Immune regulation of the L5178Y murine tumor-dormant state. I. In vivo and in vitro effects of prostaglandin E2 and indomethacin on tumor cell growth.

Immunization and intraperitoneal challenge of DBA/2 mice with L5178Y lymphoma cells results in the suppression and maintenance of the L5178Y cells in a tumor-dormant state in the peritoneal cavity for many months. Cell-mediated immune responses involving lymphocytes and macrophages are involved in maintenance of the tumor-dormant state. Macrophages that have increased immunosuppressive activity and that produce increased amounts of PGE2 appear in the peritoneal cavity of tumor-dormant mice before the breakdown of the tumor-dormant state and formation of ascitic tumors. We report here that the tumor-dormant state can be terminated with formation of ascitic tumors by treatment of tumor-dormant mice with PGE2. Treatment with indomethacin results in inhibition of tumor cell growth and elimination of all recoverable tumor cells. Cultures of peritoneal cells (PC) from mice harboring L5178Y cells in a tumor-dormant state were used to analyze the PGE2 and indomethacin effects. Tumor cells did not grow out in the high-cell density PC cultures prepared from many tumor-dormant mice, but addition of PGE2 to these cultures resulted in tumor cell growth. The tumor cell growth that did occur in the PC cultures from some tumor-dormant mice was associated with PGE2 production by the associated host cells, and the addition of indomethacin to these cultures inhibited both PGE2 synthesis and tumor cell growth. Removal of plastic-adherent cells from the PC cultures eliminated the restraint on tumor cell growth. These experiments suggest that L5178Y tumor cells are maintained in a tumor-dormant state by host peritoneal cells, which are under PGE2 regulation.

The kappa-deleting element. Germline and rearranged, duplicated and dispersed forms.

Human light chain genes are used in a kappa before lambda order. Accompanying this hierarchy is the rearrangement of a kappa-deleting element (Kde) which eliminates the kappa locus before lambda gene rearrangement. In approximately 60% of rearrangements the Kde recombines at a conserved heptamer within the J kappa-C kappa intron. We demonstrated that aberrant V/J rearrangements possessing apparent "N" nucleotides existed 5' to the J kappa-Kde rearrangements. This suggests that the Kde may selectively eliminate nonfunctional V/J alleles. A kappa-producing cell that displayed the unusual finding of lambda gene rearrangement demonstrated a rearranged Kde. This rearrangement was a V kappa/Kde recombination and the heptamer-11 bp spacer-nonamer flanking the V kappa is the target site of the Kde 40% of the time. The mouse possesses a counterpart to the Kde (recombining sequence [RS]) and the highly conserved regions surround the heptamer-spacer-nonamer signals. No complete protein product was predicted from the germline Kde near its break-point and no consistent fusion product was predicted from either the V/Kde or V/J-Kde rearrangements. A distal portion of the Kde is duplicated and is present at 2q11 as well as 2p11. The evolutionary conservation of the kappa-elimination event, the duplication and maintenance of the Kde indicates that it has a function. A portion of the Kde may still prove to encode a trans-acting factor that directly affects lambda rearrangement. A certain role for the Kde is its site-specific rearrangement, which destroys ineffective kappa genes and sets the stage for lambda gene utilization.

Neuronal plasticity in primate telencephalon: anomalous projections induced by prenatal removal of frontal cortex.

When the dorsolateral prefrontal cortex in one hemisphere of a rhesus monkey is resected 6 weeks before birth and the fetus survives to postnatal ages, neurons of the corresponding cortex in the intact hemisphere issue a greatly expanded projection to the contralateral caudate nucleus in addition to a normal projection to the ipsilateral caudate. The enhancement of the crossed prefronto-caudate pathway after prenatal neurosurgery provides direct evidence for lesion-induced neuronal rearrangement in the primate telencephalon.

Nitrate synthesis in the germfree and conventional rat.

Metabolic balance studies show that germfree and conventional Sprague-Dawley rats synthesize nitrate. Equivalent results for germfree and conventional rats eliminate the microflora as obligatory components of nitrate production. Nitrate synthesis appears to be a mammalian process.

Delayed recovery of function following orbital prefrontal lesions in infant monkeys.

Monkeys given orbital prefrontal lesions at 1, 4, or 8 weeks of age exhibited a severe learning disability when they were tested at 1 year of age, but showed substantial recovery by the time they were 2 years old. These results suggest that the protracted maturation of intact cortical regions is important in recovery of function after early brain injury.

Cognitive deficit caused by regional depletion of dopamine in prefrontal cortex of rhesus monkey.

Depletion of dopamine in a circumscribed area of association cortex in rhesus monkeys produces an impairment in spatial delayed alternation performance nearly as severe as that caused by surgical ablation of the same area. This behavioral deficit can be pharmacologically reversed with dopamine agonists such as L-dopa and apomorphine. These data provide direct evidence that dopamine plays an important role in a specific cortical function.

A metronidazole metabolite in human urine and its risk.

Metronidazole is a drug used for the treatment of trichomonal vaginitis, amebiasis, giardiasis, and certain anaerobic bacterial infections in humans. Acetamide and N-(2-hydroxyethyl)oxamic acid are metabolites of metronidazole in the rat, and we find small amounts of both metabolites in the urine of human patients taking the drug. Although acetamide is carcinogenic for rats, we do not believe that our finding further defines metronidazole's risk for humans. That risk can only be estimated from surveillance of people previously exposed to the drug.

Sex-dependent behavioral effects of cerebral cortical lesions in the developing rhesus monkey.

Male rhesus monkeys with orbital prefrontal lesions were imtpaired on behavioral tests at 2(1/2) months of age whereas similar deficits were not detected in females with comparable lesions until 15 to 18 months of age. The results suggest that the maturation of a cortical region in the primate brain proceeds at different tempos in males and females.

Expression of Bcl-2 and Bcl-2-Ig fusion transcripts in normal and neoplastic cells.

We examined the expression of the Bcl-2 gene at chromosome segment 18q21, that is translocated into the Ig heavy chain gene locus in t(14;18) bearing lymphomas. Bcl-2, while B cell associated, is expressed in a variety of hematopoietic lineages including T cells. Bcl-2 mRNA levels are high during pre-B cell development, the time at which the t(14;18) translocation occurs, but are down regulated with maturation. Like certain other oncogenes, Bcl-2 is quiescent in resting B cells but up-regulated with B cell activation. Mature B cell lymphomas with a t(14;18) have log-folds more mRNA than matched counterparts without the translocation. A sensitive S1 protection assay revealed that all transcripts in t(14;18) B cells were Bcl-2-Ig fusion mRNAs and originated from the translocated allele. Thus, there is a marked deregulation of Bcl-2 when it is introduced into the Ig locus in t(14;18) lymphomas.

T cell receptor alpha-, beta-, and gamma-genes in T cell and pre-B cell acute lymphoblastic leukemia.

We examined alpha-, beta-, and gamma-T cell receptor (TCR) gene activation within acute lymphoblastic leukemias (ALLs) that represent early stages of B and T cell development. We wished to determine if TCR rearrangement and expression was lineage restricted, showed any developmental hierarchy, or could identify new subsets of T cells. Rearrangement of gamma and beta TCR genes occurred early in development but in no set order, and most T-ALLs (22/26) were of sufficient maturity to have rearranged both genes. T-ALLs preferentially rearranged C gamma 2 versus the C gamma 1 complex; no preference within the beta locus was apparent. Once rearranged, the beta TCR continued to be expressed (11/13), whereas the gamma TCR was rarely expressed (3/14). The alpha TCR was expressed only in more mature T-ALLs (8/14) that usually displayed T3. The 3A-1 T cell associated antigen appeared earliest in development followed by T11 and T3. Within pre-B cell ALL a higher incidence of lineage spillover was noted for gamma TCR rearrangements (8/17) than for beta rearrangements (3/17). This also contrasts with the only occasional rearrangement of immunoglobulin (Ig) heavy chains (3/25) in T-ALL. However, in pre-B ALL the pattern of gamma TCR usage was distinct from that of T cells, with the C gamma 1 complex utilized more frequently. Almost all ALLs could be classified as pre-B or T cell in type by combining Ig and TCR genes with monoclonal antibodies recognizing surface antigens, although examples of lineage duality were noted. Unique subpopulations of cells were discovered including two genetically uncommitted ALLs that failed to rearrange either Ig or TCR loci. Moreover, two T lymphoblasts were identified that possessed the T3 molecule but failed to express alpha plus beta TCR genes. These T-ALLs may represent a fortuitous transformation of T cell subsets with alternative T3-Ti complexes.

The reduction of N-hydroxy-4-acetylaminobiphenyl by the intestinal microflora of the rat.

The role of the intestinal flora in the conversion of N-hydroxy-4-acetyl-aminobiphenyl (N-OH-AABP) to 4-acetylaminobiphenyl has been examined. This reaction, which reverses the metabolic activation of the parent carcinogen, can be demonstrated in cultures of some bacteria indigenous to the intestinal microflora. These include cultures of Clostridium sp., Clostridium perfringens, Peptostreptococcus productus I, and Bacteroides fragilis ss. thetaiotaomicron and ss. vulgatus. In contrast, cultures of Lactobacillus plantarum and Escherichia coli show little or no capacity for this reaction. The reduction of N-OH-AABP is also carried out by homogenates of liver, kidney, and brain. On a weight basis, the cecal flora is considerably more active in reducing N-OH-AABP than are homogenates of tissues of the gastrointestinal tract. The cecal flora also has a greater activity for reducing N-OH-AABP than the stomach flora, an observation which may relate to the induction of tumors in the forestomach but not in the cecum of rats fed this compound. The products of the metabolism of N-OH-AABP have been compared in germ-free and conventional animals. Glucuronide conjugates of N-OH-AABP are found in the cecal contents and feces only of the germ-free rats, while 4-acetylaminobiphenyl is found in the feces only of conventional rats. These results suggest that the flora, by hydrolyzing glucuronides and reducing N-OH-AABP, may influence the level of metabolities of 4-acetylaminobiphenyl which are critical for carcinogenesis.

The effects of additional flora on the response of salmonella mutants lodged in the gastrointestinal tract.

A histidine auxotroph of Salmonella typhimurium, strain TA1538, will lodge for several months in the gastrointestinal tract of otherwise germ-free rats and of rats additionally associated with bacteria characteristic of the normal flora such as Lactobacillus plantarum and Bacteroides vulgatus. In the presence of the additional flora, the concentration of strain TA1538 is diminished in the stomach but not in the lower gastrointestinal tract or in the feces. Following the ingestion of 2-nitrofluorene, there is an increase in the concentration of revertants in the feces which reflects that observed in the colon and cecum. A dose-response relationship can be demonstrated between the amount of 2-nitrofluorene ingested and the concentration of revertants in the feces. A given dose of 2-nitrofluorene, however, produces fewer revertants in the feces of rats with the additional flora than in the feces of rats associated only with strain TA1538. It is not clear whether the decreased number of revertants in the feces in the presence of the additional flora is a result of metabolic transformations of 2-nitrofluorene by B. vulgatus, which can be demonstrated in vitro, or a result of the displacement of strain TA1538 from the stomach. The rat associated with strain TA1538, or other Ames tester strains, may be useful for detecting carcinogens as mutagens within the gastrointestinal tract and for determining the influence of various constituents of the bacterial flora on the concentration of mutagenic compounds.

Age-dependency of vascular phospholipid deacylation-reacylation in spontaneously hypertensive rats.

We have studied the temporal relation of phospholipid turnover and prostaglandin synthesis to the evolution of hypertensive vascular disease in the spontaneously hypertensive rat. The incorporation of arachidonate into aortic phospholipids, its release by phospholipase A2 and its utilization for prostaglandin synthesis were compared in spontaneously hypertensive and Wistar-Kyoto rats aged 7, 20 and 42 weeks. When expressed per mg of protein in the assay medium, arachidonate incorporation into aortic phospholipids decreased, while prostaglandin synthesis increased, with age in both rat strains. No significant differences were noted between hypertensive and normotensive animals at 7 weeks of age whereas both enhanced phospholipid turnover and prostaglandin synthesis was demonstrated in hypertensive rats at 20 and 42 weeks of age. The higher phospholipase activity in hypertensive aortas was associated with a significant increase in the capacity for exogenous lysophosphatide hydrolysis. Transacylation and reacylation of lysolecithin, however, were not significantly enhanced in hypertensive aortas. These biochemical changes accompany, and may be related to, structural modifications of the aortic wall in the course of hypertension.

Cost-effectiveness of a new short-stay unit to "rule out" acute myocardial infarction in low risk patients.

OBJECTIVES: This study attempted to determine the safety and costs of a new short-stay unit for low risk patients who may be admitted to a hospital to rule out myocardial infarction or ischemia. BACKGROUND: One strategy to reduce the costs of ruling out acute myocardial infarction in low risk patients is to develop alternatives to coronary care units. METHODS: The short-term and 6-month clinical outcomes and costs for 592 patients admitted to a short-stay coronary observation unit at Brigham and Women's Hospital with a low (< or = 10%) probability of acute myocardial infarction were compared with those for 924 consecutive comparison patients who were eligible for the same unit but were admitted to other hospital settings or discharged home. Actual costs were calculated using detailed cost-accounting methods that incorporated nursing intensity weights. RESULTS: The rate of major complications, recurrent myocardial infarction or cardiac death during 6 months after the initial presentation of the 592 patients admitted to the coronary observation unit was similar to that of the 924 comparison patients before and after adjustment for clinical factors influencing triage and initial diagnoses (adjusted relative risk 0.86, 95% confidence interval 0.49 to 1.53). Their median total costs (25th, 75th percentile) at 6 months ($1,927; 1,455, 3,650) were significantly lower than for comparison patients admitted to the wards $4,712; 1,868, 11,187), to stepdown or intermediate care units ($4,031; 2,069, 9,169) or to the coronary care unit ($9,201; 3,171, 20,011) but were higher than for comparison patients discharged home from the emergency department ($403; 403,927) before and after the same adjustments (all adjusted p < 0.0001). CONCLUSIONS: These data suggest that the coronary observation unit may be a safe and cost-saving alternative to current triage strategies for patients with a low risk of acute myocardial infarction admitted from the emergency department. Its replication in other hospitals should be tested.

The effect of risk factor reductions between 1981 and 1990 on coronary heart disease incidence, prevalence, mortality and cost.

OBJECTIVES: We sought to estimate the impact and cost-effectiveness of risk factor reductions between 1981 and 1990. BACKGROUND: Coronary heart disease (CHD) mortality rates have declined dramatically, partly as a result of reductions in CHD risk factors. METHODS: We used the CHD Policy Model, a validated computer-simulation model, to estimate the effects of actual investments made to change coronary risk factors between 1981 and 1990, as well as the impact of these changes on the incidence, prevalence, mortality and costs of CHD during this period and projected to 2015. RESULTS: Observed changes in risk factors between 1981 and 1990 resulted in a reduction of CHD deaths by approximately 430,000 and overall deaths by approximately 740,000, with an estimated cost-effectiveness of about $44,000 per year of life saved during this period, based on the estimated actual costs of the interventions used. However, because much of the benefit of risk factor reductions is delayed, the estimated reductions for the 35-year period of 1981 to 2015 were 3.6 million CHD deaths and 1.2 million non-CHD deaths, at a cost of only about $5,400 per year of life saved. CONCLUSIONS: Aggregate efforts to reduce risk factors between 1981 and 1990 have led to substantial reductions in CHD and should be well worth the cost, largely because of population-wide changes in life-style and habits. Some interventions are much better investments than others, and attention to such issues could lead to better use of resources and better outcomes in the future.

The relative influence of secondary versus primary prevention using the National Cholesterol Education Program Adult Treatment Panel II guidelines.

OBJECTIVES: This study was undertaken to project the population-wide effect of full implementation of the Adult Treatment Panel (ATP) II guidelines of the National Cholesterol Education Program (NCEP). BACKGROUND: The ATP II has proposed guidelines for cholesterol reduction, but the long-term epidemiologic influence of its components has not been fully examined. METHODS: We used a calibrated, validated simulation of the U.S. population, aged 35 to 84 years to estimate the potential for the NCEP guidelines, under varying assumptions, to reduce coronary heart disease morbidity and mortality and overall mortality from the years 2000 to 2020. RESULTS: Primary prevention would yield only about half of the benefits of secondary prevention despite requiring nearly twice as many person-years of treatment. The projected increase in quality-adjusted years of life per year of treatment for secondary prevention was 3- to 12-fold higher than for primary prevention. To yield population-wide epidemiologic benefits equivalent to NCEP recommendations for secondary prevention, primary prevention would require a nearly sixfold increase in the number of persons treated compared with NCEP recommendations. All benefits of universal success of the NCEP primary prevention "screen and treat" guidelines could be achieved by a 11 mg/dl (8%) population-wide reduction in low-density lipoprotein cholesterol levels among persons without preexisting coronary heart disease. CONCLUSIONS: The NCEP guidelines for targeted primary prevention can be a useful component of a rational public health strategy, but only as a complement to the more appealing strategies of secondary prevention and "across-the-board" programs to lower all cholesterol levels.

Elevation of serum uric acid as a clue to alcohol abuse.

Hyperuricemia in unselected admissions to a general hospital was found to be a significant correlate of alcohol abuse in male subjects. Unexplained elevation of the serum uric acid level should always stimulate inquiry for the behavioral features of alcoholism.

Effect of salt on prostaglandin metabolism in hypertension-prone and -resistant Dahl rats.

The effect of high salt intake on vascular and renomedullary prostaglandin (PG) synthesis was compared in Sprague-Dawley and salt-sensitive (S) and -resistant (R) Dahl rats. Animals were given a diet containing either 0.6% or 8% NaCl starting at 5 weeks of age, and were sacrificed 6 weeks later. Systolic blood pressure of S rats increased to 220 +/- 7 mm Hg but was unaffected in R and Sprague-Dawley rats. Prostaglandin synthesis was studied in aortic rings and renomedullary microsomes using 14C-arachidonate as substrate. [3H]PGE2 degradation was measured in the renocortical cytosol. In Sprague-Dawley and R rats, aortic PGI2 synthesis was not affected by high salt intake, while a significant increase compared to animals on 0.6% NaCl (from 608 +/- 84 to 992 +/-108 pmoles/60 min, p less than 0.05) was noted in S rats. Enhancement of PGI2 synthesis in S rats may be secondary to the hypertension. Salt-loading consistently stimulated renomedullary PGE2 synthesis in all three animal groups. S rats, however, had the lowest PG synthesis in renal medullas compared to Sprague-Dawley and R rats when placed on either diet. Thus, even after 6 weeks on high salt, S rats did not reach the levels of PGE2 synthesis seen in R or Sprague-Dawley rats on regular diet. The activity of cortical 15-hydroxyprostaglandin dehydrogenase was increased by salt-loading in S and Sprague-Dawley, but not in R rats. R rats had lower dehydrogenase activity than the other two groups when placed on either diet. The observed differences in PG synthesis and catabolism will tend to maintain the net output of renal PGs highest in R and lowest in S rats. These differences correlate with the reported differences in renal papillary flow between these two rat strains and may be relevant to their susceptibility or resistance to hypertension in response to salt.

Reliability of the toxic screen in drug overdose.

To determine the reliability of the laboratory in detecting drugs taken by overdosed patients, we evaluated laboratory performance on an unbiased sample of actual clinical specimens. Replicate serum and urine samples from a series of 20 consecutive clinically overdosed patients were sent to three commercial laboratories and one academic research laboratory for identification and quantification of intoxicating agents. All laboratories used the advance analytical techniques of gas chromatography-mass spectrometry. The results suggest that laboratories do not reliably identify drugs in the serum of overdosed patients, partly because of technical limitation, partly because of laboratory error, and possibly because of inadequate specimens. Drugs judged responsible for the overdose were identified in only 50% to 70% of the cases, depending on the laboratory. Reported concentrations sometimes varied over a 10-fold range.

An intricately patterned prefronto-caudate projection in the rhesus monkey.

The distribution of prefronto-caudate fibers in the caudate nucleus was studied autoradiographically in monkeys of various ages in which tritiated amino acids had been injected into the middle one-third of the length of the dorsal bank of the principal sulcus. The results indicate that, contrary to previous reports which had suggested a projection to only the head of the caudate nucleus, area 9 of Brodmann projects to the entire length of the nucleus. In the head of the caudate nucleus the cortico-caudate fibers are distributed in a pattern which is remarkable in two respects. First, the grains are not uniformly distributed but rather are segregated into clusters separated from one another by territories in which grain density does not exceed background. Second, individual clusters of grains, circular or elliptical in shape, surround grain free cores. These patterns of fiber distribution within the head of the nucleus are more sharply defined in newborn than in older monkeys. Our findings suggest that the caudate nucleus is organized more as an anatomic and functional mosaic than as the homogeneously organized structure that it is commonly considered to be.