Dietary restriction impacts health and lifespan of genetically diverse mice.

Caloric restriction extends healthy lifespan in multiple species1. Intermittent fasting, an alternative form of dietary restriction, is potentially more sustainable in humans, but its effectiveness remains largely unexplored2-8. Identifying the most efficacious forms of dietary restriction is key for developing interventions to improve human health and longevity9. Here we performed an extensive assessment of graded levels of caloric restriction (20% and 40%) and intermittent fasting (1 and 2 days fasting per week) on the health and survival of 960 genetically diverse female mice. We show that caloric restriction and intermittent fasting both resulted in lifespan extension in proportion to the degree of restriction. Lifespan was heritable and genetics had a larger influence on lifespan than dietary restriction. The strongest trait associations with lifespan included retention of body weight through periods of handling-an indicator of stress resilience, high lymphocyte proportion, low red blood cell distribution width and high adiposity in late life. Health effects differed between interventions and exhibited inconsistent relationships with lifespan extension. 40% caloric restriction had the strongest lifespan extension effect but led to a loss of lean mass and changes in the immune repertoire that could confer susceptibility to infections. Intermittent fasting did not extend the lifespan of mice with high pre-intervention body weight, and two-day intermittent fasting was associated with disruption of erythroid cell populations. Metabolic responses to dietary restriction, including reduced adiposity and lower fasting glucose, were not associated with increased lifespan, suggesting that dietary restriction does more than just counteract the negative effects of obesity. Our findings indicate that improving health and extending lifespan are not synonymous and raise questions about which end points are the most relevant for evaluating aging interventions in preclinical models and clinical trials.

Plasma proteomic associations with genetics and health in the UK Biobank.

The Pharma Proteomics Project is a precompetitive biopharmaceutical consortium characterizing the plasma proteomic profiles of 54,219 UK Biobank participants. Here we provide a detailed summary of this initiative, including technical and biological validations, insights into proteomic disease signatures, and prediction modelling for various demographic and health indicators. We present comprehensive protein quantitative trait locus (pQTL) mapping of 2,923 proteins that identifies 14,287 primary genetic associations, of which 81% are previously undescribed, alongside ancestry-specific pQTL mapping in non-European individuals. The study provides an updated characterization of the genetic architecture of the plasma proteome, contextualized with projected pQTL discovery rates as sample sizes and proteomic assay coverages increase over time. We offer extensive insights into trans pQTLs across multiple biological domains, highlight genetic influences on ligand-receptor interactions and pathway perturbations across a diverse collection of cytokines and complement networks, and illustrate long-range epistatic effects of ABO blood group and FUT2 secretor status on proteins with gastrointestinal tissue-enriched expression. We demonstrate the utility of these data for drug discovery by extending the genetic proxied effects of protein targets, such as PCSK9, on additional endpoints, and disentangle specific genes and proteins perturbed at loci associated with COVID-19 susceptibility. This public-private partnership provides the scientific community with an open-access proteomics resource of considerable breadth and depth to help to elucidate the biological mechanisms underlying proteo-genomic discoveries and accelerate the development of biomarkers, predictive models and therapeutics1.

A data analysis framework for combining multiple batches increases the power of isobaric proteomics experiments.

We present a framework for the analysis of multiplexed mass spectrometry proteomics data that reduces estimation error when combining multiple isobaric batches. Variations in the number and quality of observations have long complicated the analysis of isobaric proteomics data. Here we show that the power to detect statistical associations is substantially improved by utilizing models that directly account for known sources of variation in the number and quality of observations that occur across batches.In a multibatch benchmarking experiment, our open-source software (msTrawler) increases the power to detect changes, especially in the range of less than twofold changes, while simultaneously increasing quantitative proteome coverage by utilizing more low-signal observations. Further analyses of previously published multiplexed datasets of 4 and 23 batches highlight both increased power and the ability to navigate complex missing data patterns without relying on unverifiable imputations or discarding reliable measurements.

Impact of regulatory variation across human iPSCs and differentiated cells.

Induced pluripotent stem cells (iPSCs) are an essential tool for studying cellular differentiation and cell types that are otherwise difficult to access. We investigated the use of iPSCs and iPSC-derived cells to study the impact of genetic variation on gene regulation across different cell types and as models for studies of complex disease. To do so, we established a panel of iPSCs from 58 well-studied Yoruba lymphoblastoid cell lines (LCLs); 14 of these lines were further differentiated into cardiomyocytes. We characterized regulatory variation across individuals and cell types by measuring gene expression levels, chromatin accessibility, and DNA methylation. Our analysis focused on a comparison of inter-individual regulatory variation across cell types. While most cell-type-specific regulatory quantitative trait loci (QTLs) lie in chromatin that is open only in the affected cell types, we found that 20% of cell-type-specific regulatory QTLs are in shared open chromatin. This observation motivated us to develop a deep neural network to predict open chromatin regions from DNA sequence alone. Using this approach, we were able to use the sequences of segregating haplotypes to predict the effects of common SNPs on cell-type-specific chromatin accessibility.

Allele-specific expression reveals interactions between genetic variation and environment.

Identifying interactions between genetics and the environment (GxE) remains challenging. We have developed EAGLE, a hierarchical Bayesian model for identifying GxE interactions based on associations between environmental variables and allele-specific expression. Combining whole-blood RNA-seq with extensive environmental annotations collected from 922 human individuals, we identified 35 GxE interactions, compared with only four using standard GxE interaction testing. EAGLE provides new opportunities for researchers to identify GxE interactions using functional genomic data.

Modeling the Impact of Transcendental Meditation on Stroke Incidence and Mortality.

OBJECTIVES: Meditation has shown promise in clinical trials in reducing systolic blood pressure, one of the main risk factors for stroke. We aim to estimate the potential benefits of popularizing meditation on stroke incidence and mortality in the United States (U.S.). METHODS: We developed a dynamic population-based microsimulation model to simulate the disease progression of each individual and compute disease burden. We calibrated the microsimulation model for stroke incidence and further validated it by comparing the stroke-related mortality for each age group generated by the model with that observed in the U.S. We used the population simulation model to estimate the effects of meditation intervention on the number of stroke cases and deaths over a course of 15 years. RESULTS: Our results show that we could avert nearly 200,000 stroke cases and 50,000 stroke-related deaths over the course of 15 years. Our sensitivity analysis reveals that most of the benefits come from applying the intervention for individuals older than 60 years. In addition, meditation acceptance and adherence rate play a critical role in its effectiveness. CONCLUSIONS: The practice of meditation, if properly utilized along with the regular antihypertensive medication, could substantially alleviate the burden of stroke in the U.S. In order to design an effective meditation program, policymakers may prioritize funding to the programs that aim to encourage older individuals to practice meditation.

Addressing Oral Health Disparities of a Tribal Population Through a Combined Implementation of Focus Group Discussion, Mobile Technology Networking, and Creating a Supportive Environment: A Prospective Study.

BACKGROUND AND OBJECTIVE:  Oral health disparities generally exist among tribal populations, prompting creative solutions to tackle these challenges. By using a combined implementation strategy of including focus group discussion (FGD), mobile technology networking (MTN), and creating a supportive environment, this study aims to assess and bring positive changes in oral health in these populations. METHODS: The current study employed a mixed-method approach on a sample of 100 tribal volunteers. Qualitative assessment included FGD conducted regularly for three months based on themes such as oral hygiene habits, access to oral health, technology in oral health, the relationship of oral health to general health, and the role of diet in oral health. Quantitative evaluation included recording of the oral hygiene index-simplified and gingival index to measure gingival status. Messages on oral health were routinely posted to mobile phones to reinforce oral health education. Appropriate use of indigenous oral hygiene aids (neem and datun) was also taught during the discussion session. Clinical examinations were compared before and after FGD. Data were analyzed using IBM SPSS Statistics for Windows, Version 25 (Released 2017; IBM Corp., Armonk, New York, United States). A paired 't' test was used to find significant differences in gingival status at p<0.05. RESULTS: The FGD sessions deduced observations such as limited access to dental care, inadequate oral hygiene practices such as usage of neem sticks and twigs, and lack of oral health awareness. The implementation of MTN facilitated the dissemination of oral health information and enhanced communication between community members and healthcare providers. The gingival index score significantly improved from pre-FGD to post-FGD with a mean difference of 0.41700 significant at p=0.000. Oral hygiene of the target population shifted from "Fair" oral hygiene status to "Good" oral hygiene status. CONCLUSION: The combined implementation of FGD, MTN, and creation of a supportive environment demonstrated promising results in addressing oral health disparities among the tribal population. The interventions led to improved gingival status and better utilization of oral hygiene practices. These findings highlight the importance of tailored interventions, community engagement, and mobile technology in addressing oral health disparities in tribal populations. Ongoing support, sustainability, and further research are necessary to ensure the long-term impact and effectiveness of these interventions.

Correlation Between BMI, Caries Prevalence, and Sugar-containing Beverage Intake in 6-10 Year Old Children.

Background: The beverages containing sugar are proven risk factors for obesity and dental caries. Therefore, owing to the shared risk factors, an interrelationship is suspected between BMI, sugar beverage consumption, and dental caries in children. Aims: The present trial was carried out to assess the interrelationship between BMI, sugar beverage consumption, and dental caries in children aged 6-10 years. Materials and Methods: Eighty-six children within the age range of 6-10 years answered the health questionnaire. The BMI was calculated, intra-oral assessment was done, the frequency of sweetened beverage consumption was recorded, and the collected data were subjected to the statistical evaluation to formulate results. Results: On evaluation, a non-significant difference was observed in BMI levels in the four groups (P = 0.12). Whole-milk intake also showed an inverse correlation with dental caries and BMI, but this correlation was statistically non-significant with the respective values of P = 0.57 and 0.55. A similar inverse relationship was seen for low-fat milk for caries and BMI with P = 0.65 and 0.45, respectively. Regarding soft drinks, 44.1% (n = 38) took soft drinks, and a non-significant relation between caries and intake as well as BMI and intake with P = 0.86 and 0.55, respectively. Conclusion: Within its limitations, the present study concludes that no correlation exists between BMI and dental caries as well as between sugar-containing beverage consumption and dental caries. Also, BMI and sugar-containing beverage consumption showed no correlation in children aged 6-10 years.

Age and diet shape the genetic architecture of body weight in diversity outbred mice.

Understanding how genetic variation shapes a complex trait relies on accurately quantifying both the additive genetic and genotype-environment interaction effects in an age-dependent manner. We used a linear mixed model to quantify diet-dependent genetic contributions to body weight measured through adulthood in diversity outbred female mice under five diets. We observed that heritability of body weight declined with age under all diets, except the 40% calorie restriction diet. We identified 14 loci with age-dependent associations and 19 loci with age- and diet-dependent associations, with many diet-dependent loci previously linked to neurological function and behavior in mice or humans. We found their allelic effects to be dynamic with respect to genomic background, age, and diet, identifying several loci where distinct alleles affect body weight at different ages. These results enable us to more fully understand and predict the effectiveness of dietary intervention on overall health throughout age in distinct genetic backgrounds.

Body weight is one trait influenced by genes, age and environmental factors. Both internal and external environmental pressures are known to affect genetic variation over time. However, it is largely unknown how all factors ­ including age ­ interact to shape metabolism and bodyweight. Wright et al. set out to quantify the interactions between genes and diet in ageing mice and found that the effect of genetics on mouse body weight changes with age. In the experiments, Wright et al. weighed 960 female mice with diverse genetic backgrounds, starting at two months of age into adulthood. The animals were randomized to different diets at six months of age. Some mice had unlimited food access, others received 20% or 40% less calories than a typical mouse diet, and some fasted one or two days per week. Variations in their genetic background explained about 80% of differences in mice's weight, but the influence of genetics relative to non-genetic factors decreased as they aged. Mice on the 40% calorie restriction diet were an exception to this rule and genetics accounted for 80% of their weight throughout adulthood, likely due to reduced influence from diet and reduced interactions between diet and genes. Several genes involved in metabolism, neurological function, or behavior, were associated with mouse weight. The experiments highlight the importance of considering interactions between genetics, environment, and age in determining complex traits like body weight. The results and the approaches used by Wright et al. may help other scientists learn more about how the genetic predisposition to disease changes with environmental stimuli and age.

Automated, high-dimensional evaluation of physiological aging and resilience in outbred mice.

Behavior and physiology are essential readouts in many studies but have not benefited from the high-dimensional data revolution that has transformed molecular and cellular phenotyping. To address this, we developed an approach that combines commercially available automated phenotyping hardware with a systems biology analysis pipeline to generate a high-dimensional readout of mouse behavior/physiology, as well as intuitive and health-relevant summary statistics (resilience and biological age). We used this platform to longitudinally evaluate aging in hundreds of outbred mice across an age range from 3 months to 3.4 years. In contrast to the assumption that aging can only be measured at the limits of animal ability via challenge-based tasks, we observed widespread physiological and behavioral aging starting in early life. Using network connectivity analysis, we found that organism-level resilience exhibited an accelerating decline with age that was distinct from the trajectory of individual phenotypes. We developed a method, Combined Aging and Survival Prediction of Aging Rate (CASPAR), for jointly predicting chronological age and survival time and showed that the resulting model is able to predict both variables simultaneously, a behavior that is not captured by separate age and mortality prediction models. This study provides a uniquely high-resolution view of physiological aging in mice and demonstrates that systems-level analysis of physiology provides insights not captured by individual phenotypes. The approach described here allows aging, and other processes that affect behavior and physiology, to be studied with improved throughput, resolution, and phenotypic scope.

Intermittent fasting and caloric restriction interact with genetics to shape physiological health in mice.

Dietary interventions can dramatically affect physiological health and organismal lifespan. The degree to which organismal health is improved depends upon genotype and the severity of dietary intervention, but neither the effects of these factors, nor their interaction, have been quantified in an outbred population. Moreover, it is not well understood what physiological changes occur shortly after dietary change and how these may affect the health of an adult population. In this article, we investigated the effect of 6-month exposure of either caloric restriction (CR) or intermittent fasting (IF) on a broad range of physiological traits in 960 1-year old Diversity Outbred mice. We found CR and IF affected distinct aspects of physiology and neither the magnitude nor the direction (beneficial or detrimental) of effects were concordant with the severity of the intervention. In addition to the effects of diet, genetic variation significantly affected 31 of 36 traits (heritabilities ranged from 0.04 to 0.65). We observed significant covariation between many traits that was due to both diet and genetics and quantified these effects with phenotypic and genetic correlations. We genetically mapped 16 diet-independent and 2 diet-dependent significant quantitative trait loci, both of which were associated with cardiac physiology. Collectively, these results demonstrate the degree to which diet and genetics interact to shape the physiological health of adult mice following 6 months of dietary intervention.

Childhood dental fear in children aged 7-11 years old by using the Children's Fear Survey Schedule-Dental Subscale.

In children, dental fear is not only associated with fear of pain or invasive procedures, but it is also correlated with separation from parents or confronting unfamiliar people or environments. The Children's Fear Survey Schedule-Dental Subscale (CFSS-DS) was developed to evaluate dental fear in children, and this scale is now used across the world for evaluating dental fear. The aim of this study was to evaluate dental fear in children between 7-11 years of age and to find out the association between caries and fear of dental treatment. A total of 300 subjects of both sexes were enrolled in the study. Prior to the oral examination, all patients' attendants were informed about the study, and the subjects were asked to fill in a questionnaire regarding the CFSS-DS scale. The data obtained through the questionnaires were analyzed using the Chi-Square test. Fear scores were highest for "injections" (3.91±0.17), "dentist drilling" (3.91±0.10) and "choking" (3.65±0.82). It was also observed that subjects who had already visited a dental clinic or those who were familiar with the dental environment at an early stage of life were less anxious than patients who were receiving dental treatment for the first time. In this study, we found that female subjects were more anxious in comparison to male patients. Once the child's fear is identified, the dentist can use various behavior modification techniques to eliminate fear, explain the steps, and use the instruments accordingly until fear has vanished.

Prevalence of vitamin D inadequacy and bone turnover markers concerning bone mineral density in older persons.

Owing to the early tooth loss in a large population, the need for removable prosthodontic therapy has widely increased. Loss of teeth can occur due to trauma, periodontal disease, dental caries, and/or oral health negligence. The removable prosthesis is preferred over other teeth replacement methods owing to its cost-effectiveness. However, due to continuous alveolar bone resorption, relining of the removable prosthesis is needed regularly. We aimed to assess the prevalence of vitamin D deficiency and bone turnover marker concerning bone mineral density. Eighty complete denture wearers either in the maxilla or in the mandible or both constituted the study sample. Before the study, all subjects were explained the procedures of the study, and informed consent was obtained before proceeding with the study. Subjects were instructed to provide an early morning sample in a fasting state as serum osteocalcin (OCN) and C-terminal telopeptide (C-Tx) have diurnal variations. Blood samples were collected, and samples were ruled out for 25-OH vitamin D, serum OCN, and C-TX. To determine the association between frequency of relining and bone turnover marker, the collected data were subjected to statistical evaluation. Significant correlations were seen between bone C-Tx, turnover markers (p<0.001), and frequency of denture relining and osteocalcin (p<0.001). No statistically significant effect of gender, vitamin D levels, age, calcium levels, vitamin D supplements, and diabetes was seen on denture relining frequency. The present study concludes that an elevated level of the bone turnover markers such as OCN and C-Tx increases the frequency of relining.

A New Zinc Reinforced Glass Ionomer Cement: A Boon in Dentistry.

BACKGROUND: Conventional glass ionomer cement (CGIC) has many beneficial properties, but it has poor physical and mechanical properties. Therefore, new glass ionomer cement (GIC) is manufactured by adding zinc to improve the mechanical properties of GIC ChemFil Rock. This material possesses better flexural tensile strength and compressive strength in comparison to conventional to CGIC. OBJECTIVES: The aim of this study was to compare four properties of ZRGI like fracture toughness, surface micro-hardness, abrasive wear, and roughness to other GIC material, which are commercially available as: resin-coated glass ionomer (EQUIA FIL). MATERIALS AND METHODS: The study was done in dual phase. In phase-1, micro-hardness surface roughness, abrasion of four GIC and a composite resin as control was analyzed and in phase-2, fracture toughness of four GIC was done at 24 h interval so that all cement achieve its peak strength. RESULTS: Micro-hardness value of ChemFil Rock was lowest among different GIC groups. All four GIC group exhibit similar abrasion capacities, while composite were more wear-resistant significantly. Roughness change was highest on ChemFil Rock compared to other GIC. EQUIA FIL has the highest fracture toughness, followed by ChemFil Rock. CONCLUSION: We can conclude that incorporating zinc in the matrix of chemfil rock increases fracture toughness and good abrasive wear, but it does not improve micro-hardness or surface roughness.

Evaluation of Dental Status in Relation to Excessive Horizontal and Vertical Overlap in North Indian Population.

INTRODUCTION: Tooth wear (attrition) is a multifactorial entity, the pathogenesis of which involves many factors but the role of malocclusion cannot be undermined. Overjet and overbite play an important role in defining occlusion, hence they should also play an important role in causing attrition. OBJECTIVES: This study aims to evaluate the relationship of dental health status between the vertical and horizontal overlap. MATERIALS AND METHODS: This study was conducted on 600 patients selected randomly from rural population which were divided into three equal groups as per the predefined criteria, namely, horizontal overlap, vertical overlap, and control group. All patients were examined for the tooth attrition. RESULTS: In Group 1 - 73 patients out of 200 showed one or more teeth attrition, in Group 2 - 38 patients out of 200 showed one or more teeth attrition, and in the control group, 22 patients out of 200 showed one or more teeth attrition. CONCLUSION: It was concluded that both vertical and horizontal abnormal relationships of teeth play an important role in the etiology of attrition, with abnormal horizontal overlap being the more detrimental as compared to the vertical one.

Dry-EEG Manifestations of Acute and Insidious Hypoxia During Simulated Flight.

INTRODUCTION: Recently, portable dry electroencephalographs (dry-EEGs) have indexed cognitive workload, fatigue, and drowsiness in operational environments. Using this technology this project assessed whether significant changes in brainwave frequency power occurred in response to hypoxic exposures as experienced in military aviation.METHODS: There were 60 (30 women, 30 men) student Naval Aviators or Flight Officers who were exposed to an intense (acute) high-altitude (25,000 ft) normobaric hypoxic exposure, and 20 min later, more gradual (insidious) normobaric hypoxic exposure up to 20,000 ft while flying a fixed-wing flight simulation and monitored with a dry-EEG system. Using MATLAB, EEG frequencies and power were quantified and analyzed. Cognitive performance was also assessed with a cognitive task validated under hypoxia. Normobaric hypoxia and O2 saturation (Spo2) were produced and monitored using the Reduced Oxygen Breathing Device (ROBD2).RESULTS: Significant Spo2 decreases were recorded at acute 25K and insidious 20K simulated altitudes. Significant power decreases were recorded in all frequencies (alpha, beta, gamma, and theta) and all channels with acute 25K exposures. Gamma, beta, and theta frequency power were significantly decreased with insidious 20K exposures at most of the channels. The frequency power decreases corresponded to significant decreases in cognitive performance and flight performance. Most importantly, frequency power suppressions occurred despite 42% of the volunteers not perceiving they were hypoxic in the acute phase, nor 20% in the insidious phase.DISCUSSION: Results suggest EEG suppression during acute/insidious hypoxia can index performance decrements. These findings have promising implications in the development of biosensors that mitigate potential in-flight hypoxic physiological episodes.Rice GM, Snider D, Drollinger S, Greil C, Bogni F, Phillips J, Raj A, Marco K, Linnville S. Dry-EEG manifestations of acute and insidious hypoxia during simulated flight. Aerosp Med Hum Perform. 2019; 90(2):92-100.

Gender Differences in Dry-EEG Manifestations During Acute and Insidious Normobaric Hypoxia.

INTRODUCTION: Prior research suggests there may be gender differences with regards to hypoxia resilience. Our study was designed to determine whether there were differences between genders in neuronal electrical activity at simulated altitude and whether those changes correlated with cognitive and aviation performance decrements.METHODS: There were 60 student Naval Aviators or Flight Officers who completed this study (30 women, 30 men). Participants were exposed to increasing levels of normobaric hypoxia and monitored with dry EEG while flying a fixed-base flight simulation. Gender differences in brainwave frequency power were quantified using MATLAB. Changes in flight and cognitive performance were analyzed via simulation tasks and with a cognitive test validated under hypoxia.RESULTS: Significant decreases in theta and gamma frequency power occurred for women compared to men with insidious hypoxic exposures to 20K, with an average frequency power decrease for women of 19.4% compared to 9.3% for men in theta, and a 42.2% decrease in gamma for women compared to 21.7% for men. Beta frequency power correlated highest between genders, with an average correlation coefficient of r = 0.95 across seven channels.DISCUSSION: Results of this study suggest there is identifiable brain wave suppression for both men and women with hypoxic exposure and, moreover, there are significant differences in this suppression between genders. Beta frequency power was most sensitive for both genders and highly correlative compared to other brainwave frequencies. The implications of these findings are important considerations for next-generation aviation helmets, which may employ this technology as an early warning mechanism.Rice GM, Snider D, Drollinger S, Greil C, Bogni F, Phillips J, Raj A, Marco K, Linnville S. Gender differences in dry-EEG manifestations during acute and insidious normobaric hypoxia. Aerosp Med Hum Perform. 2019; 90(4):369-377.

Patterns of tobacco usage among subjects with potentially malignant oral lesions or conditions in Chennai city: A comparative study.

OBJECTIVE: To determine the patterns of tobacco usage among subjects with potentially malignant oral lesions or conditions through a comparative study design. METHODS: The study was carried out in a span of 2 months on a sample of 120 subjects; 60 in case group (30 subjects with leukoplakia and oral submucous fibrosis [OSMF], respectively) and 60 subjects in control group (30 current smokers and current chewers, respectively), attending the tobacco cessation clinic at a private dental college hospital in Chennai city. Demographic data, details of tobacco usage, and Fagerstrom nicotine dependence scores (FNTD) were recorded in a prevalidated tobacco cessation intake form. RESULTS: Cases with leukoplakia had a higher mean FNTD score when compared to the control group (P = 0.0001). The most common form of smokeless tobacco used by case (OSMF) subjects was found to be mawa (53%) significantly higher than the control group (P = 0.05). Mean FNTD scores of mawa users were higher than other tobacco users in both case and control group. CONCLUSION: The current study has hence put forth the role of mawa form of tobacco in causation of OSMF at a time when implementation of tougher anti-tobacco laws is the talk of the town.

RNA splicing is a primary link between genetic variation and disease.

Noncoding variants play a central role in the genetics of complex traits, but we still lack a full understanding of the molecular pathways through which they act. We quantified the contribution of cis-acting genetic effects at all major stages of gene regulation from chromatin to proteins, in Yoruba lymphoblastoid cell lines (LCLs). About ~65% of expression quantitative trait loci (eQTLs) have primary effects on chromatin, whereas the remaining eQTLs are enriched in transcribed regions. Using a novel method, we also detected 2893 splicing QTLs, most of which have little or no effect on gene-level expression. These splicing QTLs are major contributors to complex traits, roughly on a par with variants that affect gene expression levels. Our study provides a comprehensive view of the mechanisms linking genetic variation to variation in human gene regulation.