Improved Tensor Current Limit from

A precision measurement of the ß^{+} decay of ^{8}B was performed using the Beta-decay Paul Trap to determine the ß-ν angular correlation coefficient a_{ßν}. The experimental results were combined with new ab initio symmetry-adapted no-core shell-model calculations to yield the second-most precise measurement from Gamow-Teller decays, a_{ßν}=-0.3345±0.0019_{stat}±0.0021_{syst}. This value agrees with the standard model value of -1/3 and improves uncertainties in ^{8}B by nearly a factor of 2. By combining results from ^{8}B and ^{8}Li, a tight limit on tensor current coupling to right-handed neutrinos was obtained. A recent global evaluation of all other precision ß decay studies suggested a nonzero value for right-handed neutrino coupling in contradiction with the standard model at just above 3σ. The present results are of comparable sensitivity and do not support this finding.

Direct Determination of Fission-Barrier Heights Using Light-Ion Transfer in Inverse Kinematics.

We demonstrate a new technique for obtaining fission data for nuclei away from ß stability. These types of data are pertinent to the astrophysical r process, crucial to a complete understanding of the origin of the heavy elements, and for developing a predictive model of fission. These data are also important considerations for terrestrial applications related to power generation and safeguarding. Experimentally, such data are scarce due to the difficulties in producing the actinide targets of interest. The solenoidal-spectrometer technique, commonly used to study nucleon-transfer reactions in inverse kinematics, has been applied to the case of transfer-induced fission as a means to deduce the fission-barrier height, among other variables. The fission-barrier height of ^{239}U has been determined via the ^{238}U(d,pf) reaction in inverse kinematics, the results of which are consistent with existing neutron-induced fission data indicating the validity of the technique.

Quenching of Single-Particle Strength in A=15 Nuclei.

Absolute cross sections for the addition of s- and d-wave neutrons to ^{14}C and ^{14}N have been determined simultaneously via the (d,p) reaction at 10 MeV/u. The difference between the neutron and proton separation energies, ΔS, is around -20 MeV for the ^{14}C+n system and +8 MeV for ^{14}N+n. The population of the 1s_{1/2} and 0d_{5/2} orbitals for both systems is reduced by a factor of approximately 0.5 compared with the independent single-particle model, or about 0.6 when compared with the shell model. This finding strongly contrasts with results deduced from intermediate-energy knockout reactions between similar nuclei on targets of ^{9}Be and ^{12}C. The simultaneous technique used removes many systematic uncertainties.

Search for Nova Presolar Grains: γ-Ray Spectroscopy of

The discovery of presolar grains in primitive meteorites has initiated a new era of research in the study of stellar nucleosynthesis. However, the accurate classification of presolar grains as being of specific stellar origins is particularly challenging. Recently, it has been suggested that sulfur isotopic abundances may hold the key to definitively identifying presolar grains with being of nova origins and, in this regard, the astrophysical ^{33}Cl(p,γ)^{34}Ar reaction is expected to play a decisive role. As such, we have performed a detailed γ-ray spectroscopy study of ^{34}Ar. Excitation energies have been measured with high precision and spin-parity assignments for resonant states, located above the proton threshold in ^{34}Ar, have been made for the first time. Uncertainties in the ^{33}Cl(p,γ) reaction have been dramatically reduced and the results indicate that a newly identified ℓ=0 resonance at E_{r}=396.9(13) keV dominates the entire rate for T=0.25-0.40 GK. Furthermore, nova hydrodynamic simulations based on the present work indicate an ejected ^{32}S/^{33}S abundance ratio distinctive from type-II supernovae and potentially compatible with recent measurements of a presolar grain.

Key

Detection of nuclear-decay γ rays provides a sensitive thermometer of nova nucleosynthesis. The most intense γ-ray flux is thought to be annihilation radiation from the ß^{+} decay of ^{18}F, which is destroyed prior to decay by the ^{18}F(p,α)^{15}O reaction. Estimates of ^{18}F production had been uncertain, however, because key near-threshold levels in the compound nucleus, ^{19}Ne, had yet to be identified. We report the first measurement of the ^{19}F(^{3}He,tγ)^{19}Ne reaction, in which the placement of two long-sought 3/2^{+} levels is suggested via triton-γ-γ coincidences. The precise determination of their resonance energies reduces the upper limit of the rate by a factor of 1.5-17 at nova temperatures and reduces the average uncertainty on the nova detection probability by a factor of 2.1.

Constraint of the astrophysical

The Galactic 1.809-MeV γ-ray signature from the ß decay of ^{26g}Al is a dominant target of γ-ray astronomy, of which a significant component is understood to originate from massive stars. The ^{26g}Al(p,γ)^{27}Si reaction is a major destruction pathway for ^{26g}Al at stellar temperatures, but the reaction rate is poorly constrained due to uncertainties in the strengths of low-lying resonances in ^{27}Si. The ^{26g}Al(d,p)^{27}Al reaction has been employed in inverse kinematics to determine the spectroscopic factors, and hence resonance strengths, of proton resonances in ^{27}Si via mirror symmetry. The strength of the 127-keV resonance is found to be a factor of 4 higher than the previously adopted upper limit, and the upper limit for the 68-keV resonance has been reduced by an order of magnitude, considerably constraining the ^{26g}Al destruction rate at stellar temperatures.

Halo nucleus 11Be: a spectroscopic study via neutron transfer.

The best examples of halo nuclei, exotic systems with a diffuse nuclear cloud surrounding a tightly bound core, are found in the light, neutron-rich region, where the halo neutrons experience only weak binding and a weak, or no, potential barrier. Modern direct-reaction measurement techniques provide powerful probes of the structure of exotic nuclei. Despite more than four decades of these studies on the benchmark one-neutron halo nucleus 11Be, the spectroscopic factors for the two bound states remain poorly constrained. In the present work, the 10Be(d,​p) reaction has been used in inverse kinematics at four beam energies to study the structure of 11Be. The spectroscopic factors extracted using the adiabatic model were found to be consistent across the four measurements and were largely insensitive to the optical potential used. The extracted spectroscopic factor for a neutron in an nℓj=2s(1/2) state coupled to the ground state of 10Be is 0.71(5). For the first excited state at 0.32 MeV, a spectroscopic factor of 0.62(4) is found for the halo neutron in a 1p(1/2) state.

cDNA cloning of the B cell membrane protein CD22: a mediator of B-B cell interactions.

We have cloned a full-length cDNA for the B cell membrane protein CD22, which is referred to as B lymphocyte cell adhesion molecule (BL-CAM). Using subtractive hybridization techniques, several B lymphocyte-specific cDNAs were isolated. Northern blot analysis with one of the clones, clone 66, revealed expression in normal activated B cells and a variety of B cell lines, but not in normal activated T cells, T cell lines, Hela cells, or several tissues, including brain and placenta. One major transcript of approximately 3.3 kb was found in B cells although several smaller transcripts were also present in low amounts (approximately 2.6, 2.3, and 1.6 kb). Sequence analysis of a full-length cDNA clone revealed an open reading frame of 2,541 bases coding for a predicted protein of 847 amino acids with a molecular mass of 95 kD. The BL-CAM cDNA is nearly identical to a recently isolated cDNA clone for CD22, with the exception of an additional 531 bases in the coding region of BL-CAM. BL-CAM has a predicted transmembrane spanning region and a 140-amino acid intracytoplasmic domain. Search of the National Biological Research Foundation protein database revealed that this protein is a member of the immunoglobulin super family and that it had significant homology with three homotypic cell adhesion proteins: carcinoembryonic antigen (29% identity over 460 amino acids), myelin-associated glycoprotein (27% identity over 425 amino acids), and neural cell adhesion molecule (21.5% over 274 amino acids). Northern blot analysis revealed low-level BL-CAM mRNA expression in unactivated tonsillar B cells, which was rapidly increased after B cell activation with Staphylococcus aureus Cowan strain 1 and phorbol myristate acetate, but not by various cytokines, including interleukin 4 (IL-4), IL-6, and gamma interferon. In situ hybridization with an antisense BL-CAM RNA probe revealed expression in B cell-rich areas in tonsil and lymph node, although the most striking hybridization was in the germinal centers. COS cells transfected with a BL-CAM expression vector were immunofluorescently stained positively with two different CD22 antibodies, each of which recognizes a different epitope. Additionally, both normal tonsil B cells and a B cell line were found to adhere to COS transfected with BL-CAM in the sense but not the antisense direction.(ABSTRACT TRUNCATED AT 400 WORDS)

Mitochondrial DNA damage is involved in apoptosis caused by pro-inflammatory cytokines in human OA chondrocytes.

OBJECTIVE: Pro-inflammatory cytokines play a pivotal role in cartilage destruction during the progression of osteoarthritis (OA). Additionally, these cytokines are capable to generate reactive oxygen and nitrogen species within chondrocytes. Mitochondrion is a prime target of oxidative damage and an important player in aging and degenerative processes. The purpose of the present study was to investigate whether these cytokines will alter the mitochondrial DNA (mtDNA) integrity and mitochondrial function in both normal and osteoarthritic human chondrocytes. DESIGN: Primary normal and osteoarthritic human chondrocyte cultures were exposed to various concentrations of interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) for different time. Following exposure, chondrocytes were evaluated for mitochondrial DNA damage, ATP production, changes in mitochondrial transcription, and apoptosis. Adenoviral vectors were used to deliver DNA repair enzyme hOGG1 to mitochondria. RESULTS: Pro-inflammatory cytokines IL-1beta and TNF-alpha disturb mitochondrial function in human chondrocytes by inducing mitochondrial DNA damage, decreasing energy production and mitochondrial transcription, which correlated with the induction of apoptosis. Increased NO production was the key factor responsible for accumulation of mtDNA damage after cytokine exposure. Mitochondrial superoxide production was also enhanced following pro-inflammatory cytokine exposure. OA chondrocyte mitochondria were more susceptible to damage induced by pro-inflammatory cytokines then mitochondria from normal chondrocytes. Protection of human chondrocytes from mtDNA damage by the mitochondria-targeted DNA repair enzyme hOGG1 rescued mtDNA integrity, preserved ATP levels, reestablished mitochondrial transcription, and significantly diminished apoptosis following IL-1beta and TNF-alpha exposure. CONCLUSION: Mitochondrion is an important target in pro-inflammatory cytokine toxicity, maintaining of mitochondrial DNA integrity is necessary to prevent chondrocytes from apoptosis induced by IL-1beta and TNF-alpha.

Diminished mitochondrial DNA integrity and repair capacity in OA chondrocytes.

OBJECTIVES: Osteoarthritis (OA) is characterized by the failure of chondrocytes to respond to injury and perform the cartilage remodeling process. Human articular chondrocytes actively produce reactive oxygen and nitrogen species (ROS and RNS) capable of causing cellular dysfunction and death. A growing body of evidence indicates that mitochondrial dysfunction and mitochondrial DNA (mtDNA) damage play a causal role in disorders linked to excessive generation of oxygen free radicals. The aim of this study was to determine whether mtDNA damage was present in OA chondrocytes, and whether mtDNA repair capacity is compromised in OA chondrocytes following oxidative stress, leading to chondrocyte death. METHODS: Human articular cartilage was isolated from knee joints of cadavers available through the Anatomical Gifts Program at the University of South Alabama (normal donors) or OA patients undergoing total knee replacement surgeries (OA patients). Total DNA was isolated from either chondrocytes released following collagenase digestion, or from first passage chondrocytes grown in culture and exposed to ROS or RNS. mtDNA integrity and repair capacity were analyzed by quantitative Southern blot analysis, using a mtDNA-specific radioactive probe. Cell viability was determined by the trypan blue exclusion method. RESULTS: mtDNA damage was found in chondrocytes from OA patients compared to normal donors. It was accompanied with reduced mtDNA repair capacity, cell viability, and increased apoptosis in OA chondrocytes following exposure to ROS and RNS. CONCLUSIONS: These results indicate that mtDNA damage and poor mtDNA repair capacity for removing damage caused by oxidative stress may contribute to the pathogenesis of OA.

Selective elimination of mutant mitochondrial genomes as therapeutic strategy for the treatment of NARP and MILS syndromes.

Mitochondrial diseases are not uncommon, and may result from mutations in both nuclear and mitochondrial DNA (mtDNA). At present, only palliative therapies are available for these disorders, and interest in the development of efficient treatment protocols is high. Here, we demonstrate that in cells heteroplasmic for the T8993G mutation, which is a cause for the NARP and MILS syndromes, infection with an adenovirus, which encodes the mitochondrially targeted R.XmaI restriction endonuclease, leads to selective destruction of mutant mtDNA. This destruction proceeds in a time- and dose-dependent manner and results in cells with significantly increased rates of oxygen consumption and ATP production. The delivery of R.XmaI to mitochondria is accompanied by improvement in the ability to utilize galactose as the sole carbon source, which is a surrogate indicator of the proficiency of oxidative phosphorylation. Concurrently, the rate of lactic acid production by these cells, which is a marker of mitochondrial dysfunction, decreases. We further demonstrate that levels of phosphorylated P53 and gammaH2ax proteins, markers of nuclear DNA damage, do not change in response to infection with recombinant adenovirus indicating the absence of nuclear DNA damage and the relative safety of the technique. Finally, some advantages and limitations of the proposed approach are discussed.

Palmitate induced mitochondrial deoxyribonucleic acid damage and apoptosis in l6 rat skeletal muscle cells.

A major characteristic of type 2 diabetes mellitus (T2DM) is insulin resistance in skeletal muscle. A growing body of evidence indicates that oxidative stress that results from increased production of reactive oxygen species and/or reactive nitrogen species leads to insulin resistance, tissue damage, and other complications observed in T2DM. It has been suggested that muscular free fatty acid accumulation might be responsible for the mitochondrial dysfunction and insulin resistance seen in T2DM, although the mechanisms by which increased levels of free fatty acid lead to insulin resistance are not well understood. To help resolve this situation, we report that saturated fatty acid palmitate stimulated the expression of inducible nitric oxide (NO) synthase and the production of reactive oxygen species and NO in L6 myotubes. Additionally, palmitate caused a significant dose-dependent increase in mitochondrial DNA (mtDNA) damage and a subsequent decrease in L6 myotube viability and ATP levels at concentrations as low as 0.5 mM. Furthermore, palmitate induced apoptosis, which was detected by DNA fragmentation, caspase-3 cleavage, and cytochrome c release. N-acetyl cysteine, a precursor compound for glutathione formation, aminoguanidine, an inducible NO synthase inhibitor, and 5,10,15,20-tetrakis(4-sulphonatophenyl) porphyrinato iron (III), a peroxynitrite inhibitor, all prevented palmitate-induced mtDNA damage and diminished palmitate-induced cytotoxicity. We conclude that exposure of L6 myotubes to palmitate induced mtDNA damage and triggered mitochondrial dysfunction, which caused apoptosis. Additionally, our findings indicate that palmitate-induced mtDNA damage and cytotoxicity in skeletal muscle cells were caused by overproduction of peroxynitrite.

Mitochondrial DNA repair: a critical player in the response of cells of the CNS to genotoxic insults.

Cells of the CNS are constantly exposed to agents which damage DNA. Although much attention has been paid to the effects of this damage on nuclear DNA, the nucleus is not the only organelle containing DNA. Within each cell, there are hundreds to thousands of mitochondria. Within each mitochondrion are multiple copies of the mitochondrial genome. These genomes are extremely vulnerable to insult and mutations in mitochondrial DNA (mtDNA) have been linked to several neurodegenerative diseases, as well as the normal process of aging. The principal mechanism utilized by cells to avoid DNA mutations is DNA repair. Multiple pathways of DNA repair have been elucidated for nuclear DNA. However, it appears that only base excision repair is functioning in mitochondria. This repair pathway is responsible for the removal of most endogenous damage including alkylation damage, depurination reactions and oxidative damage. Within the rat CNS, there are cell-specific differences mtDNA repair. Astrocytes exhibit efficient repair, whereas, other glial cell types and neuronal cells exhibit a reduced ability to remove lesions from mtDNA. Additionally, a correlation was observed between those cells with reduced mtDNA repair and an increase in the induction of apoptosis. To demonstrate a causative relationship, a strategy of targeting DNA repair proteins to mitochondria to enhance mtDNA repair capacity was employed. Enhancement of mtDNA repair in oligodendrocytes provided protection from reactive oxygen species- and cytokine-induced apoptosis. These experiments provide a novel strategy for protecting sensitive CNS cells from genotoxic insults and thus provide new treatment options for neurodegenerative diseases.

Detection and determination of oligonucleotide triplex formation-mediated transcription-coupled DNA repair in HeLa nuclear extracts.

Transcription-coupled repair (TCR) plays an important role in removing DNA damage from actively transcribed genes. It has been speculated that TCR is the most important mechanism for repairing DNA damage in non-dividing cells such as neurons. Therefore, abnormal TCR may contribute to the development of many age-related and neurodegenerative diseases. However, the molecular mechanism of TCR is not well understood. Oligonucleotide DNA triplex formation provides an ideal system to dissect the molecular mechanism of TCR since triplexes can be formed in a sequence-specific manner to inhibit transcription of target genes. We have recently studied the molecular mechanism of triplex-forming oligonucleotide (TFO)-mediated TCR in HeLa nuclear extracts. Using plasmid constructs we demonstrate that the level of TFO-mediated DNA repair activity is directly correlated with the level of transcription of the plasmid in HeLa nuclear extracts. TFO-mediated DNA repair activity was further linked with transcription since the presence of rNTPs in the reaction was essential for AG30-mediated DNA repair activity in HeLa nuclear extracts. The involvement of individual components, including TFIID, TFIIH, RNA polymerase II and xeroderma pigmentosum group A (XPA), in the triplex-mediated TCR process was demonstrated in HeLa nuclear extracts using immunodepletion assays. Importantly, our studies also demonstrated that XPC, a component involved in global genome DNA repair, is involved in the AG30-mediated DNA repair process. The results obtained in this study provide an important new understanding of the molecular mechanisms involved in the TCR process in mammalian cells.

Nitric oxide-induced damage to mtDNA and its subsequent repair.

Mutations in mitochondrial DNA (mtDNA) have recently been associated with a variety of human diseases. One potential DNA-damaging agent to which cells are continually exposed that could be responsible for some of these mutations is nitric oxide (NO). To date, little information has been forthcoming concerning the damage caused by this gas to mtDNA. Therefore, this study was designed to investigate damage to mtDNA induced by NO and to evaluate its subsequent repair. Normal human fibroblasts were exposed to NO produced by the rapid decomposition of 1-propanamine, 3-(2-hydroxy-2-nitroso-1-propylhydrazino) (PAPA NONOate) and the resultant damage to mtDNA was determined by quantitative Southern blot analysis. This gas was found to cause damage to mtDNA that was alkali-sensitive. Treatment of the DNA with uracil-DNA glycosylase or 3-methyladenine DNA glycosylase failed to reveal additional damage, indicating that most of the lesions produced were caused by the deamination of guanine to xanthine. Studies using ligation-mediated PCR supported this finding. When a 200 bp sequence of mtDNA from cells exposed to NO was analyzed, guanine was found to be the predominantly damaged base. However, there also was damage to specific adenines. No lesions were observed at pyrimidine sites. The nucleotide pattern of damage induced by NO was different from that produced by either a reactive oxygen species generator or the methylating chemical, methylnitrosourea. Most of the lesions produced by NO were repaired rapidly. However, there appeared to be a subset of lesions which were repaired either slowly or not at all by the mitochondria.

Base excision repair of mitochondrial DNA damage in mammalian cells.

This review of the work from our laboratory describes initial studies in which base excision repair in mtDNA was first seen. It considers the results of experiments in which the substrates for mtDNA repair were identified. The discussion then focuses on studies during which the sequence context for mtDNA damage and repair were explored. Next, it addresses factors that have been identified that influence mtDNA repair. Finally, it summarizes the results of studies that evaluated cell-specific differences in the repair of mtDNA and explored some of the biological consequences of these differences.

Heterogeneous repair of methylnitrosourea-induced alkali-labile sites in different DNA sequences.

The repair of DNA damage induced by methylnitrosourea (MNU) in restriction fragments containing the dihydrofolate reductase (DHFR) gene in Chinese hamster ovary cells was compared to that in equal size restriction fragments containing a nontranscribed flanking sequence 3' to the DHFR gene or the c-fos gene. Following exposure to 10(-3) M MNU, restriction fragments containing either the DHFR gene or the 3' flanking sequence had similar amounts of alkali labile sites, approximately 2 sites/restriction fragment. Fragments encompassing the c-fos gene had less than 2 breaks/fragment. Twenty-four h after exposure to MNU a consistent, but slight and not statistically significant, difference was seen with more adducts removed from the DHFR gene than the 3' flanking sequence. No repair was detected in the c-fos containing fragments. In addition, the repair of N7-methylguanine in the overall genome was assessed by use of a 32P end-labeling technique. Seventy % of this major alkylation product was repaired after 24 h. These findings establish that repair heterogeneity occurs in Chinese hamster ovary cells after exposure to MNU.

Defective repair of oxidative damage in the mitochondrial DNA of a xeroderma pigmentosum group A cell line.

Recent evidence has linked mitochondrial DNA (mtDNA) damage to several disease processes,including cancer and aging. An important source of such damage is reactive oxygen species. These molecules can be generated endogenously via the electron transport system or may arise from a host of exogenous sources. It has been reported that extracts from cells of individuals with xeroderma pigmentosum group A (XP-A) do not repair some types of oxidative DNA damage. The current experiments were designed to determine whether there is a correlation between the inadequate repair of oxidatively damaged nuclear DNA in XP-A cells and the capacity of such cells to repair similar damage to their mtDNA. The ability of karyotypically normal human fibroblasts (WI-38) and XP-A fibroblasts to repair alloxan-generated oxidative damage to nuclear and mtDNA was assessed using a quantitative Southern blot method in conjunction with the repair enzymes endonuclease III and formamidopyrimidine DNA glycosylase. The data indicate that both nuclear and mtDNA repair of each damage type investigated is more efficient in the WI-38 cells. These findings suggest a similarity between the process(es) used to repair oxidative damage to nuclear and mtDNA in that both are inhibited by the defect in XP-A.

Therapeutic and diabetogenic potential of two newly synthesized nitrosoureido sugars.

Two newly synthesized nitrosoureido sugars have been evaluated for their antitumor activity and diabetogenic potential in a number of in vitro and in vivo preclinical tumor model systems. 2-Amino-2-deoxy-N'-methyl-N'-nitrosoureido-1,3,4,6-tetra-O-acetyl-alpha- D- mannopyranose (MAZ), a lipophilic mannosamine derivative, and ethyl-6-deoxy-3,5-di-O-methyl-6-(3-methyl-3-nitrosoureido)-alph a- D-glucofuranoside (EDOMEN or CGP 6'809), were both found to inhibit L1210 leukemia cell growth in vitro by 50% at approximately 5.0 X 10(-5) M. At these concentrations, little effect was noted immediately on L1210 cell radiolabeled precursor incorporation; however, at higher concentrations, EDOMEN inhibited [3H]leucine and [3H]mannose incorporation, while MAZ specifically decreased L1210 cell [3H]thymidine and [3H]leucine incorporation. Inhibition of Lewis lung carcinoma and B16 melanoma cell growth by 50% in vitro was achieved at higher concentrations of these agents (10(-4) to 10(-3) M). Since the currently available nitrosoureido sugars, streptozotocin and chlorozotocin, have been observed by us to be diabetogenic, EDOMEN and MAZ were evaluated for their specific toxicity to rat pancreatic beta-cells in vitro. Cytotoxicity in beta-cell cultures was monitored both by phase-contrast microscopy and the release of insulin into the culture medium. beta-Cells were found to be 10-fold more sensitive to the toxic effects of MAZ than were pancreatic fibroblasts. EDOMEN, on the other hand, did not damage beta-cells preferentially and therefore was not considered diabetogenic. Both MAZ and EDOMEN had moderate activity as antileukemic agents in mice. At 50 mg/kg/day i.p. for 5 days, MAZ increased the life span of female DBA/2J mice with L1210 leukemia by over 50%. Similarly, doses of EDOMEN at 125 to 250 mg/kg/day i.p. for 5 days increased L1210 leukemic life span by nearly 60%. At these doses, no effect of MAZ was observed on primary Lewis lung carcinoma growth or life span of tumor-bearing C57BL/6 mice. EDOMEN, however, increased life span in Lewis lung carcinoma mice by up to 33% and caused an apparent antimetastatic effect. These studies indicate that EDOMEN may have enhanced value as a cancer chemotherapeutic agent due to its therapeutic effectiveness, lack of diabetogenic potential, and other favorable formulation properties (water solubility) as compared with other clinically available nitrosoureas.

Glial cell-specific differences in repair of O6-methylguanine.

Normal and malignant cells of the oligodendrocyte lineage show increased sensitivity to alkylating agents compared to astrocytes. One of the most mutagenic DNA lesions formed following exposure to alkylating agents is O6-alkylguanine. To determine whether the increased sensitivity to nitrosoureas seen in oligodendrocytes is due to decreased repair capacity for O6-alkylguanine, removal of this lesion from DNA was assessed in primary cultures of rat oligodendrocytes, astrocytes, and microglia. Glial cells were exposed to 1 mM N-methyl-N-nitrosourea for 1 h and allowed 8 or 24 h for repair. Repair was evaluated using an immunoslot blot technique and a monoclonal antibody which recognizes O6-methylguanine (O6MeGua). Astrocytes removed O6MeGua more efficiently (approximately 80% in 24 h) than either oligodendrocytes (approximately 20%) or microglia (approximately 4%). Determination of O6-alkylguanine-DNA-alkyltransferase (AT) activity revealed that astrocytes contain 0.4 pmol/mg protein, which is average by comparison to other cell types. Both oligodendrocytes and microglia exhibited very low levels of AT (oligodendrocytes, 0.08; microglia, 0.01 pmol/mg protein). These data are the first to show that within different populations of glial cells, O6MeGua adduct removal is substantially reduced in both oligodendrocytes and microglia. Rapid removal of O6MeGua in astrocytes coupled with persistence of this mutagenic lesion in oligodendrocytes following exposure of the developing central nervous system to nitrosoureas could contribute to the observed formation of oligodendrogliomas. Inefficient removal of O6MeGua in oligodendrogliomas might also account for their response to chemotherapeutic regimens involving alkylating agents such as procarbazine, lomustine, and carmustine. The lack of repair of O6MeGua in microglia suggests that primary lymphomas of the central nervous system might be sensitive to treatment with alkylating drugs whose toxicity depends on repair of this adduct.