Measurement in saliva from neurotypical adults of biomarkers pertinent to autism spectrum disorders.

AIM: Measure biomarkers pertinent to autism in saliva from humans. MATERIALS & METHODS: At 7:30 PM (reading instructions) and 8:30 PM (hearing instructions), neurotypical adults (6 M, 6 F) each spat into tubes containing protease inhibitors. Cells were counted, samples aliquoted, frozen and thawed. Rationale was given for choice of biomarkers. ELISA: CD26, IL-12, carnitine, C4B, GSH, GSSG, MT-2, testosterone, IFN-γ. Mass spectrometry: cystine, glutamine, glutamic acid, GABA, serotonin. Electrochemiluminescentimmunoassay: cortisol. Radioimmunoassay: melatonin. RESULTS: Cells averaged 2.16 × 106/ml. M > F: CD-26, C4B, MT-2. Testosterone, cortisol. Glutamine, glutamic acid, IFN-γ, melatonin and GSSG were measurable. Remaining biomarkers were measured in <50% of samples. Concentrations were equal at both times. CONCLUSION: Saliva can be collected by literate individuals without added instruction. Ten biomarkers were measurable.

The Chronobra identifies prevailing mammary vascularity as a candidate variable in breast cancer post-operative outcome prediction.

We previously described a menstrual heat cycle of the breast in four groups of women (healthy, family history of breast cancer, benign breast disease, 'cancer-associated') who wore a thermometric brassiere (Chronobra). We now ask if 'breast minus oral temperature', indicating 'breast-associated vascularity', could be associated with breast cancer cell vascular access around different aspects of the menstrual cycle rhythm and survival. Thirty-six pre-menopausal breast cancer patients (average age: 38.97 y) were enrolled consecutively over 15 y and followed for more than 22 y after surgery in order to compare survival and peri-operative vascularity. Each subject wore the Chronobra, which provides an internal bioassay of the vascularity of both breasts, including the operated breast, during 1 h each evening at home for one menstrual cycle, and collected saliva for "free" progesterone to confirm pre-menopausal status and ovulation. Sixty-five healthy age-matched pre-menopausal women served as controls. Both oral and breast temperatures revealed menstrual cycle oscillations, rising just before ovulation until menses onset. Breast-adjusted vascularity also showed menstrual cycle oscillations, with levels differing significantly between the 3 groups during the luteal phase only. At the end of the follow-up span, 18 post-operative breast cancer patients had died from "disseminated" breast cancer and 18 were alive and well. Median follow-up time was 22.6 y for survivors, 6.2 y for non-survivors, and 21.0 y for controls (3 died from diseases unrelated to breast cancer). Based on 'during luteal-phase breast-adjusted vascularity', breast cancer survivors (mean ± SD: -1.65 ± 0.23°C) were significantly hypo-vascular (i.e., -0.23°C cooler) compared with controls (-1.42 ± 0.09°C), while non-survivors (-1.25 ± 0.12°C) were highly significantly hyper-vascular compared with survivors (+0.41°C warmer) and controls (+0.23°C warmer). This suggests that in pre-menopausal breast cancer patients, peri-operative mammary vascularity could offer an outcome test of survival and biologically may be on the "final common pathway" of any tumor to metastatic risk and recurrence.

Menstrual cycle and respiratory symptoms in a general Nordic-Baltic population.

RATIONALE: There is little knowledge of variations in respiratory symptoms during the menstrual cycle in a general population, and potential modifying factors are not investigated. OBJECTIVES: To investigate menstrual cycle variation in respiratory symptoms in a large general population, using chronobiology methodology, and stratifying by body mass index (BMI), smoking, and asthma status. METHODS: A total of 3,926 women with regular cycles less than or equal to 28 days and not taking exogenous sex hormones answered a postal questionnaire regarding the first day of their last menstruation and respiratory symptoms in the last 3 days. Moving 4-day means were computed to smooth uneven records of daily sampling; best-fitting 28-day composite cosine curves were applied to each time series to describe rhythmicity. MEASUREMENTS AND MAIN RESULTS: Significant rhythmic variations over the menstrual cycle were found in each symptom for all subjects and subgroups. Wheezing was higher on cycle Days 10-22, with a midcycle dip near the time of putative ovulation (approximately Days 14-16) in most subgroups. Shortness of breath was higher on days 7-21, with a dip just before midcycle in many subgroups. Cough was higher just after putative ovulation for subjects with asthma, BMI greater than or equal to 23 kg/m(2), and smokers, or just before ovulation and menses onset for low symptomatic subgroups. CONCLUSIONS: Respiratory symptoms varied significantly during the menstrual cycle and were most frequent from the midluteal to midfollicular stages, often with a dip near the time of ovulation. The patterns varied by BMI, smoking, and asthma status. These relations link respiratory symptoms with hormonal changes through the menstrual cycle and imply a potential for individualized chronotherapy for respiratory diseases.

A risperidone-induced prolactinoma resolved when a woman with schizoaffective disorder switched to ziprasidone: a case report.

Antipsychotic drug therapy, e.g., risperidone, can be associated with endocrine abnormalities, including an increase in serum prolactin level (sPrl) due to a drug-induced benign pituitary tumor (prolactinoma). A few case reports have noted a resolution of hyperprolactinemia and prolactinoma after cessation of risperidone treatment. We report a similar finding for a woman with schizoaffective disorder, manic type.Due to a neurological disorder involving the tongue (tardive dyskinesia), a woman with schizoaffective disorder switched from 50mg thioridazine after 21 years to 2mg of risperidone at bedtime for 10 years. Elevated sPrl was noted in June and August 2005 (83.8 and 100.1µg/L; normal: 3.4-24.1µg/L) and a cranial magnetic resonance imaging scan showed evidence of a small area of decreased signal in the pituitary gland consistent with a microadenoma. The subject transitioned slowly to ziprasidone and off risperidone in October and November of 2005. The prolactinoma completely resolved with the switch to ziprasidone. It is recommended that sPrl be measured annually in patients taking antipsychotic drugs to test for any indication of pituitary prolactinoma that could suggest the need to switch the primary treatment to another drug.

Decadal Cycles in the Human Cardiovascular System.

Seven of the eight authors of this report each performed physiologic self-surveillance, some around the clock for decades. We here document the presence of long cycles (decadals, including circaundecennians) in the time structure of systolic (S) and diastolic (D) blood pressure (BP) and heart rate (HR). Because of the non-stationary nature in time and space of these and other physiologic and environmental periodic components that, like the wind, can appear and disappear in a given or other geographic location at one or another time, they have been called "Aeolian". The nonlinear estimation of the uncertainties of the periods (τs) of two or more variables being compared has been used to determine whether these components are congruent or not, depending on whether their CIs (95% confidence intervals) overlap or not. Among others, congruence has been found for components with τs clustering around 10 years in us and around us. There is a selective assortment among individuals, variables and cycle characteristics (mean and circadian amplitude and acrophase). Apart from basic interest, like other nonphotic solar signatures such as transyears with periods slightly longer than one year or about 33-year Brückner-Egeson-Lockyer (BEL) cycles, about 10-year and longer cycles present in 7 of 7 self-monitoring individuals are of interest in the diagnosis of Vascular Variability Anomalies (VVAs), including MESOR-hypertension, and others. Some of the other VVAs, such as a circadian overswing, i.e., CHAT (Circadian Hyper-Aplitude-Tension), or an excessive pulse pressure, based on repeated 7-day around-the-clock records, can represent a risk of severe cardiovascular events, greater than that of a high BP. The differential diagnosis of physiologic cycles, infradians (components with a τ longer than 28 hours) as well as circadians awaits the collection of reference values for the infradian parameters of the cycles described herein. Just as in stroke-prone spontaneously hypertensive rats during the weeks after weaning CHAT precedes an elevation of the BP MESOR, a decadal overswing seems to precede the occurrence of high BP in two of the subjects here examined. Only around-the-clock monitoring in health for the collection of reference values will allow on their basis the differential diagnosis of the onsets of a circadian versus a circadecadal overswing in BP and the specification whether, and if so, when to initiate hypotensive non-drug or drug treatment.

Circadian aspects of growth hormone-insulin-like growth factor axis function in patients with lung cancer.

BACKGROUND AND PURPOSE: Insulin-like growth factor 1 (IGF1) promotes cell cycle progression and inhibition of apoptosis and may have a role in carcinogenesis and cancer promotion. Growth hormone (GH) stimulates IGF1 production in liver and other tissues. The aim of our study was to evaluate differences between healthy subjects and patients with lung cancer in the GH-IGF1 axis function. PATIENTS AND METHODS: In 11 healthy male patients (mean age ± standard error [SE], 43.6 ± 1.7), and 9 male patients with non-small-cell lung cancer (mean age ± SE, 51.0 ± 2.4), GH, total IGF1, melatonin, and interleukin (IL)-2 serum levels were measured in blood samples collected every 4 hours for 24 hours. RESULTS: A clear circadian rhythm was present for melatonin and GH serum levels in the group of healthy subjects and for melatonin in the group of patients with cancer. The midline estimating statistic of rhythm (MESOR) of GH was higher in patients with lung cancer (P < .001), the MESOR of IGF1 was higher in healthy subjects (P < .001), the MESOR of melatonin was not different between subjects (P = .383), and the MESOR of IL-2 was higher in patients with cancer (P = .02). The GH/IGF1 ratio was higher in patients with lung cancer (P = .006). Linear regressions across the stages of cancer showed a significant increasing slope for IL-2 (P < .001), GH (P < .001), and the GH/IGF1 ratio (P < .001), a decreasing slope for IGF1 (P < .001), but no significant trend for melatonin (P = .430). CONCLUSION: There is evidence that in patients with lung cancer there is a severe alteration of GH-IGF1 axis function, with loss of circadian rhythmicity of hormone secretion, which may play a role in the progression of neoplastic disease and must be considered in defining the therapeutic approach.

Alteration of hypothalamic-pituitary-thyroid axis function in non-small-cell lung cancer patients.

The aim of this study was to evaluate the hypothalamic-pituitary-thyroid (HPT) axis function in patients suffering from lung cancer. Thyrotropin-releasing hormone (TRH), thyroid-stimulating hormone (TSH), free thyroxine (FT4), interleukin (IL)-2, and melatonin serum levels were measured in blood samples collected every 4 hours for 24 hours from 11 healthy participants (H; ages 35-53 years) and 9 patients suffering from non-small-cell lung cancer (C; ages 43-63 years). Relationships between hormone levels overall and over time of day were evaluated within and among groups. A prominent circadian rhythm with peaks near midnight was present for TSH and melatonin serum levels in both H and C, indicating similar synchronization of the main body clock to the 24-hour environmental light-dark cycle. As regards 24-hour means in H and C, TSH was lower in C, whereas TRH, FT4, and IL-2 were higher in C, with no difference in melatonin levels. Simple linear regression, FT4 versus TRH, showed a positive correlation in H and a negative correlation in C, whereas FT4 versus TSH showed a negative correlation in both groups. For FT4 versus IL-2, a negative correlation was found in C but not for H, whereas TSH versus TRH showed no correlation for either group. Both groups were found to be similarly synchronized to the 24-hour sleep-wake schedule, but HPT axis function was altered in patients suffering from lung cancer. When compared with healthy controls, cancer patients showed modifications of hormone serum levels overall and a negative correlation between individual TRH and FT4 levels.

Comparison of circadian characteristics for cytotoxic lymphocyte subsets in non-small cell lung cancer patients versus controls.

Lymphocyte subsets are major cellular components of the adaptive immune response and in most cases show 24-h (circadian) variations in health. In order to determine overall levels and circadian characteristics of cytotoxic natural killer (NK) and T and B lymphocyte subsets, blood samples were collected every 4 h for 24 h from eleven male controls (C) without neoplastic disease and nine men with untreated non-small cell lung cancer (NSCLC) and analyzed for 3 hormones (melatonin, cortisol, and interleukin 2 [IL2]) and for 11 lymphocyte subpopulations classified by cell surface clusters of differentiation (CD) and antigen receptors. Circadian rhythmicity for each variable was evaluated by ANOVA and 24 h cosine fitting and groups compared. Rhythms in melatonin and cortisol (peaks near 01:30 and 08:00 h) indicated identical synchronization to the light-dark schedule and probable persistent entrainment of rhythms for both groups in metabolism or proliferation of healthy tissues normally tightly coupled to the sleep-wake cycle. Twenty-four hours means were significantly higher in NSCLC for CD16, CD25, cortisol, and IL2 and lower for CD8, CD8bright, and γδTCR. A significant circadian rhythm was found in C with daytime peaks for CD8, CD8dim, CD16, Vδ2TCR, and cortisol and nighttime peaks for CD3, CD4, CD20, and melatonin, and in NSCLC, with daytime peaks for CD16, γδTCR, Vδ2TCR and cortisol, and nighttime peaks for CD4, CD25, and melatonin. Thus, NSCLC was associated with significant increases or decreases in proportions for several lymphocyte subsets that may reflect disease development, but peak times were nevertheless similar between C and NSCLC for each variable, suggesting that timed circadian administration (chronotherapy) of immunotherapy and other cancer treatments may improve efficacy due to persistent circadian entrainment of healthy tissues.

Determination of whole body circadian phase in lung cancer patients: melatonin vs. cortisol.

BACKGROUND: A quantifiable and reliable technique for the determination of body circadian phase applicable to non-laboratory studies would allow the evaluation of circadian dysregulation. In this study we evaluated feasible methodologies to individualize whole body circadian phase in lung cancer patients. METHODS: Cortisol and melatonin serum levels were measured in blood samples collected every 4 h for 24 h from eleven male controls and nine men suffering from non-small cell lung cancer. Circadian rhythmicity was evaluated and the 4-hourly fractional variations (FV) were calculated to evaluate the dynamics of the rise and fall in serum levels. RESULTS: Overall cortisol serum levels were higher in cancer patients (p<0.001), and lower for melatonin, but not significantly (p=0.261). Original serum levels of cortisol and melatonin each showed a prominent 24 h oscillation in both study groups, with highest values at night for melatonin and near awakening for cortisol. Using all data after normalization to percent of individual mean, ANOVA detected a significant time-effect (p<0.001) and cosinor analysis detected a significant 24 h rhythm (p<0.001) in each group. Overall fractional variation (FV) levels were lower for cortisol in cancer patients and higher for melatonin, but these differences were not significant. FV levels of cortisol and melatonin each showed a prominent 24 h oscillation in both study groups, with highest values prior to darkness onset for melatonin and near mid-dark for cortisol. ANOVA also detected a significant time-effect (p<0.002) and cosinor analysis detected a significant 24 h rhythm (p<0.001) for FV in each group, with maximal FV preceding maximal serum levels by ∼3 h for melatonin and ∼5 h for cortisol. CONCLUSIONS: A chronobiological evaluation of serum levels and fractional variations for cortisol and especially melatonin is a valuable methodology to define body circadian phase in lung cancer patients. It is possible to describe the complex process of hormone secretion with a methodology that allows the definition of both temporal characteristics and dynamic components. IMPACT: This kind of analysis might be useful in the study of hormone secretion(s) in cancer patients and other diseases and to guide therapeutic interventions. While lung cancer patients may have a negative prognostic value based upon stage and/or other hormonal aspects of their hypothalamus-pituitary-thyroid-adrenal axis function, patients that nevertheless maintain circadian rhythmicity in key body rhythm markers may still be eligible candidates for a chronotherapeutic approach of treatment(s).

Circadian aspects of hyperthermia in mice induced by Aconitum napellus.

BACKGROUND: Aconitum napellus (Acn) is used topically to relieve pain, itching and inflammation, and internally to reduce febrile states, among others. Any circadian time-related consequences of Acn administration are unknown. The objective of this study was to explore the effects of two doses of Acn on body temperature (BT) of mice treated at six different times over 24 hours. MATERIALS AND METHODS: BALB/c female mice were housed in six chambers (six mice each) with air temperature 24 ± 3°C, humidity 60 ± 4%, and a 12-hours light (L)/12-hours dark cycle, but with L-onset staggered by 4 hours between chambers so that study at one external test time resulted in six test times (02, 06, 10, 14, 18 and 22 hours [h] after light onset). Rectal temperature (RT; in °C) was measured at baseline (B) and 1 hour after oral treatment with placebo (P) or two doses of Acn (6C and 30C, two studies each) in six studies over an 8 day span. The difference in RT for each mouse from the respective B + P timepoint mean RT was computed following each Acn treatment, and data from each of the six studies (original RT and difference from B + P) were analyzed for time-effect by analysis of variance (ANOVA) and for circadian rhythm by 24-hour cosine fitting. RESULTS: A CIRCADIAN RHYTHM IN RT WAS FOUND AT B AND AFTER P (MEAN: 35.58°C vs. 35.69°C; peak: 15:31 h vs. 15:40 h) and after each Acn dose (30C or 6C). Acn induced hyperthermia and the overall change in BT was rhythmically significant for each dose (mean = +1.95°C vs. +1.70°C), with greatest hyperthermia observed during the L-span for each dose (peak = 08:56 h vs. 05:17 h). CONCLUSION: Acn administered around the clock induced hyperthermia overall and in a time-dependent manner, with greatest effects during the resting (L) span. Thus, time of day may significantly impact the outcome of Acn and other homeopathic treatments and should be considered in determining optimal dosing and treatment time(s) in order to increase the desired outcome and decrease undesired effects.

Sunspot dynamics are reflected in human physiology and pathophysiology.

Periodic episodes of increased sunspot activity (solar electromagnetic storms) occur with 10-11 and 5-6 year periodicities and may be associated with measurable biological events. We investigated whether this sunspot periodicity characterized the incidence of Pap smear-determined cervical epithelial histopathologies and human physiologic functions. From January 1983 through December 2003, monthly averages were obtained for solar flux and sunspot numbers; six infectious, premalignant and malignant changes in the cervical epithelium from 1,182,421 consecutive, serially independent, screening Pap smears (59°9″N, 4°29″E); and six human physiologic functions of a healthy man (oral temperature, pulse, systolic and diastolic blood pressure, respiration, and peak expiratory flow), which were measured ∼5 times daily during ∼34,500 self-measurement sessions (44°56″N, 93°8″W). After determining that sunspot numbers and solar flux, which were not annually rhythmic, occurred with a prominent 10-year and a less-prominent 5.75-year periodicity during this 21-year study span, each biological data set was analyzed with the same curve-fitting procedures. All six annually rhythmic Pap smear-detected infectious, premalignant and malignant cervical epithelial pathologies showed strong 10-year and weaker 5.75-year cycles, as did all six self-measured, annually rhythmic, physiologic functions. The phases (maxima) for the six histopathologic findings and five of six physiologic measurements were very near, or within, the first two quarters following the 10-year solar maxima. These findings add to the growing evidence that solar magnetic storm periodicities are mirrored by cyclic phase-locked rhythms of similar period length or lengths in human physiology and pathophysiology.

Circadian study of decompression sickness symptoms and response-associated variables in rats.

In order to study circadian rhythms and decompression sickness (DCS), we determined: 1) the baseline circadian time structure in noncompressed rats of potential response variables to compression/decompression (C/D), and 2) whether rats subjected to C/D display a circadian time-dependent difference in inflammatory response intensity and biological tolerance. Subgroups of male rats, standardized to a 12 h light/12 h dark schedule, were evaluated every 4 h over 24 h after they were either compressed to 683 kPa (group E) or remained at sea level (group C). During 60 min recovery, evaluation included gross DCS symptoms and pulmonary edema in all E rats, and cell counts, nitric oxide, protein, thromboxane B(2,) and leukotriene E(4) levels in survivors. Chi-square, ANOVA, and 24 h cosinor analyses were used to test for time-of-day effects. C/D exposures near the end of dark/activity or during light/resting were generally better tolerated, with lowest signs of DCS symptoms and lowest responses by most of the variables monitored. More deaths were observed in the first half of the dark/activity span. Of the 16 subsets of inflammatory-associated variables, overall increases were observed in 13 and decreases in 2. Significant or borderline significant circadian time effects were found in 14 variables in group C, 12 variables in group E, and 13 variables in response (E%C). Thus, nearly all baseline indices of DCS demonstrated circadian time-dependencies in the sea-level exposed control rats (group C), and nearly all were modified by the circadian time of C/D. Such time-of-day effects of DCS are potentially relevant to the operational concerns of occupations involving decompression exposures and the investigation of prevention and treatment intervention strategies of DCS.

Chronomics, human time estimation, and aging.

BACKGROUND: Circadian rhythm stage affects many outcomes, including those of mental aging. METHODS: Estimations of 1 minute approximately 5 times/day for a year, 25 years apart, by a healthy male biomedical scientist (RBS), are analyzed by the extended cosinor. RESULTS: Cycles of a half-week, a week, approximately 30 days, a half-year and a year, in self-assessed 1-minute estimation by RBS between 25 and 60 years of age in health, are mapped for the first time, compared and opposite effects are found. For RBS at 60 vs at 25 years of age, it takes less time in the morning around 10:30 (P < 0.001), but not in the evening around 19:30 (P = 0.956), to estimate 1 minute. DISCUSSION: During the intervening decades, the time of estimating 1 minute differed greatly, dependent on circadian stage, being a linear decrease in the morning and increase in the evening, the latter modulated by a -33.6-year cycle. CONCLUSION: Circadian and infradian rhythm mapping is essential for a scrutiny of effects of aging. A approximately 30-day and a circannual component apparent at 25 years of age are not found later; cycles longer than a year are detected. Rhythm stages await tests as markers for timing therapy in disease.

Chronobiological analysis of circadian patterns in transcription of seven key clock genes in six peripheral tissues in mice.

The molecular clock machinery in mammals consists of a number of clock genes (CGs) and their resultant proteins that form interlocking transcription-translation feedback loops. These loops generate and maintain the 24 h mRNA and protein oscillations and consequential biological and physiological rhythms. To understand whether peripheral oscillators share similarly-timed clock machinery, the temporal expression patterns of the seven recognized key CGs (mPer1, mPer2, mCry1, mCry2, mRev-erb alpha, mClock, and mBmal1) were examined simultaneously in six peripheral tissues in mice every 4 h for 24 h in synchronized light-dark conditions using real time PCR assays. Time series were analyzed for time-effect by ANOVA and for rhythm characteristics by the single cosinor fitting procedure. The expression levels of most CGs were comparable in liver, kidney, and spleen, but mBmal1 and mCry1 were more abundant in the thymus, and mPer1, mCry1, and mCry2 were more abundant in the testis. In addition, mCry2 was dramatically lower in the kidney, spleen, and thymus; mPer2 was significantly lower in the spleen, testis, and thymus; and all of the genes tested were strikingly less abundant in peripheral blood. A significant 24 h rhythmic component was found for each CG in the liver and kidney and for some CGs in other tissues. Of note, a 12 h ultradian rhythmic component was also found in mRNA expression for some CGs in several of the tissues and was the only significant oscillation observed for CGs in the testis. Ultradian oscillations were also observed for mPer1 in the testis (8 h) and thymus (12 h and 8 h) in a second study where mice were sampled every 2 h. The present results suggest that the functioning of the molecular circadian clock may be modified to some extent between peripheral tissues, as denoted by differences in amplitude and phasing, and operates differently or is less developed in tissues containing differentiating cells (i.e., testis and thymus), as denoted by the presence of ultradian patterns resulting in two or more peaks within 24 h.

Circadian variations in clock gene expression of human bone marrow CD34+ cells.

Time-dependent variations in clock gene expression have recently been observed in mouse hematopoietic cells, but the activity of these genes in human bone marrow (BM) has so far not been investigated. Since such data can be of considerable clinical interest for monitoring the dynamics in stem/progenitor cells, the authors have studied mRNA expression of the clock genes hPer1 , hPer2, hCry1, hCry2, hBmal1, hRev-erb alpha, and hClock in human hematopoietic CD34-positive (CD34( +)) cells. CD34(+) cells were isolated from the BM samples obtained from 10 healthy men at 6 times over 24 h. In addition, clock gene mRNA expression was analyzed in the whole BM in 3 subjects. Rhythms in serum cortisol, growth hormone, testosterone, and leukocyte counts documented that subjects exhibited standardized circadian patterns. All 7 clock genes were expressed both in CD34(+) cells and the whole BM, with some differences in magnitude between the 2 cell populations. A clear circadian rhythm was shown for hPer1, hPer2, and hCry2 expression in CD34(+) cells and for hPer1 in the whole BM, with maxima from early morning to midday. Similar to mouse hematopoietic cells, h Bmal1 was not oscillating rhythmically. The study demonstrates that clock gene expression in human BM stem/progenitor cells may be developmentally regulated, with strong or weaker circadian profiles as compared to those reported in other mature tissues.

Circadian expression of clock genes in purified hematopoietic stem cells is developmentally regulated in mouse bone marrow.

OBJECTIVE: Clock genes are known to mediate circadian rhythms in the central nervous system and peripheral organs. Although they are expressed in mouse hematopoietic progenitor and stem cells, it is unknown if they are related to circadian rhythms in these cells. We therefore investigated the 24-hour patterns in the activity of several clock genes in the bone marrow (BM) side population (SP) primitive stem cells, and compared these 24-hour patterns to clock gene variations in the whole BM and liver. METHODS: Cells were obtained from 84 B6D2F(1) mice in three replicate experiments on the second day after release into constant darkness from a standardizing light-dark schedule. mRNA expression of clock genes was measured with quantitative reverse transcriptase polymerase chain reaction. RESULTS: mPer2 displayed circadian rhythms in SP cells, whole BM, and liver cells. mPer1 and mRev-erb alpha showed a circadian rhythm in whole BM and liver, but not SP cells. mBmal1 was not expressed rhythmically in SP cells, nor in the whole BM, contrary to rhythms observed in the liver. CONCLUSIONS: With the exception of mPer2, most clock genes studied in primitive hematopoietic SP stem cells were not oscillating in a fully organized circadian manner, which is similar to immature cells in rapidly proliferating organs, such as the testis and thymus. These findings indicate that circadian clock gene expression variations in BM are developmentally regulated.

Sun exposure, sexual behavior and uterine cervical human papilloma virus.

We have previously observed marked seasonal fluctuations in the frequency of cervical smears positive for human papilloma virus (HPV) in a series of smears obtained in Holland, with a peak in the summer months, especially August. Here, we tested two possible mechanisms that might underlie this summer peak: (1) enhanced transmission of HPV due to increased seasonal sexual activity, or (2) suppression of immunity due to summertime population exposure to solar ultraviolet (UV) radiation. Data derived from a continuous series of >900,000 independent cervical smears obtained from 1983 to 1998 were assessed for histopathologic epithelial changes pathognomonic of HPV. The rate of HPV positivity was then compared to both the rate of sexual activity (using conception frequency as a readily available surrogate) as well as yearly and monthly fluctuations in solar-UV fluency. The rate of HPV positivity was found to be twice as high during the summer months, with a peak in August corresponding with maximal UV fluency. Furthermore, over these 16 consecutive years of continuous observation, maximum HPV detection rate and maximum UV fluency are positively correlated (r=0.59, P<0.01); the sunnier the year, the greater the rate of HPV. Likewise, there is a positive correlation of the monthly UV fluency, and monthly HPV discovery rate (r=0.16, P<0.03). In contrast, conception frequency (and, presumably, population sexual HPV transmission) was maximal near the vernal equinox, with relatively modest (<10%) seasonal fluctuation, i.e., not fully explaining this prominent August peak in HPV discovery. There is a clear relationship between the detection of HPV-positive cervical smears and sunlight exposure. We speculate that the well-known phenomenon of UV-mediated suppression of immune surveillance may be causally related to this unusual increase in cytologically defined active HPV infections during the summer months in northern countries such as Holland. Confirming this relationship elsewhere may be important, because whatever the risk conferred by sunlight is, in principle, behaviorally avoidable.

Season, sun, sex, and cervical cancer.

INTRODUCTION: Sunlight's UV B component, a known cellular immunosupressant, carcinogen, and activator of viral infections, is generally seasonally available. Venereal human papillomavirus (HPV) transmission, at least in part, causes cervical cancer. We have previously inspected the monthly rates of venereal HPV infection and sunlight fluency in Southern Holland over 16 consecutive years. Both peak in August with at least 2-fold seasonality. The amount of available sunlight and the rate of Papanicolaou (Pap) smear screen-detected HPV are positively correlated. We now investigate whether premalignant and malignant cervical epithelial changes are also seasonal and related to seasonal sunlight fluency. METHODS: We have studied >900,000 consecutive, serially independent, interpretable screening Pap smears obtained by a single cervical cancer screening laboratory in Leiden, Holland, during a continuous 16-year span from 1983 through 1998. The average monthly rates of premalignant and malignant epithelial change were inspected and the annual patterns contrasted to the annual pattern of sunlight fluency at this global location and to monthly average HPV infection rate. Because HPV is venereally transmitted, Dutch seasonal sexual behavior was evaluated by assessment of the annual pattern of Dutch conception frequency as a competing cause for cervical cancer seasonality. RESULTS: (a) Twice as many premalignant and malignant epithelial changes were found among Pap smears obtained in the summer months, with an August peak concurrent with histopathologic evidence of HPV infection and sunlight fluency in Southern Holland. (b) Monthly sunlight fluency is correlated positively with both the monthly rates of Pap smear-detected cervical epithelial dysplasia and carcinomatous histopathology, as well as HPV. (c) Conception frequency, in this location, peaks in Spring not summer, and has a 4.8% annual amplitude. CONCLUSIONS: (a) Cervical epithelial HPV infection and HPV-induced cervical epithelial dysplasia and carcinomatous change may each be novel sun exposure risks and thereby behaviorably avoidable. (b) Because screening Pap smears uncover many abnormalities that resolve spontaneously (false positives), these data may argue for screening and follow-up Pap smear examinations in seasons other than summer in the Northern Hemisphere, to diminish the false-positive smear rate. Global data are available to confirm and further test each of these conclusions.