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Regulation of mammalian siderophore 2,5-DHBA in the innate immune response to infection.

Liu Z, Reba S, Chen WD, Porwal SK, Boom WH, Petersen RB, Rojas R, Viswanathan R, Devireddy L - J. Exp. Med. (2014)

Bottom Line: Pathogens secrete small iron-binding moieties, siderophores, to acquire host iron.In support of this idea, supplementation with mammalian siderophore enhances bacterial growth in vitro.Thus, reciprocal regulation of 24p3 and mammalian siderophore is a protective mechanism limiting microbial access to iron.

View Article: PubMed Central - HTML - PubMed

Affiliation: Case Comprehensive Cancer Center; Department of Pathology; Department of Medicine, Tuberculosis Research Institute and Division of Infectious Diseases; and Department of Chemistry, Case Western Reserve University, Cleveland, OH 44106Case Comprehensive Cancer Center; Department of Pathology; Department of Medicine, Tuberculosis Research Institute and Division of Infectious Diseases; and Department of Chemistry, Case Western Reserve University, Cleveland, OH 44106.

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Derivation of bdh2 knockout mice. (A) Schematic representation of the targeting strategy and the expected patterns of Cre-mediated homologous recombination. The structure of the bdh2 locus is shown at the top, the targeting construct in the middle and the resulting targeted allele at the bottom. Numbered boxes indicate exons. Black triangles indicate LoxP sites. Arrowheads indicate PCR primer locations for genotyping (P1-P4). 5′ and 3′ external probes indicated by solid lines. TK, herpes simplex virus thymidine kinase gene. Restriction sites, HindIII (H), EcoRI (E), used for Southern blot genotyping are indicated. (B) Assessment of 2,5-DHBA levels in urine samples of control and bdh2- mice. A total of 10 age- and sex-matched WT and bdh2- mice on C57BL/6 genetic background were analyzed. Representative GC-MS spectra of the TMS-derivatized, small molecule fractions from urine specimen from WT and bdh2- mice were shown. A set of DHBA standards is shown on the top. (C and D) Bdh2 deficiency does not alter ketone body metabolism. Measurement of 3-hydroxybutyrate and acetoacetate levels in plasma of 8-wk-old female WT and bdh2- mice on C57BL/6 genetic background. Data are representative of two independent experiments, each with at least 4–9 mice per group. Statistical analysis by one-way analysis of variance followed by the Tukey HSD test for multiple comparisons. P < 0.05 was considered significant. (E–G) Evaluation of fatty acid profiles in plasma of 8-wk-old female WT and bdh2- mice on C57BL/6 genetic background. Data are representative of two independent experiments, each with at least 4–9 mice per group. Statistical analysis by one-way analysis of variance followed by the Tukey HSD test for multiple comparisons. P < 0.05 was considered significant.
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fig2: Derivation of bdh2 knockout mice. (A) Schematic representation of the targeting strategy and the expected patterns of Cre-mediated homologous recombination. The structure of the bdh2 locus is shown at the top, the targeting construct in the middle and the resulting targeted allele at the bottom. Numbered boxes indicate exons. Black triangles indicate LoxP sites. Arrowheads indicate PCR primer locations for genotyping (P1-P4). 5′ and 3′ external probes indicated by solid lines. TK, herpes simplex virus thymidine kinase gene. Restriction sites, HindIII (H), EcoRI (E), used for Southern blot genotyping are indicated. (B) Assessment of 2,5-DHBA levels in urine samples of control and bdh2- mice. A total of 10 age- and sex-matched WT and bdh2- mice on C57BL/6 genetic background were analyzed. Representative GC-MS spectra of the TMS-derivatized, small molecule fractions from urine specimen from WT and bdh2- mice were shown. A set of DHBA standards is shown on the top. (C and D) Bdh2 deficiency does not alter ketone body metabolism. Measurement of 3-hydroxybutyrate and acetoacetate levels in plasma of 8-wk-old female WT and bdh2- mice on C57BL/6 genetic background. Data are representative of two independent experiments, each with at least 4–9 mice per group. Statistical analysis by one-way analysis of variance followed by the Tukey HSD test for multiple comparisons. P < 0.05 was considered significant. (E–G) Evaluation of fatty acid profiles in plasma of 8-wk-old female WT and bdh2- mice on C57BL/6 genetic background. Data are representative of two independent experiments, each with at least 4–9 mice per group. Statistical analysis by one-way analysis of variance followed by the Tukey HSD test for multiple comparisons. P < 0.05 was considered significant.

Mentions: To test the role of the mammalian siderophore on bacterial growth in vivo, we generated mice lacking Bdh2 activity. The catalytic activity of Bdh2 is dependent on NPG and SYK motifs, which are encoded by exon 7 (Guo et al., 2006). We replaced exon 7 of the bdh2 gene with a neomycin resistance cassette (Fig. 2 A). By interbreeding bdh2 heterozygous mice, we obtained nearly 25% bdh2- mice suggesting that the bdh2 deletion did not affect the viability of the embryos.


Regulation of mammalian siderophore 2,5-DHBA in the innate immune response to infection.

Liu Z, Reba S, Chen WD, Porwal SK, Boom WH, Petersen RB, Rojas R, Viswanathan R, Devireddy L - J. Exp. Med. (2014)

Derivation of bdh2 knockout mice. (A) Schematic representation of the targeting strategy and the expected patterns of Cre-mediated homologous recombination. The structure of the bdh2 locus is shown at the top, the targeting construct in the middle and the resulting targeted allele at the bottom. Numbered boxes indicate exons. Black triangles indicate LoxP sites. Arrowheads indicate PCR primer locations for genotyping (P1-P4). 5′ and 3′ external probes indicated by solid lines. TK, herpes simplex virus thymidine kinase gene. Restriction sites, HindIII (H), EcoRI (E), used for Southern blot genotyping are indicated. (B) Assessment of 2,5-DHBA levels in urine samples of control and bdh2- mice. A total of 10 age- and sex-matched WT and bdh2- mice on C57BL/6 genetic background were analyzed. Representative GC-MS spectra of the TMS-derivatized, small molecule fractions from urine specimen from WT and bdh2- mice were shown. A set of DHBA standards is shown on the top. (C and D) Bdh2 deficiency does not alter ketone body metabolism. Measurement of 3-hydroxybutyrate and acetoacetate levels in plasma of 8-wk-old female WT and bdh2- mice on C57BL/6 genetic background. Data are representative of two independent experiments, each with at least 4–9 mice per group. Statistical analysis by one-way analysis of variance followed by the Tukey HSD test for multiple comparisons. P < 0.05 was considered significant. (E–G) Evaluation of fatty acid profiles in plasma of 8-wk-old female WT and bdh2- mice on C57BL/6 genetic background. Data are representative of two independent experiments, each with at least 4–9 mice per group. Statistical analysis by one-way analysis of variance followed by the Tukey HSD test for multiple comparisons. P < 0.05 was considered significant.
© Copyright Policy - openaccess
Related In: Results  -  Collection

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fig2: Derivation of bdh2 knockout mice. (A) Schematic representation of the targeting strategy and the expected patterns of Cre-mediated homologous recombination. The structure of the bdh2 locus is shown at the top, the targeting construct in the middle and the resulting targeted allele at the bottom. Numbered boxes indicate exons. Black triangles indicate LoxP sites. Arrowheads indicate PCR primer locations for genotyping (P1-P4). 5′ and 3′ external probes indicated by solid lines. TK, herpes simplex virus thymidine kinase gene. Restriction sites, HindIII (H), EcoRI (E), used for Southern blot genotyping are indicated. (B) Assessment of 2,5-DHBA levels in urine samples of control and bdh2- mice. A total of 10 age- and sex-matched WT and bdh2- mice on C57BL/6 genetic background were analyzed. Representative GC-MS spectra of the TMS-derivatized, small molecule fractions from urine specimen from WT and bdh2- mice were shown. A set of DHBA standards is shown on the top. (C and D) Bdh2 deficiency does not alter ketone body metabolism. Measurement of 3-hydroxybutyrate and acetoacetate levels in plasma of 8-wk-old female WT and bdh2- mice on C57BL/6 genetic background. Data are representative of two independent experiments, each with at least 4–9 mice per group. Statistical analysis by one-way analysis of variance followed by the Tukey HSD test for multiple comparisons. P < 0.05 was considered significant. (E–G) Evaluation of fatty acid profiles in plasma of 8-wk-old female WT and bdh2- mice on C57BL/6 genetic background. Data are representative of two independent experiments, each with at least 4–9 mice per group. Statistical analysis by one-way analysis of variance followed by the Tukey HSD test for multiple comparisons. P < 0.05 was considered significant.
Mentions: To test the role of the mammalian siderophore on bacterial growth in vivo, we generated mice lacking Bdh2 activity. The catalytic activity of Bdh2 is dependent on NPG and SYK motifs, which are encoded by exon 7 (Guo et al., 2006). We replaced exon 7 of the bdh2 gene with a neomycin resistance cassette (Fig. 2 A). By interbreeding bdh2 heterozygous mice, we obtained nearly 25% bdh2- mice suggesting that the bdh2 deletion did not affect the viability of the embryos.

Bottom Line: Pathogens secrete small iron-binding moieties, siderophores, to acquire host iron.In support of this idea, supplementation with mammalian siderophore enhances bacterial growth in vitro.Thus, reciprocal regulation of 24p3 and mammalian siderophore is a protective mechanism limiting microbial access to iron.

View Article: PubMed Central - HTML - PubMed

Affiliation: Case Comprehensive Cancer Center; Department of Pathology; Department of Medicine, Tuberculosis Research Institute and Division of Infectious Diseases; and Department of Chemistry, Case Western Reserve University, Cleveland, OH 44106Case Comprehensive Cancer Center; Department of Pathology; Department of Medicine, Tuberculosis Research Institute and Division of Infectious Diseases; and Department of Chemistry, Case Western Reserve University, Cleveland, OH 44106.

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Related in: MedlinePlus