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. 1995 May;63(5):1652–1657. doi: 10.1128/iai.63.5.1652-1657.1995

Detection of muramic acid in a carbohydrate fraction of human spleen.

M A Hoijer 1, M J Melief 1, C G van Helden-Meeuwsen 1, F Eulderink 1, M P Hazenberg 1
PMCID: PMC173205  PMID: 7729869

Abstract

In previous studies, we showed that peptidoglycan polysaccharides from anaerobic bacteria normally present in the human gut induced severe chronic joint inflammation in rats. Our hypothesis is that peptidoglycan from the gut flora is involved in perpetuation of idiopathic inflammation. However, in the literature, the presence of peptidoglycan or subunits like muramyl peptides in blood or tissues is still a matter of debate. We were able to stain red pulp macrophages in all six available human spleens by immunohistochemical techniques using a monoclonal antibody against gut flora-derived antigens. Therefore, these human spleens were extracted, and after removal of most of the protein, the carbohydrate fraction was investigated for the presence of muramic acid, an amino sugar characteristic for peptidoglycan. Using three different methods for detection of muramic acid, we found a mean of 3.3 mumol of muramic acid with high-pressure liquid chromatography, 1.9 mumol with a colorimetric method for detection of lactate, and 0.8 mumol with an enzymatic method for detection of D-lactate per spleen (D-lactate is a specific group of the muramic acid molecule). It is concluded that peptidoglycan is present in human spleen not as small muramyl peptides as were previously searched for by other investigators but as larger macromolecules probably stored in spleen macrophages.

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Selected References

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  1. Bahr G. M., Eshhar Z., Ben-Yitzhak R., Modabber F. Z., Arnon R., Sela M., Chedid L. Monoclonal antibodies to the synthetic adjuvant muramyl dipeptide: characterization of the specificity. Mol Immunol. 1983 Jul;20(7):745–752. doi: 10.1016/0161-5890(83)90052-4. [DOI] [PubMed] [Google Scholar]
  2. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  3. Cromartie W. J., Craddock J. G., Schwab J. H., Anderle S. K., Yang C. H. Arthritis in rats after systemic injection of streptococcal cells or cell walls. J Exp Med. 1977 Dec 1;146(6):1585–1602. doi: 10.1084/jem.146.6.1585. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Ellouz F., Adam A., Ciorbaru R., Lederer E. Minimal structural requirements for adjuvant activity of bacterial peptidoglycan derivatives. Biochem Biophys Res Commun. 1974 Aug 19;59(4):1317–1325. doi: 10.1016/0006-291x(74)90458-6. [DOI] [PubMed] [Google Scholar]
  5. Fox A., Fox K. Rapid elimination of a synthetic adjuvant peptide from the circulation after systemic administration and absence of detectable natural muramyl peptides in normal serum at current analytical limits. Infect Immun. 1991 Mar;59(3):1202–1205. doi: 10.1128/iai.59.3.1202-1205.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Hadzija O. A simple method for the quantitative determination of muramic acid. Anal Biochem. 1974 Aug;60(2):512–517. doi: 10.1016/0003-2697(74)90261-9. [DOI] [PubMed] [Google Scholar]
  7. Hazenberg M. P., Klasen I. S., Kool J., Ruseler-van Embden J. G., Severijnen A. J. Are intestinal bacteria involved in the etiology of rheumatoid arthritis? Review article. APMIS. 1992 Jan;100(1):1–9. doi: 10.1111/j.1699-0463.1992.tb00833.x. [DOI] [PubMed] [Google Scholar]
  8. Hazenberg M. P., de Visser H. Assay for N-acetylmuramyl-L-alanine amidase in serum by determination of muramic acid released from the peptidoglycan of Brevibacterium divaricatum. Eur J Clin Chem Clin Biochem. 1992 Mar;30(3):141–144. [PubMed] [Google Scholar]
  9. Johannsen L., Toth L. A., Rosenthal R. S., Opp M. R., Obal F., Jr, Cady A. B., Krueger J. M. Somnogenic, pyrogenic, and hematologic effects of bacterial peptidoglycan. Am J Physiol. 1990 Jan;258(1 Pt 2):R182–R186. doi: 10.1152/ajpregu.1990.258.1.R182. [DOI] [PubMed] [Google Scholar]
  10. Johannsen L., Wecke J., Obál F., Jr, Krueger J. M. Macrophages produce somnogenic and pyrogenic muramyl peptides during digestion of staphylococci. Am J Physiol. 1991 Jan;260(1 Pt 2):R126–R133. doi: 10.1152/ajpregu.1991.260.1.R126. [DOI] [PubMed] [Google Scholar]
  11. Kool J., De Visser H., Gerrits-Boeye M. Y., Klasen I. S., Melief M. J., Van Helden-Meeuwsen C. G., Van Lieshout L. M., Ruseler-Van Embden J. G., Van den Berg W. B., Bahr G. M. Detection of intestinal flora-derived bacterial antigen complexes in splenic macrophages of rats. J Histochem Cytochem. 1994 Nov;42(11):1435–1441. doi: 10.1177/42.11.7930525. [DOI] [PubMed] [Google Scholar]
  12. Kool J., Severijnen A. J., Klasen I. S., Gerrits-Boeye M. Y., Hazenberg M. P. Influence of decontamination on induction of arthritis in Lewis rats by cell wall fragments of Eubacterium aerofaciens. Arthropathic properties of indigenous anaerobic bacteria. Ann Rheum Dis. 1992 Apr;51(4):510–515. doi: 10.1136/ard.51.4.510. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Lichtman S. N., Bachmann S., Munoz S. R., Schwab J. H., Bender D. E., Sartor R. B., Lemasters J. J. Bacterial cell wall polymers (peptidoglycan-polysaccharide) cause reactivation of arthritis. Infect Immun. 1993 Nov;61(11):4645–4653. doi: 10.1128/iai.61.11.4645-4653.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Ogawa T., Kotani S., Kusumoto S., Shiba T. Possible chemotaxis of human monocytes by N-acetylmuramyl-L-alanyl-D-isoglutamine. Infect Immun. 1983 Jan;39(1):449–451. doi: 10.1128/iai.39.1.449-451.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Ohlsson K., Olsson I. The neutral proteases of human granulocytes. Isolation and partial characterization of granulocyte elastases. Eur J Biochem. 1974 Mar 1;42(2):519–527. doi: 10.1111/j.1432-1033.1974.tb03367.x. [DOI] [PubMed] [Google Scholar]
  16. Sen Z., Karnovsky M. L. Qualitative detection of muramic acid in normal mammalian tissues. Infect Immun. 1984 Mar;43(3):937–941. doi: 10.1128/iai.43.3.937-941.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Severijnen A. J., Kool J., Swaak A. J., Hazenberg M. P. Intestinal flora of patients with rheumatoid arthritis: induction of chronic arthritis in rats by cell wall fragments from isolated Eubacterium aerofaciens strains. Br J Rheumatol. 1990 Dec;29(6):433–439. doi: 10.1093/rheumatology/29.6.433. [DOI] [PubMed] [Google Scholar]
  18. Severijnen A. J., van Kleef R., Hazenberg M. P., van de Merwe J. P. Cell wall fragments from major residents of the human intestinal flora induce chronic arthritis in rats. J Rheumatol. 1989 Aug;16(8):1061–1068. [PubMed] [Google Scholar]
  19. Severijnen A. J., van Kleef R., Hazenberg M. P., van de Merwe J. P. Chronic arthritis induced in rats by cell wall fragments of Eubacterium species from the human intestinal flora. Infect Immun. 1990 Feb;58(2):523–528. doi: 10.1128/iai.58.2.523-528.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Smith S. M., Eng R. H., Campos J. M., Chmel H. D-lactic acid measurements in the diagnosis of bacterial infections. J Clin Microbiol. 1989 Mar;27(3):385–388. doi: 10.1128/jcm.27.3.385-388.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Timmerman C. P., Mattsson E., Martinez-Martinez L., De Graaf L., Van Strijp J. A., Verbrugh H. A., Verhoef J., Fleer A. Induction of release of tumor necrosis factor from human monocytes by staphylococci and staphylococcal peptidoglycans. Infect Immun. 1993 Oct;61(10):4167–4172. doi: 10.1128/iai.61.10.4167-4172.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Todome Y., Ohkuni H., Mizuse M., Furuya M., Fujikawa S., Tanaka S., Watanabe N., Fujii K., Zabriskie J. B. Detection of antibodies against streptococcal peptidoglycan and the peptide subunit (synthetic tetra-D-alanyl-bovine serum albumin complex) in rheumatic-diseases. Int Arch Allergy Immunol. 1992;97(4):301–307. doi: 10.1159/000236137. [DOI] [PubMed] [Google Scholar]
  23. WEIDEL W., PELZER H. BAGSHAPED MACROMOLECULES--A NEW OUTLOOK ON BACTERIAL CELL WALLS. Adv Enzymol Relat Areas Mol Biol. 1964;26:193–232. doi: 10.1002/9780470122716.ch5. [DOI] [PubMed] [Google Scholar]
  24. Wilkinson B. J., Kim Y., Peterson P. K. Factors affecting complement activation by Staphylococcus aureus cell walls, their components, and mutants altered in teichoic acid. Infect Immun. 1981 Apr;32(1):216–224. doi: 10.1128/iai.32.1.216-224.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]

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