaglandin D2. Data are expressed as means ± S.E. (n = 10)

Full size image
Fig. 3
figure 3

Levels of histamine, tryptase, LTC4 and PGD2 in peritoneal lavage of an ovalbumin-induced anaphylaxis model. *: Significant changes (P < 0.05) when compared to the normal control group. NC: normal control group; OVA: ovalbumin-induced systemic anaphylaxis group; LTC4: Leukotriene C4; PGD2: Prostaglandin D2. Data are expressed as means ± S.E. (n = 10)

Full size image

Cytokines and H2O2 levels in peritoneal lavage of experimental groups

The peritoneal lavage of TNF-α, IL-4, IL-12 and IL-1β levels were increased significantly (P < 0.05) in both C48/80 and OVA anaphylaxis models as compared with their corresponding normal controls (Fig. 4). In OVA-induced anaphylaxis model, TGF-β and IL-8 verified significant (P < 0.05) increases compared to their corresponding normal controls, whereas the C48/80-induced anaphylaxis model did not exhibit any significant changes. Regarding H2O2 levels in peritoneal lavage, there was a significant elevation (P < 0.05) in both C48/80, and OVA anaphylaxis groups compared to their NC groups (Figs. 4, 5).

Fig. 4
figure 4

Levels of TNF-α, IL-4, IL-12, IL-1β, TGF-β, IL-8 and H2O2 in peritoneal lavage of the compound 48/80-induced anaphylaxis model. *: Significant changes (P < 0.05) compared to the normal control group. NC: normal control group; C48/80: compound 48/80 induced systemic anaphylaxis group; Data are expressed as means ± S.E. (n = 10)

Full size image
Fig. 5
figure 5

Levels of TNF-α, IL-4, IL-12, IL-1β, TGF-β, IL-8 and H2O2 in peritoneal lavage of an ovalbumin-induced anaphylaxis model. *: Significant changes (P < 0.05) compared to the normal control group. NC: normal control group; OVA: ovalbumin-induced systemic anaphylaxis group; Data are expressed as means ± S.E. (n = 10)

Full size image

Lung IL-33 and autophagy mediators of experimental groups

In the current study, lung tissue IL-33 levels displayed significant (p < 0.05) increases of 8.67 and 4.28-fold in the C48/80 and OVA groups respectively, compared to normal control mice. Similarly, lung tissue IGFBP-3 levels manifested a significant increase (p < 0.05) in both anaphylaxis models in relation to the NC animals. Alternatively, lung tissue ATF-6 levels proved a significant (p < 0.05) decrease of 6.17 and 2.18- fold in the C48/80 and OVA groups respectively compared to their corresponding normal control animals. The C48/80 group verified a significant (p < 0.05) decline in lung tissue ATG4B and DRAM1 gene expressions compared to NC group. Conversely, the gene expressions of ATG4B and DRAM1 established a significant (p < 0.05) rise in the OVA group in relation to the NC group. Lung tissue of LC3-II and p62 levels displayed significant (p < 0.05) 6.07 and 3.7-fold increases respectively in the C48/80 compared to their normal control mice. Also, lung tissue of LC3-II and p62 levels displayed significant (p < 0.05) increases of 2.65 and 2.15-fold in the OVA group, respectively, compared to the normal control mice (Figs. 6, 7).

Fig. 6
figure 6

Levels of IL-33, IGFBP-3, ATF-6, ATG4B, LC3-II, p62 and DRAM1 in lung tissue of the compound 48/80-induced anaphylaxis model. *: Significant changes (P < 0.05) compared to the normal control group. NC: normal control group; C48/80: compound 48/80 induced systemic anaphylaxis group; IGFBP-3: insulin-like growth factor-binding protein-3; ATF-6: activating transcription factor-6; ATG4B: autophagy related gene4B; LC3-II: microtubule-associated protein light chain3-II; DRAM1: DNA damage-regulated autophagy modulator 1; Data are expressed as means ± S.E. (n = 10). Expression levels of DRAM1 and ATG4B were decided by qPCR

Full size image
Fig. 7
figure 7

Levels of IL-33, IGFBP-3, ATF-6, ATG4B, LC3-II, p62 and DRAM1 in lung tissue of an ovalbumin-induced anaphylaxis model. *: Significant changes (P < 0.05) compared to the normal control group. NC: normal control group; OVA: ovalbumin-induced systemic anaphylaxis group; IGFBP-3: insulin-like growth factor-binding protein-3; ATF-6: activating transcription factor-6; ATG4B: autophagy related gene4B; LC3-II: microtubule-associated protein light chain3-II; DRAM1: DNA damage-regulated autophagy modulator 1; Data are expressed as means ± S.E. (n = 10). Expression levels of DRAM1 and ATG4B were decided by qPCR

Full size image

Discussion

Mast cell degranulation results in the release of mediators that have angiogenic, vasodilatory and pro-inflammatory effects, causing an increase in heart rate thus may lead to death (Rosas et al., 2022). This expresses the elevated mortality rates in the C48/80, and OVA groups compared to the NC group. Compound 48/80 increases the permeability of the cell membrane causing its disturbance thus triggering the release of mast cells mediators (Yu et al., 2018). Additionally, sensitization with ovalbumin enhances the hypersensitivity reaction pathway via secretion of Th2 cytokines and crosslinking of IgE on mast cells; causing their degranulation (Wan et al., 2022). MC mediators are divided into pre-formed and newly synthesized mediators. Pre-formed mediators make up histamines, serotonin, heparin, proteoglycans, and proteases (Dileepan et al., 2023). The present study elucidates a significant increase in histamine and tryptase levels of peritoneal lavage in C48/80 and OVA models compared to the NC group. Histamine is a major mediator of anaphylaxis which handles many anaphylaxis symptoms. When histamine binds to its receptors, it generates nitric oxide, increases vascular permeability, causes bronchospasm, and cardiac contraction (Nguyen et al., 2021). These changes may result in T-cell cytokines synthesis, eosinophil recruitment and cell adhesion molecule expression (Schirmer & Neumann, 2021). Tryptase is the most abundant secretory granule protein in lung mast cells. Mast cell-derived tryptase can damage microvascular tissue and promote protease-activated receptors resulting in the inflammatory response (Theoharides et al., 2023). Furthermore, the present study confirmed that MCs newly synthesized mediators as leukotrienes, prostaglandin, TNF-α, IL-4, IL-12, IL-1β, TGF-β, IL-8 and H2O2 were increased in the C48/80, and OVA models compared to their corresponding NC groups. Leukotriene is a lipid mediator that promotes acute allergic reactions (Butola et al., 2021). PGD2 is another pro-inflammatory lipid mediator that regulates airway hyperreactivity, mucus production and Th2 cytokine levels (Lee et al., 2020). A more severe variant of hypersensitivity reaction is the cytokine storm which is characterized by the expression of large quantities of cytokines such as TNFα, IL-1β, IL-8, IL-12, IL-4, TGF-β (Solimando et al., 2022). IL-1β and TNF-α can be synthesized and secreted from MCs. They exert a pro-inflammatory response, causing increased vascular permeability at the inflamed site associated with leukocyte migration (de Oliveira et al., 2021). Moreover, MCs produce a proinflammatory cytokine IL-12 that contributes to the development of CD4 + T cells into various helper T-cell subsets (Vignali & Kuchroo, 2012). Similarly, the synthesis and expression of chemokine IL-8 (CXCL8) by mast cells take part in various mechanisms, incorporating mitogenesis, angiogenesis, inflammation, chemotaxis and neutrophil degranulation (Matsushima et al., 2022). In addition, mast cells release IL-4 which stimulates T cell adhesion, triggers Th2 lymphocyte polarization and promotes B lymphocyte IgE class switching (León & Ballesteros-Tato, 2021). TGF-β1 is a potent chemotaxis for several different cell types, including mast cells, neutrophils, and monocytes (Olsson et al., 2000). In hypersensitivity, TGF- β1 contributes to the generation of Treg which is necessary to limit the immunopathological effects of unrestricted effector T-cell stimulation (Fallegger et al., 2022). Reactive oxygen species might have a role in mast cell activation in allergic subjects. Mast cells and other immune cells produce H2O2 which is an essential tool that enhances mast cell activation (Suzuki et al., 2005). This reflects the significant elevation in H2O2 levels in both allergic models in the present study. The alarming cytokine IL-33 is the focus of this study, where the C48/80 and OVA groups showed a significant increase in lung IL-33 content compared to the NC group. IL-33 is released by fibroblasts, endothelial cells, epithelial cells and mast cells upon stress induced by systemic anaphylaxis (West et al., 2021). Following lung stress, IL-33 is released from lung epithelial cells and exerts its pro-inflammatory functions. IL-33 induces lung cell metaplasia and fibrosis producing structural remodeling of the lung airways (Du et al., 2020). Autophagy, a highly maintained catabolic mechanism that is related to inflammatory disorders and airway remodeling (López et al., 2006). However, the relation between IL-33 and lung autophagy in anaphylaxis mouse models is not yet fully elucidated. In this study, a significant increase in IL-33 parallel with IGFBP-3 elevation were seen in both models. This means that IGFBP-3 may be triggered by IL-33 stress (Wu et al., 2021). IGFBP-3 stimulates lung epithelial autophagy through interaction with the ER protein that is involved in the regulation of cellular stress (Yin et al., 2017). Then, UPR is initiated when unfolded proteins accumulate in the ER (Read & Schröder, 2021). The activation of the UPR involves ATF6 signaling pathway, which plays a vital role in restoring protein homeostasis to the levels observed in non-stressed cells (Shen et al., 2021). ATF6 stands for a transmembrane ER stress sensor that is inactive in normal cells by its relation to the ER chaperone protein glucose-regulated protein 78 (GRP78) (Schröder & Kaufman, 2005). When unfolded proteins concentrate in the ER lumen, ATF6 is delivered to the Golgi apparatus in the COPII vesicle (Aridor, 2022). At the Golgi, ATF6 gets processed by proteases, which remove the luminal and transmembrane anchor that gives rise to N-terminal ATF6 (ATF6-N) (Tungkum, 2023). This might explain the significant decrease in lung ATF6 levels in both anaphylaxis mouse models compared to the NC groups in the present study. In the nucleus, ATF6-N enhances UPRER gene transcription GRP78. Then, GRP78 separates and binds unfolded or misfolded proteins, allowing the sensors to stimulate signaling pathways that rebuild protein folding and secretion (Lei et al., 2024). More to the point, autophagy is governed by autophagy related proteins (ATGs). Autophagy-related 4 (ATG4) is the special cysteine protease that promotes autophagy by facilitating autophagosome maturation by reversible lipidation and delipidation of microtubule associated protein 1-light chain 3 (LC-3) (Herhaus et al., 2020). ATG4B homologs contain an LC3-interacting region motif, which is essential for binding to and processing LC3 (Park et al., 2022). ATG4B splits the carboxyl terminus of newly synthesized pro-LC3 to generate LC3-I, allowing its conjugation to make membrane-bound LC3-II, which is essential for auto phagosome elongation (Gray et al., 2021). This proves the significant parallel increase of gene expression of ATG4B, and LC3-II content in lung tissue in the OVA-induced anaphylaxis model compared to the NC group in the present study. However, there was a negative correlation between gene expression of ATG4B and LC3-II content in lung tissue in the C48/80-induced anaphylaxis model. This decreased ATG4B expression was associated with the aggressive severity reaction against compound 48/80. This abrupt reaction may limit the phosphorylation of ATG4B thus decreasing its expression (Bortnik et al., 2020). The present results again elucidated a parallel increase in LC3-II and p62 in both models compared to normal control animals. p62 is used as an autophagic marker. p62 helps to deliver ubiquitin proteins to the proteasome for degradation. p62 possesses a less diffuse localization pattern than LC3, making it crucial to recognize the small autophagic vesicles by this marker. Nevertheless, p62 performs an architectural function in the formation of inclusion bodies connecting these structures to the autophagy machinery via the close interaction with LC3 (Komatsu & Ichimura, 2010). DRAM (DNA damage-regulated autophagy modulator) is a p53 target gene that encodes a lysosomal protein whichf1 provokes macro-autophagy (Crighton et al., 2006). In this study, the gene expression of DRAM1 in lung tissue was significantly reduced in the C48/80 group compared to the NC group. This might be related to the autophagy mechanism. Autophagy can inhibit apoptosis by clearing misfolded/unfolded proteins and damaged organelles and inhibiting caspase activation (Song et al., 2017). In ovalbumin-induced anaphylaxis mouse model, a prolonged duration of 21 days induces sustained ER stress that activates apoptosis. Here, the UPR is abortive in reducing the load of unfolded proteins in the ER. Then, ATF6-N enhances proapoptotic signals. This may explain the elevated gene expression of DRAM1 in the OVA group compared to the NC group.

Conclusion

This study investigated the role of IL-33 in triggering lung autophagy in anaphylaxis mouse models induced by compound 48/80 or ovalbumin. This study may open new insights to study the mechanism of autophagy in anaphylaxis mouse models. However, a deeper understanding of the particular role of IL-33 on this pathway is needed.

Availability of data and materials

All data and materials generated and analyzed during the current study are included in this manuscript.

Abbreviations

ANOVA:

Analysis of variance

ARRIVE:

Animal Research Reporting In Vivo Experiments

ATF-6:

Activating transcription factor-6

ATG4B:

Autophagy related gene 4B

C48/80:

Compound 48/80

DRAM1:

DNA damage-regulated autophagy modulator 1

ER:

Endoplasmic reticulum

H2O2 :

Hydrogen peroxide

IL-33:

Interleukin-33

IGFBP-3:

Insulin-like growth factor-binding protein-3.

IL-:

Interleukin-

LC3-II:

Microtubule-associated protein light chain3-II

LTC4:

Leukotriene C4

MCs:

Mast cells

OVA:

Ovalbumin

PGD2:

Prostaglandin D2

SPSS:

Statistical package for the social sciences

TNF-α:

Tumor necrosis factor-α

TGF-β:

Transforming growth factor-beta

UPR:

Unfolded protein response

References

  • Aridor, M. (2022). A tango for coats and membranes: New insights into ER-to-Golgi traffic. Cell Reports, 38(3), p110258.

    Article  Google Scholar 

  • Bortnik, S., Tessier-Cloutier, B., Leung, S., Xu, J., Asleh, K., Burugu, S., Magrill, J., Greening, K., Derakhshan, F., Yip, S., & Ng, T. (2020). Differential expression and prognostic relevance of autophagy-related markers ATG4B, GABARAP, and LC3B in breast cancer. Breast Cancer Research and Treatment, 183, 525–547.

    Article  CAS  PubMed  Google Scholar 

  • Butola, L. K., Dhok, A., Ambad, R., Kanyal, D., & Jha, R. K. (2021). Leukotrienes and inflammation–A review. Indian Journal of Forensic Medicine & Toxicology, 15(2), 295–301.

    Article  CAS  Google Scholar 

  • Cianferoni, A. (2021). Non–IgE-mediated anaphylaxis. Journal of Allergy and Clinical Immunology, 147(4), 1123–1131.

    Article  CAS  PubMed  Google Scholar 

  • Crighton, D., Wilkinson, S., O’Prey, J., Syed, N., Smith, P., Harrison, P. R., Gasco, M., Garrone, O., Crook, T., & Ryan, K. M. (2006). DRAM, a p53-induced modulator of autophagy, is critical for apoptosis. Cell, 126(1), 121–134.

    Article  CAS  PubMed  Google Scholar 

  • DÄ…browska-Bouta, B., Sulkowski, G., Gewartowska, M., & StrużyÅ„ska, L. (2022). Endoplasmic reticulum stress underlies nanosilver-induced neurotoxicity in immature rat brain. International Journal of Molecular Sciences, 23(21), 13013.

    Article  PubMed  PubMed Central  Google Scholar 

  • de Oliveira, R. G., Damazo, A. S., Antonielli, L. F., Miyajima, F., Pavan, E., Duckworth, C. A., da Silva Lima, J. C., Arunachalam, K., & de Oliveira Martins, D. T. (2021). Dilodendron bipinnatum Radlk. extract alleviates ulcerative colitis induced by TNBS in rats by reducing inflammatory cell infiltration, TNF-α and IL-1β concentrations, IL-17 and COX-2 expressions, supporting mucus production and promotes an antioxidant effect. Journal of Ethnopharmacology, 269, 113735.

    Article  PubMed  Google Scholar 

  • Dileepan, K. N., Raveendran, V. V., Sharma, R., Abraham, H., Barua, R., Singh, V., Sharma, R., & Sharma, M. (2023). Mast cell-mediated immune regulation in health and disease. Frontiers in Medicine, 10, 1213320.

    Article  PubMed  PubMed Central  Google Scholar 

  • Du, X., Li, C., Wang, W., Huang, Q., Wang, J., Tong, Z., Huang, K., Chen, Y., Yuan, H., Lv, Z., & Corrigan, C. J. (2020). IL-33 induced airways inflammation is partially dependent on IL-9. Cellular Immunology, 352, 104098.

    Article  CAS  PubMed  Google Scholar 

  • Fallegger, A., Priola, M., Artola-Borán, M., Núñez, N. G., Wild, S., Gurtner, A., Becher, B., Yousefi, S., Simon, H. U., Arnold, I. C., & Müller, A. (2022). TGF-β production by eosinophils drives the expansion of peripherally induced neuropilin− RORγt+ regulatory T-cells during bacterial and allergen challenge. Mucosal Immunology, 15(3), 504–514.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Gray, J. P., Uddin, M. N., Chaudhari, R., Sutton, M. N., Yang, H., Rask, P., Locke, H., Engel, B. J., Batistatou, N., Wang, J., & Grindel, B. J. (2021). Directed evolution of cyclic peptides for inhibition of autophagy. Chemical Science, 12(10), 3526–3543.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Herhaus, L., Bhaskara, R. M., Lystad, A. H., Gestal-Mato, U., Covarrubias-Pinto, A., Bonn, F., Simonsen, A., Hummer, G., & Dikic, I. (2020). TBK1-mediated phosphorylation of LC3C and GABARAP-L2 controls autophagosome shedding by ATG4 protease. EMBO Reports, 21(1), e48317.

    Article  CAS  PubMed  Google Scholar 

  • Je, I. G., Kim, D. S., Kim, S. W., Lee, S., Lee, H. S., Park, E. K., Khang, D., & Kim, S. H. (2015). Tyrosol suppresses allergic inflammation by inhibiting the activation of phosphoinositide 3-kinase in mast cells. PLoS ONE, 10(6), e0129829.

    Article  PubMed  PubMed Central  Google Scholar 

  • Khandia, R., Dadar, M., Munjal, A., Dhama, K., Karthik, K., Tiwari, R., Yatoo, M. I., Iqbal, H. M., Singh, K. P., Joshi, S. K., & Chaicumpa, W. (2019). A comprehensive review of autophagy and its various roles in infectious, non-infectious, and lifestyle diseases: Current knowledge and prospects for disease prevention, novel drug design, and therapy. Cells, 8(7), p674.

    Article  Google Scholar 

  • Khoza, L. J., Kumar, P., Dube, A., Demana, P. H., & Choonara, Y. E. (2022). Insights into innovative therapeutics for drug-resistant tuberculosis: Host-directed therapy and autophagy inducing modified nanoparticles. International Journal of Pharmaceutics, 622, 121893.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Komatsu, M., & Ichimura, Y. (2010). Physiological significance of selective degradation of p62 by autophagy. FEBS Letters, 584(7), 1374–1378.

    Article  CAS  PubMed  Google Scholar 

  • Kunimura, K., Akiyoshi, S., Uruno, T., Matsubara, K., Sakata, D., Morino, K., Hirotani, K., & Fukui, Y. (2023). DOCK2 regulates MRGPRX2/B2-mediated mast cell degranulation and drug-induced anaphylaxis. Journal of Allergy and Clinical Immunology, 151(6), 1585–1594.

    Article  CAS  PubMed  Google Scholar 

  • Lee, K., Lee, S. H., & Kim, T. H. (2020). The biology of prostaglandins and their role as a target for allergic airway disease therapy. International Journal of Molecular Sciences, 21(5), 1851.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lei, M., Wang, C. J., Yu, F., Xie, K., Lin, S. H., & Xu, F. (2019). Different intensity of autophagy regulate interleukin-33 to control the uncontrolled inflammation of acute lung injury. Inflammation Research, 68, 665–675.

    Article  CAS  PubMed  Google Scholar 

  • Lei, Y., Yu, H., Ding, S., Liu, H., Liu, C., & Fu, R. (2024). Molecular mechanism of ATF6 in unfolded protein response and its role in disease. Heliyon.

  • León, B., & Ballesteros-Tato, A. (2021). Modulating Th2 cell immunity for the treatment of asthma. Frontiers in Immunology, 12, 637948.

    Article  PubMed  PubMed Central  Google Scholar 

  • Li, J., Ullah, M. A., Jin, H., Liang, Y., Lin, L., Wang, J., Peng, X., Liao, H., Li, Y., Ge, Y., & Li, L. (2021). ORMDL3 functions as a negative regulator of antigen-mediated mast cell activation via an ATF6-UPR-autophagy–dependent pathway. Frontiers in Immunology, 12, 604974.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • López, E., del Pozo, V., Miguel, T., Sastre, B., Seoane, C., Civantos, E., Llanes, E., Baeza, M. L., Palomino, P., Cárdaba, B., & Gallardo, S. (2006). Inhibition of chronic airway inflammation and remodeling by galectin-3 gene therapy in a murine model. The Journal of Immunology, 176(3), 1943–1950.

    Article  PubMed  Google Scholar 

  • Matsushima, K., Yang, D., & Oppenheim, J. J. (2022). Interleukin-8: An evolving chemokine. Cytokine, 153, 155828.

    Article  CAS  PubMed  Google Scholar 

  • Nguyen, S. M. T., Rupprecht, C. P., Haque, A., Pattanaik, D., Yusin, J., & Krishnaswamy, G. (2021). Mechanisms governing anaphylaxis: Inflammatory cells, mediators, endothelial gap junctions and beyond. International Journal of Molecular Sciences, 22(15), 7785.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Oh, S. S., & Narver, H. L. (2024). Mouse and rat anesthesia and analgesia. Current Protocols, 4(2), e995.

    Article  CAS  PubMed  Google Scholar 

  • Olsson, N., Piek, E., ten Dijke, P., & Nilsson, G. (2000). Human mast cell migration in response to members of the transforming growth factor-β family. Journal of Leukocyte Biology, 67(3), 350–356.

    Article  CAS  PubMed  Google Scholar 

  • Park, N. Y., Jo, D. S., & Cho, D. H. (2022). Post-translational modifications of ATG4B in the regulation of autophagy. Cells, 11(8), 1330.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Piper, P. J. (1977). Anaphylaxis and the release of active substances in the lungs. Pharmacology & Therapeutics. Part B: General and Systematic Pharmacology, 3(1), 75–98.

    CAS  Google Scholar 

  • Read, A., & Schröder, M. (2021). The unfolded protein response: An overview. Biology, 10(5), 384.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Rosas, E. P., Paz, S. T., Costa, R. F., da Silva, A. A., da Silva, R. L., da Silva, A. P., da Silva, S. R., de Medeiros, P. L., de Freitas, M. F., & Valença, M. M. (2022). Histomorphometry of mast cells in the convexity of human intracranial dura mater. Journal of Anatomy, 240(4), 724–734.

    Article  CAS  PubMed  Google Scholar 

  • Schirmer, B., & Neumann, D. (2021). The function of the histamine H4 receptor in inflammatory and inflammation-associated diseases of the gut. International Journal of Molecular Sciences, 22(11), 6116.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Schröder, M., & Kaufman, R. J. (2005). The mammalian unfolded protein response. Annual Review of Biochemistry, 74, 739–789.

    Article  PubMed  Google Scholar 

  • Shakerian, L., Kolahdooz, H., Garousi, M., Keyvani, V., Kheder, R. K., Faraj, T. A., Yazdanpanah, E., & Esmaeili, S. A. (2022). IL-33/ST2 axis in autoimmune disease. Cytokine, 158, 156015.

    Article  CAS  PubMed  Google Scholar 

  • Shen, Y., Li, R., Yu, S., Zhao, Q., Wang, Z., Sheng, H., & Yang, W. (2021). Activation of the ATF6 (activating transcription factor 6) signaling pathway in neurons improves outcome after cardiac arrest in mice. Journal of the American Heart Association, 10(12), e020216.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Singh, A., Handa, M., Ruwali, M., Flora, S., Shukla, R., & Kesharwani, P. (2021). Nanocarrier mediated autophagy: An emerging trend for cancer therapy. Process Biochemistry, 109, 198–206.

    Article  CAS  Google Scholar 

  • Solimando, A. G., Desantis, V., & Ribatti, D. (2022). Mast cells and interleukins. International Journal of Molecular Sciences, 23(22), 14004.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Song, S., Tan, J., Miao, Y., Li, M., & Zhang, Q. (2017). Crosstalk of autophagy and apoptosis: Involvement of the dual role of autophagy under ER stress. Journal of Cellular Physiology, 232(11), 2977–2984.

    Article  CAS  PubMed  Google Scholar 

  • Suzuki, Y., Yoshimaru, T., Inoue, T., Niide, O., & Ra, C. (2005). Role of oxidants in mast cell activation. Mast Cells in Allergic Diseases, 87, 32–42.

    Article  CAS  Google Scholar 

  • Theoharides, T. C., Perlman, A. I., Twahir, A., & Kempuraj, D. (2023). Mast cell activation: Beyond histamine and tryptase. Expert Review of Clinical Immunology, 19(6), 639–654.

    Article  CAS  PubMed  Google Scholar 

  • Tungkum, W. (2023). Regulating the trafficking of the unfolded protein response (UPR) reporter ATF6. University of Glasgow.

  • Vignali, D. A. A., & Kuchroo, V. K. (2012). IL-12 family cytokines: Immunological playmakers. Nature Immunology, 13(8), 722–728.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Waheed, G., Ramadan, G., & Mohammed, H. A. (2021). Sodium R-lipoate and enzymatically-modified isoquercitrin suppressed IgE-independent anaphylactic reactions and stress-induced gastric ulceration in mice. International Immunopharmacology, 97, 107735.

    Article  CAS  PubMed  Google Scholar 

  • Wan, X., Zhang, Y., Tang, H., Li, M., Jiang, T., He, J., Bao, C., Wang, J., Song, Y., Xiao, P., & Liu, Y. (2022). IL-27 signaling negatively regulates FcÉ›RI-mediated mast cell activation and allergic response. Journal of Leukocyte Biology, 112(3), 411–424.

    Article  CAS  PubMed  Google Scholar 

  • Wang, Y. F., Ma, R. X., Zou, B., Li, J., Yao, Y., & Li, J. (2023). Endoplasmic reticulum stress regulates autophagic response that is involved in Saikosaponin a-induced liver cell damage. Toxicology in Vitro, 88, 105534.

    Article  CAS  PubMed  Google Scholar 

  • West, P. W., Bahri, R., Garcia-Rodriguez, K. M., Sweetland, G., Wileman, G., Shah, R., Montero, A., Rapley, L., & Bulfone-Paus, S. (2021). Interleukin-33 amplifies human mast cell activities induced by complement anaphylatoxins. Frontiers in Immunology, 11, 615236.

    Article  PubMed  PubMed Central  Google Scholar 

  • Wu, M. X., Wang, S. H., Xie, Y., Chen, Z. T., Guo, Q., Yuan, W. L., Guan, C., Xu, C. Z., Huang, Y. N., Wang, J. F., & Zhang, H. F. (2021). Interleukin-33 alleviates diabetic cardiomyopathy through regulation of endoplasmic reticulum stress and autophagy via insulin-like growth factor-binding protein 3. Journal of Cellular Physiology, 236(6), 4403–4419.

    Article  CAS  PubMed  Google Scholar 

  • Yin, H., Zhang, S., Sun, Y., Li, S., Ning, Y., Dong, Y., Shang, Y., & Bai, C. (2017). MicroRNA-34/449 targets IGFBP-3 and attenuates airway remodeling by suppressing Nur77-mediated autophagy. Cell Death & Disease, 8(8), e2998–e2998.

    Article  CAS  Google Scholar 

  • Yu, X., Sun, X., Zhao, M., Hou, Y., Li, J., Yu, J., & Hou, Y. (2018). Propofol attenuates myocardial ischemia reperfusion injury partly through inhibition of resident cardiac mast cell activation. International Immunopharmacology, 54, 267–274.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

All authors acknowledge Research Laboratory Unit of Immunology, Assiut University, Faculty of Science for supporting with severa biochemical assessments employed in this study.

Funding

The present work did not receive specific funds or grants from any agencies either in the public or not-for-profit sectors.

Author information

Authors and Affiliations

  1. Zoology Department, Faculty of Women for Arts, Science and Education, Ain Shams University, Asmaa Fahmy St., Heliopolis, Cairo, 11757, Egypt

    Nawal Zakaria Haggag, Nashwa Ahmed El-Shinnawy & Sahar Sobhy Abd-Elhalem

  2. Laboratory of Immunology and Molecular Physiology, Zoology Department, Faculty of Science, Assiut University, Assiut, 71516, Egypt

    Gamal Badr

  3. Department of Biological and Geological Sciences, Faculty of Education, Ain Shams University, Cairo, 11566, Egypt

    Hany N. Yousef

Authors
  1. Nawal Zakaria Haggag
    View author publications

    Search author on:PubMed Google Scholar

  2. Nashwa Ahmed El-Shinnawy
    View author publications

    Search author on:PubMed Google Scholar

  3. Gamal Badr
    View author publications

    Search author on:PubMed Google Scholar

  4. Hany N. Yousef
    View author publications

    Search author on:PubMed Google Scholar

  5. Sahar Sobhy Abd-Elhalem
    View author publications

    Search author on:PubMed Google Scholar

Contributions

NZH, NAF and SSA: Conceptualization, Methodology, Validation, Formal analysis, Investigation, Writing-Original Draft, Funding acquisition, Writing-Review & Editing; GB: Methodology, Validation, Funding acquisition, Writing-Review and Supervision; HNY: Software, Funding acquisition.

Corresponding author

Correspondence to Nawal Zakaria Haggag.

Ethics declarations

Ethics and consent to participate

The protocol and animal procedures were followed the Animal Research: Reporting of In Vivo Experiments (ARRIVE) guidelines and Animals approved by the ethics committee of Faculty of Women, Ain Shams University (Code: Sci1332403002).

Consent for publication

All authors authorize the publication of this paper in this journal.

Competing interests

The authors declare that they have no any competing interests.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Haggag, N.Z., El-Shinnawy, N.A., Badr, G. et al. IL-33 triggers lung autophagy in anaphylaxis mice models. JoBAZ 85, 55 (2024). https://doi.org/10.1186/s41936-024-00410-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1186/s41936-024-00410-9

Keywords