• Toxicological Research
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• v.29 no.2
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• pp.129-135
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• 2013

Salmonella is one of the major pathogenic bacteria that cause food poisoning. This study investigated whether heat-killed as well as live Lactobacillus protects host animal against Salmonella infection. Live and heat-killed Lactobacillusacidophilus was administered orally to Sprague-Dawley rats for 2 weeks before the rats were inoculated with Salmonella. Rise in body temperature was moderate in the group that was treated with heat-killed bacteria as compared to the Salmonella control group. The mean amount of feed intake and water consumption of each rat in the heat-killed bacteria group were nearly normal. The number of fecal Salmonellae was comparable between the live and the heat-killed L. acidophilus groups. This finding shows that L. acidophilus facilitates the excretion of Salmonella. Moreover, the levels of pro inflammatory cytokines, including tumor necrosis factor (TNF)-alpha and interleukin (IL)-1 beta, in the heat-killed L. acidophilus group were significantly lower when compared to the levels in the Salmonella control group. These results indicate that nonviable lactic acid bacteria also could play an important role in preventing infections by enteric pathogens such as Salmonella.

• Journal of Microbiology and Biotechnology
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• v.32 no.5
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• pp.638-644
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• 2022

Probiotics modulate the gut microbiota, which in turn regulate immune responses to maintain balanced immune homeostasis in the host. However, it is unclear how probiotic bacteria regulate immune responses. In this study we investigated the immunomodulatory effects of heat-killed probiotics, including Lactiplantibacillus plantarum KC3 (LP3), Lactiplantibacillus plantarum CKDB008 (LP8), and Limosilactobacillus fermentum SRK414 (LF4), via phagocytosis, nitric oxide (NO), and pro-inflammatory cytokine production in macrophages. We thus found that heat-killed LP8 could promote the clearance of foreign pathogens by enhancing the phagocytosis of macrophages. Treatment with heat-killed LP8 induced the production of NO and pro-inflammatory cytokines, including TNF-α, IL-6, and IL-1β. In addition, heat-killed LP8 suppressed the production of NO and cytokines in LPS-induced RAW264.7 cells, suggesting that heat-killed LP8 exerts immunomodulatory effects depending on the host condition. In sum, these results indicate that heat-killed LP8 possesses the potential for immune modulation while providing a molecular basis for the development of functional probiotics prepared from inactivated bacterial cells.

• Journal of Microbiology and Biotechnology
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• v.33 no.9
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• pp.1189-1196
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• 2023

This study proposed to demonstrate the neuroprotective effects of heat-killed Levilactobacillus brevis KU15152. Heat-killed L. brevis KU15152 showed antioxidant activity similar to that of Lacticaseibacillus rhamnosus GG, in terms of radical scavenging activity. To evaluate the neuroprotective effects, conditioned medium (CM) obtained by incubating heat-killed bacteria in intestinal cells (HT-29) was used through gut-brain axis. CM from L. brevis KU15152 protected neuroblastoma cells (SH-SY5Y) against H₂O₂-induced oxidative stress. Pretreatment with CM significantly alleviated the morphological changes induced by H₂O₂. Heat-killed L. brevis KU15152 showed an increased brain-derived neurotrophic factor (BDNF) expression in HT-29 cells. L. brevis KU15152-CM remarkably downregulated the Bax/Bcl-2 ratio, while upregulating the expression of BDNF and tyrosine hydroxylase (TH) in SH-SY5Y cells. Furthermore, L. brevis KU15152-CM reduced caspase-3 activity following H₂O₂ treatment. In conclusion, L. brevis KU15152 can be potentially used as food materials to avoid neurodegenerative diseases.

• Journal of Animal Science and Technology
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• v.63 no.3
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• pp.520-530
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• 2021

This experiment was performed to verify whether dietary heat-killed Lactobacillus rhamnosus (LR) improves growth performance and modulates immune responses of weaned pigs. Ninety-six weaned pigs ([Landrace × Yorkshire] × Duroc; 6.95 ± 0.25 kg body weight [BW]; 28 d old) were randomly allocated to four treatments: 1) a basal diet without heat-killed LR (CON), 2) T1 (CON with 0.1% heat-killed LR), 3) T2 (CON with 0.2% heat-killed LR), and 4) T3 (CON with 0.4% heat-killed LR). Each treatment had six pens with four pigs (6 replicates per treatment) in a randomized completely block design. The heat-killed LR used in this study contained 1 × 10⁹ FU/g of LR in a commercial product. Pigs were fed each treatment for four weeks using a two-phase feeding program to measure growth performance and frequency of diarrhea. During the last week of this study, all diets contained 0.2% chromic oxide as an indigestible marker. Fecal sampling was performed through rectal palpation for the consecutive three days after the four adaptation days to measure apparent total tract digestibility (ATTD) of dry matter, crude protein, and gross energy (GE). Blood sampling was also performed on day 1, 3, 7, and 14 after weaning to measure immune responses such as serum tumor necrosis factor-α (TNF-α), transforming growth factor-β1 (TGF-β1), C-reactive protein (CRP), and cortisol. The heat-killed LR increased (p < 0.05) growth rate, feed efficiency, and ATTD of GE for overall experimental period compared with CON, but reduced (p < 0.05) post-weaning diarrhea. In addition, pigs fed diets contained heat-killed had lower concentrations of serum TNF-α (d 7; p < 0.05), TGF-β1 (d 7; p < 0.10), and cortisol (d 3 and 7; p < 0.05) than pigs fed CON. In conclusion, dietary heat-killed LR improved growth rate, modified immune responses of weaned pigs, and alleviated post-weaning diarrhea.

• Journal of Microbiology and Biotechnology
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• v.31 no.8
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• pp.1134-1143
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• 2021

In the present study, we investigated the inhibitory effect of heat-killed Enterococcus faecalis (E. faecalis) and live E. faecalis on benign prostatic hyperplasia (BPH). The BPH rat model was established by administering male rats with testosterone propionate (TP, 5 mg/kg, in corn oil) via subcutaneous injections daily for four weeks after castration. The rats were divided into five groups: Con, corn oil-injected (s.c.) + DW administration; BPH, TP (5 mg/kg, s.c.) + DW administration; BPH+K_EF, TP (5 mg/kg, s.c.) + heat-killed E. faecalis (7.5 × 10¹² CFU/g, 2.21 mg/kg) administration; BPH+L_EF, TP (5 mg/kg, s.c.) + live E. faecalis (1 × 10¹¹ CFU/g, 166 mg/kg) administration; BPH+Fi, TP (5 mg/kg, s.c.) + finasteride (1 mg/kg) administration. In both of BPH+K_EF and BPH+L_EF groups, the prostate weight decreased and histological changes due to TP treatment recovered to the level of the Con group. Both of these groups also showed regulation of androgen-signaling factors, growth factors, and apoptosis-related factors in prostate tissue. E. faecalis exhibited an inhibitory effect on benign prostatic hyperplasia, and even heat-killed E. faecalis showed similar efficacy on the live cells in the BPH rat model. As the first investigation into the effect of heat-killed and live E. faecalis on BPH, our study suggests that heat-killed E. faecalis might be a food additive candidate for use in various foods, regardless of heat processing.

• Restorative Dentistry and Endodontics
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• v.25 no.4
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• pp.606-618
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• 2000

Dental pulp infection is most commonly caused by extensive dental caries, and some bacterial species invade root canals; bacterial components and products are thought to be associated with the pathogenesis of periapical periodontitis. A principle driving force behind pulpal disease response appears to lie in the host immune system's to bacteria and their products. We examined the production of interleukin $1{\beta}$ (IL-$1{\beta}$) and tumor necrosis factor ${\alpha}$(TNF-${\alpha}$) from human peripheral mononuclear cells, lymphocytes and monocytes stimulated by heat-killed Acitnobacillus actinomycetemcomitans (ATCC 29523), Porphyromonas gingivalis (ATCC 33277) and Prevotella intermedia (ATCC 25611), and also by their sonicated bacterial extracts (SBE), respectively. The effects of three strains of heat-killed bacteria and their SBEs on the morphology of cultured blood cell lines HL-60 (KCLB 10240) and J774A.1 (KCLB 40067) were observed under the inverted microscope. Ultrastructural changes of J774A.1 exposed to heat-killed P. intermedia and its SBE were investigated using transmission electron microscopy. Production of IL-$1{\beta}$ was reduced in human peripheral mononuclear cells after stimulation by sonic bacterial extracts of A. actinomycetemcomitans, P. gingivalis, and P. intermedia. Heat-killed and sonic extract of P. gingivalis inhibited the production of TNF-${\alpha}$ in peripheral mononuclear cells. Production of TNF-${\alpha}$ was inhibited in peripheral monocytes after stimulation by sonic extracts of A. actinomycetemcomitans, P. gingivalis, and P. intermedia. HL-60 and J 774A.1 cells showed granular degeneration after treatment with heat-killed and sonic extracts of A. actinomycetemcomitans, P. gingivalis, and P. intermedia Chromatin margination and shrinkage were observed in 774A.1 treated with heat-killed P. intermedia. Cell wall structure and organelles were destroyed and vacuoles were formed in cytoplasm in J774A.1 treated with P. intermedia sonic extract. These results suggest that A actinomycetemcomitans, P gingivalis and P intermedia may have an important role in the formation and progression of pulpal diseases via both modulation of production of IL-$1{\beta}$ and TNF-${\alpha}$ from blood mononuclear cells and cytopathic effects.

• Korean Journal of Microbiology
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• v.51 no.3
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• pp.241-248
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• 2015

Although studies on probiotics have been performed mostly with viable microbes, the beneficial functions of dead or heat-killed form of probiotic strains have also been examined. In this study, live and heat-killed forms of Lactobacillus acidophilus CBT LA1 were investigated in vitro and in vivo to evaluate the properties necessary for gut barrier protection. Cell surface hydrophobicity (CSH), autoaggregation (AA), cell adhesion, and lipopolysaccharide (LPS)-binding properties were evaluated. In addition, the suppressive effect on LPS-induced interleukin (IL)-8 expression was investigated in HT-29 cells. To identify optimal conditions for CBT LA1 to adhere to HT-29 cells, CBT LA1 cells were heat-treated at 80, 85, 90, 95, 100, or $121^{\circ}C$ for 10 min; cells treated at $80^{\circ}C$ for 10 min showed the highest adhesion. Heat-killed bacteria at $80^{\circ}C$ showed higher levels of LPS-binding, CSH, AA, adhesion to HT-29, and suppression of IL-8 expression than did live CBT LA1. In vivo imaging was performed to evaluate the ability of live or heat-killed CBT LA1 to remove LPS from the intestine in a rat model of infection. At 16 h after infection, fluorescence from FITC-conjugated LPS had mostly disappeared from the intestine of the rats administered with live or heat-killed CBT LA1; the effect was greater with heat-killed CBT LA1 at $80^{\circ}C$. These results suggest that heat-killed CBT LA1 as well as its live form can be applied as a pharmabiotic for protection of the gut barrier.

• Journal of Dairy Science and Biotechnology
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• v.40 no.1
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• pp.48-56
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• 2022

Enterococcus species have been reported to produce unique flavoring ingredients in fermented dairy products. Generally, they were found in cheese and fermented dairy products. Enterococcus spp. produce extracellular polysaccharides and reduce blood cholesterol levels in humans. This study used heat-killed E. faecalis and E. faecium in yogurt production to increase safety during consumption. The addition of heat-killed E. faecalis and E. faecium to milk did not affect the fermentation time of yogurt production, the growth of starter cultures, and the viscosity of yogurt. These results concluded that heat-killed Enterococcus, rather than live Enterococcus, is sufficiently possible and even safer to be added to milk products. Enterococcus species could be used as a safe and functional food additive to fermented milk products and supplements in health foods.

• Journal of Microbiology and Biotechnology
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• v.29 no.12
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• pp.1894-1903
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• 2019

The purpose of this study was to determine the probiotic properties of Lactobacillus brevis KCCM 12203P isolated from the Korean traditional food kimchi and to evaluate the antioxidative activity and immune-stimulating potential of its heat-killed cells to improve their bio-functional activities. Lactobacillus rhamnosus GG, which is a representative commercial probiotic, was used as a comparative sample. Regarding probiotic properties, L. brevis KCCM 12203P was resistant to 0.3% pepsin with a pH of 2.5 for 3 h and 0.3% oxgall solution for 24 h, having approximately a 99% survival rate. It also showed strong adhesion activity (6.84%) onto HT-29 cells and did not produce β-glucuronidase but produced high quantities of leucine arylamidase, valine arylamidase, β-galactosidase, and N-acetyl-β-glucosaminidase. For antioxidant activity, it appeared that viable cells had higher radical scavenging activity in the 2,2-diphenyl-1-picryl-hydrazyl (DPPH) assay, while in the 2-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) assay, heat-killed cells had higher antioxidant activity. Additionally, L. brevis KCCM 12203P showed higher lipid oxidation inhibition ability than L. rhamnosus GG; however, there was no significant difference (p < 0.05) between heat-killed cells and control cells. Furthermore, heat-killed L. brevis KCCM 12203P activated RAW 264.7 macrophage cells without cytotoxicity at a concentration lower than 10⁸ CFU/ml and promoted higher gene expression levels of inducible nitric oxide synthase, interleukin-1β, and interleukin-6 than L. rhamnosus GG. These results suggest that novel L. brevis KCCM 12203P could be used as a probiotic or applied to functional food processing and pharmaceutical fields for immunocompromised people.

• Proceedings of the KWS Conference
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• 2009.11a
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• pp.197-201
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• 2009

A lot of work is carried out concerning to acicular ferrite formation in the weld metal of high strength and low-alloy steel. Those results are suggesting that oxides that contain titanium elements provides nucleation site of intragranular ferrite, referred as acicular ferrite. Thus, when intragranular ferrite is expected to form in heat-affected zone, oxide containing titanium element should be formed in the steel. However, normal steel is deoxidized by using aluminum element (Al-killed steel) with little oxygen content. It means almost oxygen is deoxidized with aluminum elements. In the present work, in order to form the acicular ferrite in the heat affected zone, with the same concept in the case of weld metal, the steel deoxidized with titanium element (titanium killed-steel) is prepared and the acicular ferrite formation is observed in detail by using laser-conforcal microscopy technique. The confocal technique makes it possible that the morphological change along the phase transformation from austenite to ferrite is in-situ tracked. Thus, the inclusion that stimulated the ferrite nucleation could be directly selected from the observed images, in the HAZ of the Ti-killed steel. The chemical composition of the selected inclusion is analyzed and the nucleation potential is discussed by changing the nucleation site with boron element. The potency for the ferrite nucleation is summarized and the existence of effective and ineffective manganese sulfide for nucleation is made clear.