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Caseation of human tuberculosis granulomas correlates with elevated host lipid metabolism.

Kim MJ, Wainwright HC, Locketz M, Bekker LG, Walther GB, Dittrich C, Visser A, Wang W, Hsu FF, Wiehart U, Tsenova L, Kaplan G, Russell DG - EMBO Mol Med (2010)

Bottom Line: Biochemical analysis of the lipid species within the caseum identified cholesterol, cholesteryl esters, triacylglycerols and lactosylceramide, which implicated low-density lipoprotein-derived lipids as the most likely source.M. tuberculosis infection in vitro induced lipid droplet formation in murine and human macrophages.Furthermore, the M. tuberculosis cell wall lipid, trehalose dimycolate, induced a strong granulomatous response in mice, which was accompanied by foam cell formation.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, NY, USA.

ABSTRACT
The progression of human tuberculosis (TB) to active disease and transmission involves the development of a caseous granuloma that cavitates and releases infectious Mycobacterium tuberculosis bacilli. In the current study, we exploited genome-wide microarray analysis to determine that genes for lipid sequestration and metabolism were highly expressed in caseous TB granulomas. Immunohistological analysis of these granulomas confirmed the disproportionate abundance of the proteins involved in lipid metabolism in cells surrounding the caseum; namely, adipophilin, acyl-CoA synthetase long-chain family member 1 and saposin C. Biochemical analysis of the lipid species within the caseum identified cholesterol, cholesteryl esters, triacylglycerols and lactosylceramide, which implicated low-density lipoprotein-derived lipids as the most likely source. M. tuberculosis infection in vitro induced lipid droplet formation in murine and human macrophages. Furthermore, the M. tuberculosis cell wall lipid, trehalose dimycolate, induced a strong granulomatous response in mice, which was accompanied by foam cell formation. These results provide molecular and biochemical evidence that the development of the human TB granuloma to caseation correlates with pathogen-mediated dysregulation of host lipid metabolism.

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Thin-layer chromatographic analysis of lipids from the caseum and normal lung tissuesDual solvent separation of granuloma lipids by TLC. Total lipids were extracted from normal lung tissues (lane 4–6, 120 µg) and caseous TB lung granulomas (lane 7–9, 120 µg), and then equivalent amounts were run in parallel with total lipids from Mtb CDC1551 (lane 1, 400 µg), and standard lipids, cardiolipin (lane 2, 50 µg), phosphatidylcholine (lane 3, 50 µg), CHOL (lane 10, 20 µg), CE (lane 11, 25 µg), TAG (lane 12, 20 µg). The lipids from the TB caseum exhibit an abundance of CHOL, CE, and TAG.Single solvent separation of lipid species with similar migration to the sphingolipids, illustrating the enrichment of LacCer in the caseum extract. Total lipids from the caseum (lane 2, 120 µg) were run together with sphingomyelin standard (lane 1, 5 µg) and neutral glycosphingolipids standard (lane 3, 30 µg). Caseous and fibrocaseous granuloma samples were derived from seven independent tissue samples, and normal lung tissue samples.
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fig06: Thin-layer chromatographic analysis of lipids from the caseum and normal lung tissuesDual solvent separation of granuloma lipids by TLC. Total lipids were extracted from normal lung tissues (lane 4–6, 120 µg) and caseous TB lung granulomas (lane 7–9, 120 µg), and then equivalent amounts were run in parallel with total lipids from Mtb CDC1551 (lane 1, 400 µg), and standard lipids, cardiolipin (lane 2, 50 µg), phosphatidylcholine (lane 3, 50 µg), CHOL (lane 10, 20 µg), CE (lane 11, 25 µg), TAG (lane 12, 20 µg). The lipids from the TB caseum exhibit an abundance of CHOL, CE, and TAG.Single solvent separation of lipid species with similar migration to the sphingolipids, illustrating the enrichment of LacCer in the caseum extract. Total lipids from the caseum (lane 2, 120 µg) were run together with sphingomyelin standard (lane 1, 5 µg) and neutral glycosphingolipids standard (lane 3, 30 µg). Caseous and fibrocaseous granuloma samples were derived from seven independent tissue samples, and normal lung tissue samples.

Mentions: We extracted total lipids from the caseum of caseous or fibrocaseous human pulmonary TB granulomas and from uninvolved lung parenchyma and analyzed them by thin-layer chromatography (TLC). There were noticeable differences in lipid profiles; the caseum had markedly increased amounts of lipids that co-migrated with the CE, CHO and TAG standards, when compared to lipids isolated from normal lung tissues (Fig 6A). The identities of CE, CHO and TAG were confirmed by mass spectrometric analysis. Electron impact gas chromatography/mass spectrometry (EI-MS) and total ion current (TIC) chromatogram generated a peak at 12.23 min and an EI mass spectrum that confirmed the presence of CHO (Fig S3A and B). The tandem mass spectrum contained the major [M + Na]+ ions at m/z 671.5 and m/z 673.8, corresponding to CE (Fig S3C). The dominating [M + NH4]+ ions were at m/z 876.8 corresponding to TAG possessing a total of C52 with the presence of two total unsaturated bonds situated on the fatty acyl chains (52:2) (Fig S3D).


Caseation of human tuberculosis granulomas correlates with elevated host lipid metabolism.

Kim MJ, Wainwright HC, Locketz M, Bekker LG, Walther GB, Dittrich C, Visser A, Wang W, Hsu FF, Wiehart U, Tsenova L, Kaplan G, Russell DG - EMBO Mol Med (2010)

Thin-layer chromatographic analysis of lipids from the caseum and normal lung tissuesDual solvent separation of granuloma lipids by TLC. Total lipids were extracted from normal lung tissues (lane 4–6, 120 µg) and caseous TB lung granulomas (lane 7–9, 120 µg), and then equivalent amounts were run in parallel with total lipids from Mtb CDC1551 (lane 1, 400 µg), and standard lipids, cardiolipin (lane 2, 50 µg), phosphatidylcholine (lane 3, 50 µg), CHOL (lane 10, 20 µg), CE (lane 11, 25 µg), TAG (lane 12, 20 µg). The lipids from the TB caseum exhibit an abundance of CHOL, CE, and TAG.Single solvent separation of lipid species with similar migration to the sphingolipids, illustrating the enrichment of LacCer in the caseum extract. Total lipids from the caseum (lane 2, 120 µg) were run together with sphingomyelin standard (lane 1, 5 µg) and neutral glycosphingolipids standard (lane 3, 30 µg). Caseous and fibrocaseous granuloma samples were derived from seven independent tissue samples, and normal lung tissue samples.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2913288&req=5

fig06: Thin-layer chromatographic analysis of lipids from the caseum and normal lung tissuesDual solvent separation of granuloma lipids by TLC. Total lipids were extracted from normal lung tissues (lane 4–6, 120 µg) and caseous TB lung granulomas (lane 7–9, 120 µg), and then equivalent amounts were run in parallel with total lipids from Mtb CDC1551 (lane 1, 400 µg), and standard lipids, cardiolipin (lane 2, 50 µg), phosphatidylcholine (lane 3, 50 µg), CHOL (lane 10, 20 µg), CE (lane 11, 25 µg), TAG (lane 12, 20 µg). The lipids from the TB caseum exhibit an abundance of CHOL, CE, and TAG.Single solvent separation of lipid species with similar migration to the sphingolipids, illustrating the enrichment of LacCer in the caseum extract. Total lipids from the caseum (lane 2, 120 µg) were run together with sphingomyelin standard (lane 1, 5 µg) and neutral glycosphingolipids standard (lane 3, 30 µg). Caseous and fibrocaseous granuloma samples were derived from seven independent tissue samples, and normal lung tissue samples.
Mentions: We extracted total lipids from the caseum of caseous or fibrocaseous human pulmonary TB granulomas and from uninvolved lung parenchyma and analyzed them by thin-layer chromatography (TLC). There were noticeable differences in lipid profiles; the caseum had markedly increased amounts of lipids that co-migrated with the CE, CHO and TAG standards, when compared to lipids isolated from normal lung tissues (Fig 6A). The identities of CE, CHO and TAG were confirmed by mass spectrometric analysis. Electron impact gas chromatography/mass spectrometry (EI-MS) and total ion current (TIC) chromatogram generated a peak at 12.23 min and an EI mass spectrum that confirmed the presence of CHO (Fig S3A and B). The tandem mass spectrum contained the major [M + Na]+ ions at m/z 671.5 and m/z 673.8, corresponding to CE (Fig S3C). The dominating [M + NH4]+ ions were at m/z 876.8 corresponding to TAG possessing a total of C52 with the presence of two total unsaturated bonds situated on the fatty acyl chains (52:2) (Fig S3D).

Bottom Line: Biochemical analysis of the lipid species within the caseum identified cholesterol, cholesteryl esters, triacylglycerols and lactosylceramide, which implicated low-density lipoprotein-derived lipids as the most likely source.M. tuberculosis infection in vitro induced lipid droplet formation in murine and human macrophages.Furthermore, the M. tuberculosis cell wall lipid, trehalose dimycolate, induced a strong granulomatous response in mice, which was accompanied by foam cell formation.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, NY, USA.

ABSTRACT
The progression of human tuberculosis (TB) to active disease and transmission involves the development of a caseous granuloma that cavitates and releases infectious Mycobacterium tuberculosis bacilli. In the current study, we exploited genome-wide microarray analysis to determine that genes for lipid sequestration and metabolism were highly expressed in caseous TB granulomas. Immunohistological analysis of these granulomas confirmed the disproportionate abundance of the proteins involved in lipid metabolism in cells surrounding the caseum; namely, adipophilin, acyl-CoA synthetase long-chain family member 1 and saposin C. Biochemical analysis of the lipid species within the caseum identified cholesterol, cholesteryl esters, triacylglycerols and lactosylceramide, which implicated low-density lipoprotein-derived lipids as the most likely source. M. tuberculosis infection in vitro induced lipid droplet formation in murine and human macrophages. Furthermore, the M. tuberculosis cell wall lipid, trehalose dimycolate, induced a strong granulomatous response in mice, which was accompanied by foam cell formation. These results provide molecular and biochemical evidence that the development of the human TB granuloma to caseation correlates with pathogen-mediated dysregulation of host lipid metabolism.

Show MeSH
Related in: MedlinePlus