CLOCK Protein Analysis

* It is thought that the microbes regulate the expression of the gut protein fasting-induced adipocyte protein (FIAF). This protein is an inhibitor of lipoprotein lipase (LPL).

In this study, we used one adult male Sprague-Dawley rat weighing 283 g. The subject was on a 6 am turn on, 6 pm turn off light cycle and was fed and given water regularly. The subject had also already gone through a rotometer test to find the subject’s preferred rotational direction.

The RER of both TRF groups was significantly higher during the feeding phase and significantly reduced during the fasting phase than their ad-lib counterparts. Using 45 mice, Chung and colleagues12 studied an 8-hour high-fat TRF diet over 27 weeks. RER was measured from 6 p.m. to 10 p.m., 10 p.m. to 6 a.m. (TRF period), and 6 a.m. to 6 p.m. RER for the ad-lib group was approximately 0.75 throughout the study. RER for the TRF group was 0.70 from 6 a.m. to 10 p.m. and increased during the TRF period. Comparing the two groups, the RER of the TRF group was significantly lower than the ad-lib group from 6 p.m. to 10 p.m., significantly higher than the ad-lib group during the TRF period, and similar to the ad-lib group from 6 a.m. to 6 p.m. In Moro et al.’s18 8-hour TRF study with 34 men, RER was measured between 6 a.m. and 8 a.m. At the end of the study, the RER decreased significantly in the TRF group compared to baseline (0.83 ± 0.02 to 0.81 ± 0.01). Yet the RER in the TRF group was the same as the ad-lib group (0.83 ± 0.02). Sundaram and Yan13 tested the impact of an 8- and 12-hour high fat TRF diet on RER in mice. The diurnal rhythms of RER oscillations of the TRF groups (ranging from about 0.9 to 0.75 between the feeding and fasting cycles) were improved compared to the high-fat ad-lib mice.

Over the past years, there have been many conflicting views over weather or not rest has an effect on controlling cell fate. There has been previous studies that have indicated that the rest gene influences gene expression. Researchers at the State University of New York at Stony Brook have taken it upon themselves to determine if it is in fact true. By using methods like zinc finger nuclease and quantitative PCR, they were able to find reliable evidence that: rest in zebrafish does have an influencing affect on neural-specific gene expression but it fails to have many significant effects on zebrafish neurogenesis.

Chronotype (CT) is part of the circadian routine and it is defined as an individual’s inclination in rise and sleep timing (Horne & Ostberg, 1977). Chronotypes comprise three different groups; morning, intermediate, and evening group (Adal et al, 2012). Specifically, the early group rise and sleep early, while individuals who belong to the evening group rise and sleep later than the morning group, whereas the intermediate group does not follow a specific pattern (Adal et al, 2012). Interestingly, around 40% of the individuals belong to one of the two poles, morning or evening group (Adal et al,

Circadian rhythm refers to the twenty-four-hour cycle that results in physical, mental and behavioral changes in living things. Animals and plants respond differently to the lightness and darkness depending on the circadian rhythm or the internal biological clock. The body clock works responsibly in making sure that the pattern of mental alertness of a person is enhanced. A DNA molecule is transmitted to pass the message for opening up of the RNA, and the mRNA is transformed into a protein which takes some time for the DNA to be turned into a molecule. The time of conversion is determined and kept by the body’s biological clock (Alice, 2012). If the clock fails to work the level of alertness can be affected, and this can make a person to cause traffic accidents or even cause injuries at the place of work. Also, some people can suffer from chronic diseases if the biological clock is altered with.

The rodents were placed in a controlled roomed with five of them receiving time restricting calories while the other five rodents receive intermittent fasting. Researchers report that energy-restricted rats fed a “synthetic” diet consisting of 48% sucrose, 32% casein, 6% corn oil, and defined mineral and salt mixes were given to five mice. While the other rats consumed ad libitum of the identical diet of 1 kcal= 4.186kJ of healthy eating and proper macronutrients. Rodents consume ad libitum or reduced calorie diets throughout the research. However, caloric restriction (CR) limited total food consumption, feeding time of 12 hours every day, four –

Romijn, J. A., Coyle, E. F., Sidossis, L. S., Gastaldelli, A., Horowitz, J. F., Endert, E & Wolfe, R. R. (1993). Regulation of endogenous fat and carbohydrate metabolism in relation to exercise intensity and duration. American Journal of Physiology - Endocrinology and Metabolism, 265(3), 380-391. Wu, C., Nicholas, C., Williams, C., Took, A., & Hardy, L. (2003).

We defined perinatal (p) period as the time including gestation and lactation. Male offspring were used as there is an inherent difference between genders and their response to diet induced-NASH, with estrogen likely playing a protective role in females (17), male mice have more fidelity to the human NASH phenotype, and appear to be more responsive to the FFC diet as suggested by the significant increase in ALT level, body weight and liver to body weight in the males when compared to the age-matched female breeders (Figure S1). Offspring were weaned to either FFC or chow diet generating 4 experimental groups pFFC-FFC [n=12], pFFC-Chow [n=9], pChow-FFC [n=11], pChow-Chow [n=12]) (Figure 1A). Mice were sacrificed at 10 weeks of age

Circadian rhythm is most known for the effect on sleep and wake, but it also affects eating, hormone secretion, urination, and much more (Kalat, 2013). Circadian rhythms are endogenous, which means they are generated from within the body. When broken down into its latin roots “circum” and “dies”, circadian means about a day. This name was coined as animal circadian rhythms last about a day, or around 24 hours (Kalat, 2013). Though rhythms are around 24 hours, each individual differs in their exact rhythm. This can be seen in the difference between “morning people”, who are alert and awake early in the day, and “night owls”, who awaken at a slower pace and reach their peak alertness later in the day (Taillard, Philip, Coste, Sagaspe, & Bioulac,

Another key study, performed by Darlington et al, analyzed the function of dCLOCK, the CLOCK protein in Drosophila. This study confirmed the identity of dclock, investigated the function of dCLOCK in regulation of the circadian genes per and tim, and demonstrated the function of PER and TIM in an inhibitory feedback loop (Darlington et al., 1998). First, a screen for a homolog of the mouse clock gene in Drosophila was conducted using the cDNA library of adult Drosophila heads and a probe of the mClock gene. A high yield of overlap in the clones confirmed the presence of a homolog of the mClock gene in Drosophila (Darlington et al., 1998). A Southern blot of Drosophila genomic DNA with a dclock probe produced only one band, indicating that dclock is in fact a single-copy gene (Darlington et al., 1998). Dclock was then used in a Northern blot to probe RNA from the entire body of Drosophila. Results showed that the probe hybridized to RNA in the head, the body, and the appendages of the fly (Darlington et al., 1998). This showed that the expression of dclock occurs at a variety of locations (Darlington et al., 1998). The expression of dclock was then analyzed in terms of the light-dark cycle. Expression patterns during the 24 hour time period confirmed that dclock oscillates with the circadian rhythm (Darlington et al., 1998). After the initial tests to establish dclock as a circadian gene were completed, the function of the protein dCLOCK in the regulation of transcription of

Bremner WJ, Vitiello MV, Prinz PN 1983 Loss of circadian rhythmicity in blood testosterone levels with aging in normal men. J Clin Endocrinol Metab 56:1278-1281

CPT activities are high compared to other vertebrates and HOAD are similar-> fatty acid oxidation is important energy source

A large population study found that insomniacs tend to be physically smaller than normal sleepers: they have smaller skeletons and less lean body mass. Incidence of obesity is also higher. Insomnias are more often underweight and overweight. They also have a different blood analysis profile: levels of albumin are lower and insulin and triglycerides are higher. The glutamyl transferase enzyme, frequently used as a diagnostic marker, is also higher in insomniacs, and this incidence, along with high insulin levels, can suggest diabetes. The study found insomniacs have diabetes in higher numbers than normal sleepers. Insomniacs also tend to have high calcium levels in the blood – which has been shown to be associated with cognitive decline in the long run. (“Insomnia” Tuck). The PER3 gene is known to influence the circadian system, and one variant is associated with a lower homeostatic sleep drive. While scientists do not say there is a direct connection, the findings imply people with this variant PER3(4/4) are vulnerable for insomnia. Scientists found a gene mutation named hDEC2-P385R which is a transcriptional repressor and has been found to cause individuals to be short sleepers. ( “Insomnia” Tuck). Linkage studies have shown some hypotheses that are very promising for finding a genetic link in the

Sleep is an essential part of any person’s health, however few people seem to realize that not receiving enough sleep can double the risk of becoming obese.“The ‘epidemic’ of obesity is paralleled by a ‘silent epidemic’ of reduced sleep duration with short sleep duration linked to the increased risk of obesity both in adults and in children”(Professor Cappuio). Someone who has developed a sleeping disorder such as, sleep apnea, may not have the motivation to diet or exercise, due to daytime sleepiness. Scientists at the University of Chicago conucted a study in 1999, which restricted eleven healthy young adults to four hours of sleep for six nights. The scientist discovered that restricting individuals to this amount of sleep impaired metabolism functions and disrupted hormone levels. It was also found that the subject’s abilities to process glucose in the blood had declined, in some cases to the level of diabities. ” Not only does obesity contribute to sleep problems such as sleep apnea, but sleep problems can also contribute to obesity.”(National Sleep Foundation)