Circannual variation of cell proliferation in lymphoid organs and bone marrow of BDF1 male mice on three lighting regimens.

BDF1 male mice, which had been raised for several generations on a lighting regimen of LD 12:12, were studied. Experiments were conducted over 24 h spans during winter, spring, summer, and fall. For 3-4 weeks prior to each study, one-third of the animals were kept on a lighting regimen of 8 h of light alternating with 16 h of darkness (LD 8:16), one-third was kept on a regimen of LD 12:12, and one-third was kept on a regimen of LD 16:8. Subgroups of mice on all three lighting regimens were killed at 4 h intervals over a 24 h span. At 20 minutes prior to sacrifice, the animals received 5 microCi of 3H-thymidine/0.2 ml/20 gm body weight intraperitoneally. The thymidine uptake in DNA (DPM[3H]/microgram DNA) was studied as an index of cell proliferation in the thymus, inguinal lymph node, spleen, femur, and a segment of the lumbar vertebral column. A circannual variation of 3H-thymidine uptake in DNA was found in all organs irrespective of the lighting regimen under which the animals were kept. The timing of the circannual variation, however, varied among the organs. In the thymus, the lowest thymidine uptake occurred during summer, with higher thymidine uptake during the other three seasons. In the inguinal lymph node, the peak in thymidine uptake was in the spring, with lower values during the other three seasons, the lowest during summer. In the spleen, the highest thymidine uptake occurred in the mice on all three lighting regimens during fall, with lower uptake during winter, spring, and summer. In the bone marrow of both the femur and the vertebral column, the thymidine uptake was high in winter and fall and low in spring and summer. Serum corticosterone measurements were available in winter, spring, and fall, and they showed statistically significant lower values in winter and fall than in spring. The conclusion was drawn that circannual rhythms of 3H-thymidine uptake in the DNA of the thymus, spleen, lymph nodes, and bone marrow are found in mice reared for generations under a LD 12:12 lighting regimen and persist if the animals are kept under a regimen of LD 8:16 or LD 16:8 for 3-4 weeks prior to sacrifice.

Cancer marker assessment: case report on salivary and urinary CEA.

A circadian rhythm is demonstrated for salivary CEA in a clinically healthy man who collected unstimulated saliva samples around the clock for 4 days. Its acrophase occurs around 07:00, slightly later than for patients with colon cancer. A circadian rhythm of borderline statistical significance is found for the urinary excretion rate of CEA determined during the same span by this patient. It has an acrophase occurring around 15:00, differing from that of salivary CEA. Although CEA may have only limited value to assess tumor burden, even when determined in blood, rhythm characteristics of tumor markers such as CEA await applications for guiding treatment timing and for detecting earliest chronome alterations not only in the case of an overt cancer but as a feature of predisease and/or disease risk elevation.

Comparison of gentamicin pharmacokinetic parameters determined by fluorescence polarization immunoassay and latex agglutination methods.

Fluorescence polarization immunoassay (FPIA) (TDx, Abbott Laboratories Diagnostics, Irvin, TX, U.S.A.) is commonly utilized for quantitative determination of gentamicin serum concentrations. Recently, automated homogeneous latex agglutination (LA) (Technicon Immuno-1, Miles Diagnostics Division, Tarrytown, NY, U.S.A.) for the quantitative determination of gentamicin serum concentrations has been approved for commercial use. The purpose of this study was to determine whether gentamicin serum concentration-time data from the same patients assayed by FPIA and LA would produce the same estimates for half-life, elimination rate constant, distribution volume, drug clearance, dosage interval, and dose. A total of 70 pre- and postinfusion serum samples were obtained from 19 patients. Each sample was divided into two aliquots; one was assayed by FPIA and the other by LA. The correlation coefficient between the two assay methods was 0.99 (y = 1.03x - 0.05). The mean differences for half-life, volume of distribution, elimination rate constant, total body clearance, and gentamicin dosage were 0.13, 0.32, 0.66, -0.99, and 0.03%, respectively. No statistically significant differences were seen in calculated gentamicin pharmacokinetic parameters (p < 0.05). Pharmacokinetic parameters and dosage recommendations derived from FPIA and the LA assay using pre- and postinfusion serum concentration-time data appear interchangeable and do not result in differences between gentamicin dosing regimens.

Chronobiology in laboratory medicine.

A critical amount of information has accumulated over the last decades to allow the application of chronobiology to clinical and laboratory medicine. The tasks faced in laboratory medicine include the quantitative measurement of the multifrequency human time structure in health and disease. For this purpose, it is essential to choose an adequate sample size in order to obtain meaningful results and quantitative endpoints which can be interpreted by inferential statistical techniques. No statistical technique is applicable for all purposes and it is essential that the assumptions underlying each technique and its limitation are well known to the investigator. The multifrequency nature of the human time structure has to be kept in mind in order to avoid erroneous results. Time qualified reference ranges have to be established for high amplitude rhythms. Circadian and/or circannual rhythm alterations have been described as group phenomenon in subjects with epidemiologically determined risk states for common diseases, but will require much further studies for the application to individual subjects. Rhythm parameters are new endpoints in the evaluation of the human time structure in health. Alterations of these parameters may occur as cause or as consequence of disease. Recognition of rhythm abnormalities in disease are critical for a meaningful application of chronopharmacology. Time dependent changes in pharmacokinetics and pharmacodynamics have to be taken into account in the interpretation of drug level determinations. A considerable degree of individuality of timing has been documented in some frequencies. This individuality and the rhythm abnormalities found in disease require the study of reference or marker rhythms. If the complexity of the human time structure is clearly understood and its study pursued in a critical manner with quantitative endpoints, chronobiology opens a new dimension in laboratory and clinical medicine.

Chronobiology of pituitary-thyroid functions.

One hundred ninety four children, 11 +/- 1.5 years of age and 166 elderly men and women, 77 +/- 8 years of age were studied over one or (in the case of some of the elderly subjects) over several (up to 4) 24-hours spans. All subjects were diurnally active and rested at night and followed their regular three meal pattern. The subjects were studied in subgroups of 20-25 during all four seasons of the year. During each study, blood was collected at 4 hour intervals over one 24-hour span (6 samples). Circadian and circannual variations were found and described by cosinor analysis in the children as well as in the elderly subjects. The children with endemic goiter (134) as compared to those without endemic goiter (60) showed a slight circadian phase advance in plasma total and free T3, a lower circadian amplitude of total T4 concentrations and the absence of a detectable circadian rhythm in free T4. The children with goiter showed a phase delay in serum TBG. There was no difference between the children with and without goiter in the circadian MESOR of any thyroid parameter or of TSH. The children with endemic goiter in the region of Dimbovita, Romania, are in clinical and biochemically euthyroid condition with some slight poral abnormalities of thyroid function. Seasonal variations in children and elderly patients showed the highest values of TSH during summer and fall, while the highest values in the plasma concentrations of thyroid hormones were found during the cold season of the years. Thyroglobulin in the children showed a circadian rhythm but no seasonal variation.

Human mesor-hypertensive chronorisk.

Twelve endocrine variables in blood from a small number of clinically healthy adult women were sampled systematically around the clock and the seasons. Pattern discrimination methods singled out certain hormone values in certain seasons as classifiers for a high vs low risk of developing diseases associated with a high blood pressure. Further evidence in support of such classifiers is obtained on data from adolescent, menstrually cycling young adults and post-menopausal women, here analyzed as pool of series, with the scope of the data from any one age group greatly extended by a resampling procedure, namely, by bootstrapping. This mathematical approach was carried out on data series around the clock and seasons on several hormones as well as systolic and diastolic blood pressure. Classifier roles were strongly supported for plasma aldosterone and thyroid stimulating hormone, originally by an analysis of variance and, in the case of aldosterone, by circannual cosinor analysis and by numerical resampling. Circannual bootstrapping, a procedure recommended for broad routine use as a safeguard for hypothesis testing, was also done for plasma cortisol, dehydro-epi-androsterone sulfate and prolactin, variables for which (parametric) analyses of variance and cosinors did not reveal any difference between groups at high and low cardiovascular risk. In these instances, bootstrapping results are tentative and await further analyses. Results show the ability of circannual bootstrapping to detect outliers. Identification of classifiers provides cost-effective endocrine checks complementing the targeted automatic monitoring of blood pressure. Circannual indices for risk evaluation are, however, costly in several ways since it takes at least a year and quite a few samples to estimate them reliably. Accordingly, we also extended the scope of previous results by the application of an added procedure for circadian bootstrapping. With circadian as well as circannual bootstrapping, we here illustrate a major potential component of a system of chrono-engineering for health maintenance. This system should start with focus on the newborn. The results on adults here analyzed are likely to be more prominent in the neonate, to the extent that they are genetic in origin, yet amenable to modification by the extra-uterine environment.

Circadian rhythm parameters of endocrine functions in elderly subjects during the seventh to the ninth decade of life.

The circadian rhythm of 17 endocrine parameters (ACTH, aldosterone, cortisol, C-peptide, DHEA-S, FSH, growth hormone, insulin, LH, 17-OH progesterone, prolactin, testosterone, total T3, total T4 and TSH and estradiol and progesterone in women only) were studied in 63 clinically apparently healthy men (124 profiles) and 86 women (154 profiles) during the 7th to 9th decade of life. The subjects lived under very uniform conditions in a home for the aged with their daily schedule standardized by institutional routine with rest at night on the average from 21:30 to 06:30 local time and 3 daily meals at 08:30, 13:00 and 18:30. Blood was drawn over a 24-h span at 4-h intervals. Circadian periodicity was ascertained and the rhythm parameters quantified by cosinor analysis. In clinically healthy elderly subjects, circadian periodicity persisted in most parameters studied well into the 9th decade of life. The timing of the circadian rhythm was comparable between subjects in their 7th decade and 9th decade of life with the exception of cortisol and DHEA-S, which showed a phase advance with advancing age. A decrease in circadian amplitude is limited during this part of the human life span to only a few of the functions investigated and with the exception of prolactin in the women, a decrease in amplitude did accompany a decrease in MESOR.

Circadian and seasonal variations in iodine excretion in children with and without endemic goiter.

The urinary iodine excretion was measured in 193 children 11 +/- 1.5 years of age living in the endemic goiter area of Dîmbovita, Romania. One hundred and thirty four of the children showed some degree of endemic goiter, 59 showed none. All children followed a diurnal activity pattern with rest during the night. They received their usual iodine supplement of 1 gm potassium iodide once a week during the school year (which included the time of all measurements made). Urine was collected in six 4-hour samples over a 24-hour span. The examinations were conducted during the months of March, June, September and December. Iodine was determined by an automated ceric ion arsenic acid method using a Technicon Autoanalyzer. Circadian and seasonal variations of urine volume and iodine excretion were statistically verified by the cosinor technique and the seasonal variations also by one way analysis of variance using the circadian means as input. A comparable circadian rhythm of iodine excretion was found in the children with and without endemic goiter, with an acrophase during the evening (20:16 with a 95% C.I., from 19:32 to 21:04). The circadian rhythm in iodine excretion has to be taken into account whenever an estimate of the 24-hour excretion is attempted from a sample covering less than the entire 24-hour span. There was a statistically significant seasonal variation of the 24-hour iodine excretion in the boys with and without endemic goiter and in the group as a whole. The 24-hour iodine excretion during March was 102 +/- 6 mcg, during June 81 +/- 4 mcg, during September 79 +/- 3 mcg and during December 102 +/- 7 mcg. The average 24-hour iodine excretion pooled over all seasons was 91 +/- 3 mcg/24 hrs in the children with and 91 +/- 5 mcg/24 hrs in the children without endemic goiter. During March and December the iodine excretion indicates an iodine intake not usually associated with a high prevalence of endemic goiter. However, during the months of June and September (and presumably even more during the months of July and August when during summer vacation no iodine supplementation was given in school) the 24-hour iodine excretion indicates some degree of iodine deficiency. The seasonal variation in urinary iodine excretion thus points to a time when increased iodine prophylaxis may be of value.

Chronobiology, growth hormone and healthy and malignant growth.

Chronobiology, the computer-aided science (logos) of life (bios) in time (chronos), provides novel concepts, tools and facts for those concerned with mitosis, growth and growth hormone (GH). GH concentrations in human plasma demonstrate a statistically significant circadian rhythm on a 6h as well as on a 24h rest-activity cycle. On a 24h routine of light (L) and darkness (D), alternating at 12h intervals, and in continuous D, circadian mitotic rhythms in mice persist as a feature of growth or regeneration. A circadian cell cycle commences with an increase in phospholipid labeling, followed by an increase in cytoplasmic RNA formation, preceding, in regular sequences, an increase in nuclear DNA formation and the next mitotic peak in those cells that are dividing in a growing or regenerating (reversibly "post-mitotic') rodent liver. About 5-day (presumably estral) and about 7-day (circaseptan) components as well as circadians are resolved as a spectrum of mitotic rhythms in rodent cornea. The effects of hormones such as GH or a synthetic ACTH analogue, ACTH 1-17, depend upon the circadian cell cycle stages when the agent is administered. No effect or statistically significant effect can be the result only of 1) the timing of a fixed dose of GH or 2) of the timing of the samples taken to investigate any effect. For both the timing of administration and the assessment of effects, a multifrequency spectrum of rhythms, if taken into account, can provide (in lieu of a considerable and often formidable source of variation) a new critical dimension of growth and development.

Non-immune fetal hydrops with hepatic hemangioendothelioma and Kasabach-Merritt syndrome: a case report.

A premature infant presented with non-immune hydrops fetalis, a liver mass, thrombocytopenia, and hypofibrinogenemia. Histologic examination of the liver tumor showed an infantile hemangioendothelioma. The clinical features of this case can be explained by anemia, hypoalbuminemia, and coagulopathy. The association with Kasabach-Merritt syndrome, the pathophysiology of non-immune hydrops fetalis, and primary hepatic neoplasms of the neonate are discussed.

Circadian hematologic time structure in the elderly.

Twenty-three clinically healthy, diurnally active elderly subjects, 71 +/- 5 years of age were studied over a 24-hr span (six samples). Complete blood counts and differential counts were done (Ortho ELT-8, Wright stained smears). The circadian rhythm parameters of the hematologic variables in the elderly subjects were compared with reference values obtained from a larger group of clinically healthy young adult and adult subjects studied independently. The data were analyzed by cosinor and the Bingham test. Circadian rhythms in the number of circulating formed elements in the peripheral blood persist in the aged. In comparison with the young adult, the elderly subjects show differences in the timing (phase advance) of the circadian rhythms in circulating neutrophil leukocytes and lymphocytes, a decrease in the circadian amplitude of circulating platelets, a decrease in circadian rhythm adjusted mean (mesor) in the red cell count, and in the neutrophil band forms.

Natural killer cell activity: age, estrous- and circadian-stage dependence and inverse correlation with metastatic potential.

The timing within the estrous cycle of surgical removal of a transplanted murine mammary tumor profoundly influences the frequency of pulmonary metastases. We investigated the potential role of the immune response in this phenomenon by measuring splenic natural killer (NK) cell activity and interleukin-2 (IL-2) production in syngeneic tumor-free mice of two age groups at each of two circadian times and in each of four estrous stages. Estrous stage was determined by assessment of vaginal smear cellularity immediately prior to killing and spleen harvest. In a single-cell splenocyte preparation, NK cytotoxicity against a standard tumor cell target was assessed using a radiolabeled chromium release assay while IL-2 activity was determined in a bioassay utilizing the IL-2-dependent CTLL-2 cell line. Mice from the younger group were found to have eight-fold higher NK activity and 35% greater IL-2 production. After normalization of NK and IL-2 values for age, a highly statistically significant difference in NK activity was found among the four estrous and between the two circadian stages of sacrifice. NK activity was greater during the daily resting span across every estrous stage. IL-2 values were highest in diestrus and proestrus when sampled in the light span and in estrus-metestrus when sampled in the dark. The stages within the fertility cycle associated with lowest metastatic potential (proestrus/estrus) correspond precisely with those of highest splenocyte NK activity. These results indicate that an important component of the cellular immune response varies rhythmically both during the fertility and circadian cycles of the host. The rhythmic changes in NK activity may be in part responsible for the similarly rhythmic frequency of postsurgical metastatic dissemination.

Splenocyte natural killer cell activity and metastatic potential are inversely dependent on estrous stage.

We have previously shown that the timing of surgical removal of an estrogen-receptor-bearing mammary adenocarcinoma within the estrous cycle of the female C3HeB/FeJ mouse profoundly influences the frequency of subsequent tumor cell metastasis. In order to investigate the role of the immune response in this phenomenon, we measured splenic natural killer (NK) cell activity and interleukin-2 (IL-2) production in 80 female cycling mice, 16 to 18 weeks old, assigned to one of four estrous stages as determined by relative quantity of vaginal cellularity; proestrus, estrus, metestrus, and diestrus. After prolonged synchronization on 12-hours-on, 12-hours-off light-dark circadian schedules, daily vaginal smears were obtained for 2 weeks to characterize estrous cycling. On the day the animals were killed, vaginal smears were performed and single-cell suspensions were prepared from the harvested spleens. Direct cytotoxicity of spleen cells against the YAC tumor target was assessed immediately in a 3 1/2 hour 51Cr release assay and expressed as NK activity in lytic units (LU 20%). IL-2 production was determined in a bioassay with the IL-2-dependent CTLL-2 cell line. Significant differences in NK activity among estrous stages mimicking the variation found in frequency of surgical cure from mammary adenocarcinoma were observed (p = 0.035; one-way analysis of variance), with the time of lowest metastatic potential corresponding precisely with the time of highest splenocyte NK activity. These both occurred during the proestrus and estrus stages, characterized by high fertility, ovulation, and peak FSH, LH, and estrogen concentrations. In addition, NK activity was found to correlate significantly with IL-2 production (r = 0.4, p less than 0.0005). These results indicate that important components of the cellular immune response to cancer vary rhythmically with hormonal changes in the host and may represent one of the factors affecting the delicate balance between host and tumor that alters the frequency of postsurgical metastatic dissemination.

Plasma and urinary catecholamines in salt-sensitive idiopathic hypertension.

Nineteen patients with normal renin idiopathic hypertension were arbitrarily classified as salt-sensitive or salt-resistant depending on whether their mean arterial pressure did or did not increase by 8% or more when sodium intake was increased. The responses of the two subsets and of five normal subjects to sodium intakes of 9, 109, and 249 mEq/day given for 7 days were as follows: The salt-sensitive subjects retained more sodium than normal and plasma or urinary norepinephrine did not decrease when they were given a high sodium intake; urinary dopamine was normal but did not increase normally when sodium intake was increased. The salt-resistant subjects excreted sodium normally and plasma and urinary norepinephrine was decreased by 30 and 37%, respectively, when they were given a high sodium intake; urinary dopamine was supernormal and did not increase further when sodium intake was increased. Cumulative sodium retention during the high sodium intake was directly related to the percentage of change in plasma norepinephrine in the hypertensive subjects, suggesting that renal adrenergic activity was a factor in the impaired sodium excretion in the salt-sensitive patients. Cumulative sodium retention and the percentage of change in plasma norepinephrine were inversely related to urinary dopamine in the hypertensive subjects, suggesting that increased formation of dopamine in renal and neural tissue in the salt-resistant subjects may have been responsible for the differences between the subsets in renal and adrenergic responses to a high sodium intake.(ABSTRACT TRUNCATED AT 250 WORDS)