Relative Cell Counts Research Articles

Comparison of Neutrophil CD64 Expression, Manual Myeloid Immaturity Counts, and Automated Hematology Analyzer Flags as Indicators of Infection or Sepsis

There is a clear need for improved indicators of infection or sepsis to increase the sensitivity and specificity of both diagnosis and therapeutic monitoring. One of the effects of inflammatory cytokines on the innate immune response is the rapid up-regulation of CD64 expression on the neutrophil membrane. We and others have hypothesized that the measurement of neutrophil CD64 expression might represent an improved diagnostic indicator of infection and sepsis. In this study we assessed the relative ability of flow cytometric neutrophil CD64 measurements, neutrophil counts, myeloid immaturity differential counts, and flagging on an automated hematology analyzer to correlate with the presence of infection, as determined by a retrospective clinical scoring system of infection or sepsis. A total of 160 blood samples were randomly selected to derive equal proportions of the 3 categories of flags on a Coulter STKS blood counter that indicate the presence of a myeloid left shift. The patients for these samples were scored by retrospective chart review and placed into 4 groups on the basis of likelihood of infection, sepsis, or severe tissue injury. Neutrophil CD64 expression demonstrated a superior sensitivity (94.1%), specificity (84.9%), and positive predictive likelihood ratio (6.24), compared with neutrophil counts (sensitivity, 79.4%; specificity, 46.8%; positive predictive likelihood ratio, 1.49), band counts (sensitivity, 87.5%; specificity, 43.5%; positive predictive likelihood ratio, 1.55), myeloid immaturity fraction (sensitivity, 94.6%; specificity, 84.5%; positive predictive likelihood ratio, 2.12), and flagging on an automated hematology analyzer (sensitivity, 94.1%; specificity, 40.5%; positive predictive likelihood ratio, 1.58). Relative to the other laboratory parameters, the neutrophil CD64 parameter also provided the best separation of the 4 clinical groups. The findings indicate that neutrophil CD64 expression as determined by quantitative flow cytometry is an improved diagnostic indicator of infection/sepsis relative to current laboratory indicators of relative or absolute myeloid cell counts or hematology analyzer flagging algorithms.

The adaptive immune system and long-term outcome in patients with stable coronary disease

Components of the adaptive immune system, in particular lymphocytes and immunoglobulin, play a major role in advanced atherosclerotic lesions. We sought to determine whether routine, measurements of the relative number of circulating lymphocytes (%L) and gamma-globulin (%G) reflecting immunoglobulin are related to event-free survival in patients with stable coronary artery disease (CAD). We prospectively studied the combined endpoint all-cause mortality, myocardial infarction and coronary revascularization procedures in 141 patients after successful percutaneous coronary intervention during a median follow-up time of 13.2 years. Using Cox regression, we found a significant influence of %L on event-free survival (P=0.007) with a relative risk of 2.21 comparing third to first tertile. Subjects with higher %G values likewise had a shorter event-free survival (P=0.008) with a relative risk of 1.67 comparing third to first tertile. The predictive value of %L and %G remained significant after adjustment for demographic data, cardiovascular risk factors, extent of CAD and other inflammatory markers. We conclude that the fraction of gamma-globulin and in particular the relative lymphocyte cell count may serve as readily available and reliable prognostic tools for the long-term outcome in patients with stable CAD.

Absolute and relative cell counts for synovial fluid from clinically normal shoulder and stifle joints in cats

To determine absolute and relative cell counts for synovial fluid from grossly, radiographically, and histologically normal shoulder and stifle joints in healthy cats. Clinical study. 52 cats scheduled to be euthanatized for unrelated reasons. Arthrocentesis of the shoulder and stifle joints was performed bilaterally, and synovial fluid was analyzed for absolute WBC count, WBC morphology, and percentages of neutrophils and mononuclear cells. Joints were examined grossly and radiographically, and synovial membrane specimens were submitted for histologic examination. Synovial fluid samples that were contaminated with blood and samples from joints with any gross, radiographic, or histologic abnormalities were excluded. 82 of the 208 synovial fluid samples were excluded because abnormalities were identified during physical examination; the volume of fluid obtained was insufficient for analysis; there was evidence of blood contamination; or the joint had gross, radiographic, or histologic abnormalities. Median WBC count for the remaining 126 synovial fluid samples was 91 cells/microL (96.4% mononuclear cells and 3.6% neutrophils); WBC count was not significantly different between left and right joint samples or between shoulder and stifle joint samples. Body weight was associated with synovial fluid WBC count, with WBC count increasing as body weight increased. Sixteen of the 52 (30%) cats had radiographic evidence of osteoarthritis involving at least 1 joint. Results suggest that synovial fluid can be obtained reliably from shoulder and stifle joints in cats.

Role of Cftr genotype in the response to chronic Pseudomonas aeruginosa lung infection in mice.

Patients with cystic fibrosis have a lesion in the cystic fibrosis transmembrane conductance regulator gene (CFTR), which is associated with abnormal regulation of other ion channels, abnormal glycosylation of secreted and cell surface molecules, and vulnerability to bacterial infection and inflammation in the lung usually leading to the death of these patients. The exact mechanism(s) by which mutation in CFTR leads to lung infection and inflammation is not clear. Mice bearing different mutations in the murine homolog to CFTR (Cftr) (R117H, S489X, Y122X, and DeltaF508, all backcrossed to the C57BL/6J background) were compared with respect to growth and in their ability to respond to lung infection elicited with Pseudomonas aeruginosa-laden agarose beads. Body weights of mice bearing mutations in Cftr were significantly smaller than wild-type mice at most ages. The inflammatory responses to P. aeruginosa-laden agarose beads were comparable in mice of all four Cftr mutant genotypes with respect to absolute and relative cell counts in bronchoalveolar lavage fluid, and cytokine levels (TNF-alpha, IL-1beta, IL-6, macrophage inflammatory protein-2, and keratinocyte chemoattractant) and eicosanoid levels (PGE2 and LTB4) in epithelial lining fluid: the few small differences observed occurred only between cystic fibrosis mice bearing the S489X mutation and those bearing the knockout mutation Y122X. Thus we cannot implicate either misprocessing of CFTR or failure of CFTR to reach the plasma membrane in the genesis of the excess inflammatory response of CF mice. Therefore, it appears that any functional defect in CFTR produces comparable inflammatory responses to lung infections with P. aeruginosa.

Human leukocyte antigen-DR expression in peripheral blood mononuclear cells from healthy donors influenced by the sera of injured patients prone to severe sepsis.

To study the influence of sera from severely injured patients on the human leukocyte antigen (HLA)-DR expression of normal peripheral blood mononuclear cells (PBMC). In vitro study. University hospital. Sera from 34 patients were obtained within 8 h after trauma. Seventeen of these patients developed posttraumatic sepsis (sepsis group) and 17 recovered without infectious complications. Sera from ten healthy individuals served as controls. Phytohemagglutinin (PHA)-activated PBMC from 44 healthy donors were used to study the effects of a patient's serum. Medium containing 5% of serum from the sepsis group significantly ( p<0.05) reduced the HLA-DR expression (channels, mean +/- standard error of the mean) on monocytes (patients 883+/-22, controls 962+/-15), B (patients 922+/-14, controls 972+/-7) and T cells (patients 932+/-13, controls 968+/-5) of PHA-activated PBMC. Significantly increased accumulation of TNFalpha on (1.8+/-0.4% of PBMC) and within T cells (0.98+/-0.26% of PBMC) was observed by flow cytometry after incubation with medium containing sera of the sepsis group compared with controls (on 0.5+/-0.1%, within 0.27+/-0.05% of PBMC). A significant negative correlation between relative cell counts of intracellular TNFalpha-positive T cells with HLA-DR expression was observed for monocytes ( r= -0.61), B cells ( r= -0.57) and proliferation ( r= -0.68) as estimated by (3)H-thymidine uptake [patients 139971+/-12844 counts per minute (cpm), controls 198973+/-19347 cpm, p<0.05] in the presence of sera from the sepsis group. Reduced cellular immunity and, therefore, immunodeficiency after trauma appears to be caused by soluble factors influencing T cell function in particular.

Therapeutic immune reconstitution in HIV-1-infected children is independent of their age and pretreatment immune status.

To evaluate long-term immune reconstitution of children treated with highly active antiretroviral therapy (HAART). The long-term immunological response to HAART was studied in 71 HIV-1-infected children (aged 1 month to 18 years) in two prospective, open, uncontrolled national multicentre studies. Blood samples were taken before and after HAART was initiated, with a follow-up of 96 weeks, and peripheral CD4 and CD8 T cells plus naive and memory subsets were identified in whole blood samples. Relative cell counts were calculated in relation to the median of the age-specific reference. The absolute CD4 cell count and percentage and the CD4 cell count as a percentage of normal increased significantly (P < 0.001) to medians of 939 x 106 cells/l (range, 10-3520), 32% (range, 1-50) and 84% (range, 1-161), respectively, after 48 weeks. This increase was predominantly owing to naive CD4 T cells. There was a correlation between the increase of absolute naive CD4 T cell counts and age. However, when CD4 T cell restoration was studied as percentage of normal values, the inverse correlation between the increase of naive CD4 T cell count and age was not observed. In addition, no difference in immunological reconstitution was observed at any time point between virological responders and non-responders. Normalization of the CD4 cell counts in children treated with HAART is independent of age, indicating that children of all age groups can meet their CD4 T cell production demands. In general, it appears that children restore their CD4 T cell counts better and more rapidly than adults, even in a late stage of HIV-1 infection.

Clinical and virologic response to combination treatment with indinavir, zidovudine, and lamivudine in children with human immunodeficiency virus-1 infection: A multicenter study in The Netherlands

<b>Objective:</b> To evaluate the clinical, immunologic, and virologic response to indinavir, zidovudine, and lamivudine in children with human immunodeficiency virus-1 (HIV-1) infection. <b>Study design:</b> Twenty-eight HIV-1-infected children (3 months to 16 years of age) with or without prior treatment with reverse-transcriptase inhibitors and a HIV-1 RNA >5000 copies/mL and/or a CD4 cell count less than the lower limit of the age-specific reference value were treated with indinavir, zidovudine, and lamivudine. Pharmacokinetics of indinavir were determined in each child. <b>Results:</b> The combination treatment was well tolerated in the majority of patients. Clinical improvement was seen in all patients. After 6 months of therapy, 70% of the patients had an HIV-1 RNA load below 500 copies/mL, whereas 48% of the children had a viral load below 40 copies/mL. Relative CD4 cell counts in relation to the lower limit of the age-specific reference value increased significantly from a median value of 79% at baseline to 106% after 6 months of therapy. The doses of indinavir necessary to achieve area under the curve values comparable to adult values varied from 1250 mg/m²/d to 2450 mg/m²/d. <b>Conclusions:</b> Highly active antiretroviral therapy consisting of indinavir, zidovudine, and lamivudine in children reduced HIV-1 RNA to less than 500 copies/mL in 70% of the children within 6 months. Improved CD4 cell counts were observed in most patients, as was a better clinical condition (no invasive or opportunistic infections, increased weight gain). Side effects of the triple therapy were mild. Highly active antiretroviral therapy can be used as successfully in children as in adults.

HEALTH EFFECTS OF SULFUR-RELATED ENVIRONMENTAL AIR POLLUTION. II. Cellular and Molecular Parameters of Injury

Recently, concern has been raised about effects related to environmental sulfur and/ or acidic aerosols. To assess long-term effects on nonrespiratory lung function, 8 beagle dogs were exposed over a period of 13 mo for 16.5 h/day to a neutral sulfite aerosol at a sulfur(IV) concentration of 0.32 mg m-3 and for 6 h/day to an acidic sulfate aerosol providing a hydrogen concentration of 15.2 mumol m-3 for inhalation. Prior to exposure the dogs were kept under clean air conditions for 16 mo to establish physiological baseline values for each animal. A second group of eight dogs (control) was kept for the entire study under clean air conditions. No clinical symptoms were identified that could be related to the combined exposure. Biochemical and cellular parameters were analyzed in sequential bronchoalveolar lavage (BAL) fluids. The permeability of the alveolocapillary membrane and diethylenetriaminepentaacetic acid (DTPA) clearance was not affected. Similarly, oxidant burden of the epithelial lining fluid evaluated by levels of oxidation products in the BAL fluid protein fraction remained unchanged. Both the lysosomal enzyme- N -acetylglucosaminidase and the alpha-1-AT were increased (p < .05). In contrast, the cytoplasmic marker lactate dehydrogenase remained unchanged, indicating the absence of severe damages to epithelial cells or phagocytes. Various surfactant functions were not altered during exposure. Three animals showed elevated levels of the type II cell-associated alkaline phosphatase (AP), indicating a nonuniform response of type II cells. Significant correlations were found between AP and total BAL protein, but not between AP and lactate dehydrogenase, suggesting proliferation of alveolar type II cells. Absolute and relative cell counts in the BAL fluid were not influenced by exposure. Alveolar macrophages showed no alterations with regard to their respiratory burst upon stimulation with opsonized zymosan. The percentage of alveolar macrophages capable of phagocytozing latex particles was significantly decreased (p < .05), while the phagocytosis index was not altered. In view of the results of this and previous studies, we conclude that there is no synergism of effects of these two air pollutants on nonrespiratory lung functions. It is hypothesized that antagonistic effects of these air pollutants on phospholipase A2-dependent pathways account for compensatory physiological mechanisms. The results emphasize the complexity of health effects on lung functions in response to the complex mixture of air pollutants and disclose the precariousness in the risk assessment of air pollutants for humans.

Prognostic value of the relative lymphocyte concentration and white blood cell count in heart failure: Rayburn BK, Rodeheffer RJ, Young JB, Mullen GM, Torre G, Tolman DE, Gibbons RJ, Brown RN, Bourge RC, Kirklin JK for the Cardiac Transplant Research Database Investigators, Birmingham, Alabama

Prognostic value of the relative lymphocyte concentration and white blood cell count in heart failure: Rayburn BK, Rodeheffer RJ, Young JB, Mullen GM, Torre G, Tolman DE, Gibbons RJ, Brown RN, Bourge RC, Kirklin JK for the Cardiac Transplant Research Database Investigators, Birmingham, Alabama

CD4 and CD8 counts are associated with interactions of gender and psychosocial stress.

This study examined relationships of gender, psychosocial stress/distress (caregiving, hassles, depressed mood), and the relative percentage and absolute cell counts of CD4 and CD8 cells in two samples of older adults (mean age = 69.4)--spouse caregivers of persons with Alzheimer's disease (N = 78) and age- and gender-matched spouses of nondemented controls (N = 72). Counts and percentages of CD4 and CD8 cells and psychosocial variables were assessed twice (Time 1, Time 2) over a 15- to 18-month period. Several covariates were examined in the analyses, including body mass index (BMI), medication use, alcohol use, exercise, and illness history. Caregiver men had fewer CD4 cell counts at Times 1 and 2 than did control men (p < .05). At Times 1 and 2, both CD8 cell counts and percentages were positively associated with hassles in men (p < .05), but not in women. Although interactions of hassles and gender were present for CD8 percentages at both times, interactions and main effects were not present for CD4 percentages at either time. When the ratio of CD4 to CD8 levels was analyzed, hassles by gender interactions were present at both Times 1 and 2-hassles were negatively associated with the CD4/CD8 ratio in men (p < .05), but unrelated in women. From Time 1 to Time 2, change analyses showed that increases in hassles scores were associated with decreases in CD4 counts (p < .05), whereas increases in Hamilton Depression Scores were related to increases in both CD8 counts and percentages (p < .05). Caregiver status, hassles, and depressed mood had cross-sectional and/or longitudinal associations with CD4 and CD8 counts, but such relationships occurred primarily in men. Moreover, absolute cell counts were more related to psychosocial factors than were percentages.

Electron microscopic microanalysis of bronchoalveolar lavage: a way to identify exposure to silica and silicate dust.

OBJECTIVES: The diagnostic implications of finding non-fibrous inorganic particles in bronchoalveolar lavage (BAL) fluid has not been fully assessed. The aim of this study has been to measure the silica...

Milk differential cell counts in relation to total counts in Sardinian ewes

Milk samples from 40 primiparous Sardinian ewes were examined during a whole lactation period in order to evaluate the dynamics of somatic cell counts (SCC), differential cell counts (DCC) and bacteriological status of the mammary gland. Only four samples out of 240 were bacteriologically positive and this was confirmed by the geometrical mean of SCC being always below 200 000 cells per ml. DCC during lactation showed two different dynamics: one concerning polymorphonuclear neutrophil leucocytes (PMN) and macrophages, and the other one lymphocytes and epithelial cells. Macrophages and PMNs in particular showed a progressive increase as lactation went on, while lymphocytes and epithelial cells showed a uniform decrease during lactation. The relationship between SCC and DCC indicated that PMN was the only cell type increasing when SCC rose. The progressive increase of SCC during lactation in absence of infection was probably an effect of cell concentration owing to the physiological reduction of milk yield, while the increase of macrophages and PMN was very likely a consequence of immune mechanisms approaching the dry period. The results confirmed that DCC and particularly PMN rate could provide a more sensible method of identifying mammary inflammations, particularly in well managed herds with low levels of infections.

Impact of oxygenator design on hemolysis, shear stress, and white blood cell and platelet counts

To determine whether relative pressure drop, shear stress, hemolysis, and white blood cell and platelet counts are influenced by different oxygenator designs. To compare the oxygenator results with the average shear stress over an arterial cannula. Prospective; patients enrolled consecutively. University hospital. Three groups of 12 adults patients, scheduled for routine cardiac surgery. Each group was submitted to a different oxygenator design: group 1 to a high-pressure hollow-fiber membrane oxygenator (Sarns Turbo); group 2 to a medium-pressure hollow-fiber membrane oxygenator (Cobe optima); and group 3 to a flat-sheet membrane oxygenator (Cobe Duo). Although the investigated oxygenators have important differences in pressure drop and shear stress, no statistical differences were found in hemolysis generation or blood handling among the different groups. Actually, the study shows much higher shear stress levels over an average arterial cannula than over any of the evaluated oxygenators. The pressure drop over an oxygenator does not correlate well with shear stress and hemolysis because the dimensions of the system (radius and length) must be included in the calculation of shear stress from pressure drop.

The estrogen antagonist tamoxifen inhibits carrageenan induced inflammation in LEW/N female rats

Carrageenan induces a measurable inflammatory response in susceptible animals, and mature females are more responsive to carrageenan, than males. In the present study, we tested whether the estrogen antagonist tamoxifen influences carrageenan-induced inflammatory responses. Female LEW/N rats were treated with tamoxifen and compared to a control group of animals injected with vehicle. Tamoxifen significantly reduced estrous phase of estrous cycle during treatment, consistent with its functional anti-estrogen effects. Moreover, tamoxifen significantly decreased exudate volume but did not significantly influence relative white blood cell counts in the exudate. Interestingly, tamoxifen induced differential dose-dependent alterations in peripheral blood lymphocyte subpopulations. Low dose of tamoxifen increased CD25 cells. The high tamoxifen dose significantly increased CD8 blood lymphocyte counts. Our data indicate that tamoxifen treatment decreases carrageenan-induced inflammatory response in female LEW/N rats and suggest therefore that this inflammatory response is, at least in part, estrogen related. Moreover, our results suggest a possible role for tamoxifen in treatment of inflammatory disorders.

Inflammatory responses to carrageenan injection in LEW/N and F344/N rats: LEW/N rats show sex- and age-dependent changes in inflammatory reactions.

We studied the inflammatory responses of LEW/N and F344/N inbred rat strains after peripheral injection of carrageenan. The inflammatory responses were assessed in terms of volume, relative and total white blood cell counts of the exudates. Moreover, in both strains, blood CD4, CD8, CD25, naive CD4 (CD4/CD45RC) cell and B (CD45R) cell counts and plasma corticosterone levels, constituents of systemic inflammatory responses to carrageenan were evaluated. In general, LEW/N rats are highly responsive to challenge with carrageenan, whereas F344 rats are not. The strong local inflammatory responses to carrageenan are primarily exhibited by female LEW/N rats. The intensity of local inflammatory responses of LEW/N rats changes with the rat age, the highest exhibited by LEW/N rats up to 3 months of age, thereafter the carrageenan-induced inflammatory responses decline. Our results indicate that peripheral injection of carrageenan induces strong systemic immune component. After carrageenan injection, increases in CD8 and naive CD4 blood lymphocytes are seen. Although the carrageenan challenge does not change CD4 blood lymphocytes in both LEW/N and F344/N rat strains, LEW/N rats exhibit higher levels of CD4 cells than F344/N rats. Additionally, LEW/N rats demonstrated lower levels of B cells and higher naive CD4 lymphocytes. Carrageenan challenges induce significant increases in plasma corticosterone response in F344/N rats, as well as increases in LEW/N rats 1 h after injection. Our data stress the importance of rat age and gender in experiments studying inflammatory responses.

Toxicity studies of paclitaxel. (III)--Six-month intermittent intravenous toxicity in rats

Paclitaxel, an antineoplastic agent, was intravenously given to Crj:CD (SD) rats of both sexes at 0 (saline), 0 (vehicle), 0.3 (low dose), 1.0 (intermediate dose) and 3.3 (high dose) mg/kg at seven-day interval over a six-month period (total of 27 doses) to investigate its repeated dose toxicity. The results obtained are summarized as follows: 1. No deaths occurred in this study. Vehicle-related salivation was seen for some high dose and vehicle control rats. Soiling of the perigenital region was observed for some high dose females. 2. Red blood cell count, hemoglobin, hematocrit and white blood cell count were decreased for high dose rats. Reticulocyte count was increased and relative neutrophil count was decreased for high dose males. 3. Relative erythroid and myeloid cell count were decreased for high dose rats in bone marrow smear examinations. 4. Bone marrow hypoplasia and splenic hemosiderosis were observed for high dose rats, and thymic atrophy and lymphoid depletion were seen for some high dose males. Based on the these results, the no-toxic effect dose of paclitaxel was estimated to be 1.0 mg/kg in rats under this study condition.

Bronchoalveolar lavage in ponies with heaves during disease remission.

Bronchoalveolar ravage (BAL) fluid was collected from control ponies and from affected ponies with heaves which were in clinical remission. Significant differences were not detected between the two groups in total nucleated cell count or relative percentages (relative cell counts) of small lymphocytes, large mononuclear cells, eosinophils, or mast cells. The mean relative neutrophil count of BAL fluid from the affected group was statistically greater than that of controls; however, the difference between the two groups was not sufficient to be of diagnostic significance.

Somatic cell counts in relation to infection status of the goat udder.

Bacteriological analyses, cell counts using the Fossomatic method and California Mastitis Test were performed on 1523 goat milk samples taken aseptically at monthly intervals throughout lactation from three goat herds. Of the goat udders, 81.4% were infected, minor pathogens being the most frequent isolates (65.7%). Differences in the level of infection by minor pathogens were found between herds. Cell counts were influenced by stage of lactation and intramammary infection. Cell counts greater than 10(6) cells/ml were found in 80% of milk samples infected by major pathogens and in 45% infected by minor pathogens. About 81% of udders infected with major pathogens gave California Mastitis Test scores of 2 and 3, compared with 20% for uninfected goats. A high proportion (65%) of udders infected with minor pathogens also produced scores of 2 and 3. A significant positive correlation was found between the California Mastitis Test and cell counts. The use of cell counts for the detection of abnormal goat milk is discussed.

The associations of race, cigarette smoking, and smoking cessation to measures of the immune system in middle-aged men

To estimate the association between the immunologic responses of the cell-mediated and humoral systems and race or tobacco smoking, we used data from the Vietnam Experience Study conducted by the Centers for Disease Control. That study, done from 1985 to 1986, was based on a random sample of 4462 male, Vietnam-era, U.S. veterans. Racial groups were white, black, Hispanic, Asian, and American Indian. We used linear regression to evaluate how (i) the race of the subjects, (ii) the number of pack-years of cigarettes the subjects smoked, and (iii) the smoking cessation of certain subjects were associated with their relative and absolute T, B, CD4, and CD8 lymphocyte counts and immunoglobulin A (IgA), IgM, and IgG levels. The results indicated that immune status was associated with race and smoking history and that, generally, the associations remained after adjustment for covariates. For example, the average IgA level and absolute CD8 lymphocyte count for blacks were, respectively, 19 and 16% higher than those for whites. On the other hand, smokers had lower immunoglobulin levels and relative CD8 cell counts and higher counts for other lymphocytes of the cell-mediated system than nonsmokers. For example, the average absolute B count of heavy smokers was 37% higher than that of nonsmokers. The pattern after cigarette smoking cessation was consistent with a reversible effect of smoking and a return toward immune levels of nonsmokers.

Flow cytometric evaluation of cell‐cycle progression in ethyl methanesulfonate and methyl methanesulfonate‐exposed P3 Cells: Relationship to the induction of sister‐chromatid exchanges and cellular toxicity

In order to determine the relationships among the reduction in relative cloning efficiency (RCE), sister-chromatid exchange (SCE) formation, and interference with progression through the cell-cycle, human teratocarcinoma-derived (P3) cells were exposed to either ethyl methanesulfonate or to methyl methanesulfonate. The relationship between SCE and toxicity was quantified, the progression through the cell-cycle was evaluated with flow cytometric methods, and the effects of these chemicals on cell growth and average generation time (AGT) were determined. A strong correlation existed between RCE and SCE (r = -0.978, p less than .001) which was accompanied by an inhibition of growth as evidenced by a significant (p less than .0001) negative linear effect of concentration on the relative cell count from 24 to 72 hours after exposure and by a concentration-dependent increase (p less than .0001) in the AGT. Delays in the transit through S-phase were evident 4 hours after exposure to toxic concentrations of either carcinogen and by 8 to 12 hours post-exposure at the lower concentrations. Increases in the percentage of nuclei in G2 + M, indicative of G2 arrest, occurred from 12 to 24 hours after exposure. One interpretation of these results is that those effects of EMS and MMS exposure which result in S-phase delay and G2 arrest may be those elements common to the induction of SCE and cellular toxicity.