Search

Background and Objective: The first circadian study of the 361st Medical Laboratory, USAR, was conducted in May 1969 during the Annual Military Training at Brook Army Hospital, Fort Sam Houston, Texas. The study was approved by the Surgeon General, 5th US Army, and was designed to establish a circadian database for 63 medically relevant variables of 13 young members of the Unit. The subsequent studies, all in the month of May, in 1979, 1988, 1993,1998, and 2003, followed the same protocol and were conducted at Edward Hines Jr., Veterans Administration Hospital, after approval by Human Studies Subcommittees. Since a reduction in Creatinine Clearance (CrCl) to the level of 60 ml/min/1.73m2 signals the onset of kidney malfunction and since a concurrent increase in blood pressure (BP) >140/90 mm Hg, contributes greatly to an unfavorable cardiovascular prognosis, it seemed prudent to examine possible changes in these and in other relevant variables in a group of young Army men, which may have developed over a 34 year period of time., Material and Methods: Thirteen US Army male volunteers (23-27y of age) served as subjects in the 1969 study. A majority of these men, two additional Army men and two non-military subjects, participated in subsequent studies: 1979 (7,2,1), 1988 (8,2,1), 1993 (5,4,1), 1998 (7,2,2), 2003 (7,2,1). In each study, subjects were admitted to a hospital ward, were given medical examination including a 12-lead electrocardiogram and followed the same Protocol. Lights "OUT" at 22:30h and "ON" at 06:30h. The meals, hospital 2400-calorie diets, were served at 17:30, 07:30 and at 13:30h. Vital signs were measured immediately after each 3h urine collections, around the clock, and bloods were collected every 3h. Blood, plasma, serum, saliva and urines were analyzed for numerous analytes including creatinine, using automated laboratory systems. Kidney functions were assessed using the measured and estimated glomerular filtration rates., Results: Over the 34y study span, 16 men provided sixty-one 24h profiles for CrCl-related variables (urine volume, creatinine, and serum creatinine) and fifty-eight profiles for BP. Using all normalized data, a significant circadian rhythm was found for each of these variables. Significant circadian variations in SBP, DBP, serum and urine creatinine, and urine volume, were evident with peak levels, on average, occurring in the evening hours., Conclusions: In healthy subjects, age was associated with an increase in SBP and urine volume and with a decrease in urine creatinine. In diabetic subjects, aging was associated with increases in both blood pressure and Creatinine Clearance. It is interesting to note that for the 3 subjects who at a later date developed diabetes, the CrCl levels were higher than the 5 age-matched controls during each study year, over the entire 34y observation span, including the period prior to diagnosis. Clin Ter 2008; 159(6):409-417.

Objective: To monitor the around-the-clock distribution of serum and urine concentrations of calcium, magnesium and eight trace elements and of those same elements in urine after their dialysis, and to statistically describe their circadian characteristics by chronobiological procedures., Materials and Methods: Serum and urine samples were collected every 3h over a single 24h period from eleven clinically-healthy male subjects, 41-60 years of age, and were analyzed for calcium (Ca), magnesium (Mg), iron (Fe), copper (Cu), zinc (Zn), lead (Pb), cadmium (Cd), cobalt (Co), chromium (Cr), and nickel (Ni). Urines were also sequentially dialyzed against ammonium-barbituric acid buffer at pH 7.35+/-0.02 using a 12.000-14.000 molecular weight exclusion sieve and then reanalyzed for the same elements. Urine concentrations were adjusted by urine volume to reflect a 3h excretion rate. Time-series were analyzed for circadian time-effect by ANOVA and for rhythm characteristics by the single cosinor fitting procedure., Results: The dialysis effectively removed 90% of total solids, 97% of urea, 92% creatinine, 72% uric acid, and essentially all of glucose. It also removed 99% of potassium (K), 96% of sodium (Na), 65% of Ca and P, 55% of Mg, 41% of Zn and 88% of Ni. A significant or borderline-significant 24h rhythm in serum was detected for Ca, Mg, Fe, Cu, Zn, Cd and Cr; in untreated urine for Ca, Fe, Cu, Zn, Ni, creatinine and volume; and in dialyzed urine for Ca, Fe, Cu, Zn, Pb, Cr, Cd and Ni. A 12h component was significant or borderline-significant in serum for Mg, Fe, Zn, and Cd; in untreated urine for volume, creatinine, Ca, Mg, Cu, and Ni; and in dialyzed urine for Ca, Mg, Fe, Cu, Zn, and Cr. In general, values in serum were lowest near the onset of sleep and highest in the first half of the day (between 02:28 and 13:56 h), while highest values in untreated or dialyzed urine were found several hours later in the day and at night., Conclusions: Significant circadian variations were found in levels of nearly every element that was measured in blood and urine of 11 healthy men, but with highest and lowest levels occurring at different times. This suggests not only that urine concentrations need to be adjusted for collection time interval and urine volume, but that different biological limits at different times of the 24h day should be applied for serum and urinary monitoring of trace elements. We also found that the non-dialyzable segments of these elements in urine represent metallo-moieties composed of proteinacious matter greater than 12,000-14,000 Daltons. Further studies would be of interest to reveal time specificity for metabolic functions associated with any of these trace elements.

Objective: The purpose of this study was to examine the circadian distribution of creatinine and uric acid clearances in subjects with Multiple Sclerosis., Materials and Methods: Eleven subjects with MS, 6 women (48+/-7y) and 5 men (58+/-5y) volunteered for this circadian study. Thirteen healthy females (39+/-11y) served as controls. Data of seven healthy male controls (64+/-8 y) were extracted from a similar circadian study conducted previously. Each MS patient, and each male control had blood samples drawn around the clock, at 3h intervals (8/24h), and each collected urines over 3h periods (8/24h). Each female control contributed only one blood sample and one complete 24h urine collection. Blood and urine samples were analyzed for a number of relevant analytes: ELAM, IL-6, NO, insulin, ACTH, aldosterone, cortisol, electrolytes, lymphocytes, monocytes including creatinine and uric acid clearances. Those were standardized to an average body surface area of 1.73 m2., Results: The relevant analytes demonstrated increased synthesis of insulin, IL-6, ELAM, monocytes, and reduced concentrations of serum NO. The creatinine clearances were significantly lower in MS females than in female controls, 63+/-22 vs.108+/-18 ml/min. They were also lower than those of MS males and male controls, 107.8+/-17, 97.5+/-8.2 ml/min. Uric acid clearances in MS females were also lower 6.9+/-2.4 vs. 10.5+/-4.4 ml/min. The uric acid clearance in MS males was higher than in male controls, 7.0+/-4.5 vs. 4.0+/-1.0 ml/min., Conclusions: The alterations in selected relevant analytes and the reduced creatinine and uric acid clearances in females but not in males, suggest a renal dysfunction in MS females. These observations may contribute to understanding better the mechanism of renal dysfunction in female patients and perhaps this may be an additional factor contributing to greater frequency of MS in females than in male subjects.

Objectives: To examine the circadian distribution and total 24h levels of urinary zinc (Zn) in same male subjects over an extended period of time in order to ascertain their relationship with aging., Materials and Methods: Eight young army volunteers served as subjects over a period of 29 years: 1969, 1979, 1988, 1998. By 1979 three of them became latent diabetics. Complete physical examination, anthropometric measurements and same procedural protocol was followed in each study. Samples were collected over 3 hour periods for 24 hours in the middle of each month of May. Urine aliquots were analyzed for creatinine, using conventional laboratory procedure. Zn was analyzed using Atomic Absorption Spectrophotometry in 1969, and 1979 and by Inductively Coupled Plasma, in 1988 and 1998., Results: Over the course of 29 years the circadian distribution of Zn was altered by decrease in amplitude in Zn levels, while the 24h concentrations of Zn decreased progressively with increasing age in healthy and diabetic subjects: Healthy; 966+/-130 microg at age of 29; 666+/-14 microg at 39; 511+/-80 microg at 48; and 555+/-71 microg at age of 58y; Diabetics exhibited similar trend; 1757+/-60 microg at age 28; 1253+/-40 microg at age 38, 1132+/-31 microg at 47, and 1025+/-11 microg at the age of 57. Anthropometric measurements in each study period revealed significant increases in diabetic subjects for body weight, body surface area, BMI and significant decrease in body heights of both groups., Conclusions: The daily excretion of urinary Zn over the 29 years period decreased by 42% in healthy and diabetic subjects. Although there appears to be a lack of a reliable index of intracellular Zn status to accurately monitor and control zinc deficiency in younger and older populations, the present data suggest that depletions of Zn are also evident in healthy aging subjects whose daily diet was not deficient in zinc.

Aims: The purpose of this study was to examine circadian distribution of selected cytokine levels (IL-2, IL-10, GM-CSF, TNF-alpha, and IFN-gamma) in serum of subjects with active Multiple Sclerosis (MS) and non-MS subjects., Materials and Methods: Six females (36-56y) and five males (52-68y) with active MS volunteered and consented for the study conducted at Special Diagnostic Ward of this hospital. All subjects gave their medical history and were given complete physical examination. Low purine meals were served at 16:30, 07:30 and 13:00 h. Lights were "OFF' at 22:30 hr and "ON" at 06:30h. Blood collections were made at 3h intervals over a 24h period of time. Six healthy male subjects (53-76y) subjects' data were obtained from a study conducted 3 years previously using the same procedural protocol. Cytokine assays were assessed using commercial enzyme-linked immuno-absorbent procedure. Time series of average data and the range of change between the highest and lowest concentrations are presented for MS subjects along with data from non-MS subjects., Results: IL-2, IL-10, and GM-CSF levels were significantly reduced in females with MS when compared with levels of healthy subjects while their IL-6 levels were increased. The IL-6, GM-CSF and TNF-alpha levels in males with MS were below detection limits. The TNF-alpha levels were essentially similar in MS females and healthy subjects., Conclusions: These preliminary studies, although with very small number of patients and healthy male controls appear to suggest that the circadian analysis of cytokines and other markers of immunity may have utility in understanding the pathogenesis of diseases like MS.

Objective: To evaluate associations between intraocular pressure (IOP) and blood pressure (BP), heart rate (HR), serum nitric oxide (NO), diurnal variations, diabetes and aging in data collected during 24h studies of men conducted over 34y., Materials and Methods: As part of the Medical Chronobiology Aging Project, male Army veterans, ages 22 to 81y, without a history of eye disease, were studied around-the-clock in May 1969 (n = 13), 1979 (n = 11), 1988 (n = 11), 1993 (n = 11), 1998 (n =12) and 2003 (n = 10). Measurements of IOP (R & L eyes, supine position), BP and HR (sitting position), and collection of blood were obtained every 3h (8 readings/24h) from 19:00h to 16:00h the next day. Individual time series were analyzed for circadian characteristics by the least-squares fit of a 24& 12h cosine. After normalizing all data to percent of mean to reduce inter-subject variability in levels, grouped data were analyzed for time-effect by ANOVA and for circadian rhythm by multiple component (24h&12h) cosine fitting. Individual 24h averages were analyzed by simple and multiple regression for relationships between IOP and systemic variables, diabetic status and age., Results: Over the 34y study span, 22 men provided sixty-three 24h profiles for IOP & HR, 61 for BP, and 21 for NO. Using all normalized data, a significant circadian rhythm was found for each variable at p <0.001. Circadian peaks (orthophases) are located in the late morning for IOP-R (10:20h) and IOP-L (10:52h), and in the evening for HR (18:52h), NO (20:00h), SBP (20:40h) and DBP (21:44h). An out-of-phase relationship of about 10h is noted on a group basis between IOP vs BP, HR and NO. The locations of individual circadian peaks for IOP-R were found around the clock, but with a significant predominance between 10:00 and 16:00h (day type), and 04:00-10:00h (morning type). In contrast, BP, HR and NO showed a significant clustering of evening type or night type peaks. The overall mean IOP for the right eye was slightly, but not significantly, higher than the left eye (17.60+/-0.21 vs 17.34+/-0.18 mmHg; p = 0.385), with a strong positive correlation between both eyes (R = 0.952, p <0.0001). IOP showed a significant positive correlation with SBP (R = 0.49, p <0.001), diabetic status (R = 0.47, p <0.001), age (R = 0.32, p = 0.011), and HR (R = 0.28, p = 0.031). A multiple regression using SBP, DBP, HR, age and diabetic status (5 men became diabetic over the 34y study span) as independent variables resulted in SBP being the strongest predictor of IOP (p = 0.0001), followed by DBP (p = 0.0103). After adjustment for BP, independent effects of age (p = 0.187), HR (p = 0.789) and diabetic status (p = 0.153) were eliminated from the prediction equation., Conclusions: The results of these studies reveal significant circadian variations in IOP, BP, HR and NO, with peak levels, on average, near noon for IOP and in the evening for BP, HR and NO. An increase in SBP was associated with an increase in IOP. While SBP and DBP are significant predictors of IOP levels, single measurements during regular clinic hours may not reveal the full functional relationship between the variables measured in our studies. Therefore, circadian information on total 24h patterns may contribute to the reliability of diagnosis and guide proper individualized timing of optimal patient management (e.g., for glaucoma, hypertension, diabetes, among other conditions).

A 7-year-old patient with Stage III-c ovarian cancer was subjected to 8 cycles, approximately four weeks apart, of chronobiologically-optimized treatments with combination of three anti-cancer agents: Four cycles at AM, Cytoxan and PM, cis-Platinum; four cycles at AM, Adriamycin and PM, cis-Platinum. A second look laporoscopy revealed clean intestines, no definite masses in the pelvis area although there was an apparent mass in the right upper pelvis and several slightly enlarged lymph nodes in the base of mesentery. Six cycles of Taxol were administered at about Noon. Seven months remission appeared evident as judged by no changes in monthly examinations, in blood work or in CA-125 marker levels which remained below 12 U/ml. During the eight month the CA-125 marker began to rise, 36 then to 52 U/ml. A second 6 cycle series of Taxol was initiated but the CA-125 marker continued to rise, 57, 65, 72, 86, and 87 U/ml level. The patient declined in spirit, in well-being and expired 2 weeks later, 31 months after the initial diagnosis of cancer. Blood hematology, chemistry, and cytokines variables were analyzed at about weekly intervals. Significant reductions in total WBC, neutrophiles and platelet levels were evident during the second week of all cycle treatments, while increases were noted in serum levels of IL-2, IL-6 and IL-10 following Cytoxan-cis-Platinum-Adriamycin, but not Taxol. After each infusion moderate and temporary increases in RBC levels were noted. The treatments impact on hematology, chemistry, cytokine variables and on the integrity of the patient, are presented and briefly discussed.

Hematology variables were measured in blood samples obtained every 3h (8/24h) from 10 multiple sclerosis (MS) patients and 34 healthy subjects and analyzed for circadian characteristics using the population multiple-components method. Red blood cell (RBC) and hemoglobin levels as well as hematocrits exhibited circadian rhythms with minimal amplitudes in healthy individuals and insignificant variability in the smaller group of MS patients. In contrast the total white blood cell (WBC) and platelet counts for MS patients and healthy individuals both showed significant circadian characteristics while the mean 24h WBC and platelet levels did not significantly differ between the two groups. When the different WBC subsets were examined independently, statistically significant circadian rhythms were seen for lymphocytes and eosinophils for both MS patients and healthy individuals and for neutrophils only in the latter. Moreover, the 24h mean levels of lymphocytes, basophils, and eosinophils were significantly higher for the healthy controls while those of monocytes were higher for the MS patients. However, of all the variables tested with significant circadian rhythms in both groups of individuals, only those of lymphocyte numbers exhibited different patterns with somewhat higher amplitude in healthy individuals and a peak level occurring over an hour after that of MS patients. These changes may be the reflection of a disturbance in the regulation of patterns of lymphocyte activity and migration in MS patients. In addition, the elevation in circulating monocytes in MS patients is consistent with the inflammatory nature of the disease.

Objective: To investigate circadian rhythm (CR) of urinary creatinine and 8-hydroxy-2-deoxyguanosine (8-OHdG) in patients with Multiple Sclerosis (MS) and to present concentrations of this DNA damage marker, 5 years prior to mastectomy, in one MS study subject, and 2 years prior to biopsy confirmed a carcinoma (CA) of the prostate in one non-MS subject., Materials and Methods: Eleven subjects with MS (6 women 36-52 years of age and 5 men 51-68 years) volunteered for this study, carried out at Edward Hines Jr., Medical Center. Subjects were offered a general hospital diet (2400 cal in total/24h) at 16:30h, 07:30h and 13:00h. The dark (sleep) phase of the light-dark cycle extended from 22:30h to 06:30h with brief awakening for sampling at 01:00h, and 04:00h. Urine samples were collected for consecutive 3h spans beginning at 16:00-19:00h and were analyzed for creatinine and 8-OHdG. Twelve men (including 3 with type 2 diabetes) provided 21 profiles according to the same protocol used for comparison. In addition, 10 healthy women provided 24h urine samples. Statistical analysis of data was performed using the Single-Cosinor and Population-Mean Cosinor., Results: A CR was detected for creatinine in healthy men (p < 0.001) but not for MS patients. Urinary creatinine concentrations were lower in MS women than in healthy women (p = 0.015) and were lower in MS women than in men healthy or with MS (p < 0.001): Women; MS 655 +/- 76; H 1381 +/- 316; Men, MS 1830 +/- 285; H 1532 +/- 265 mg/24h vol. A CR was evident in 8-OHdG in MS (p = 0.007) and in non-MS subjects (p < 0.001) with highest values occurring at about 16:45h. The average concentrations of 8-OHdG in MS patients were similar to those in healthy subjects: Women, MS 589 +/- 125; H 794 +/- 318; Men, MS 504 +/- 156; H 591 +/- 134 picomoles/kg bw/24h vol. The 8-OHdG concentrations of a MS patient, later diagnosed with breast cancer, were found to exceed the upper 95% prediction limit in health. An increased 8-OHdG level was also noted in a non-MS subject who 2 years later received a biopsy-confirmed diagnosis of prostate CA., Conclusions: Despite the small number of subjects in this study, a statistically significant CR was documented for 8-OHdG in urine of subjects with MS. Interestingly, the increased concentrations of DNA damage marker, the 8-OHdG, 5 years prior to mastectomy and the 2 years prior to affirmative diagnosis of prostate CA, could be the most significant clinical observations of this study. Follow-up studies of a larger population of subjects would, thus, be required to ascertain the predictive validity of such challenging observation.

Aim: We examined the circulating levels of iron and ferritin in serum of seven healthy and three insulin non-dependent diabetic (Type 2) males in order to compare their circadian characteristics., Methods: Blood samples were collected every 3h over a 24h period and were analyzed for serum iron and ferritin., Results: The mean Fe level was significantly higher in healthy than in diabetic subjects: 80.0 +/- 3.3 vs. 63.0 +/- 3.7 microg/dL. The ferritin level was significantly lower in healthy than in diabetic men: 79.8 +/- 4.7 vs. 186.3 +/- 110.5 microg/L. A significant (p < 0.001) time-effect was found by ANOVA and circadian rhythm was detected at p < 0.001 in all data sets when a 24h cosine was fitted to the normalized data. Acrophases were located in mid to late morning for Fe (11:30, vs. 09:22h) and for ferritin (11:10 vs. 11:46h)., Discussion: We concluded that there is significant circadian variation in both serum Fe and ferritin, with predictable peaks in the mid to late morning.

The free radical nitric oxide (NO*) is involved in a variety of diverse biological processes from acting as a vasodilator in the cardiovascular system to being the rate-limiting component in the production of peroxynitrite (ONOO-), a contributor to neurodegenerative disorders such as multiple sclerosis (MS). Uric acid (UA), the end product of purine metabolism in humans and a selective inhibitor of toxic reactions attributed to radicals formed by the interaction of ONOO- and CO2, is generally low in MS patients. We investigated the relationship between serum ONOO-, CO2, and UA in MS patients and normal controls by comparing the circadian characteristics of the NO* metabolites nitrite/ nitrate (NO), CO2, and UA. In this preliminary study, we found the functional relationship ascribed to the circadian timing of the peak and trough levels of NO, CO2, and UA in healthy subjects to be clearly altered in MS patients. These findings suggest that alterations in the temporal relationship between the 24h pattern in serum ONOO- formation and UA may either contribute to or reflect the disease processes in MS.

Seven clinically healthy, nondiabetic (ND) and four Type II diabetic (D) men were assessed for circadian rhythms in oxidative "stress markers." Blood samples were collected at 3h intervals for approximately 27 h beginning at 19:00h. Urine samples were collected every 3 h beginning with the 16:00h-19:00h sample. The dark (sleep) phase of the light-dark cycle extended from 22:30h to 06:30h, with brief awakening for sampling at 01:00h and 04:00h. Subjects were offered general hospital meals at 16:30h, 07:30h, and 13:30h (2400 cal in total/24h). Serum samples were analyzed for uric acid (UA) and nitrite (NO) concentrations, and urine samples were assayed for 8-hydroxydeoxyguanosine (8-OHdG), malondialdehyde (MDA), and 8-isoprostane (ISP). Data were analyzed statistically both by the population multiple-components method and by the analysis of variance (ANOVA). The 24h mean level of UA and NO was greater in D than in ND subjects (424 vs. 338 micromol/L and 39.2 vs. 12.7 microM, respectively). A significant circadian rhythm in UA (p = 0.001) and NO (p = 0.048) was evident in ND but not in D (p = 0.214 and 0.065). A circadian rhythm (p = 0.004, amplitude = 8.6 pmol/kgbw/3h urine vol.) was also evident in urine 8-OHdG of ND but not of D. The 24h mean levels of ND and D were comparable (76.8 vs. 65.7 pmol/kgbw/3h urine vol.). No circadian rhythm by population multiple-components was evident in MDA and ISP levels of ND subjects, or in 8-OHdG, MDA, and ISP in D. However, a significant time-effect was demonstrated by ANOVA in all variables and groups. The 24h mean of MDA and ISP in D was significantly greater than in ND (214 vs. 119 nmol/3h urine vol. and 622 vs. 465 ng/3h urine vol.). The peak concentrations of the three oxidative "stress markers" in urine, like those of serum NO, occurred early in the evening in both groups of men. This observation suggests a correlation between increased oxidative damage and increased rate of anabolic-catabolic events as evidenced by similarities in the timing of peak NO production and in parameters relevant to metabolic functions.

Circadian (8/24 hours) variations in serum nitric oxide (NO), total tissue factor pathway inhibitor (T-TFPI). and E-selectin levels were studied in healthy adults and in subjects with type II diabetes. We postulated a possibility a functional relationship between them because vascular endothelium is the primary site of their synthesis and functions. NO is released by the action of eNO synthase isoform and modulates physiologic responses (e.g., vascular dilation, relaxation, increasing blood flow, inhibition of platelet and white blood cell adhesion); T-TFPI, a coagulation inhibitor, is also released from endothelial cells, and is bound to plasma lipoproteins and to glycosaminoglycans; E-selectin is expressed on endothelial cells after activation by inflammatory cytokines (interleukin-1beta and tumor necrosis factor-alpha) and elevated levels have been reported in a variety of pathologic conditions, including diabetes. We found that obese diabetic subjects had greater mean concentrations of NO and E-selectin than healthy men, 39.25 versus 12.71 microM and 81.51 versus 26.03 ng/mL, respectively. The T-TFPI levels were essentially similar in both groups of men, 47.10 versus 48.76 ng/mL. We observed that the time of peak concentrations of T-TFPI and E-selectin was similar to the timing of NO trough levels, suggesting a possible functional relationship. It may be hypothesized, therefore, that the higher concentrations of NO, unbalanced by increases in T-TFPI and E-selectin, may result in increased vascular wall uptake of lipoproteins in diabetic subjects, who are at greater risk than healthy men for developing diffuse atherosclerosis.

Leptin, from the Greek leptos, meaning thin (in reference to its ability to reduce body fat stores), is a hormone secreted primarily by adipocytes. At one time, leptin was portrayed as a potential means of combating obesity. Recently, leptin has been identified as a potent inhibitor of bone formation, acting through the central nervous system. Since numerous studies clearly show that bone remodeling is circadian rhythmic with peak activity during sleep, it is of interest to explore circadian variability in serum leptin. Accordingly, circadian characteristics of serum leptin were examined in 7 clinically healthy men and 4 obese men with type II diabetes. Blood samples were collected for 24 h at 3 h intervals beginning at 19:00. The dark (sleep) phase of the light-dark cycle extended from 22:30 to 06:30, with brief awakening for sampling at 01:00 and 04:00. Subjects consumed general hospital meals (2400 calories) at 16:30, 07:30, and 13:30. Serum leptin levels were determined by a R&D Systems enzyme immunoassay technique. Data were analyzed by linear least-squares estimation using the population multiple components method. A statistically significant (P < .018) circadian rhythm modeled by a single 24 h cosine curve characterized the data of each group. The 24 h mean leptin level was statistically greater (P < .001) in the obese diabetic men than in the healthy men (9.47 +/- 0.66 ng/mL vs. 24.07 +/- 1.71 ng/mL, respectively). Higher leptin levels occurred between midnight and roughly 02:30, and lowest leptin levels occurred between noon and the early afternoon. The phasing of this rhythm is similar to the circadian rhythm in bone remodeling previously described. Our results suggest the findings from a single morning blood sampling for leptin may be misleading since it may underestimate the mean 24 h and peak concentrations of the hormone.

Serum homocysteine levels were examined in a 24-hour study of 7 healthy and 5 diabetic men, revealing a statistically significant circadian rhythm (p = 0.030), normal concentrations of 11.83 +/- 1.2 vs 12.99 +/- 1.2 micromol/L, with peak values occurring during the evening (10:37 P.M.) and lowest levels occurring during the morning. These findings imply that increased atherosclerotic risk in insulin-resistant diabetics during morning hours does not appear to be explained by differences in homocysteine levels in the normal population.

Background: A correlation has been reported between lipoprotein(a) [Lp(a)] concentration and risk for coronary artery disease. High concentrations of Lp(a) might be markers for vascular or tissue injury or might be associated with other genetic or environmental factors that can cause acute myocardial infarction., Methods: We measured the circadian characteristics of circulating Lp(a), fibrinogen, platelets, cholesterol, triglycerides, high-density lipoprotein cholesterol, and low-density lipoprotein cholesterol for a group of adult male volunteers who had no clinical symptoms. We obtained samples every 3 hours around the clock to assess the normal degree of variation within a 24-hour period and to test for similarities in circadian patterns and correlations with level of Lp(a)., Results: Each variable displayed a highly significant circadian rhythm. Lp(a), fibrinogen, high-density lipoprotein cholesterol, and low-density lipoprotein cholesterol peaked in the morning. Cholesterol and platelets peaked in the late afternoon, and triglycerides peaked in the evening., Conclusions: Although peak levels of Lp(a) and fibrinogen coincide with reported morning peak frequencies of myocardial infarction and stroke, the platelet peak appears to coincide with late afternoon peak frequencies of sudden cardiac death and fatal stroke. The data suggest that proper timing of single samples may improve the usefulness and accuracy of diagnosis, risk assessment, and therapy.

Circadian (24 h) rhythms of fibrinogen, interleukin-6 (IL-6), and platelet levels were studied in 11 males ages 46 to 72 years. Since there is a known circadian rhythm for fibrinogen and IL-6, we postulated that the peak level (acrophase) of fibrinogen would follow the acrophase of IL-6, based on the fact that IL-6 is the stimulus for fibrinogen production in the liver. Platelet levels were measured to show whether there was any correlation with the IL-6 acrophase because it has been reported that IL-6 affects megakaryocytes and platelets in dogs. We found that the acrophase for IL-6 occurred at 02:03 h and the acrophase for fibrinogen occurred at 09:16 h. Platelet counts peaked at 16:56 h. Thus, there was a positive correlation between IL-6 and fibrinogen acrophases and a negative correlation of each with the acrophase for platelets. The positive linkage of IL-6 with fibrinogen in this study suggests that suppression of IL-6 production would lower those peak fibrinogen levels that occur in the morning in association with arterial ischemic events. This could result in fewer arterial ischemic events, especially in the morning.

Long-acting natriuretic peptide (LANP), vessel dilator (VSDL), and atrial natriuretic factor (ANF) consisting of amino acids (aa) 1 to 30, 31 to 67, and 99 to 126, respectively, of the 126-aa ANF prohormone circulate in humans. Among the biologic properties of these peptides is the ability of ANF to decrease intracellular calcium concentrations. To determine if atrial natriuretic peptides are directly related to serum calcium and/or phosphate in healthy normocalcemic humans, we examined 21 24-hour profiles of VSDL, LANP, ANF, and serum calcium and phosphate in 14 healthy humans. VSDL, LANP, and ANF each had significant (P < .001) circadian rhythms, with peak concentrations late during sleep (at 4:00 AM) being nearly twice the concentrations in the afternoon and evening. Serum calcium and phosphate also had significant circadian rhythms (P < .001) with troughs nearly opposite to those of the atrial natriuretic peptides, suggesting that atrial peptides may be important in the modulation of the circadian rhythms of calcium and phosphate. The nearly identical circadian rhythms of the atrial natriuretic peptides and of parathyroid hormone (PTH) reported by others, along with evidence that PTH may increase atrial peptide release, suggest that some of the effects attributed to PTH may be mediated by atrial natriuretic peptides.

Long-acting natriuretic peptide (LANP), vessel dilator (VSDL), and atrial natriuretic factor (ANF) consisting of amino acids 1-30, 31-67, and 99-126 of the 126 amino acid ANF prohormone, respectively, circulate in humans and have potent natriuretic properties. To determine whether these peptides have a direct relationship to serum Na and/or Cl, we examined 21 24-hour profiles of these peptides and Na and Cl in 14 healthy humans. LANP, VSDL, ANF, and Cl had significant (p < 0.001) circadian rhythms with peak concentrations at 04.00 h. The circadian rhythm of serum Na was exactly opposite. Sodium correlated negatively with LANP (p = 0.021) and ANF (p = 0.007), while Cl correlated positively with LANP (p = 0.003) and VSDL (p = 0.001). These data suggest that the atrial peptides may be important for the maintenance of serum Na and Cl within their normal ranges and in the modulation of their daily circadian rhythms.

The variation in plasma fibrinogen level demonstrating prominent circaseptan and circannual cycles is clinically relevant. There is a correlation between increasing level of fibrinogen and other hemostatic factors and risk of myocardial infarction and sudden death. The circaseptan and circannual cycles in fibrinogen concentration described in this study may help to explain further the variation in frequency of coronary events. Furthermore, the recent demonstration of a circadian pattern in the efficacy of tissue plasminogen activator, with peak efficacy occurring at 2000 hours--10 hours after the peak incidence of myocardial infarction--implies that further patterns to coronary artery syndromes may be predicted and the treatment efficacy may rely on demonstrated circaseptan and circannual cycles of these events.

Objectives: To study the circadian relationship between serum prostate-specific antigen (PSA) and total testosterone in men without clinically evident prostate disease., Methods: Blood samples were collected every 3 hours for 24 hours (eight per subject) from 11 clinically healthy men, ages 46 to 72 years. PSA was also monitored once a week for 6 weeks in 16 additional healthy men. PSA, testosterone, and age were correlated by linear regression, and 3-hourly PSA and testosterone values normalized to percent of individual mean were analyzed for circadian rhythm by the least squares fit of a 24-hour cosine., Results: Mean PSA correlated positively (P < 0.001) and testosterone correlated negatively (P = 0.014) with age and inversely with each other (P < 0.001). The mean circadian range of change (ROC) from lowest to highest values for PSA was 0.37 +/- 0.07 ng/mL (28 +/- 9%), and for testosterone it was 202 +/- 23 ng/dL (53 +/- 7%). The mean ROC over 6 weeks was 0.32 +/- 0.04 ng/mL. A significant circadian rhythm was found for PSA (P = 0.011, amplitude = 5.4 +/- 1.8%, acrophase = 5:02 AM; 95% limits, 2:40 to 7:24 PM) and testosterone (P < 0.001, amplitude = 9.4 +/- 1.8%, acrophase = 8:38 AM; 95% limits, 7:12 to 10:04 AM)., Conclusions: The temporal relationship between circadian rhythms in PSA and testosterone suggests different physiologic states over the 24 hours, which may be of chronopharmacologic interest with regard to dosing time of drugs or hormonal treatments intended to affect prostate growth and function. Within-day variation in PSA is of little diagnostic significance and does not prevent accurate clinical classification when a single specimen is used.

The cytokine interleukin-6 (IL-6) is a multi-functional small peptide molecule that is produced by various types of lymphoid and non-lymphoid cells and plays a central role in hematopoiesis, host defense mechanisms, and acute phase reactions, including regulation of inflammatory and immune responses. A high-sensitivity ELISA assay was applied to serum (S) and urine (U) samples available from 10 men (median age = 50y, range = 46-71y) in order to compare circadian characteristics of IL-6 between assays and in 2 biological fluids. S and U samples were collected at 3-h intervals for 24hrs beginning at 19:00h on May 14, 1993 (8 samples/subj) and frozen at -25 degrees C until analysis. IL-6 in U was adjusted for time & volume (pg/hr) and assigned to midpoint of collection interval. A significant time-effect was found by ANOVA and a high-amplitude circadian rhythm was detected by the least-squares fit of a 24-hr &/or 24+12-hr cosine for each assay. Higher serum IL-6 values were detected throughout the night, with a peak at 01:00h, and lower values throughout the day, with a nadir at 10:00h. In contrast, IL-6 values in urine were highest during the day, with a major peak in the afternoon at 17:30h and a minor peak at 08:30h, and lowest values overnight, with a nadir at 23:30h. Of interest, the rhythm in urinary IL-6 concentration (pg/ml) was more prominent than hourly excretion rate (pg/hr). Thus, endogenous IL-6 (and possibly other cytokine) levels may be significantly influenced by their large and predictable day-night variations and the biological fluid used.

Background: The cytokine interleukin-6 (IL-6) is a multifunctional small-peptide molecule that is produced by various types of lymphoid and nonlymphoid cells. It plays a central role in hematopoiesis, host defense mechanisms, and acute-phase reactions, including regulation of inflammatory and immune responses., Methods: Because a circadian time structure has been shown to characterize nearly every biologic function tested in human beings, including some cytokines, we sought to investigate the 24-hour pattern of circulating IL-6 in a group of 11 clinically symptom-free men (median age, 50 years; range, 46 to 72 years). Blood samples were obtained every 3 hours for 24 hours (eight samples per subject), and serum was frozen until analysis for IL-6 with a solid-phase ELISA., Results: Average IL-6 values ranged from 1.66 to 5.38 pg/ml, with lowest to highest values within 24 hours ranging from 1.20 to 7.58 pg/ml (120% to 531%) between subjects. On average, values were greater than the mean throughout the night, with a peak at 01:00 hours and less than the mean throughout the day, with a nadir at 10:00 hours. A significant time effect was found by analysis of variance; and a high-amplitude circadian rhythm was described by the least-squares fit of a 24-hour cosine (p < 0.001; amplitude, 41% +/- 5%; acrophase, 02:16 +/- 00:28 hours). In addition, a positive correlation between mean IL-6 levels and age was found (r = 0.63, p = 0.037)., Conclusions: Because monitoring of endogenous cytokine levels is suggested for assessing immune function and pathologic condition, clinical decisions and immunotherapies may be significantly influenced by the large and predictable day-night variations in endogenous cytokine production and bioactivity.

Vessel dilator consisting of amino acids (a.a.) 31-67 and atrial natriuretic factor (ANF) composed of a.a. 99-126 of the 126 a.a. ANF prohormone circulate in humans and have potent vasodilatory properties. To determine whether these atrial natriuretic peptides are directly related to blood pressure in healthy normotensive humans, we recently had the unique opportunity to examine the circadian rhythms of vessel dilator, ANF, and blood pressure in seven individuals in 1988 and again in 1993. The changes in mean arterial pressure and systolic and diastolic blood pressure in these individuals during this 5-year hiatus allows comparison in the same individual, if circulating concentrations of atrial natriuretic peptides directly correlate with naturally occurring changes in blood pressure. In both 1988 and in 1993 vessel dilator and ANF each had significant (p < 0.001) circadian rhythms with their peak concentrations at 4:00 AM being nearly twice their concentrations at 4:00 PM. Mean arterial pressure, systolic blood pressure, and diastolic blood pressure also had significant circadian rhythms with peaks and troughs that were exactly opposite to those of ANF and vessel dilator. A significant inverse correlation between 24-hour averages of mean arterial blood pressure and 24-hour averages of vessel dilator (p = 0.05) and ANF (p = 0.02) was also found. These data suggest that vessel dilator and ANF are important for the maintenance of blood pressure within the normotensive range.

Long-acting natriuretic peptide, vessel dilator, and atrial natriuretic factor consisting of amino acids (a.a.) 1 to 30, 31 to 67, and 99 to 126 of the 126-a.a. atrial natriuretic factor (ANF) prohormone, respectively, circulate in humans and have potent vasodilatory properties. To determine if these atrial natriuretic peptides are directly related to blood pressure in clinically healthy normotensive humans, we obtained 24-h profiles of vessel dilator, long-acting natriuretic peptide, ANF, and blood pressure in 10 men in 1988 and 11 men in 1993 (seven men were studied twice) to compare circulating concentrations of atrial natriuretic peptides with naturally occurring changes in blood pressure. Overall, vessel dilator, long-acting natriuretic peptide, and ANF each had significant (p<0.001) circadian rhythms, with peak concentrations late during sleep (at 04:00 h) being nearly twice their concentrations in the afternoon and evening. This high-amplitude circadian change allowed for the refinement of normal limits for ANF peptides by computing 3-hourly tolerance intervals (chronodesms) against which to compare time-specified single samples for normality. Systolic, diastolic, and mean arterial blood pressure also had significant circadian rhythms (p<0.001) with peaks and troughs that were exactly opposite those of the ANF peptides. In addition to this inverse temporal relationship, there was a significant inverse correlation between absolute values for blood pressure and each ANF peptide (p<0.001), implying a functional relationship. These data suggest that in addition to other well-established neurochemical factors, the ANF peptides (vessel dilator, long-acting natriuretic peptide, and ANF) are important for the maintenance of blood pressure and modulation of its circadian rhythm.

Serum levels of four cytokines [tumor necrosis factor-alpha (TNF-alpha), interleukin-2 (IL-2), granulocyte-macrophage colony-stimulating factor (GM-CSF), and interleukin-10 (IL-10)] were measured in nine diurnally active healthy adult male subjects at 3-h intervals during a 24-h period. Statistical evaluation by analysis of variance and/or the least- squares fit of a cosine model revealed significant 24-h rhythms for each cytokine. Although the amount of IL-2 in the serum was low, the levels fluctuated to form a single peak at approximately noon. In contrast, the other three cytokines exhibited a biphasic temporal pattern. In subjects with detectable TNF-alpha levels, the first peak occurred at 07:30 and the second at 13:30. IL-10 levels also exhibited a biphasic pattern, with one peak at 07:30 and the second 12 h later at 19:30. GM-CSF levels were last to rise, first peaking at approximately 13:30 and then again at 19:30. These results suggest temporal patterns that are unique for each cytokine, generally with daytime highs and nighttime lows.

Recent progress in the treatment of coronary artery disease is reviewed from the standpoint of changes in lifestyle, surgical techniques to revascularize the myocardium and a variety of medical interventions. Among the medical modalities, heparin appears to have a greater potential than any other agent tested to neutralize the atherogenic process at most of its stages. This potential is supported by success in clinical trials of heparin administered by intravenous, subcutaneous, pulmonary, sublingual and topical routes. The suggested self-administration of low-dose heparin by inhalation appears to be well justified and easily adaptable to home therapy. The summarized evidence suggests the need for further clinical trials to test the use of heparin in the prophylaxis of atherosclerotic disease.

Reports on clinical trials with subcutaneous and intrapulmonary administration of low-dose heparin suggest that it may be an attractive therapeutic modality for the treatment of coronary artery disease because of unprecedented reduction in mortality of treated subjects. As a preliminary to a clinical trial with low-dose intrapulmonary heparin, a pilot study was conducted on three subjects. It compares overall circadian responses of 37 blood variables following intrapulmonary administration of heparin (10,500-18,800 U) in the morning (0800 h) and in the evening (2000 h). After each of these times, blood samples, mostly at 3 h intervals for the ensuing 27 h, were analyzed for heparin, APTT, TT, functional fibrinogen, CBC, enzymes, lipids, electrolytes, and hormones. Each time series was analyzed for circadian rhythm by the least-squares fit of a 24 h cosine and circadian mesors were compared by the Bingham test of rhythm parameters. Following heparin in the evening, but not in the morning, a statistically significant increase in circulating heparin levels, as well as directional increases in APTT and TT and decreases in fibrinogen, were observed in all three subjects. Same direction changes in several other variables were also observed. It is concluded that inhalation of heparin in low-dose levels results in variable circadian effects on blood parameters measured, ranging from no changes in their levels to minimal within normal range changes, and that these effects are dependent upon the timing of dose administration. It is suggested that the timed self-administration of low-dose heparin by inhalation be seriously considered for long-term clinical trials in the treatment and prevention of atherosclerosis.

These results are meant to serve as chrono-biologically-defined reference values for healthy adult men in their fifth decade of life. A thorough statistical analysis of these data is being prepared for presentation elsewhere. A rigorous comparison with results obtained when these same men were in their 20's and again in their 30's may help us to document the kinds and extent of changes in circadian rhythm characteristics (MESOR, amplitude and/or acrophase) which may occur with aging.

Circadian variation in the circulating concentrations of the N-terminal and C-terminal portions of the atrial natriuretic factor prohormone (pro ANF) was evaluated in 8 men, ages 41-47, who have been followed for 19 years with respect to circadian variation in physiological variables including blood pressure and clinical chemistries. The N-terminus of the ANF prohormone contains two peptides consisting of amino acids 1-30 and 31-67 while the C-terminus contains 1 peptide (amino acids 99-126) of this 126 amino acid prohormone which lower blood pressure and have natriuretic properties. To determine if either the N-terminus and/or the C-terminus of the prohormone have a circadian variation in their circulating plasma concentrations these 8 men had blood samples obtained for radiommunoassay every 3 hr during a 24-hr period. Three radiommunoassays which immunologically recognize (1) the whole N-terminus (i.e. amino acids 1-98), (2) the midportion of the N-terminus (amino acids 31-67) and (3) the C-terminus (amino acids 99-126) of the ANF prohormone were utilized. The whole N-terminus, the midportion of the N-terminus which circulates after being proteolytically cleaved from the rest of the N-terminus, and the C-terminus each had a peak circulating concentration between 0400 and 0700 which were significantly (P less than 0.001) higher than their concentrations at any other time throughout the 24-hr period.(ABSTRACT TRUNCATED AT 250 WORDS)

Nine clinically healthy men, 41-47 yr of age, served as subjects in a 24-hr study conducted at the Edward Hines Jr Veterans Administration Hospital in the Chicago area in May 1988. Physiologic measurements, and blood and urine samples were collected at 3-hr intervals over a single 24-hr period beginning at 1900. The number of variables measured or calculated (total = 98) included: 6 vital signs (oral temperature, pulse, blood- and intraocular pressures); 16 in whole blood (counts and differentials); 50 in serum (SMAC-24, lipids, hormones, electrophoresis of LDH and proteins); and 26 in urine (solids, proteins, creatinine, catecholamines, melatonin, cortisol, electrolytes and metals). Data were analyzed for time effect by analysis of variance (ANOVA) and for circadian rhythm by single cosinor. Individual rhythm characteristics for each variable were summarized for the group by population mean cosinor. The vast majority of variables revealed statistically significant within-day changes in values as validated by one-way ANOVA. All vital signs (except for intraocular pressures) and all serum hormones displayed a prominent circadian rhythm for the group, as did most variables in whole blood, while only about half of the variables in urine demonstrated a significant group rhythm. The results obtained are meant to: (a) document the circadian time structure; and (b) serve as reference values for circadian rhythm characteristics (range of change, mesor, amplitude and acrophase) for a defined group of individuals: clinically-healthy adult men in the prime of life.

Urine samples were collected at 3-hr intervals over a single 24-hr period from each of seven clinically healthy men who ranged in age from 21-25 years. Urines at each collection time were subsequently pooled using 20% of each volume and serially dialyzed against ammonium-barbituric acid buffer (pH 7.35 +/- 0.02), using a cellulose membrane permeable to compounds of less than 12,000-14,000 molecular weight (mw). When the dialyzed portions were then analyzed for total proteins, the sum of proteins in eight pools amounted to 74 mg. A 1 ml aliquot of each pool, representing approximately 50 micrograms of proteins, was concentrated and reconstituted. Approximately 20 micrograms of reconstituted proteins were then subjected to polyacrylamide gel electrophoresis. The stained gel was then scanned by laser densitometry and planimetry. Each aliquot revealed eight segments as identified by Coomassie and silver staining. Their molecular weights, estimated by extrapolation from concurrently run protein standards, and their total protein amounts were: 116,000 mw (9.44 mg), 91,000 mw (3.3 mg), 68,000 mw (11.58 mg), 53,000 mw (2.58 mg), 43,000 mw (9.12 mg), 32,000 mw (7.13 mg), 24,000 mw (4.52 mg) and 20,000 mw (5.27 mg). A statistically significant rhythm (P = 0.022 from ANOVA and 0.011 from Single Cosinor) was found for the excretion of total proteins, with an acrophase in the afternoon (1537) for these diurnally-active subjects.

Radioimmunoassay studies were conducted on 13 clinically healthy male subjects. Ten ranged in age between 23 and 44 years, two were age 51, and one was age 58. Blood samples were collected at 3-hr intervals over a period of 27 hr. Each serum sample was analyzed for the following hormones: insulin, gastrin, melatonin, prolactin, triiodothyronine (uptake), thyroid-stimulating hormone, triiodothyronine, thyroxine, luteinizing hormone, growth hormone, and follicle-stimulating hormone. Group data for each hormone were fitted to a 24-hr cosine curve. A statistically significant fit to this curve was evident in the six italicized variables. Those that did not yield a statistically significant fit frequently revealed a statistically significant variation along the 27-hr span. Chronogram and cosinor plots are presented.

Seven clinically-healthy men ranging in age from 21 to 25 years participated in this study. Urine samples were collected at 3 hr intervals over a single 24 hr span. Urines were pooled by using 20% of the total volume collected from each subject over a 3 hr collection span. The resulting 8 pools were analyzed for pH, specific gravity, osmolality, urea N, creatine, uric acid, glucose, phosphorus, chlorides, sodium, potassium, calcium, magnesium, silicon, aluminum, zinc and total solids. Each of the 8 pools was serially dialyzed at pH 7.35 against ammonium-barbituric acid buffer. The non-dialyzable portions were then re-analyzed for the remaining solids, sodium, potassium, calcium, magnesium, silicon, aluminum and zinc. Aliquots of the non-dialyzable fraction were examined by high performance liquid chromatography. Up to twelve discernable fractions were observed in each 3 hr urine by monitoring ultraviolet light absorbance at 280 nm wavelength. Range of change throughout the 24 hr (lowest to highest value) for most variables was 100% or more. In the eight 3 hr pooled urine samples, statistically-significant circadian variation could be described for volume, pH, osmolality, urea nitrogen, creatinine, uric acid, glucose, phosphorus, chlorides, for five of eight non-dialyzed (total) components (Na, K, Ca, Si and solids) and for five of twelve non-dialyzable solid fractions, as well as for total non-dialyzable solids. Single cosinor analysis resulted in description of a significant circadian rhythm in osmolality, urea nitrogen, creatinine, glucose, phosphorus, chlorides, total Na, K, Si and solids; non-dialyzable Si and solids; dialyzable Na, K, Si and solids; and for total solids, as well as their fractions at 23.4 and 25.9 min. These observations are furnished in order to further document the extreme circadian rhythmicity in all aspects of kidney function and as reference for future work which uses any of the investigated urinary endpoints whose circadian time structure is herein described.

Three-hour urine specimens were collected over a period of 27 hours from 11 healthy adult male subjects. Each specimen was analyzed for Na, K, Ca, Mg, and Zn using atomic absorption spectrophotometry. Each sample was also dialyzed, pH 7.35, and subsequently analyzed for Na, K, P, Ca, Mg, Zn, Fe, Pb, Al, Ni, Cu, Mo, Hg, Cr, Cd, and Mn using a multielemental argon-plasma emission system. The data were evaluated on conventional time plots (chronograms) and as computer-determined "cosinor" plots. A population circadian rhythm with a statistical significance was detected for total Na, K, Ca, and Mg, and for nondialyzable Na, K, P, Ca, Zn, and Mo. For almost every element studied the increase from lowest to highest 3-hour group mean along the 24-hour time scale was more than 100%. The 24-hour excretion of Na, K, Ca, Mg, and Zn appeared in good agreement with the so-called "normals." The nondialyzable levels of Fe, Pb, Al, Ni, Cu, Mo, Hg, Cr, Cd, and Mn were similar to the total urinary excretions reported in the literature.

Human thumbnails were analyzed for trace elements by instrumental analysis using thermal neutron activation technique. The average concentration of metals studied in clinically symptom-free adult female and male subjects were: zinc, 184 vs. 153 ppm; chromium, 6.8 vs. 4.2; selenium, 0.9 vs. 0.6; gold, 2.6 vs. 0.4; mercury, 1.9 vs. 0.4; silver, 0.7 vs. 0.3; cobalt, 0.07 vs. 0.04. A summary of literature reported concentration of metals in human nail is also presented.

Urinary levels of epinephrine and norepinephrine were studied in three populations: presumably healthy young men who were studied twice, 8 years apart, and in the same month to avoid seasonal influences; blinded but otherwise healthy males and females; and patients with leprosy. Determinations of epinephrine and norepinephrine were done on urine samples collected at 3-hr intervals for 72 hr from the first two groups and for 10 days from patients with leprosy. The same chemist, method, and protocol were used in all studies. In one study of the group of healthy young men, the effect of meal timing and fasting on the rhythms was investigated. From these studies several generalizations can be made. Highly statistically significant population circadian rhythms characterize both epinephrine and norepinephrine in all three groups. In general, the rhythm in norepinephrine was "noisier" than that in epinephrine; in the blind subjects, both variables were characterized by noisier rhythms than in the sighted. The overall levels of epinephrine were much higher in blind than in sighted subjects. The overall levels of norepinephrine were much higher in blind males than in blind females. In the studies done with the group of healthy young men, the overall levels of epinephrine and norepinephrine increased significantly over a 10-year period. The amplitude of the rhythm also increased significantly with time. The average age of the group was 27 years at the time of the first study and 40 years at the time of the second. The patients with leprosy also showed strong circadian variation in both epinephrine and norepinephrine, but, because many of the subjects were on medication, comparisons with the other groups are questionable. In general the data raise some questions about, but do not refute, the commonly held view that the amplitude and mesor of the circadian rhythm in epinephrine decrease with age. Additional work is needed to resolve this question completely.

At 3-hr intervals over a 24-hr span, 36 systemic, serologic and urinary variables were examined in 7 men in their mid 20's in the Spring of 1969, and again in the same 7 men in the Spring of 1979 under a similar chronobiologic protocol, using the same chemical and numerical analytical procedures. The variables examined for rhythms by cosinor were: vital signs--blood pressure (systolic, diastolic, pulse pressure and mean arterial pressure), heart rate, intraocular pressure (left and right), oral temperature; serum components--albumin, albumin/globulin ratio, total bilirubin, calcium, carbon dioxide, chlorides, bilirubin, cholesterol, globulin, glucose, potassium, sodium, sodium/potassium ratio, transaminase, triglycerides, total protein, urea nitrogen; and urine components--calcium, calcium/magnesium ratio, creatinine, magnesium, pH, potassium, sodium, sodium/potassium ratio, urea clearance, urea nitrogen, volume and zinc. Although all subjects appeared clinically healthy in 1969 and in 1979, certain inter-study differences were observed in a number of rhythm parameters of different variables. Statistically significant increases in mesor for the group as a whole were observed for serum Ca, cholesterol, Cl, CO2, K, Na, and while statistically significant mesor decreases for a group as a whole were noted in serum glucose and transaminase. Statistically significant increases in amplitude for the group as a whole were observed in serum chloride and urinary Na/K ratio, while statistically significant decreases were observed in amplitude for blood pressure, heart rate, serum albumin, A/G ratio, globulin, glucose, protein, sodium and transaminase.(ABSTRACT TRUNCATED AT 250 WORDS)