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The preventive effects of Lactobacillus casei 03 on Escherichia coli-induced mastitis in vitro and in vivo

Journal of Inflammation volume 21, Article number: 5 (2024) Cite this article

Abstract

Background

Lactobacillus casei possesses many kinds of bioactivities, such as anti-inflammation and anti-oxidant, and has been applied to treating multiple inflammatory diseases. However, its role in mastitis prevention has remained ambiguous.

Methods

This study aimed to examine the mechanisms underlying the preventive effects of L. casei 03 against E. coli- mastitis utilizing bovine mammary epithelial cells (BMECs) and a mouse model.

Results

In vitro assays revealed pretreatment with L. casei 03 reduced the apoptotic ratio and the mRNA expression levels of IL1β, IL6 and TNFα and suppressed phosphorylation of p65, IκBα, p38, JNK and ERK in the NF-κB signaling pathway and MAPK signaling pathway. Furthermore, in vivo tests indicated that intramammary infusion of L. casei 03 relieved pathological changes, reduced the secretion of IL1β, IL6 and TNFα and MPO activity in the mouse mastitis model.

Conclusions

These data suggest that L. casei 03 exerts protective effects against E. coli-induced mastitis in vitro and in vivo and may hold promise as a novel agent for the prevention and treatment of mastitis.

Background

Mastitis is a common disease associated with dairy cattle and causes pathological alterations in the mammary gland tissue [1]. It is characterized by high economic losses because of high morbidity and impaired production [2]. Some studies revealed that mastitis in dairy cows could affect the physiological status of newborn calves during pregnancy [3], and the growth and the acquisition of immune antibodies in calves may be influenced due to drinking low-quality milk from mastitis udders [4]. Also, the pathogen can be vertically transmitted to calves via contaminated milk and colostrum, causing gastrointestinal diseases and even death [5].

Dairy cow mastitis is mainly caused by pathogen infection [6]. A recent report shows that Escherichia coli is the main pathogen of acute mastitis in dairy farming and it is generally considered an environmental pathogen that can enter the mammary gland through the teat canal under appropriate conditions [7, 8]. Invasive E. coli release endotoxins such as LPS and induce strong responses from the cows’ immune systems, which in turn activates the blood clotting system, leading to local microcirculation disturbances and tissue damage [9]. Currently, antimicrobials have been widely used to prevent and control mastitis and other bacterial diseases in intensive food animal production [10]. Reliance on antibiotics inevitably leads to the onset of antibiotic resistant bacterial strains, which threaten animal production and human health.

As a possible alternative to antibiotics, probiotics caught wide attention and have elicited numerous related research in recent years [11]. One such probiotic, Lactobacillus casei, is a commensal bacterium generally found in human and animal intestines which can enhance animal immune capacity and promote animal growth and development [12]. With the deepening of research, more and more researchers have begun to focus on the potential anti-inflammatory effects of L. casei. Zhang et al. [13] showed that L. casei Zhang might prevent experimental colitis and rapamycin-induced inflammation in the mice. Haro et al. [14] demonstrated that L. casei could modulate inflammation-coagulation interactions in an experimental model of pneumococcal pneumonia. The study by Chen et al. [2] suggested that L. plantarum KLDS 1.0344 showed good antibacterial properties and may be developed into a probiotic formulation for inflammatory disease control and prevention. These findings herald that L. casei may be an effective prevention strategy for dairy cow mastitis. However, the exact mechanism of its action remains unclear. In this study, we investigated the preventive effect of Lactobacillus casei 03 on the E. coli -induced inflammatory model and its mechanism of action in vitro and in vivo, in order to provide a theoretical basis for developing novel drugs for mastitis prevention and treatment in cows.

Materials and methods

Bacterial and cultural conditions

The Lactobacillus casei 03 strain obtained from American Type Culture Collection (ATCC393; Manassas, VA, USA) and was cultivated in de Man, Rogosa, and Sharpe (MRS) broth (Aobox, Beijing, China) under microaerobic conditions at 37℃ for 48 h. Escherichia coli O111:K58 (CVCC1450, provided by China Constitute of Veterinary Drug Centre, Beijing, China) was grown overnight in LB medium (Aobox, Beijing, China) with shaking at 37℃. Bacterial colony counts (CFU) were calculated and recorded after three generations.

Cells culture and treatment

Bovine mammary epithelial cells (BMECs) were provided by Animal Clinical Laboratory of Hebei Agricultural University (Baoding, China) and grown in Dulbecco’s modified Eagle’s medium/Ham’s F12 nutrient mixture (DMEM/F12, Gibco, Grand Island, USA) supplemented with 15% FBS (Gibco), 0.1% hydrocortisone (Sigma-Aldrich, MO, USA), 100 U/mL penicillin-streptomycin (Solarbio, Beijing, China) and 0.025 M HEPES (Solarbio) at 37℃ with 5% CO2. When cells were 70–80% confluence, they were treated as follows in six-well plates: the CON group (DMEM/F12), the ECOL group (107 CFU/mL E. coli), pretreated with L. casei 03 (104, 105 and 106 CFU/mL) for 3 h before the addition of E. coli, the LC group (106 CFU/mL L. casei 03). Eight hours after E. coli (107 CFU/mL) infection, cells were washed with PBS and collected for subsequent assays.

Cell viability assay

The effect of different doses of L. casei 03 on BMECs viability was determined using the CCK-8 assay. In brief, cells were plated in 96-well plates and cultured to approximately 80% confluence. Subsequently, the cells were treated with various concentrations of L. casei 03 (103 to 108 CFU/mL) for 3 h and CCK-8 reagent was added to each well to incubate at 37℃ for another 2 h. The absorbance of the wells was read at 450 nm on a microplate reader.

Cell immunofluorescence staining

Treated cells were stained using the Annexin V-FITC/Propidium Iodide Apoptosis Detection Kit (#C1062M, Beyotime Shanghai, China) according to the manufacturer’s instructions. Images were visualized using a fluorescence microscope. Five visual fields were selected randomly for cell counting. Annexin V-positive cells were labeled as the apoptotic cells and the apoptotic rate (%) was calculated as the number of apoptotic cells/total number of cells×100%.

qRT–PCR analysis

Total RNA extraction was performed using Ultrapure RNA extraction kit (CWBio, Beijing, China). The purity, concentration and integrity of the RNA samples were assessed by spectrophotometry and electrophoresis. Subsequently, RNA samples were reverse transcribed into cDNA using reverse transcription kit (US Everbright Inc, CA, USA) for quantitative Real-Time PCR (qRT-PCR) analyses. The reaction program was 95℃ for 2 min, 95℃ for 15 s, 58 ℃ for 30 s, and 72℃ for 30 s for 40 cycles. The expression level of target genes was normalized to GAPDH and β-actin and calculated with the 2−ΔΔCt method. The primer sequences were given in Table 1.

Table 1 Sequence of primers used in qRT-PCR
Full size table

Western blot analysis

The total protein from cells was extracted using RIPA lysis buffer (Solarbio, Beijing, China) and quantified with the BCA protein assay kit (Solarbio, Beijing, China). Protein samples (25 µg) were separated by 10% SDS-PAGE, electrotransferred onto a nitrocellulose membrane and blocked with 5% milk. The membranes were incubated against NF-κB p65 (1:1000), NF-κB phospho-p65 (1:1000), phospho-IκBα (1:500) and β-actin (1:1000) from Bioss Biotech Limited Company (Beijing, China) and antibodies against p38 (1:1000), phospho-p38 (1:1000), ERK (1:1000), phospho-ERK (1:2000), JNK (1:1000), phospho-JNK (1:1000) and IκBα (1:1000) from Cell Signaling Technology (MA, USA) at 4℃ with primary antibodies overnight and then with the corresponding secondary antibodies (1:2000, Zhongshan Golden Bridge, Beijing, China) for 1 h. Finally, the blots were stained using BCIP/NBT color development kit (Solarbio, Beijing, China) and analyzed for grayscale values using Image J software.

Animals and experiment design

SPF-grade male and female KM mice (6–8 weeks old) were purchased from Liaoning Changsheng Biotechnology Corporation (Benxi, China). After 3 days of adaptive diet feeding, male and female mice (1:3 ratio) were placed in the same cage until the female mice became pregnant and were able to drink and eat freely during the experiment. The lactating mice were randomly divided into four groups: the CON group (PBS), the ECOL group (107 CFU/100 µL), the LC + ECOL group: pretreated with L. casei 03 (106 CFU/100 µL) for 3 h before the addition of E. coli and the LC group (106 CFU/100 µL L. casei 03). Mice were anesthetized by ether and their fourth pair of nipples were sterilized with 75% ethanol. Then, the mice were injected with 100 µL bacterium or PBS (50 µL/side) into their nipples with 32G needle. After 24 h of infection, mice were sacrificed and the mammary gland tissues were collected for subsequent experiments. Animal assays were approved by the Animal Ethics Committee of Hebei Agricultural University (Protocol number 2,020,044).

Histopathologic analysis

Tissue samples were fixed in 4%-buffered paraformaldehyde solution, dehydrated with alcohol, and embedded in paraffin. Subsequently, paraffin-embedded tissues were cut into 5 μm sections, which were routinely stained with HE, followed by observation under a light microscope. Histological scores were performed according to the criteria previously described [15]. Briefly, the scores were graded on edema, inflammatory cell infiltration, hyperemia and necrosis. Scores range from 1 to 5, with higher scores indicating greater inflammation.

MPO activity determination and cytokines analysis

Mammary gland tissues were homogenized and centrifuged (2500 rpm for 10 min) to collect supernatants. The MPO activity in mammary gland tissues was detected according to the manufacturer’s instructions (Nanjing Jiancheng Bioengineering Institute, Nanjing, China). Cytokine concentrations in supernatants were measured using ELISA kits from Shanghai Enzyme-linked Biotechnology (Shanghai, China).

Statistical analysis

The data were presented as means ± standard error of the mean (SEM). One-way analysis of variance and Tukey’s or Dunnett’s T3 test were used for the comparison among groups. P < 0.05 were considered significant.

Results

Effects of L. casei 03 on the viability of BMECs

The CCK-8 assay showed that BMECs viability was not affected upon treatment with 103 to 108 CFU/mL L. casei 03 (Fig. 1). The concentrations (104, 105 and 106 CFU/mL) of L. casei 03 were accordingly selected for subsequent experiments.

Fig. 1
figure 1

Effects of L. casei 03 on the cell viability in BMECs. Cells were cultured with different concentrations of L. casei 03 (103 to 108) for 3 h by CCK-8 assay. The data were presented as the means ± SEM of five independent experiments

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Effects of L. casei 03 on BMECs apoptosis

The cell apoptosis was analyzed using Annexin-V/PI double staining. As shown in Fig. 2A, B and E. coli treatment apparently increased the rate of Annexin-V positive cells compared with that in the CON group (P < 0.05). In contrast, pretreatment with different doses of L. casei 03 significantly reduced the rate of apoptotic cells (P < 0.05).

Fig. 2
figure 2

L. casei 03 inhibited BMECs apoptosis induced by E. coli. (A) Apoptosis was evaluated by measuring Annexin/PI fluorescent staining. Green, Annexin V-positive; Red, PI-positive; DIC, ordinary light; scale bar: 200 μm. (B) The apoptotic rate was quantified by counting the green-positive cells. Groups with different letters above the bar indicate significant differences (P < 0.05). These same conventions were used in subsequent figures

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Effects of L. casei 03 on the mRNA expression of pro-inflammatory genes in E. coli-induced BMECs

The mRNA expression levels of IL1β, IL6 and TNFα were detected by qRT-PCR (Fig. 3A-C). E. coli challenge caused a significant increase the expression levels of three pro-inflammatory genes compared with those in the CON group (P < 0.05). Pretreatment with different doses of L. casei 03 inhibited the expression in levels of IL1β, IL6 and TNFα to varying degrees.

Fig. 3
figure 3

Effects of L. casei 03 on the mRNA expression of pro-inflammatory cytokines in E. coli-induced BMECs. The mRNA expression levels of IL1β, IL6 and TNFα (A-C) in E. coli-induced BMECs were detected by qRT-PCR. Values were obtained from three independent experiments

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Effects of L. casei 03 on the protein expression of the NF-κB and MAPK signaling pathways in E. coli-induced BMECs

We further evaluated the effects of L. casei 03 on the activity of the NF-κB and MAPK signaling pathways by Western Blot. As shown in Fig. 4A and B, the phosphorylation levels of p65 and IκBα in the NF-κB signaling pathway and the phosphorylation levels of p38, ERK and JNK in the MAPK signaling pathway were significantly increased in the ECOL group. However, all of them were significantly decreased after pretreatment with L. casei 03 (P < 0.05). Additionally, there were no significant changes in these protein expressions in the LC group compared with the CON group.

Fig. 4
figure 4

Effects of L. casei 03 on protein expression involving in NF-κB (A) and MAPK (B) Signaling Pathways in E. coli-induced BMECs. β-actin was used as a control. Values were presented as mean ± SEM from three independent experiments

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Effects of L. casei 03 on mammary gland tissue histopathological changes in E. coli-induced mastitis

Various degrees of redness, bleeding or swelling was seen in the mammary tissues in the ECOL and LC + ECOL groups. In parallel, there were no apparent histopathologic changes in the CON and LC groups (Fig. 5A). The histological characteristics of mice mammary tissue were assessed by H&E staining (Fig. 5B). There were no obvious inflammatory changes in mice mammary tissue from the CON and LC groups. In contrast, the mammary tissue in the ECOL group presented serious histopathological changes, such as thickened alveolar walls, edema, hyperemia and inflammatory cell infiltration. However, these pathological changes were ameliorated by L. casei 03 pretreatment. The same conclusion was obtained in the inflammation score (Fig. 5C). Pretreatment with L. casei 03 significantly reduced the score rise induced by E. coli in mice mammary tissue (P < 0.05).

Fig. 5
figure 5

A The morphology of the mammary glands from CON group (a), ECOL group (b), LC + ECOL group (c) and LC group (d). B The histological characteristics of mice mammary tissue in E. coli-induced Mastitis. Representative images from CON group (a), ECOL group (b), LC + ECOL group (c) and LC group (d). scale bar: 200 μm. C The inflammation score of representative images from each group. Scores range from 1 to 5, with higher scores indicating more severe inflammatory change. Values were presented as mean ± SEM (n = 5)

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Effects of L. casei 03 on MPO activity in mice mammary tissue

The MPO activity in mice mammary tissues from each group was shown in Fig. 6. The results showed that E. coli challenge significantly increased MPO activity in mammary tissue compared with that in the CON group (P < 0.05). However, these elevations were inhibited after pretreatment with L. casei 03.

Fig. 6
figure 6

MPO activity in the homogenate of mice mammary tissues from the control group, E. coli group, pretreatment with L. casei 03 and L. casei 03 alone group. Data were presented as mean ± SEM (n = 4)

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Effects of L. casei 03 on proinflammatory cytokine levels in mice mammary tissue

The secretion levels of IL1β, IL6 and TNFα in mice mammary tissues were detected by ELISA. As shown in Fig. 7, after E. coli treatment, the levels of IL1β, IL6 and TNFα in mammary tissue increased significantly compared with those in the CON group (P < 0.05). Pretreatment with L. casei 03 suppressed these increases by varying degrees.

Fig. 7
figure 7

The levels of IL1β (A), IL6 (B) and TNFα (C) in the homogenate of mice mammary tissues including the control group, E. coli group, and pretreatment with L. casei 03 and L. casei 03 alone groups. Data expressed as mean ± SEM (n = 4)

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Discussion

Dairy cow mastitis has always been a focus of veterinary personnel and is a major problem that hinders earnings growth in dairy farms [16]. Antibiotics, the most commonly used method to control this disease, obtain a good clinical response while also leading to the emergence of multidrug-resistant (MDR) bacteria [17]. MDR bacteria not only increase the difficulty of treating disease but brings a potential threat to human health [18]. Therefore, Therefore, seeking effective alternative antimicrobial agents for mastitis is urgent and essential. E. coli is a major pathogen responsible for clinical mastitis in dairy cows, which usually causes severe inflammatory reactions and economic losses [19, 20]. Recent studies show that some lactic acid bacteria exert direct suppressive effects on E. coli isolated from dairy cow mastitis and avoid the emergence of resistant bacteria. These properties provide an excellent rationale for alternative antibiotic strategies in mastitis [21, 22]. Therefore, we screened several strains of probiotics based on previous trials and L. casei 03 gradually entered our field of vision with its potential anti-inflammatory properties.

Mammary epithelial cells form an important component of milk synthesis in mammary tissue and are involved in the local innate immune responses [23]. When pathogens gain access to mammary tissues, many pathogens associated molecular patterns are recognized by the Toll-like receptor (TLR) of surface proteins on cells, which in turn can activate intracellular signaling pathways and promote the expression of the other inflammatory cytokines and enlarges the inflammatory response [24, 25]. It is widely accepted that overproduction of these inflammatory mediators and cytokines is highly associated with inflammatory diseases [26]. IL6, TNFα and IL1β are the common pro-inflammatory cytokines, they can play a cytotoxic role in the inflammatory process and accelerate the inflammatory processes [27]. In this study, we first used in vitro inflammatory models established with E. coli-treated BMECs to simulate the pathogenesis of mastitis. The results showed that L. casei 03 pretreatment reduced the elevated mRNA expression of pro-inflammatory genes caused by E. coli. This suggests that L. casei 03 can exert an inflammatory ameliorating effect by inhibiting the expression of pro-inflammatory factors. Many studies have shown that NF-κB and MAPK pathways play key roles in inflammation modulation [28,29,30]. We next investigated the effect of L. casei 03 on the activity of NF-κB and MAPK signaling pathways. Our results showed that L. casei 03 could inhibit the elevated phosphorylation levels of key pathway proteins in signaling pathways such as p65 and p38 caused by E. coli and exert anti-inflammatory effects via inhibiting NF-κB and MAPK activity, this matches the result of Zheng et al. [31]. Apoptosis is inextricably linked with excessive inflammatory cascades [32]. Some endotoxins in E. coli can trigger inflammatory responses and induce cell apoptosis. Meanwhile, apoptotic cells release cytokines and other inflammatory mediators, thereby exacerbating inflammation [33]. Previous studies showed that E. coli treatment increased the apoptosis rate in bovine endometrial epithelial cells, and Lactobacillus rhamnosus can inhibit this increase [34]. Similarly, our study also showed that L. casei 03 could inhibit E. coli-induced apoptosis in BMECs and had positive effects on anti-inflammation.

Based on the anti-inflammatory effects of L. casei 03 in vitro, we next investigated the protective effects of L. casei 03 in E. coli-induced mastitis in the mouse model. The characteristics of mastitis mainly manifest as mammary swelling, bleeding, inflammatory cell infiltration and acinar injury [35, 36]. Simultaneously, MPO activity in tissue is increased due to BMEC damage [37]. After injecting E. coli into mice mammary tissue, we observed a series of histopathological changes, as well as a significantly increased pro-inflammatory factor and MPO activity in mammary tissue. These inflammatory symptoms were significantly alleviated with L. casei 03 pretreatment. These results indicate that L. casei 03 has protective effects on E. coli-induced mastitis in vivo.

Notably, some academics have expressed concerns about the safety of the application of probiotics treatment in mastitis, noting that intramammary infusion with live probiotics can cause or aggravate mastitis [38]. However, some scholarly studies dispelled this concern to some extent. Martín et al. successfully isolated probiotics from healthy breast tissue to demonstrate the presence of probiotics in normal mammary tissue [39]. Zheng et al. also showed the feasibility of intramammary infusion of L. casei in mastitis treatment [31]. In this study, we investigated the effect of L. casei 03 on the viability of BMECs in vitro and selected safe doses for subsequent experiments. We subsequently set up the L. casei 03 alone treatment group and did not detect appreciable toxic effects in vitro models. This demonstrates the application safety and reliability of L. casei 03 and provides the basis for its subsequent use in mastitis treatment.

Conclusions

In conclusion, L. casei 03 could inhibit the NF-κB and MAPK signaling pathway activity and reduce the pro-inflammatory gene expression to resist the inflammatory response induced by E. coli in BMECs. The in vivo results suggested that L. casei 03 alleviated pathological changes and pro-inflammatory factors secretion in mammary tissue and exerted anti-inflammatory effects in E. coli-induced mice mastitis. The present study demonstrates that L. casei 03 has preventive effects on E. coli-induced mastitis in vitro and in vivo and this provides new insights for the prevention and treatment of dairy cow mastitis.

Data availability

All data used to support the findings in this study are included within the article. The original, full-length western blot blots are listed in the supplementary information.

Abbreviations

BMECs:

Bcovine mammary epithelial cells

FITC:

Fluorescein isothiocyanate

MPO:

Myeloperoxidase

NF-κB:

Nuclear factor kappa B

MAPK:

Mitogen-activated protein kinase

MDR:

Multidrug-resistant

TLR:

Toll-like receptor

References

  1. Li Y, Gong Q, Guo W, Kan X, Xu D, Ma H, et al. Farrerol Relieve Lipopolysaccharide (LPS)-Induced Mastitis by inhibiting AKT/NF-κB p65, ERK1/2 and P38 Signaling Pathway. Int J Mol Sci. 2018;19(6):1770.

    Article  PubMed  PubMed Central  Google Scholar 

  2. Chen Q, Wang S, Guo J, Xie Q, Evivie SE, Song Y, et al. The Protective effects of Lactobacillus plantarum KLDS 1.0344 on LPS-Induced Mastitis in vitro and in vivo. Front Immunol. 2021;12:770822.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Petzer IM, Karzis J, Lesosky M, Watermeyer JC, Badenhorst R. Host adapted intramammary infections in pregnant heifers which were co-housed and reared on fresh milk as calves. BMC Vet Res. 2013;9:49.

    Article  PubMed  PubMed Central  Google Scholar 

  4. Xi H, He D, Li D, Liu SS, Wang G, Ji Y, et al. Bacteriophage protects against Aerococcus viridans infection in a murine Mastitis Model. Front Vet Sci. 2020;7:588.

    Article  PubMed  PubMed Central  Google Scholar 

  5. Zhang D, Zhang Z, Huang C, Gao X, Wang Z, Liu Y, et al. The phylogenetic group, antimicrobial susceptibility, and virulence genes of Escherichia coli from clinical bovine mastitis. J Dairy Sci. 2018;101(1):572–80.

    Article  CAS  PubMed  Google Scholar 

  6. Guo W, Liu J, Li W, Ma H, Gong Q, Kan X, et al. Niacin alleviates dairy cow mastitis by regulating the GPR109A/AMPK/NRF2 signaling pathway. Int J Mol Sci. 2020;21(9):3321.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Gao J, Barkema HW, Zhang L, Liu G, Deng Z, Cai L, et al. Incidence of clinical mastitis and distribution of pathogens on large Chinese dairy farms. J Dairy Sci. 2017;100(6):4797–806.

    Article  CAS  PubMed  Google Scholar 

  8. Luoreng ZM, Wang XP, Mei CG, Zan LS. Expression profiling of peripheral blood miRNA using RNAseq technology in dairy cows with Escherichia coli-induced mastitis. Sci Rep. 2018;8(1):12693.

    Article  ADS  PubMed  PubMed Central  Google Scholar 

  9. Liu C, Tang X, Zhang W, Li G, Chen Y, Guo A, et al. 6-Bromoindirubin-3’-Oxime suppresses LPS-Induced inflammation via inhibition of the TLR4/NF-κB and TLR4/MAPK signaling pathways. Inflammation. 2019;42(6):2192–204.

    Article  PubMed  Google Scholar 

  10. Oliver SP, Murinda SE. Antimicrobial resistance of mastitis pathogens. Vet Clin North Am Food Anim Pract. 2012;28(2):165–85.

    Article  PubMed  Google Scholar 

  11. Wang K, Dong H, Qi Y, Pei Z, Yi S, Yang X, et al. Lactobacillus casei regulates differentiation of Th17/Treg cells to reduce intestinal inflammation in mice. Can J Vet Res. 2017;81(2):122–8.

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Nagpal R, Kumar A, Kumar M, Behare PV, Jain S, Yadav H. Probiotics, their health benefits and applications for developing healthier foods: a review. FEMS Microbiol Lett. 2012;334(1):1–15.

    Article  CAS  PubMed  Google Scholar 

  13. Zhang Y, Hou Q, Ma C, Zhao J, Xu H, Li W, et al. Lactobacillus casei protects dextran sodium sulfate- or rapamycin-induced colonic inflammation in the mouse. Eur J Nutr. 2020;59(4):1443–51.

    Article  CAS  PubMed  Google Scholar 

  14. Haro C, Villena J, Zelaya H, Alvarez S, Agüero G. Lactobacillus casei modulates the inflammation-coagulation interaction in a pneumococcal pneumonia experimental model. J Inflamm (Lond). 2009;6:28.

    Article  PubMed  Google Scholar 

  15. Elinav E, Strowig T, Kau AL, Henao-Mejia J, Thaiss CA, Booth CJ, et al. NLRP6 inflammasome regulates colonic microbial ecology and risk for colitis. Cell. 2011;145(5):745–57.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Song W, Sheng L, Chen F, Tian Y, Li L, Wang G, et al. C. Sakazakii activates AIM2 pathway accompanying with excessive ER stress response in mammalian mammary gland epithelium. Cell Stress Chaperones. 2020;25(2):223–33.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Javadmanesh A, Mohammadi E, Mousavi Z, Azghandi M, Tanhaiean A. Antibacterial effects assessment on some livestock pathogens, thermal stability and proposing a probable reason for different levels of activity of thanatin. Sci Rep. 2021;11(1):10890.

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  18. Klaper K, Hammerl JA, Rau J, Pfeifer Y, Werner G. Genome-based analysis of Klebsiella spp. Isolates from animals and Food products in Germany, 2013–2017. Pathogens. 2021;10(5):573.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Botrel MA, Haenni M, Morignat E, Sulpice P, Madec JY, Calavas D. Distribution and antimicrobial resistance of clinical and subclinical mastitis pathogens in dairy cows in Rhône-Alpes, France. Foodborne Pathog Dis. 2010;7(5):479–87.

    Article  CAS  PubMed  Google Scholar 

  20. Jerjomiceva N, Seri H, Völlger L, Wang Y, Zeitouni N, Naim HY, et al. Enrofloxacin enhances the formation of neutrophil extracellular traps in bovine granulocytes. J Innate Immun. 2014;6(5):706–12.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Pellegrino M, Berardo N, Giraudo J, Nader-Macías MEF, Bogni C. Bovine mastitis prevention: humoral and cellular response of dairy cows inoculated with lactic acid bacteria at the dry-off period. Benef Microbes. 2017;8(4):589–96.

    Article  CAS  PubMed  Google Scholar 

  22. Fukuyama K, Islam MA, Takagi M, Ikeda-Ohtsubo W, Kurata S, Aso H, et al. Evaluation of the Immunomodulatory ability of lactic acid Bacteria isolated from Feedlot Cattle against Mastitis Using a bovine mammary epithelial cells in vitro assay. Pathogens. 2020;9(5):410.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Rainard P, Riollet C. Innate immunity of the bovine mammary gland. Vet Res. 2006;37(3):369–400.

    Article  CAS  PubMed  Google Scholar 

  24. Chang H, Yu DS, Liu XQ, Zhang QY, Cheng N, Zhang SQ, et al. Clinical significance of TLR3 and TLR4 in peripheral blood mononuclear cells from children with Henoch-Schönlein purpura nephritis. Exp Ther Med. 2014;7(6):1703–7.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. He C, Zhou Y, Liu F, Liu H, Tan H, Jin S, et al. Bacterial nucleotidyl cyclase inhibits the host Innate Immune response by suppressing TAK1 activation. Infect Immun. 2017;85(9):e00239–17.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Tang J, Luo K, Li Y, Chen Q, Tang D, Wang D, et al. Capsaicin attenuates LPS-induced inflammatory cytokine production by upregulation of LXRα. Int Immunopharmacol. 2015;28(1):264–9.

    Article  CAS  PubMed  Google Scholar 

  27. Shah KN, Valand P, Nauriyal DS, Joshi CG. Immunomodulation of IL-1, IL-6 and IL-8 cytokines by Prosopis juliflora alkaloids during bovine sub-clinical mastitis. 3 Biotech. 2018;8(10):409.

    Article  PubMed  PubMed Central  Google Scholar 

  28. Baldwin AS. Jr. The NF-kappa B and I kappa B proteins: new discoveries and insights. Annu Rev Immunol. 1996;14:649–83.

    Article  CAS  PubMed  Google Scholar 

  29. Wang JW, Chen XY, Hu PY, Tan MM, Tang XG, Huang MC, et al. Effects of Linderae radix extracts on a rat model of alcoholic liver injury. Exp Ther Med. 2016;11(6):2185–92.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Huang HW, Yang CM, Yang CH. Fibroblast growth factor type 1 ameliorates high-glucose-Induced oxidative stress and neuroinflammation in retinal pigment epithelial cells and a Streptozotocin-Induced Diabetic Rat Model. Int J Mol Sci. 2021;22(13):7233.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Zheng Y, Liu G, Wang W, Wang Y, Cao Z, Yang H, et al. Lactobacillus casei Zhang counteracts blood-milk barrier disruption and moderates the inflammatory response in Escherichia coli-Induced Mastitis. Front Microbiol. 2021;12:675492.

    Article  PubMed  PubMed Central  Google Scholar 

  32. Dhami R, He X, Schuchman EH. Acid sphingomyelinase deficiency attenuates bleomycin-induced lung inflammation and fibrosis in mice. Cell Physiol Biochem. 2010;26(4–5):749–60.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Yang Y, Ding Z, Wang Y, Zhong R, Feng Y, Xia T, et al. Systems pharmacology reveals the mechanism of activity of Physalis alkekengi L. var. Franchetii against lipopolysaccharide-induced acute lung injury. J Cell Mol Med. 2020;24(9):5039–56.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Liu M, Wu Q, Wang M, Fu Y, Wang J. Lactobacillus rhamnosus GR-1 limits Escherichia coli-Induced inflammatory responses via attenuating MyD88-Dependent and MyD88-Independent pathway activation in bovine endometrial epithelial cells. Inflammation. 2016;39(4):1483–94.

    Article  CAS  PubMed  Google Scholar 

  35. Nicholas RA, Fox LK, Lysnyansky I. Mycoplasma mastitis in cattle: to cull or not to cull. Vet J. 2016;216:142–7.

    Article  PubMed  Google Scholar 

  36. Sadek K, Saleh E, Ayoub M. Selective, reliable blood and milk bio-markers for diagnosing clinical and subclinical bovine mastitis. Trop Anim Health Prod. 2017;49(2):431–7.

    Article  PubMed  Google Scholar 

  37. Aratani Y, Myeloperoxidase. Its role for host defense, inflammation, and neutrophil function. Arch Biochem Biophys. 2018;640:47–52.

    Article  CAS  PubMed  Google Scholar 

  38. Camperio C, Armas F, Biasibetti E, Frassanito P, Giovannelli C, Spuria L, et al. A mouse mastitis model to study the effects of the intramammary infusion of a food-grade Lactococcus lactis strain. PLoS ONE. 2017;12(9):e0184218.

    Article  PubMed  PubMed Central  Google Scholar 

  39. Martín R, Olivares M, Marín ML, Fernández L, Xaus J, Rodríguez JM. Probiotic potential of 3 lactobacilli strains isolated from breast milk. J Hum Lact. 2005;21(1):8–17.

    Article  PubMed  Google Scholar 

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Acknowledgements

We thank the Animal Clinical Laboratory of Hebei Agricultural University for supplying experimental places and materials.

Funding

This work was supported by the Hebei Key Research and Development Program (19226611D).

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Authors and Affiliations

  1. College of Veterinary Medicine, Shandong Agricultural University, 271018, Taian, Shandong, China

    Ke Li

  2. College of Veterinary Medicine, Hebei Agricultural University, 2596 Lekai South Street, 071001, Baoding, Hebei, China

    Ke Li, Ming Yang, Li Jia, Yinghao Wu, Jinliang Du, Lining Yuan, Lianmin Li & Yuzhong Ma

  3. College of Life Science and Food Engineering, Hebei University of Engineering, 056038, Handan, Hebei, China

    Mengyue Tian

  4. Key Laboratory of Freshwater Fisheries and Germplasm Resources Utilization, Freshwater Fisheries Research Center, Ministry of Agriculture, Chinese Academy of Fishery Sciences, 214081, Wuxi, China

    Jinliang Du

Authors
  1. Ke Li
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  2. Ming Yang
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  4. Li Jia
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  8. Lianmin Li
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Contributions

Ke Li designed the study. Ming Yang and Li Jia prepared materials. Mengyue Tian, Yinghao Wu, Lining Yuan, and Lianmin Li performed all experiments. Ke Li, Ming Yang, and Li Jia analyzed the data. Mengyue Tian, Jinliang Du, and Yuzhong Ma supervised the project. Ke Li drafted the manuscript, and Yuzhong Ma revised it. All authors read and approved the final manuscript.

Corresponding author

Correspondence to Yuzhong Ma.

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Ethics approval and consent to participate

All the animal experiments were approved by the guidelines of the Animal Care and Use Committee of Hebei Agricultural University (Protocol number 2020044).

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Not applicable.

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The authors declare no competing interests.

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The original, full length blots of western blot

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Li, K., Yang, M., Tian, M. et al. The preventive effects of Lactobacillus casei 03 on Escherichia coli-induced mastitis in vitro and in vivo. J Inflamm 21, 5 (2024). https://doi.org/10.1186/s12950-024-00378-x

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Keywords

Journal of Inflammation

ISSN: 1476-9255

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