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Extraction of Peptidoglycan from L. paracasei subp. Paracasei X12 and Its Preliminary Mechanisms of Inducing Immunogenic Cell Death in HT-29 Cells.

Tian PJ, Li BL, Shan YJ, Zhang JN, Chen JY, Yu M, Zhang LW - Int J Mol Sci (2015)

Bottom Line: X12-PG could induce the production of apoptotic bodies observed by transmission electron microscopy (TEM).X12-PG could significantly induced the translocation of calreticulin (CRT) and the release of high mobility group box 1 protein (HMGB1), the two notable hallmarks of immunogenic cell death (ICD), with the endoplastic reticulum (ER) damaged and subsequently intracellular [Ca(2+)] elevated.Our findings implied that X12-PG could induce the ICD of HT-29 cells through targeting at the ER.

View Article: PubMed Central - PubMed

Affiliation: School of Food Science and Engineering, Harbin Institute of Technology, No. 73 Huanghe Road, Harbin 150000, China. tianpei0202@gmail.com.

ABSTRACT
L. paracasei subp. paracasei X12 was previously isolated from a Chinese traditional fermented cheese with anticancer activities and probiotic potential. Herein, the integral peptidoglycan (X12-PG) was extracted by a modified trichloroacetic acid (TCA) method. X12-PG contained the four representative amino acids Asp, Glu, Ala and Lys, and displayed the similar lysozyme sensitivity, UV-visible scanning spectrum and molecular weight as the peptidoglycan standard. X12-PG could induce the production of apoptotic bodies observed by transmission electron microscopy (TEM). X12-PG could significantly induced the translocation of calreticulin (CRT) and the release of high mobility group box 1 protein (HMGB1), the two notable hallmarks of immunogenic cell death (ICD), with the endoplastic reticulum (ER) damaged and subsequently intracellular [Ca(2+)] elevated. Our findings implied that X12-PG could induce the ICD of HT-29 cells through targeting at the ER. The present results may enlighten the prospect of probiotics in the prevention of colon cancer.

No MeSH data available.


Related in: MedlinePlus

Quantitative and qualitative analysis of X12-PG. (A) The degradation models of X12-PG and X12 when exposed to lysozyme. A450 nm of X12-PG declined faster than that of X12; (B) The UV-visible scanning spectrum of X12-PG and peptidoglycan standard. The standards and samples were scanned between 190 and 700 nm in a UV/VIS spectrophotometer; and (C) The molecular weight (kDa) of X12-PG quantified by SDS-PAGE. The gels were stained with Coomassie Brilliant Blue. The molecular weight was 14 kDa, corresponding to the peptidoglycan standard. Each graph represents the average of more than three replications.
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ijms-16-20033-f002: Quantitative and qualitative analysis of X12-PG. (A) The degradation models of X12-PG and X12 when exposed to lysozyme. A450 nm of X12-PG declined faster than that of X12; (B) The UV-visible scanning spectrum of X12-PG and peptidoglycan standard. The standards and samples were scanned between 190 and 700 nm in a UV/VIS spectrophotometer; and (C) The molecular weight (kDa) of X12-PG quantified by SDS-PAGE. The gels were stained with Coomassie Brilliant Blue. The molecular weight was 14 kDa, corresponding to the peptidoglycan standard. Each graph represents the average of more than three replications.

Mentions: Peptidoglycan is chiefly composed of chains of N-acetylglucosamine (GlcNAc) and N-acetylmuramic acid (MurNAc) attached to stem peptides. The total carbohydrate content of X12-PG is 199.16 mg/g, the protein content was calculated as 20.31 mg/g. Lysozyme assay is generally used as a qualitative test for peptidoglycan. Because the β-1,4-linked GlcNAc and MurNAc structure in X12-PG can be exclusively hydrolyzed by lysozyme, the absorbance values (A450 nm) sharply declined during the first two incubation hours (Figure 2A), whereas no obvious and fast dropping was observed in X12 solution during the whole process. The results suggested that X12-PG possessed the characteristics of peptidoglycan.


Extraction of Peptidoglycan from L. paracasei subp. Paracasei X12 and Its Preliminary Mechanisms of Inducing Immunogenic Cell Death in HT-29 Cells.

Tian PJ, Li BL, Shan YJ, Zhang JN, Chen JY, Yu M, Zhang LW - Int J Mol Sci (2015)

Quantitative and qualitative analysis of X12-PG. (A) The degradation models of X12-PG and X12 when exposed to lysozyme. A450 nm of X12-PG declined faster than that of X12; (B) The UV-visible scanning spectrum of X12-PG and peptidoglycan standard. The standards and samples were scanned between 190 and 700 nm in a UV/VIS spectrophotometer; and (C) The molecular weight (kDa) of X12-PG quantified by SDS-PAGE. The gels were stained with Coomassie Brilliant Blue. The molecular weight was 14 kDa, corresponding to the peptidoglycan standard. Each graph represents the average of more than three replications.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4581339&req=5

ijms-16-20033-f002: Quantitative and qualitative analysis of X12-PG. (A) The degradation models of X12-PG and X12 when exposed to lysozyme. A450 nm of X12-PG declined faster than that of X12; (B) The UV-visible scanning spectrum of X12-PG and peptidoglycan standard. The standards and samples were scanned between 190 and 700 nm in a UV/VIS spectrophotometer; and (C) The molecular weight (kDa) of X12-PG quantified by SDS-PAGE. The gels were stained with Coomassie Brilliant Blue. The molecular weight was 14 kDa, corresponding to the peptidoglycan standard. Each graph represents the average of more than three replications.
Mentions: Peptidoglycan is chiefly composed of chains of N-acetylglucosamine (GlcNAc) and N-acetylmuramic acid (MurNAc) attached to stem peptides. The total carbohydrate content of X12-PG is 199.16 mg/g, the protein content was calculated as 20.31 mg/g. Lysozyme assay is generally used as a qualitative test for peptidoglycan. Because the β-1,4-linked GlcNAc and MurNAc structure in X12-PG can be exclusively hydrolyzed by lysozyme, the absorbance values (A450 nm) sharply declined during the first two incubation hours (Figure 2A), whereas no obvious and fast dropping was observed in X12 solution during the whole process. The results suggested that X12-PG possessed the characteristics of peptidoglycan.

Bottom Line: X12-PG could induce the production of apoptotic bodies observed by transmission electron microscopy (TEM).X12-PG could significantly induced the translocation of calreticulin (CRT) and the release of high mobility group box 1 protein (HMGB1), the two notable hallmarks of immunogenic cell death (ICD), with the endoplastic reticulum (ER) damaged and subsequently intracellular [Ca(2+)] elevated.Our findings implied that X12-PG could induce the ICD of HT-29 cells through targeting at the ER.

View Article: PubMed Central - PubMed

Affiliation: School of Food Science and Engineering, Harbin Institute of Technology, No. 73 Huanghe Road, Harbin 150000, China. tianpei0202@gmail.com.

ABSTRACT
L. paracasei subp. paracasei X12 was previously isolated from a Chinese traditional fermented cheese with anticancer activities and probiotic potential. Herein, the integral peptidoglycan (X12-PG) was extracted by a modified trichloroacetic acid (TCA) method. X12-PG contained the four representative amino acids Asp, Glu, Ala and Lys, and displayed the similar lysozyme sensitivity, UV-visible scanning spectrum and molecular weight as the peptidoglycan standard. X12-PG could induce the production of apoptotic bodies observed by transmission electron microscopy (TEM). X12-PG could significantly induced the translocation of calreticulin (CRT) and the release of high mobility group box 1 protein (HMGB1), the two notable hallmarks of immunogenic cell death (ICD), with the endoplastic reticulum (ER) damaged and subsequently intracellular [Ca(2+)] elevated. Our findings implied that X12-PG could induce the ICD of HT-29 cells through targeting at the ER. The present results may enlighten the prospect of probiotics in the prevention of colon cancer.

No MeSH data available.


Related in: MedlinePlus