PLK1 protects intestinal barrier function during sepsis by targeting mitochondrial dynamics through TANK-NF-κB signalling

Cao, Ying-Ya; Zhang, Yuan; Gerile, Wuyun; Guo, Yan; Wu, Li-Na; Wu, Li-Li; Song, Kai; Lu, Wei-Hua; Yu, Jian-Bo

doi:10.1186/s10020-022-00597-z

PLK1 protects intestinal barrier function during sepsis by targeting mitochondrial dynamics through TANK-NF-κB signalling

Research Article
Open access
Published: 29 December 2022

Volume 28, article number 163, (2022)
Cite this article

Download PDF

You have full access to this open access article

Molecular Medicine Aims and scope Submit manuscript

PLK1 protects intestinal barrier function during sepsis by targeting mitochondrial dynamics through TANK-NF-κB signalling

Download PDF

Ying-Ya Cao^1,2^na1,
Yuan Zhang¹^na1,
Wuyun Gerile¹^na1,
Yan Guo¹,
Li-Na Wu¹,
Li-Li Wu¹,
Kai Song¹,
Wei-Hua Lu² &
…
Jian-Bo Yu ORCID: orcid.org/0000-0002-8674-4435¹

2381 Accesses
3 Citations
Explore all metrics

Abstract

Background

Intestinal barrier integrity in the pathogenesis of sepsis is critical. Despite an abundance of evidence, the molecular mechanism of the intestinal barrier in sepsis pathology remains unclear. Here, we report a protective role of polo-like kinase 1 (PLK1) in intestinal barrier integrity during sepsis.

Methods

Mice with PLK1 overexpression (CAG-PLK1 mice) or PLK1 inhibition (BI2536-treated mice) underwent caecal ligation and puncture (CLP) to establish a sepsis model. The intestinal barrier function, apoptosis in the intestinal epithelium, mitochondrial function and NF-κB signalling activity were evaluated. To suppress the activation of NF-κB signalling, the NF-κB inhibitor PDTC, was administered. The Caco-2 cell line was chosen to establish an intestinal epithelial injury model in vitro.

Results

Sepsis destroyed intestinal barrier function, induced excessive apoptosis in the intestinal epithelium, and disrupted the balance of mitochondrial dynamics in wild-type mice. PLK1 overexpression alleviated sepsis-induced damage to the intestinal epithelium by inhibiting the activation of NF-κB signalling. PLK1 colocalized and interacted with TANK in Caco-2 cells. Transfecting Caco-2 cells with TANK-SiRNA suppressed NF-κB signalling and ameliorated mitochondrial dysfunction, apoptosis and the high permeability of cells induced by lipopolysaccharide (LPS). Furthermore, TANK overexpression impaired the protective effect of PLK1 on LPS-induced injuries in Caco-2 cells.

Conclusion

Our findings reveal that the PLK1/TANK/NF-κB axis plays a crucial role in sepsis-induced intestinal barrier dysfunction by regulating mitochondrial dynamics and apoptosis in the intestinal epithelium and might be a potential therapeutic target in the clinic.

PLK1 protects against sepsis-induced intestinal barrier dysfunction

Article Open access 18 January 2018

A Novel Role for Programmed Cell Death Receptor Ligand-1 in Sepsis-Induced Intestinal Dysfunction

Article Open access 25 October 2016

mTOR signaling pathway regulation HIF-1 α effects on LPS induced intestinal mucosal epithelial model damage

Article Open access 23 April 2024

Introduction

Sepsis develops due to a dysregulated host response to infection and can lead to life-threatening organ dysfunction (Evans et al. 2021). Of note, this syndrome is a challenge for public health for its increasing morbidity and approximately 30% mortality (Vincent et al. 2019; Bauer et al. 2020). The intestine is vulnerable during sepsis presented as disrupted mucus integrity and increased mucosal permeability, thus allows the transport of toxic mediators to the blood circulation and leads to distant organ dysfunction (Mittal et al. 2014; Klingensmith et al. 2016).

The intestinal mucosa consists of a single-layered epithelium that is highly dynamic and precisely regulated (Peterson et al. 2014; Odenwald et al. 2017). The balance between proliferation and apoptosis in the epithelium maintains intestinal homeostasis. Excessive apoptosis in the intestinal epithelium can damage epithelial integrity, resulting in intestinal hyperpermeability (Yoseph et al. 2016). As energy-generating organelles, mitochondria are also involved in regulating apoptosis, calcium homeostasis, and oxidative metabolism (Nunnari et al. 2012). Mitochondria are dynamic organelles that undergo fusion and fission, which not only affects their morphology but is also linked with their function. During sepsis, mitochondria undergo excessive fission and insufficient fusion, which damages mitochondrial quality and function, causing cellular dysfunction (Parikh et al. 2015; Chan 2020). Treating the imbalance in mitochondrial fusion and fission will improve therapeutic options for sepsis.

Sepsis is a series of systemic inflammatory responses accompanied by the activation of nuclear factor-κB (NF-κB) signalling. The role of NF-κB signalling in sepsis is controversial for its activation contributes to inflammatory injury in tissues whereas it also mediates host defence and other protective cellular responses (Stone et al. 2017; Ren et al. 2020; Yang et al. 2020; **s of the mice

Full size table

Enzyme-linked immunosorbent assay (ELISA)

At 24 h after the surgery, the mice were anaesthetized, and blood was collected from the heart. Then, the mice were euthanized, and the blood was centrifuged at 3000×g for 15 min at 4 °C. The serum was collected, and the indicated parameters were analysed by ELISA kits (Cloud-clone, Wuhan, China) according to the manufacturer’s instructions.

Haematoxylin and eosin (H&E) staining

Small intestine samples were collected 24 h after CLP and fixed in 10% paraformaldehyde overnight. The samples were then embedded in paraffin and sliced into 5 μm-thick sections. For pathological examinations, the sections were stained with H&E as previously described (Cao et al. 2018). The severity of mucosal injury was assessed with Chiu’s scoring system as previously described (Chiu et al. 1970).

Terminal deoxynucleotidyl transferase (TdT)-mediated dUDP-biotin nick end labelling (TUNEL) staining

To examine apoptotic cells in the intestinal epithelium, a TUNEL assay was performed as previously described (Cao et al. 2018). The tissue sections were incubated in a permeabilization solution, stained with the reaction mixture and subsequently incubated with DAPI. TUNEL-positive cells were characterized by dark brown staining of the nucleus and nuclear membrane.

Cell culture and treatments

The human intestinal Caco-2 cell line was purchased from Cellcook Biotech (Guangzhou, China) and cultured in DMEM supplemented with 10% foetal bovine serum (FBS) (Invitrogen, San Diego, CA, USA), 100 U/ml penicillin and 100 mg/ml streptomycin at 37 °C in a humidified atmosphere of 5% CO₂. When the cells reached 70–80% confluence, the cells were exposed to various treatments according to various objectives.

Transfection with plasmids and siRNA

The plasmids pCDNA3.1-PLK1, pCDNA3.1-TANK and Si-TANK (5’- GAACUAUGAGCAGAGAAUATT-3’) were constructed by Keygen Biotech (Nan**g, China), along with the corresponding controls. Caco-2 cells were cultured in 6-well plates, and when they reached 60–70% confluence, the original media was replaced with serum-free Opti-MEM. Subsequently, the plasmids or siRNA was transfected into the cells using Lipofectamine 3000 (Invitrogen, Carlsbad, CA, USA) following the instructions.

Apoptosis analysis

Apoptotic cells were double-labelled with Annexin V–APC and 7-AAD using an Annexin V-APC/7-AAD kit (KGA1024, Keygen Biotech, China) and analysed with a BDTM LSRΙΙ flow cytometer (BD Biosciences). Annexin V-positive cells were counted and defined as apoptotic cells.

Measurement of reactive oxygen species (ROS) production

To measure ROS generation, tissue slides or cells were incubated with 10 µM DCFH-DA (KGT010-1, Keygen Biotech, China), which is a fluorescent ROS probe at 37 °C for 20 min. Then, the cells were washed with phosphate-buffered saline (PBS) 3 times, and the nuclei were stained with Hoechst. All procedures were performed in the dark, and the samples were observed by a laser scanning confocal microscope (Olympus, Tokyo, Japan).

Measurement of mitochondrial membrane potential (MMP)

MMP was measured using a JC-1 MMP assay kit (KGA602, Keygen Biotech, China). Fluorescence images were acquired using a fluorescence microscope (Olympus, Tokyo, Japan) with an excitation wavelength of 488 nm and emission wavelengths of 529 and 590 nm for JC-1 monomers and aggregates, respectively. The loss of MMP was reflected by the ratio of aggregates (red fluorescence) to monomers (green fluorescence).

Measurement of transepithelial electrical resistance (TEER)

TEER was measured as described previously (Zheng et al. 2020). Briefly, Caco-2 cells were seeded on the upper layer of a Transwell 6-well plate (0.4 μm, Corning, America), and different treatments were added according to the experimental design. Then, an epithelial volt–ohm metre with a chopstick electrode (World Precision Inc, America) was used to measure the TEER every 1 h.

Immunoprecipitation assay

To examine the endogenous interaction between PLK1 and TANK, Caco-2 cells were washed with ice-cold PBS and lysed in IP lysis buffer (Beyotime, China) supplemented with PMSF (Beyotime, China). The lysates were then centrifuged and incubated with Protein A/G agarose beads (Santa Cruz, CA, USA), after which the beads were cleared by centrifugation. Lysates and immunoprecipitates were incubated with the indicated primary antibodies and the appropriate secondary antibodies, followed by separation by SDS–PAGE and detection.

Transmission electron microscopy (TEM)

TEM was used to evaluate the ultrastructure of the intestine and cells according to a previous study (Shi et al. 2021). The small intestine or cells were fixed in 2.5% glutaraldehyde for 3 h and treated with 1% osmium tetroxide for 1 h. After being dehydrated in a graded ethanol series, the tissues were embedded in epoxy resin, sliced with an ultramicrotome, and subsequently stained with uranyl acetate and lead citrate. Finally, the samples were observed and analysed under a transmission electron microscope.

Immunofluorescence staining

For immunofluorescence staining, intestine tissue slices and cells were fixed with 4% paraformaldehyde for 10 min, washed with PBS, incubated with primary antibodies overnight at 4 °C, incubated with secondary antibodies at room temperature in the dark, washed with PBS three times and imaged using a fluorescence microscope (Olympus, Tokyo, Japan).

Cell lysis and protein extraction

The small intestinal epithelium was isolated as previously described (Peuker et al. 2016), and then total proteins were extracted with a protein extraction kit (Beyotime, China). To extract total proteins from the cells, the harvested cells were washed with cold PBS and lysed with lysis buffer containing protease and phosphatase inhibitor cocktails. A nuclear extraction kit (Sigma–Aldrich, MO, USA) was used for nuclear-cytoplasmic fractionation. The protein concentration was quantified with a BCA protein assay kit (Beyotime, China).

Immunoblotting

To determine the levels of the indicated proteins, equal amounts of protein were separated by SDS–PAGE and transferred to PVDF membranes (Millipore, Bedford, MA, USA). Subsequently, the membranes were blocked with 5% milk in Tris-buffered saline-Tween 20 at room temperature for 1 h and then probed with the indicated primary antibodies overnight at 4 °C. After being washed with PBS three times, the membranes were incubated with the corresponding secondary antibodies. Finally, the membranes were analysed using the super ECL detection reagent (Tanon, Guangzhou, China).

Statistical analysis

Prism 5.0 (GraphPad Software Inc, San Diego, CA, USA) was used for data analysis. Quantitative data are presented as the mean ± SD, and the data were statistically evaluated using Student’s t tests or one-way ANOVA. A value of p < 0.05 was considered statistically significant.

Results

Sepsis induces intestinal injury and reduces PLK1 expression in the intestinal epithelium

To confirm the effect of CLP on intestinal barrier function in mice, intestine slides were stained with H&E and evaluated by Chiu’s scoring. The intestinal villi in CLP mice were atrophic, and the scores were increased (Fig. 1A, B). Furthermore, the serum levels of diamine oxidase (DAO), which reflects intestinal permeability, were increased in CLP mice (Fig. 1C). The expression levels of ZO-1 and Occludin, which are two tight junction proteins in the intestinal epithelium, were markedly decreased in CLP mice (Fig. 1D). The function of the intestinal barrier largely depends on the balance in proliferation and apoptosis in the intestinal epithelium. Next, apoptosis in the intestinal epithelium was measured by TUNEL staining. CLP mice had elevated numbers of positively stained cells (Fig. 1E), along with increased expression of the proapoptotic proteins cleaved-Caspase3 and Bax and decreased level of antiapoptotic protein Bcl2 (Fig. 1F).As mitochondria coordinate cellular homoeostasis for their various roles in energy production, oxidative metabolism and apoptosis induction, mitochondrial function plays a critical role in intestinal barrier integrity (Rath et al. 2018). In this study, the production of ROS increased with decreasing MMP in CLP mice (Fig. 1G, H). The mitochondrial structure observed by electron microscopy shown that the mitochondrial ultrastructure was deformed and intact cristae were lost in the intestinal tissue of CLP mice (Fig. 1I). Moreover, Mitochondria are dynamic organelles, and the balance between mitochondrial fusion and fission is crucial for their function. We then measured the expression of related mitochondrial fusion/fission proteins in intestinal tissues. Compared with those in the sham group, the levels of mitochondrial fusion proteins (Mfn1, Mfn2, OPA1) were reduced, while the levels of mitochondrial fission proteins (Drp1, Fis1) were enhanced in mice with CLP (Fig. 1J), indicating a shift towards fission. Though our previous study found the decreased expression of PLK1 in sepsis (Cao et al. 2018, 2021), we also examined the level of PLK1 in intestinal epithelium during sepsis. Immunofluorescence staining and immunoblotting both showed lower expression of PLK1 in intestinal tissues in CLP mice (Fig. 1K, L).

PLK1 overexpression ameliorates sepsis-induced intestinal injury

To further explore the role of PLK1 in sepsis-induced intestinal barrier dysfunction, CAG-PLK1 mice were constructed and subjected to CLP. Compared with that in the control groups, the integrity of the intestinal barrier in septic CAG-PLK1 mice was ameliorated, and these mice had lower Chiu’s scores (Fig. 2A), decreased serum DAO levels (Fig. 2B) and increased expression of tight junction proteins (ZO-1 and Occludin) (Fig. 2C). Furthermore, the role of PLK1 in enterocyte apoptosis and mitochondrial function was also examined. Compared with that of control mice, the intestinal epithelia of septic CAG-PLK1 mice exhibited fewer TUNEL-positive cells, lower expression of cleaved-Caspase3 and Bax, and higher level of Bcl2 (Fig. 2D, E). Moreover, the damage to mitochondrial ultrastructure was ameliorated (Fig. 2F), ROS production was decreased (Fig. 2G), the MMP increased (Fig. 2H), the expression of mitochondrial fusion proteins (Mfn1, Mfn2, OPA1) increased, and the levels of mitochondrial fission proteins (Drp1, Fis1) decreased (Fig. 2I).To further verify the protective effect of PLK1, the selective PLK1 inhibitor BI2536 was intraperitoneally injected 24 h before CLP, and intestinal barrier function was assessed as previously described. The effects of the different treatments on PLK1 expression were also examined (Additional file 1: Fig. S1A, B). The results showed that the integrity of the intestinal barrier was worsened by PLK1 inhibition, as indicated by increased Chiu scores (Additional file 1: Fig. S1C), enhanced serum DAO levels (Additional file 1: Fig. S1D) and decreased expression of tight junction proteins (ZO-1 and Occludin) (Additional file 1: Fig. S1E). Moreover, the apoptosis in the intestinal epithelium (Additional file 1: Fig. S1F, G) and the mitochondrial function (Additional file 1: Fig. S1H–K) worsened in BI2536-treated CLP mice. These results indicate the beneficial effect of PLK1 on sepsis-induced intestinal injury.

NF-κB signalling activation induces intestinal injury during sepsis

As a classic regulator of inflammation, NF-κB activity was measured during sepsis. We first investigated the localization of NF-κB (p65) in the intestinal epithelium, and the results showed that NF-κB (p65) was mainly located in the cytoplasm in sham mice, while most NF-κB (p65) translocated to the nucleus in CLP mice (Fig. 3A). Consistently, the protein level of NF-κB (p65) was increased in the nucleus and decreased in the cytoplasm in the CLP group (Fig. 3B). NF-κB activation involves the phosphorylation of IKKα/β, which phosphorylates IκBα, leading to the subsequent ubiquitination and degradation of IκBα and the translocation of NF-κB into the nucleus. The protein expression of p-IKKα/β, IKKα/β, p-IκBα, IκBα and p-P65 in the intestine was analysed by Western blotting. The results showed an upwards trend in p-IKK α/β,p-IκBα and p-p65 expression, while IκBα was decreased (Fig. 3B). These results showed activation of the IKK/IκB/NF-κB pathway in the intestine during sepsis. Subsequently, mice that underwent CLP were pre-treated with PDTC, an inhibitor of the NF-κB pathway, and the results showed that the integrity of the intestinal barrier was improved, as indicated by the restoration of the Chiu’s scores (Fig. 3C), serum DAO levels (Fig. 3D) and expression of tight junction proteins (ZO-1 and Occludin) (Fig. 3E). Moreover, PDTC pre-treatment reduced the number of apoptotic cells in intestinal epithelium (Fig. 3F), rescued the elevated expression of cleaved-Caspase3 and Bax, as well as the dropped level of Bcl2 (Fig. 3G). In addition, the impaired mitochondrial function presented as disrupted mitochondrial ultrastructure, increased ROS production, decreased MMP level, uplifted expression of mitochondrial fusion proteins (Mfn1, Mfn2, OPA1) and declined level of mitochondrial fission proteins (Drp1, Fis1) were all meliorative in CLP mice with PDTC pre-treatment (Fig. 3H–K).

PLK1 negatively regulates the activity of NF-κB signalling in the intestine during sepsis

The relationship between PLK1 and NF-κB was also investigated in this study. In the CAG-PLK1 + CLP group, enterocyte nuclear expression of p65 was decreased compared with that in the WT + CLP group (Fig. 4A, B). Moreover, the protein expression of p-IKKα/β,p-IκBα and p-p65 decreased, while IκBα protein expression increased in the CAG-PLK1 + CLP group compared to the WT + CLP group (Fig. 4B), indicating that NF-κB pathway activity declined in the CAG-PLK1 + CLP group. Consistently, in the BI2536 + CLP group, the results were reversed. The expression of p65 in the nucleus and p-IKKα/β,p-IκBα and p-p65 in the total protein fraction increased, while the expression of p65 in the cytoplasm and IκBα in the total protein fraction decreased (Fig. 4C, D), indicating excessive activation of the NF-κB pathway. These results suggest that NF-κB pathway activity is negatively regulated by PLK1.

PLK1 physically interacts with TANK and inhibits TANK expression in the intestinal epithelium

Previous studies revealed that TANK was an upstream regulator of NF-κB that interacts with PLK1 in HEK293 cells; thus, we hypothesized that PLK1 regulated NF-κB pathway activity via TANK in the intestinal epithelium. To test this hypothesis, we first performed Co-IP to examine the physical interaction of PLK1 with TANK in Caco-2 cells and found that endogenous PLK1 and TANK bound to each other in Caco-2 cells (Fig. 5A). Immunofluorescence analysis revealed the colocalization of PLK1 and TANK in the intestinal epithelium and Caco-2 cells (Fig. 5B, C). Furthermore, TANK expression was examined in the intestines of septic mice and LPS-treated Caco-2 cells, and the results showed increased expression of TANK in vivo and in vitro (Fig. 5D, E). However, the increase in TANK expression was inhibited in septic CAG-PLK1 mice (Fig. 5F, G) but was increased in BI2536-pretreated CLP mice (Fig. 5H, I), indicating that PLK1 inhibited the expression of TANK.

Inhibiting TANK blocks NF-κB signalling and rescues LPS-induced mitochondrial dysfunction, apoptosis and high permeability in Caco-2 cells

To verify the effect of TANK on NF-κB activity, Caco-2 cells were transfected with Si-TANK and then incubated with LPS. The efficiency of Si-TANK silencing in Caco-2 cells was also measured (Additional file 1: Fig. S2A). The NF-κB pathway was markedly activated in LPS-treated Caco-2 cells, as evidenced by the increased nuclear translocation of p65, elevated expression of p-IKKα/β,p-IκBα and p-p65 and decreased expression of IκBα in the total protein fraction (Fig. 6A, B). However, the nuclear expression p65, p-IKKα/β,p-IκBα, and p-p65 in the cells decreased expression and IκBα expression increased in LPS-treated Caco-2 cells transfected with Si-TANK (Fig. 6A, B), indicating the suppression of NF-κB activity. Moreover, compared with unsilenced transfected cells, Si-TANK-transfected cells that were treated with LPS exhibited recovery of mitochondrial structure (Fig. 6C), reduced production of ROS (Fig. 6D), improved levels of MMP (Fig. 6E), increased expression of mitochondrial fusion proteins (Mfn1, Mfn2, OPA1) and decreased levels of mitochondrial fission proteins (Drp1, Fis1) (Fig. 6F), indicating the recovery of impaired mitochondrial function. Moreover, with Si-TANK transfection, the percentage of LPS-induced apoptotic cells declined (Fig. 6G), the levels of cleaved-Caspase3 and Bax decreased, and the expression of Bcl2 increased (Fig. 6H). The levels of tight junction proteins (ZO-1 and Occludin) and TEER were also improved in Si-TANK transfected Caco-2 cells (Fig. 6I, J). These results suggest that TANK knockdown inhibited NF-κB activity and ameliorated mitochondrial dysfunction, apoptosis and high permeability in LPS-treated Caco-2 cells.

TANK overexpression impairs the protective effect of PLK1 on LPS-induced mitochondrial dysfunction, apoptosis and high permeability in Caco-2 cells

The protective effect of PLK1 on sepsis-induced intestinal barrier dysfunction was shown in CAG-PLK1 CLP mice. These findings were verified in vitro by transfecting Caco-2 cells with pCDNA3.1-PLK1 (a PLK1 knock-in plasmid) and incubating the cells with LPS. After pCDNA3.1-PLK1 transfection, the activity of NF-κB was suppressed, as shown by the reduced nuclear translocation of p65, decreased expression of p-IKKα/β,p-IκBα and p-p65 and elevated expression of IκBα in the total protein fraction (Fig. 7A, B). Moreover, mitochondrial structure, ROS production, MMP and the balance of mitochondrial dynamics were improved in PLK1-overexpressing Caco-2 cells (Fig. 7C–F). The percentage of apoptotic cells was reduced, the levels of cleaved-Caspase3 and Bax decreased, and the expression of Bcl2 increased (Fig. 7G, H). The permeability of cells decreased,which was associated with reduced TEER levels and increased expression of tight junction proteins (ZO-1 and Occludin) (Fig. 7I, J). Subsequently, Caco-2 cells were co-transfected with pCDNA3.1-PLK1 and pCDNA3.1-TANK (a TANK knock-in plasmid) and then treated with LPS. The transfection efficiency of pCDNA3.1-PLK1 and pCDNA3.1-TANK in Caco-2 cells were also measured (Additional file 1: Fig. S2B, C).Compared with cells that were transfected with pCDNA3.1-PLK1 alone, cells in the co-transfection group showed increased NF-κB signal activity (Fig. 7A, B), worsened mitochondrial impairments (Fig. 7C–F), increased apoptosis (Fig. 7G, H) and increased cell permeability (Fig. 7I, J), indicating that the protective effect of PLK1 was inhibited in co-transfected LPS-treated Caco-2 cells.

Discussion

In this study, we showed that PLK1 protects against sepsis-induced intestinal barrier dysfunction by improving the imbalance in mitochondrial fusion and fission and reducing apoptosis in the intestinal epithelium. Moreover, our findings demonstrated that the protective effect of PLK1 relies on its negative regulation of NF-κB signalling. Further examination revealed that PLK1 interacts with TANK, an upstream regulator of NF-κB, and inhibits the expression of TANK to regulate NF-κB signalling (Fig. 8). Thus, the PLK1/TANK/NF-κB axis was shown to play a crucial role in sepsis-induced intestinal barrier dysfunction and might be a potential therapeutic target in the clinic.

Mitochondria are crucial organelles in sepsis because their dysfunction can lead to loss of function in different organs, such as the lung, kidney and heart (Shi et al. 2019; Li et al. 2021; van der Slikke et al. 2021). The present study revealed the sepsis-induced mitochondrial damage was characterized by increased ROS, decreased MMP and disruption of mitochondrial dynamic balance in the intestinal epithelium. Mitochondrial dynamic balance depends on fission and fusion, which are mediated by conserved dynamin-related GTPase proteins including the fusion proteins optic atrophy 1 (OPA1), mitofusin1 (Mfn1) and mitofusin2 (Mfn2), as well as the fission protein dynamin-related protein 1 (Drp1) and its receptor mitochondrial fission protein 1 (Fis1) (Hoppins et al. 2007; Whitley et al. 2019). Our results showed that the expression of Drp1 and Fis1 was increased, while OPA1, Mfn1 and Mfn2 were decreased in the intestinal epithelium during sepsis, indicating increased mitochondrial fission and insufficient mitochondrial fusion.

Excessive fission and deficient fusion in mitochondria during sepsis result in the accumulation of mitochondrial fragmentation, which induces the production of ROS and permeabilizes the outer membrane, damaging mitochondrial function (Chan 2012).Furthermore, impaired mitochondria release cytochrome c, an essential component of the respiratory chain, into the cytosol to trigger apoptosis (Borutaite 2010; Kalpage et al. 2019). The B-cell lymphoma 2 (Bcl2) family mediates cytochrome c release, and cytochrome c in the cytosol can bind to apoptotic protease factor 1 (Apaf1), forming the apoptosome complex, activating caspase 3 and caspase 9, and resulting in apoptotic features such as DNA fragmentation and chromatin condensation (Shakeri et al. 2017). This study also verified excessive apoptosis in the intestinal epithelium, which was characterized by elevated expression of Bax and cleaved caspase 3, decreased Bcl2 expression and an increase in TUNEL-positive cells, which was accompanied by mitochondrial dysfunction during sepsis. Because an efficient intestinal barrier relies on structural integrity that is maintained by properly regulated rates of epithelial proliferation and death, dysregulated apoptosis in the intestinal epithelium can damage the tight junctions of cells, increase the permeability of the intestine and result in intestinal barrier dysfunction.

NF-κB signalling is known to play a pivotal role in inflammation by mediating the expression of inflammatory cytokines and chemokines (Lawrence 2009). The NF-κB family is composed of five members known as RelA/p65, RelB, c-Rel, NF-κB1 (p105/p50) and NF-κB2 (p100/p52), among which p50/p65 is the most representative dimer (Napetschnig et al. 2013). One sign of NF-κB signalling activation is the translocation of dimers from the cytoplasm into the nucleus. In the canonical pathway, NF-κB dimers are located in the cytoplasm bound to IκBα, an inhibitory protein in the IκB family. In response to stimulation, IκBα is phosphorylated by the IκB kinase (IKK) complex and degraded, and NF-κB dimers are released and translocate to the nucleus (Oeckinghaus et al. 2009). NF-κB signalling controls mitochondrial dynamics in various diseases. In insulin-treated cardiomyocytes, NF-κB regulates OPA1 expression and mitochondrial fusion (Parra et al. 2014). In fuel-oversupplied muscle cells, activation of the IKK–NF-κB axis promotes the expression of Drp1 and suppresses MFN2 levels, thus inducing mitochondrial fragmentation (Nisr et al. 2019). Incremental mitochondrial fragmentation can in turn increase NF-κB activity by phosphorylating IKK and IκBα through ROS accumulation (Huang et al. 2016). Similarly, the present results demonstrated that pharmacological inhibition of the NF-κB pathway not only ameliorated the changes in fission proteins (Drp1 and Fis1) and fusion proteins (OPA1, Mfn1 and Mfn2) but also improved mitochondrial function by rescuing MMP and ameliorating ROS production in the intestinal epithelium during sepsis.

TANK was identified as a TRAF-binding protein that possesses opposite regulatory properties in innate immune activation (Rothe et al. 1996). Its effect on the NF-κB pathway is also controversial because TANK can activate NF-κB signalling in cells expressing TRAF2, while TANK inhibits NF-κB activation by interacting with TRAFs (Cheng et al. 1996; Maruyama et al. 2012). Because the expression of TANK differs with different stimuli (Wang et al. 2015; Feng et al. 2018), we examined the level of TANK in the intestinal epithelium and verified that TANK expression was markedly increased in vitro and in vivo during sepsis. Furthermore, the relationship between TANK and the NF-κB pathway was examined in the intestinal epithelium by transfecting Caco-2 cells with Si-TANK. The results showed that TANK knockdown inhibited the activity of the IKK–NF-κB axis and ameliorated LPS-induced apoptosis and mitochondrial dysfunction, suggesting that TANK acts as a positive regulator of NF-κB signalling in the intestinal epithelium during sepsis.

PLK1 was shown to partly rescue LPS-induced apoptosis in vitro (Cao et al. 2018, 2021). In this study, inducible PLK1 knock-in mice were subjected to CLP, and the results showed the amelioration of mitochondrial dysfunction and apoptosis in the intestinal epithelium and improvements in intestinal permeability compared with the controls. Considering that the serine/threonine protein kinase PLK1 plays a critical role in mitosis and that mitochondria supply fuel during the cell cycle (Salazar-Roa et al. 2017), it is not surprising that a specific link exists between PLK1 and mitochondrial function. Mitochondrial structure is regulated by cyclin-dependent kinase (CDK) and the anaphase-promoting complex/cyclosome (APC/C), which promote mitotic entry and sister chromatid separation during cytokinesis (Mishra et al. 2014). Overexpression of PLK1, an upstream regulator of the cyclin B1/Cdk1 complex, could restore the cell cycle, which was disrupted by Fis1 depletion (Lee et al. 2014). The mitotic kinase Aurora A has also been reported to regulate Drp1 function by phosphorylating RalA, inducing mitochondrial fission and leading to improper segregation of mitochondria (Kashatus et al. 2011; Park et al. 2012). Our results revealed that inhibiting PLK1 with BI2536, an ATP binding domain inhibitor of PLK1, could damage mitochondrial dynamics in CLP mice, increasing the expression of fission proteins (Drp1 and Fis1) and decreasing the expression of fusion proteins (OPA1, Mfn1 and Mfn2).

Previous studies revealed a paradoxical relationship between PLK1 and NF-κB signalling in different types of cells (Higashimoto et al. 2008; Lin et al. 2011; Wu et al. 2019). The present findings showed that PLK1 acts as a negative regulator of NF-κB signalling by inhibiting the phosphorylation of IKKα/β and IκBα, thus inhibiting P65 translocation to the nucleus. Further examination identified the physical interaction and colocalization of PLK1 and TANK in the intestinal epithelium, which provides structural evidence that PLK1 controls the activity of NF-κB signalling via TANK. Research on the relationship between PLK1 and TANK is limited. Zhang reported that PLK1 was recruited to the IKK complex through TANK in HEK293 cells (Higashimoto et al. 2008; Lin et al. 2011; Wu et al. 2019). In this study, we found that the expression of TANK was suppressed in septic PLK1 knock-in mice. TANK overexpression abolished the protective effect of PLK1 on LPS-induced mitochondrial dysfunction, apoptosis and hyperpermeability in the intestinal epithelium by activating NF-κB signalling. Therefore, we hypothesize that PLK1 negatively regulates NF-κB signalling by binding and inhibiting TANK in the intestinal epithelium during sepsis.

In conclusion, our work demonstrates that PLK1 protects against sepsis-induced intestinal barrier dysfunction via TANK/NF-κB signalling by rescuing mitochondrial dysfunction and inhibiting apoptosis in the intestinal epithelium. The PLK1/TANK/NF-κB pathway might serve as a novel therapeutic target for the treatment or prevention of intestinal dysfunction. Further study should be paid attention on the application of PLK1 as the diagnostic or prognostic indicator of intestinal dysfunction in clinic.

Data availability

All data needed to evaluate the conclusions in the paper are present in the paper and/or the Additional Materials. Additional data related to this paper may be requested from the authors.

Abbreviations

PLK1:: Polo-like kinase 1
NF-κB:: Nuclear factor-κB
TANK:: TRAF family member-associated NF-κB activator
TRAF:: TNF receptor associated factor
CLP:: Caecal ligation and puncture
LPS:: Lipopolysaccharide
PDTC:: Pyrrolidinedithiocarbamate ammonium
WT:: Wild-type
SPF:: Specific pathogen-free
ELISA:: Enzyme-linked immunosorbent assay
HE staining:: Haematoxylin and eosin staining
TUNEL:: Terminal deoxynucleotidyl transferase (TdT)-mediated dUDP-biotin nick end labelling
FBS:: Foetal bovine serum
ROS:: Reactive oxygen species
MMP:: Mitochondrial membrane potential
TEER:: Transepithelial electrical resistance
TEM:: Transmission electron microscopy
DAO:: Diamine oxidase
IKK:: IκB kinase
Bcl2:: B-cell lymphoma 2
Bax:: BCL2-Associated X
CDK:: Cyclin-dependent kinase
APC/C:: Anaphase-promoting complex/cyclosome

References

Bauer M, Gerlach H, Vogelmann T, Preissing F, Stiefel J, Adam D. Mortality in sepsis and septic shock in Europe, North America and Australia between 2009 and 2019- results from a systematic review and meta-analysis. Crit Care. 2020;24:239.
Article Google Scholar
Borutaite V. Mitochondria as decision-makers in cell death. Environ Mol Mutagen. 2010;51:406–16.
Google Scholar
Cao Y, Chen Q, Wang Z, Yu T, Wu J, Jiang X, et al. PLK1 protects against sepsis-induced intestinal barrier dysfunction. Sci Rep. 2018;8:1055.
Article Google Scholar
Cao YY, Wang Z, Lu LM, Xu ZX, Li JJ, Jiang XG, et al. Polo-like kinase 1 protects intestinal epithelial cells from apoptosis during sepsis via the nuclear factor-kappaB pathway. Chin Med J (engl). 2020;133:1886–8.
Article Google Scholar
Cao YY, Wang Z, Yu T, Zhang Y, Wang ZH, Lu ZM, et al. Sepsis induces muscle atrophy by inhibiting proliferation and promoting apoptosis via PLK1-AKT signalling. J Cell Mol Med. 2021;25:9724–39.
Article Google Scholar
Chan DC. Fusion and fission: interlinked processes critical for mitochondrial health. Annu Rev Genet. 2012;46:265–87.
Article Google Scholar
Chan DC. Mitochondrial dynamics and its involvement in disease. Annu Rev Pathol. 2020;15:235–59.
Article Google Scholar
Cheng G, Baltimore D. TANK, a co-inducer with TRAF2 of TNF- and CD 40L-mediated NF-kappaB activation. Genes Dev. 1996;10:963–73.
Article Google Scholar
Chiu CJ, McArdle AH, Brown R, Scott HJ, Gurd FN. Intestinal mucosal lesion in low-flow states. I. A morphological, hemodynamic, and metabolic reappraisal. Arch Surg. 1970;101:478–83.
Article Google Scholar
Ciardo D, Haccard O, Narassimprakash H, Chiodelli V, Goldar A, Marheineke K. Polo-like kinase 1 (Plk1) is a positive regulator of DNA replication in the Xenopus in vitro system. Cell Cycle. 2020;19(14):1817–32.
Article Google Scholar
Evans L, Rhodes A, Alhazzani W, Antonelli M, Coopersmith CM, French C, et al. Surviving sepsis campaign: international guidelines for management of sepsis and septic shock 2021. Intensive Care Med. 2021;47:1181–247.
Article Google Scholar
Feng C, Zhang Y, Li J, Liu J, Wu H, **ao J, et al. Molecular cloning and characterization of TANK of black carp Mylopharyngodon piceus. Fish Shellfish Immunol. 2018;81:113–20.
Article Google Scholar
Gutteridge RE, Ndiaye MA, Liu X, Ahmad N. Plk1 inhibitors in cancer therapy: from laboratory to clinics. Mol Cancer Ther. 2016;15:1427–35.
Article Google Scholar
Higashimoto T, Chan N, Lee YK, Zandi E. Regulation of I (kappa)B kinase complex by phosphorylation of (gamma)-binding domain of I (kappa)B kinase (beta) by Polo-like kinase 1. J Biol Chem. 2008;283:35354–67.
Article Google Scholar
Hoppins S, Lackner L, Nunnari J. The machines that divide and fuse mitochondria. Annu Rev Biochem. 2007;76:751–80.
Article Google Scholar
Huang Q, Zhan L, Cao H, Li J, Lyu Y, Guo X, et al. Increased mitochondrial fission promotes autophagy and hepatocellular carcinoma cell survival through the ROS-modulated coordinated regulation of the NFKB and TP53 pathways. Autophagy. 2016;12:999–1014.
Article Google Scholar
Huang W, Man Y, Gao C, Zhou L, Gu J, Xu H, et al. Short-chain fatty acids ameliorate diabetic nephropathy via GPR43-mediated inhibition of oxidative stress and NF-kappaB Signaling. Oxid Med Cell Longev. 2020;2020:4074832.
Article Google Scholar
Kalpage HA, Bazylianska V, Recanati MA, Fite A, Liu J, Wan J, et al. Tissue-specific regulation of cytochrome c by post-translational modifications: respiration, the mitochondrial membrane potential, ROS, and apoptosis. FASEB J. 2019;33:1540–53.
Article Google Scholar
Kashatus DF, Lim KH, Brady DC, Pershing NL, Cox AD, Counter CM. RALA and RALBP1 regulate mitochondrial fission at mitosis. Nat Cell Biol. 2011;13:1108–15.
Article Google Scholar
Klingensmith NJ, Coopersmith CM. The gut as the motor of multiple organ dysfunction in critical illness. Crit Care Clin. 2016;32:203–12.
Article Google Scholar
Laforge M, Rodrigues V, Silvestre R, Gautier C, Weil R, Corti O, et al. NF-kappaB pathway controls mitochondrial dynamics. Cell Death Differ. 2016;23:89–98.
Article Google Scholar
Lawrence T. The nuclear factor NF-kappaB pathway in inflammation. Cold Spring Harb Perspect Biol. 2009;1: a1651.
Article Google Scholar
Lee S, Park YY, Kim SH, Nguyen OT, Yoo YS, Chan GK, et al. Human mitochondrial Fis1 links to cell cycle regulators at G2/M transition. Cell Mol Life Sci. 2014;71:711–25.
Article Google Scholar
Li Y, Feng YF, Liu XT, Li YC, Zhu HM, Sun MR, et al. Songorine promotes cardiac mitochondrial biogenesis via Nrf2 induction during sepsis. Redox Biol. 2021;38: 101771.
Article Google Scholar
Lin DC, Zhang Y, Pan QJ, Yang H, Shi ZZ, **e ZH, et al. PLK1 Is transcriptionally activated by NF-kappaB during cell detachment and enhances anoikis resistance through inhibiting beta-catenin degradation in esophageal squamous cell carcinoma. Clin Cancer Res. 2011;17:4285–95.
Article Google Scholar
Maruyama K, Kawagoe T, Kondo T, Akira S, Takeuchi O. TRAF family member-associated NF-kappaB activator (TANK) is a negative regulator of osteoclastogenesis and bone formation. J Biol Chem. 2012;287:29114–24.
Article Google Scholar
Mishra P, Chan DC. Mitochondrial dynamics and inheritance during cell division, development and disease. Nat Rev Mol Cell Biol. 2014;15:634–46.
Article Google Scholar
Mittal R, Coopersmith CM. Redefining the gut as the motor of critical illness. Trends Mol Med. 2014;20:214–23.
Article Google Scholar
Napetschnig J, Wu H. Molecular basis of NF-kappaB signaling. Annu Rev Biophys. 2013;42:443–68.
Article Google Scholar
Nisr RB, Shah DS, Ganley IG, Hundal HS. Proinflammatory NFkB signalling promotes mitochondrial dysfunction in skeletal muscle in response to cellular fuel overloading. Cell Mol Life Sci. 2019;76:4887–904.
Article Google Scholar
Nunnari J, Suomalainen A. Mitochondria: in sickness and in health. Cell. 2012;148:1145–59.
Article Google Scholar
Odenwald MA, Turner JR. The intestinal epithelial barrier: a therapeutic target? Nat Rev Gastroenterol Hepatol. 2017;14:9–21.
Article Google Scholar
Oeckinghaus A, Ghosh S. The NF-kappaB family of transcription factors and its regulation. Cold Spring Harb Perspect Biol. 2009;1: a34.
Article Google Scholar
Parikh SM, Yang Y, He L, Tang C, Zhan M, Dong Z. Mitochondrial function and disturbances in the septic kidney. Semin Nephrol. 2015;35:108–19.
Article Google Scholar
Park YY, Cho H. Mitofusin 1 is degraded at G2/M phase through ubiquitylation by MARCH5. Cell Div. 2012;7:25.
Article Google Scholar
Parra V, Verdejo HE, Iglewski M, Del CA, Troncoso R, Jones D, et al. Insulin stimulates mitochondrial fusion and function in cardiomyocytes via the Akt-mTOR-NFkappaB-Opa-1 signaling pathway. Diabetes. 2014;63:75–88.
Article Google Scholar
Peterson LW, Artis D. Intestinal epithelial cells: regulators of barrier function and immune homeostasis. Nat Rev Immunol. 2014;14:141–53.
Article Google Scholar
Peuker K, Muff S, Wang J, Kunzel S, Bosse E, Zeissig Y, et al. Epithelial calcineurin controls microbiota-dependent intestinal tumor development. Nat Med. 2016;22:506–15.
Article Google Scholar
Rath E, Moschetta A, Haller D. Mitochondrial function - gatekeeper of intestinal epithelial cell homeostasis. Nat Rev Gastroenterol Hepatol. 2018;15:497–516.
Article Google Scholar
Ren J, Su D, Li L, Cai H, Zhang M, Zhai J, et al. Anti-inflammatory effects of Aureusidin in LPS-stimulated RAW2647 macrophages via suppressing NF-kappaB and activating ROS- and MAPKs-dependent Nrf2/HO-1 signaling pathways. Toxicol Appl Pharmacol. 2020;387:114846.
Article Google Scholar
Rittirsch D, Huber-Lang MS, Flierl MA, Ward PA. Immunodesign of experimental sepsis by cecal ligation and puncture. Nat Protoc. 2009;4:31–6.
Article Google Scholar
Rothe M, **ong J, Shu HB, Williamson K, Goddard A, Goeddel DV. I-TRAF is a novel TRAF-interacting protein that regulates TRAF-mediated signal transduction. Proc Natl Acad Sci U S A. 1996;93:8241–6.
Article Google Scholar
Salazar-Roa M, Malumbres M. Fueling the cell division cycle. Trends Cell Biol. 2017;27:69–81.
Article Google Scholar
Shakeel I, Basheer N, Hasan GM, Afzal M, Hassan MI. Polo-like Kinase 1 as an emerging drug target: structure, function and therapeutic implications. J Drug Target. 2021;29:168–84.
Article Google Scholar
Shakeri R, Kheirollahi A, Davoodi J. Apaf-1: Regulation and function in cell death. Biochimie. 2017;135:111–25.
Article Google Scholar
Shi J, Yu J, Zhang Y, Wu L, Dong S, Wu L, et al. PI3K/Akt pathway-mediated HO-1 induction regulates mitochondrial quality control and attenuates endotoxin-induced acute lung injury. Lab Invest. 2019;99:1795–809.
Article Google Scholar
Shi J, Yu T, Song K, Du S, He S, Hu X, et al. Dexmedetomidine ameliorates endotoxin-induced acute lung injury in vivo and in vitro by preserving mitochondrial dynamic equilibrium through the HIF-1a/HO-1 signaling pathway. Redox Biol. 2021;41: 101954.
Article Google Scholar
Stone S, Jamison S, Yue Y, Durose W, Schmidt-Ullrich R, Lin W. NF-kappaB activation protects oligodendrocytes against inflammation. J Neurosci. 2017;37:9332–44.
Article Google Scholar
van der Slikke EC, Star BS, van Meurs M, Henning RH, Moser J, Bouma HR. Sepsis is associated with mitochondrial DNA damage and a reduced mitochondrial mass in the kidney of patients with sepsis-AKI. Crit Care. 2021;25:36.
Article Google Scholar
Vincent JL, Jones G, David S, Olariu E, Cadwell KK. Frequency and mortality of septic shock in Europe and North America: a systematic review and meta-analysis. Crit Care. 2019;23:196.
Article Google Scholar
Wang W, Huang X, **n HB, Fu M, Xue A, Wu ZH. TRAF Family Member-associated NF-kappaB Activator (TANK) Inhibits Genotoxic Nuclear Factor kappaB Activation by Facilitating Deubiquitinase USP10-dependent Deubiquitination of TRAF6 Ligase. J Biol Chem. 2015;290:13372–85.
Article Google Scholar
Whitley BN, Engelhart EA, Hoppins S. Mitochondrial dynamics and their potential as a therapeutic target. Mitochondrion. 2019;49:269–83.
Article Google Scholar
Wu K, Wang W, Chen H, Gao W, Yu C. Insulin promotes proliferation of pancreatic ductal epithelial cells by increasing expression of PLK1 through PI3K/AKT and NF-kappaB pathway. Biochem Biophys Res Commun. 2019;509:925–30.
Article Google Scholar
**ao N, Zhao Y, He W, Yao Y, Wu N, Xu M, et al. Egg yolk oils exert anti-inflammatory effect via regulating Nrf2/NF-kappaB pathway. J Ethnopharmacol. 2021;274: 114070.
Article Google Scholar
Yang K, Xu J, Fan M, Tu F, Wang X, Ha T, et al. Lactate suppresses macrophage pro-inflammatory response to LPS stimulation by inhibition of YAP and NF-kappaB Activation via GPR81-mediated signaling. Front Immunol. 2020;11: 587913.
Article Google Scholar
Yoseph BP, Klingensmith NJ, Liang Z, Breed ER, Burd EM, Mittal R, et al. Mechanisms of intestinal barrier dysfunction in sepsis. Shock. 2016;46(1):52–9.
Article Google Scholar
Zhang W, Wang J, Zhang Y, Yuan Y, Guan W, ** C, et al. The scaffold protein TANK/I-TRAF inhibits NF-kappaB activation by recruiting polo-like kinase 1. Mol Biol Cell. 2010;21(14):2500–13.
Article Google Scholar
Zheng D, Zhang J, Zhang Z, Kuang L, Zhu Y, Wu Y, et al. Endothelial microvesicles induce pulmonary vascular leakage and lung injury during sepsis. Front Cell Dev Biol. 2020;8:643.
Article Google Scholar

Download references

Acknowledgements

We express our gratitude to the staff of the Anaesthesia Laboratory & Training Centre of Wannan Medical College for their kind help during the study.

Funding

This study was supported by the National Natural Science Foundation of China (82002092 and 81803899) and the Anhui Province Natural Science Foundation (2108085MH300).

Author information

Ying-Ya Cao, Yuan Zhang and Wuyun Gerile contributed equally to this work

Authors and Affiliations

Department of Anaesthesiology and Critical Care Medicine, Tian** Nankai Hospital, Tian** Medical University, Tian**, 300100, China
Ying-Ya Cao, Yuan Zhang, Wuyun Gerile, Yan Guo, Li-Na Wu, Li-Li Wu, Kai Song & Jian-Bo Yu
Department of Critical Care Medicine, The First Affiliated Hospital of Wannan Medical College, Wuhu, 241001, Anhui, China
Ying-Ya Cao & Wei-Hua Lu

Authors

Ying-Ya Cao
View author publications
You can also search for this author in PubMed Google Scholar
Yuan Zhang
View author publications
You can also search for this author in PubMed Google Scholar
Wuyun Gerile
View author publications
You can also search for this author in PubMed Google Scholar
Yan Guo
View author publications
You can also search for this author in PubMed Google Scholar
Li-Na Wu
View author publications
You can also search for this author in PubMed Google Scholar
Li-Li Wu
View author publications
You can also search for this author in PubMed Google Scholar
Kai Song
View author publications
You can also search for this author in PubMed Google Scholar
Wei-Hua Lu
View author publications
You can also search for this author in PubMed Google Scholar
Jian-Bo Yu
View author publications
You can also search for this author in PubMed Google Scholar

Contributions

CYY and YJB design the study. CYY and ZY carried out the study. CYY and WG drafted the manuscript. GY, WLN and WLL collected important background information. SK and LWH performed the statistical analysis. All authors read and approved the final manuscript.

Corresponding author

Correspondence to Jian-Bo Yu.

Ethics declarations

Ethics approval and consent to participate

Our studies did not include human participates, human data or human tissues. All animal experiments conducted were compliant were approved by the Institutional Animal Care and Ethics Committee of Wannan Medical College (LLSC-2020–074).

Consent for publication

Not applicable.

Competing interests

The authors declare no conflicts of interest regarding this paper.

Additional information

Publisher's Note

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

Supplementary Information

Additional file 1.

The effiencey of PLK1 inhibition on sepsis-induced intestinal injury and the details of primary antibodies for Western blots.

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

Cite this article

Cao, YY., Zhang, Y., Gerile, W. et al. PLK1 protects intestinal barrier function during sepsis by targeting mitochondrial dynamics through TANK-NF-κB signalling. Mol Med 28, 163 (2022). https://doi.org/10.1186/s10020-022-00597-z

Download citation

Received: 29 October 2022
Accepted: 22 December 2022
Published: 29 December 2022
DOI: https://doi.org/10.1186/s10020-022-00597-z

PLK1 protects intestinal barrier function during sepsis by targeting mitochondrial dynamics through TANK-NF-κB signalling

Abstract

Background

Methods

Results

Conclusion

Similar content being viewed by others

PLK1 protects against sepsis-induced intestinal barrier dysfunction

A Novel Role for Programmed Cell Death Receptor Ligand-1 in Sepsis-Induced Intestinal Dysfunction

mTOR signaling pathway regulation HIF-1 α effects on LPS induced intestinal mucosal epithelial model damage

Introduction

Enzyme-linked immunosorbent assay (ELISA)

Haematoxylin and eosin (H&E) staining

Terminal deoxynucleotidyl transferase (TdT)-mediated dUDP-biotin nick end labelling (TUNEL) staining

Cell culture and treatments

Transfection with plasmids and siRNA

Apoptosis analysis

Measurement of reactive oxygen species (ROS) production

Measurement of mitochondrial membrane potential (MMP)

Measurement of transepithelial electrical resistance (TEER)

Immunoprecipitation assay

Transmission electron microscopy (TEM)

Immunofluorescence staining

Cell lysis and protein extraction

Immunoblotting

Statistical analysis

Results

Sepsis induces intestinal injury and reduces PLK1 expression in the intestinal epithelium

PLK1 overexpression ameliorates sepsis-induced intestinal injury

NF-κB signalling activation induces intestinal injury during sepsis

PLK1 negatively regulates the activity of NF-κB signalling in the intestine during sepsis

PLK1 physically interacts with TANK and inhibits TANK expression in the intestinal epithelium

Inhibiting TANK blocks NF-κB signalling and rescues LPS-induced mitochondrial dysfunction, apoptosis and high permeability in Caco-2 cells

TANK overexpression impairs the protective effect of PLK1 on LPS-induced mitochondrial dysfunction, apoptosis and high permeability in Caco-2 cells

Discussion

Data availability

Abbreviations

References

Acknowledgements

Funding

Author information

Authors and Affiliations

Contributions

Corresponding author

Ethics declarations

Ethics approval and consent to participate

Consent for publication

Competing interests

Additional information

Publisher's Note

Supplementary Information

Additional file 1.

Rights and permissions

About this article

Cite this article

Share this article

Keywords

Search

Navigation