Little is known about endogenous systemic lupus erythematosus (SLE) plasma DNA even though it is the presumed precursor of DNA-containing immune complexes, thought to play a central role in lupus glomerulonephritis. DNA purified from SLE plasma formed discrete bands, corresponding to sizes of about 150-200, 400, 600, and 800 bp, closely resembling the characteristic 200 bp "ladder" found with oligonucleosomal (ON) DNA. By radiolabeling DNA while in whole plasma, the very small amounts present could be further characterized. All of 24 such specimens formed two or more discrete bands on 6% PAGE. Detergent treatment of plasma resulted in a DNA migration pattern similar to that of purified DNA, suggesting disruption of DNA-protein complexes. DNA purified from authentic ON and detergent-treated ON behaved similarly. A significant portion of DNA, labeled in SLE plasma could be specifically immunoprecipitated with monoclonal antihistone antibody as was the case with ON. These immunoprecipitates, when redissolved, exhibited the expected size distribution upon PAGE. It is concluded that DNA in SLE plasma occurs as a series of multimeric complexes, at least a portion of which is noncovalently bound to histone. These results are consistent with an ON-like structure for SLE plasma DNA as had been suggested by theoretical considerations and may have important implications for its immunologic behavior in SLE and perhaps other disorders. (J. Clin. Invest. 1990.86:69-74.)

SUMMARYThe rale of clearance of exiracellular plasma DNA in man ha.s importani inipiitalions for palhogenelie mechanisms in systemic lupus erythemalosus (SLE), as well as for eeriain oihcr clinical states. Present knowledge of this parameter is derived exelusively from studies of injected, naked DNA in animals. Recent informalion indicates that the physiologic form of plasma DNA in SLE is that of oligonucleosome-like molecules rather than of naked DNA and eonsists of multimcric complexes of DNA bound to histone, probably arising from an apoplotic process. In order to study the rale at which ihese oligonucleosome-like complexes are removed from plasma and to do so in man rather than experimental animals, we exploited the observation that during haemodialysis large amounts of DNA are released, apparently within the dialysis coil, inlo the paiieni's plasma. Since this release appears lo cease promptly with termination of the procedure, it offered the potential for estimating the rate of removal of such DNA from human plasma. Moreover, if that DNA. as postulated, were shown to possess an oligonucleosome-like structure resembling that fotind endogenously in human SLE, the relevance of such information to the human disease state would be further enhanced. The present results support the conclusion that DNA released into plasma during haemodialysis possesses such an oligonucleosome-like structure. The plasma half-life of ihat DNA in man was found nol lo exceed 4 min. The highly dynamic state thus implied for extracellular endogenous plasma DNA in man has important implications for pathogenetic mechanisms dependent on dsDNA in SLE. Moreover, individuals undergoing chronic hacmodialysi,s, who are thereby exposed to a very large cumulative amount of such DNA., might serve as models for studying its long-term sequelae.

In this report we describe and characterize two oligomer probes that are broadly homologous to conserved eubacterial 16S ribosomal RNA (rRNA) sequences not present in human 18 rRNA or human mitochondrial 12S rRNA. One or both of the probes can detect all of 23 phylogenetically diverse eubacterial nucleic acids against which they were tested by dot blot hybridization. A sensitivity of about 1 bacterium per 10 eukaryotic cells was achieved. By using these oligomer sequences or their complements as primers in the polymerase chain reaction (PCR), the equivalent of 1 pg of E. coli DNA was detected in the presence of a large excess of eukaryotic DNA. Information useful for partial phylogenetic classification of detected organisms may be obtained by direct sequence analysis of the amplified DNA and comparison with known sequences or catalogs. Such broadly homologous probes offer advantages over more narrowly specific probes for detecting organisms whose identity is unknown. They could thus be employed for recognizing infection by organisms that cannot be cultured as may occur, for example, in tissue culture or in plant or animal diseases of unknown cause, provided the probes fail to hybridize with host nucleic acids.

In this report we describe and characterize two oligomer probes that are broadly homologous to conserved eubacterial 16S ribosomal RNA (rRNA) sequences not present in human 18 rRNA or human mitochondrial 12S rRNA. One or both of the probes can detect all of 23 phylogenetically diverse eubacterial nucleic acids against which they were tested by dot blot hybridization. A sensitivity of about 1 bacterium per 10 eukaryotic cells was achieved. By using these oligomer sequences or their complements as primers in the polymerase chain reaction (PCR), the equivalent of 1 pg of E. coli DNA was detected in the presence of a large excess of eukaryotic DNA. Information useful for partial phylogenetic classification of detected organisms may be obtained by direct sequence analysis of the amplified DNA and comparison with known sequences or catalogs. Such broadly homologous probes offer advantages over more narrowly specific probes for detecting organisms whose identity is unknown. They could thus be employed for recognizing infection by organisms that cannot be cultured as may occur, for example, in tissue culture or in plant or animal diseases of unknown cause, provided the probes fail to hybridize with host nucleic acids.