The sex-dependent impact of PER2 polymorphism on sleep and activity in a novel mouse model of cranial-irradiation-induced hypersomnolence.

Background: Hypersomnolence is a common and disruptive side effect of cranial radiotherapy and is associated with fatigue and disturbances in mood and cognition in primary brain tumor (PBT) patients. The biological underpinnings of this effect are not understood. Our laboratory has previously found that the presence of a single nucleotide polymorphism (rs934945, G-E mutation) in the PERIOD2 (PER2) clock gene was associated with a decreased likelihood of fatigue in PBT patients. Here, we aim to understand the effects of PER2 polymorphism on radiation susceptibility within a murine model of cranial-irradiation-induced hypersomnolence (C-RIH). Methods: Male and female transgenic mice were generated using CRISPR-Cas9, replacing the endogenous mouse PER2:CRY1 binding domain with its human isoform with (hE1244 KI) or without the SNP rs934945 (hG1244 KI). Activity and sleep were monitored continuously 10 days before and after cranial irradiation (whole brain, 15Gy, single fraction). Behavioral assessments measuring anxiety, depression, and working memory were used to assess mood and cognitive changes 2 months postradiation. Results: During their active phase, hE1244 knock-ins (KIs) had less radiation-induced suppression of activity relative to hG1244 KIs and female hE1244 KIs saw a reduction of hypersomnolence over 10 days. hE1244 KIs displayed less anxiety behavior and were more ambulatory within all behavioral tests. Conclusions: The PER2 rs934945 polymorphism had long-lasting behavioral effects associated with radiation toxicity, particularly in sleep in females and the activity of all animals. Our findings shed light on biological mechanisms underlying C-RIH.

Adaptation to short photoperiods augments circadian food anticipatory activity in Siberian hamsters.

This article is part of a Special Issue "Energy Balance". Both the light-dark cycle and the timing of food intake can entrain circadian rhythms. Entrainment to food is mediated by a food entrainable circadian oscillator (FEO) that is formally and mechanistically separable from the hypothalamic light-entrainable oscillator. This experiment examined whether seasonal changes in day length affect the function of the FEO in male Siberian hamsters (Phodopus sungorus). Hamsters housed in long (LD; 15 h light/day) or short (SD; 9h light/day) photoperiods were subjected to a timed-feeding schedule for 10 days, during which food was available only during a 5h interval of the light phase. Running wheel activity occurring within a 3h window immediately prior to actual or anticipated food delivery was operationally-defined as food anticipatory activity (FAA). After the timed-feeding interval, hamsters were fed ad libitum, and FAA was assessed 2 and 7 days later via probe trials of total food deprivation. During timed-feeding, all hamsters exhibited increases FAA, but FAA emerged more rapidly in SD; in probe trials, FAA was greater in magnitude and persistence in SD. Gonadectomy in LD did not induce the SD-like FAA phenotype, indicating that withdrawal of gonadal hormones is not sufficient to mediate the effects of photoperiod on FAA. Entrainment of the circadian system to light markedly affects the functional output of the FEO via gonadal hormone-independent mechanisms. Rapid emergence and persistent expression of FAA in SD may reflect a seasonal adaptation that directs behavior toward sources of nutrition with high temporal precision at times of year when food is scarce.

Impaired leukocyte trafficking and skin inflammatory responses in hamsters lacking a functional circadian system.

The immune system is under strong circadian control, and circadian desynchrony is a risk factor for metabolic disorders, inflammatory responses and cancer. Signaling pathways that maintain circadian rhythms (CRs) in immune function in vivo, and the mechanisms by which circadian desynchrony impairs immune function, remain to be fully identified. These experiments tested the hypothesis that the hypothalamic circadian pacemaker in the suprachiasmatic nucleus (SCN) drives CRs in the immune system, using a non-invasive model of SCN circadian arrhythmia. Robust CRs in blood leukocyte trafficking, with a peak during the early light phase (ZT4) and nadir in the early dark phase (ZT18), were absent in arrhythmic hamsters, as were CRs in spleen clock gene (per1, bmal1) expression, indicating that a functional pacemaker in the SCN is required for the generation of CRs in leukocyte trafficking and for driving peripheral clocks in secondary lymphoid organs. Pinealectomy was without effect on CRs in leukocyte trafficking, but abolished CRs in spleen clock gene expression, indicating that nocturnal melatonin secretion is necessary for communicating circadian time information to the spleen. CRs in trafficking of antigen presenting cells (CD11c(+) dendritic cells) in the skin were abolished, and antigen-specific delayed-type hypersensitivity skin inflammatory responses were markedly impaired in arrhythmic hamsters. The SCN drives robust CRs in leukocyte trafficking and lymphoid clock gene expression; the latter of which is not expressed in the absence of melatonin. Robust entrainment of the circadian pacemaker provides a signal critical to diurnal rhythms in immunosurveilliance and optimal memory T-cell dependent immune responses.

Photoperiodic regulation of the orexigenic effects of ghrelin in Siberian hamsters.

Animals living in temperate climates with predictable seasonal changes in food availability may use seasonal information to engage different metabolic strategies. Siberian hamsters decrease costs of thermoregulation during winter by reducing food intake and body mass in response to decreasing or short-day lengths (SD). These experiments examined whether SD reduction in food intake in hamsters is driven, at least in part, by altered behavioral responses to ghrelin, a gut-derived orexigenic peptide which induces food intake via NPY-dependent mechanisms. Relative to hamsters housed in long-day (LD) photoperiods, SD hamsters consumed less food in response to i.p. treatment with ghrelin across a range of doses from 0.03 to 3 mg/kg. To determine whether changes in photoperiod alter behavioral responses to ghrelin-induced activation of NPY neurons, c-Fos and NPY expression were quantified in the arcuate nucleus (ARC) via double-label fluorescent immunocytochemistry following i.p. treatment with 0.3 mg/kg ghrelin or saline. Ghrelin induced c-Fos immunoreactivity (-ir) in a greater proportion of NPY-ir neurons of LD relative to SD hamsters. In addition, following ghrelin treatment, a greater proportion of ARC c-Fos-ir neurons were identifiable as NPY-ir in LD relative to SD hamsters. Changes in day length markedly alter the behavioral response to ghrelin. The data also identify photoperiod-induced changes in the ability of ghrelin to activate ARC NPY neurons as a possible mechanism by which changes in day length alter food intake.

Genetic expression profile during acute cellular rejection in clinical intestinal transplantation.

Small bowel transplantation is a successful treatment for irreversible intestinal failure. Acute cellular rejection (ACR) represents the major cause of graft loss. However, little is known regarding the mechanisms of ACR and no data exist on the genetic response occurring during ACR. We report a genetic expression profile determined using oligo-microarrays of the intestinal graft during an ACR episode. Mucosal biopsies were obtained from the graft at weeks 0, 1, 2, 3, 6, and 52 posttransplant. We observed a statistically significant increase in transcript levels from 51 genes between biopsy samples from before ACR compared with during ACR. Functional analysis of these genes revealed an interferon (IFN)-alpha signature associated with a type I IFN immune response from dendritic cells or association with cellular proliferation and division. These genetic data support that dendritic cell activation was ongoing during ACR and suggest that IFNalpha production as a potential immunosuppressive target for transplantation.

Mutations occur in the Ig Smu region but rarely in Sgamma regions prior to class switch recombination.

Nucleotide substitutions are found in recombined Ig switch (S) regions and also in unrecombined (germline, GL) Smicro segments in activated splenic B cells. Herein we examine whether mutations are also introduced into the downstream acceptor S regions prior to switch recombination, but find very few mutations in GL Sgamma3 and Sgamma1 regions in activated B cells. These data suggest that switch recombination initiates in the Smicro segment and secondarily involves the downstream acceptor S region. Furthermore, the pattern and specificity of mutations in GL and recombined Smicro segments differ, suggesting different repair mechanisms. Mutations in recombined Smicro regions show a strong bias toward G/C base pairs and WRCY/RGYW hotspots, whereas mutations introduced into the GL Smicro do not. Additionally, induction conditions affect mutation specificity within the GL Smicro segment. Mutations are most frequent near the S-S junctions and decrease rapidly with distance from the junction. Finally, we find that mice expressing a transgene for terminal deoxynucleotidyl transferase (TdT) have nucleotide insertions at S-S junctions, indicating that the recombining DNA ends are accessible to end-processing enzyme activities.