Analysis and visualization of quantitative proteomics data using FragPipe-Analyst.

The FragPipe computational proteomics platform is gaining widespread popularity among the proteomics research community because of its fast processing speed and user-friendly graphical interface. Although FragPipe produces well-formatted output tables that are ready for analysis, there is still a need for an easy-to-use and user-friendly downstream statistical analysis and visualization tool. FragPipe-Analyst addresses this need by providing an R shiny web server to assist FragPipe users in conducting downstream analyses of the resulting quantitative proteomics data. It supports major quantification workflows including label-free quantification, tandem mass tags, and data-independent acquisition. FragPipe-Analyst offers a range of useful functionalities, such as various missing value imputation options, data quality control, unsupervised clustering, differential expression (DE) analysis using Limma, and gene ontology and pathway enrichment analysis using Enrichr. To support advanced analysis and customized visualizations, we also developed FragPipeAnalystR, an R package encompassing all FragPipe-Analyst functionalities that is extended to support site-specific analysis of post-translational modifications (PTMs). FragPipe-Analyst and FragPipeAnalystR are both open-source and freely available.

Region fine-grained attention network for accurate bone age assessment.

Bone age assessment plays a vital role in monitoring the growth and development of adolescents. However, it is still challenging to obtain precise bone age from hand radiography due to these problems: 1) Hand bone varies greatly and is always masked by the background; 2) the hand bone radiographs with successive ages offer high similarity. To solve such issues, a region fine-grained attention network (RFGA-Net) was proposed for bone age assessment, where the region aware attention (RAA) module was developed to distinguish the skeletal regions from the background by modeling global spatial dependency; then the fine-grained feature attention (FFA) module was devised to identify similar bone radiographs by recognizing critical fine-grained feature regions. The experimental results demonstrate that the proposed RFGA-Net shows the best performance on the Radiological Society of North America (RSNA) pediatric bone dataset, achieving the mean absolute error (MAE) of 3.34 and the root mean square error (RMSE) of 4.02, respectively.

Origins of the Ediacaran Doushantuo High-Grade Primary Phosphorites at Kaiyang, Guizhou Province, China.

The Ediacaran Doushantuo phosphate deposit in Kaiyang, Guizhou Province, China, contains thick phosphate ores. Most of the ores are reconstituted phosphorite, and there have been few studies of the primary phosphorites, which has led to controversy regarding the origins and nature of mineralization of these phosphate-rich deposits. We identified high-grade primary phosphorites in the Kaiyang area and undertook a stratigraphic, petrological, sedimentological, geochemical, and isotopic study of these rocks. Moving up-section, the Longshui phosphate ore deposit comprises granular, micritic, stromatolitic, honeycomb, and sandy phosphorites. The first four types of phosphorite contain abundant biological structures, such as spherical, lobe-like, and amorphous forms. These are mainly fossils of benthic multicellular red algae, along with other types of algae. These fossils comprise >70% of the phosphorites, indicating that these are protist phosphorites. The ores are massive, unstratified, and contain numerous layered cavity structures, indicating that the ore bed was originally a reef. The phosphorites have P2O5 contents of 38.6-40.2 wt %, with an average of 38.9 wt %. The Al2O3 + TiO2 values are 0.02-0.44 wt %. The δ18O values of the samples vary from 13.76 to 16.57‰, with an average of 14.60‰, and δ13C values range from -15.789 to -8.697‰, with an average of -13.133‰. The samples exhibit rare-earth element patterns that are enriched with middle rare-earth elements and have strongly negative Ce anomalies. The geochemical features show that the reef was deposited in clear and oxidized waters. The discovery of this high-grade protist phosphorite shows that the involvement of algae was key to the formation of the Kaiyang phosphate-rich deposit.

Bone age assessment from articular surface and epiphysis using deep neural networks.

Bone age assessment is of great significance to genetic diagnosis and endocrine diseases. Traditional bone age diagnosis mainly relies on experienced radiologists to examine the regions of interest in hand radiography, but it is time-consuming and may even lead to a vast error between the diagnosis result and the reference. The existing computer-aided methods predict bone age based on general regions of interest but do not explore specific regions of interest in hand radiography. This paper aims to solve such problems by performing bone age prediction on the articular surface and epiphysis from hand radiography using deep convolutional neural networks. The articular surface and epiphysis datasets are established from the Radiological Society of North America (RSNA) pediatric bone age challenge, where the specific feature regions of the articular surface and epiphysis are manually segmented from hand radiography. Five convolutional neural networks, i.e., ResNet50, SENet, DenseNet-121, EfficientNet-b4, and CSPNet, are employed to improve the accuracy and efficiency of bone age diagnosis in clinical applications. Experiments show that the best-performing model can yield a mean absolute error (MAE) of 7.34 months on the proposed articular surface and epiphysis datasets, which is more accurate and fast than the radiologists. The project is available at https://github.com/YameiDeng/BAANet/, and the annotated dataset is also published at https://doi.org/10.5281/zenodo.7947923.

Estimating the CO2 emissions of Chinese cities from 2011 to 2020 based on SPNN-GNNWR.

Global warming is a serious threat to human survival and health. Facing increasing global warming, the issue of CO2 emissions has attracted more attention. China is a major contributor of anthropogenic CO2 emissions and so it is essential to accurately estimate China's CO2 emissions and analyze their changing characteristics. This study recalculates CO2 emissions from Chinese cities from 2011 to 2020 using the SPNN-GNNWR model and multiple factors to reduce the uncertainty in emission estimates. The SPNN-GNNWR model has excellent predictions (R2: 0.925, 10-fold CV R2: 0.822) when cross-validation is used. The results indicate that the total CO2 emissions in China calculated by the model are close to those accounted for by other authorities in the world, with the total CO2 emissions increasing from 9.122 billion tonnes in 2011 to 9.912 billion tonnes in 2020. The city with the largest increase in CO2 emissions is Tianjin, and the city with the largest decrease is Beijing. The study also reveals the regional differences in CO2 emissions in Chinese mainland, including emissions, emission intensity and per capita emissions. Capturing and understanding the emissions and the related socioeconomic characteristics of different cities can help to develop effective emission mitigation strategies.

ASW-Net: Adaptive Spectral Wavelet Network for Accurate Fetal ECG Extraction.

Noninvasive fetal ECG (FECG) is of great significance for monitoring fetal health. However, it is challenging to extract FECG signals from the abdominal ECG signal (AECG) due to the complexity of the task: 1) FECG signals are routinely mixed with noise; 2) FECG signals are aliased with maternal ECG signals in the time and frequency domain. To solve such problems, an adaptive spectral wavelet network (ASW-Net) is proposed for FECG extraction, where the adaptive spectral wavelet module, which can improve the computational efficiency by replacing convolution operation with element-wise Hadamard product in the frequency domain, is first developed to extract FECG components with different frequencies; then, the residual attention module is devised to distinguish FECG signals from noise by capturing waveform details; finally, the inverse spectral wavelet module is designed to reconstruct FECG signals from multi-resolution FECG components. Experiments conducted on the benchmarks demonstrate that the proposed ASW-Net outperforms the state-of-the-art methods.

Modeling the distribution of malachite green in zebrafish using matrix-assisted laser desorption/ionization mass spectrometry imaging.

Understanding the spatial distribution of bioactive small molecules is indispensable for elucidating their biological or pharmaceutical roles. Here, a rapid and effective analysis strategy was introduced to study the distribution of veterinary drugs in aquatic products. Malachite green (MG), one of the most widely used veterinary drugs in aquaculture, was selected as the targeted compound. Zebrafish (Danio rerio) was used as a model organism. After an exposure test, the matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) technique was applied to directly analyze the content changes of malachite green in zebrafish tissues. The reliable relationship of exposure time and content change of MG was described precisely by the extended Freundlich equation. The process of modeling was discussed in detail, and some important parameters or trend information was obtained, including the maximum content of MG in different fish tissues, time to maximum content, elimination time, equilibrium content, and so on. With a simplification of sample pretreatment, this research strategy can be used for monitoring the spatial distribution of veterinary drugs and related metabolites of laboratory-exposed fish. The obtained model can provide a perspective for rational drug use in aquaculture and precise drug residue detection in production activities.

Development and assessment of a mental health preliminary screening questionnaire for cancer patients.

BACKGROUND: The mental health of cancer patients is attracting increasing attention. Thus, an efficient mental health preliminary screening questionnaire (MHPSQ) for cancer patients is required. This study sought to develop an MHPSQ, test its reliability and validity, and administer it among cancer patients. METHODS: A literature review and interviews with 8 patients were conducted to determine the questionnaire item pool, and experts were consulted to confirm the first draft. Next, 150 cancer patients were selected for the project analysis and validity assessment to develop the final document; 400 patients were then selected for the reliability and validity tests. After which, 1,000 patients were assessed using the Self-Rating Anxiety Scale, Self-Rating Depression Scale (SDS), and MHPSQ, and the completion times for each scale were evaluated. Finally, the specificity and sensitivity of the MHPSQ were then determined. RESULTS: Four factors (i.e., mood, sleep, social function, and interpersonal relationships) were ultimately retained as MHPSQ items. MHPSQ's Cronbach's α and test-retest reliability were 0.76 and 0.92, respectively. The time required to complete the MHPSQ was 83.90±19.00 s. Its sensitivity and specificity were 92.4% and 98.5%, respectively. CONCLUSIONS: The MHPSQ has good reliability and validity. Comprising only 4 items, it is easy to operate and effectively identifies patients with psychological abnormalities.

The distribution and changes of glycoalkaloids in potato tubers under different storage time based on MALDI-TOF mass spectrometry imaging.

Glycoalkaloids (GAs) are toxic secondary metabolites in potatoes, which are harmful to human body. The storage time has a great influence on the biosynthesis and distribution of GAs. In present study, an imaging mass microscope (iMScope) was used to investigate the distribution and changes of GAs in potato tubers under different storage time (0, 10, 15, 20, 30, 40 and 60 days). We established a growth model with logistic equation to evaluate the growth trends of four major GAs in sprout, periderm and medulla. The results showed that the growth rate and relative contents of four GAs in sprout and periderm were significantly higher than that in medulla. In addition, four GAs also presented different change trends. For dehydrosolanine and α-solanine, rapid growth period of these two GAs in sprout (about at the day 23, similar to these in medulla) was later than which period in periderm (about at the day 17), while rapid growth of dehydrochaconine and α-chaconine appeared at almost the same time (about at the day 20). Based on the biosynthesis and metabolism of GAs, we have made possible explanations for these results. This study is useful for comprehending the metabolism of GAs in different parts and monitoring food safety in potatoes.

Targeted analysis of six emerging derivatives or metabolites together with 25 common macrolides in milk using Quick, Easy, Cheap, Effective, Rugged and Safe extraction and ultra-performance liquid chromatography quadrupole/electrostaticfield orbitrap mass spectrometry.

An analytical method for the determination of six emerging derivatives or metabolites together with 25 common macrolides antibiotics in milk by ultra-performance liquid chromatography quadrupole/electrostaticfield orbitrap mass spectrometry was established. The samples were purified with optimized Quick, Easy, Cheap, Effective, Rugged, Safe methods. The amounts of primary-secondary amine, C18, and sodium acetate adsorbent materials were optimized by response surface method to obtain the best purification effect. The chromatographic separation was carried out using the XBridge-C18 (2.1 × 100 mm, 3.5 µm, Waters) column with mobile phase of acetonitrile with 0.1% v/v formic acid-water solutions (containing 10 mmol/L ammonium acetate), separated by gradient elution. The instrument was operated in the detection mode of electrospray positive and negative ions with Full MS/data dependent MS2 acquisition mode detection, external standard method was used for quantitative analysis. The limits of detection and limits of quantitation of 31 compounds were 0.1-0.5 µg/L and 0.5-2.0 µg/L, respectively. A total of 31 compounds performed a good linearity in the range of 1 to 200 µg/L, and the correlation coefficient was greater than 0.990. The spiked recoveries in milk samples were 81.07-110.1% and the relative standard deviation was less than 5.1%. The method was successful applied to actual sample testing in the market.

Pathway attenuation of fatty acid beta-oxidation in the skeletal muscle of a type 2 diabetic mouse model.

RATIONALE: Whether catabolic abnormalities of fatty acids exist in the skeletal muscle of type 2 diabetes mellitus (T2DM) has not been determined. In this study, we postulated that a systematic evaluation of the protein abundance and metabolic activity related to fatty acids in the skeletal muscle tissues of a T2DM mouse model was feasible to address this question. METHODS: Mitochondria were extracted from wild-type (WT) and db/db mice followed by quantitative analysis of the proteins involved in mitochondrial fatty acid oxidation (mFAO). The pathway activity of mFAO in skeletal muscle tissues was monitored in vitro using mass spectrometry, and tissue lipidomic analysis was conducted in profiling and target mode to distinguish the levels of long-chain acylcarnitines between WT and db/db mice. RESULTS: Two proteins related to the mFAO pathway were significantly downregulated in the skeletal muscle mitochondria of db/db mice. The measurement of mFAO pathway activity in vitro revealed that the abundance of long-chain acylcarnitines (C14 to C18) in db/db mice was lower than that in WT mice, and the determination of acylcarnitines in skeletal muscle tissues in vivo revealed that most long-chain acylcarnitines were decreased in db/db mice. CONCLUSIONS: The findings of lower abundance of ACAD9 and CPT1B, reduced activity of the mFAO pathway in vitro and decreased acylcarnitines in vivo firmly support that the mFAO pathway in the skeletal muscle of diabetic mice is attenuated, possibly resulting in cell/tissue dysfunction in diabetes.

[Determination of 15 lipid regulators in fish meat by QuEChERS-ultra performance liquid chromatography-quadrupole/ electrostatic field orbitrap mass spectrometry].

An analytical method was established for the determination of 15 lipid regulators in fish meat by QuEChERS combined with ultra performance liquid chromatography-quadrupole/electrostatic field orbitrap mass spectrometry (UPLC-Q/Orbitrap-MS). The samples were purified by optimized QuEChERS methods. The amounts of the adsorbent materials (primary secondary amine (PSA) (20, 60, 100, 140 and 180 mg), C18 (40, 100, 160, 220 and 280 mg) and sodium acetate (0.2, 0.6, 1.0, 1.4 and 1.8 g)) were optimized by the response surface method to obtain the best purification effect. The target compounds were separated on an XBridge-C18 column (100 mm×2.1 mm, 3.5 µm) using acetonitrile-0.1% (v/v) formic acid aqueous solution (containing 1.5 mmol/L ammonium acetate) as the mobile phases by a gradient elution program. Qualitative and quantitative analysis of the target compounds were performed in the full scan and secondary mass spectrometry scan (dd-MS2) modes with positive and negative ionization. The target compounds showed good linear relationships in their respective ranges, with correlation coefficients (R2) greater than 0.99. The limits of detection (LOD, S/N=3) and limits of quantification (LOQ, S/N=10) were in the range of 0.2-1.0 µg/kg and 0.3-3.1 µg/kg, respectively. The average recoveries were 76.4%-116.0% at LOQ, 2-fold LOQ, and 10-fold LOQ levels. The intra-day relative standard deviations (RSDs) were 1.0%-7.9%, and the inter-day RSDs were 1.7%-18.4%. The method is simple, sensitive and accurate, and it is suitable for the determination and quantification of lipid regulators in fish meat.

[Research advances in imaging technology for food safety and quality control].

Food quality and safety are issues of concern to the government, food industry, and consumers; hence, it is imperative to detect harmful substances in foodstuff. Traditional techniques for this purpose include biochemical methods and instrumental analysis methods such as chromatography and chromatography-mass spectrometry. These methods, however, are time-consuming and unable to obtain the spatial distribution of the analytes. Therefore, the development of rapid, non-destructive, real-time, and visual detection technologies has emerged as a hotspot in the field of food research. In recent years, hyperspectral imaging, which combines imaging and spectral technology, is rapidly gaining ground. This technique allows one to determine the geometrical characteristics and chemical composition of samples. Compared with traditional spectral technologies, hyperspectral imaging has the advantages of wide detection ranges, in addition to being real-time and non-destructive. At present, hyperspectral imaging is widely used in meat quality evaluation, detection of adulteration, and meat classification. In addition, Raman imaging is mainly used for the detection of illegal additives in food and for adulteration detection. This technology is fast, non-destructive, and low cost; furthermore, spectral and spatial information of the targets can be simultaneously obtained. Mass spectrometry imaging allows for the visualization and high-throughput analysis of sample tissues, without the need for complex sample preparation steps such as labeling and staining. Compared with other imaging technologies, mass spectrum information of substances can be obtained by mass spectrometry imaging. As a molecular visualized technology, it helps obtain the spatial distribution of nutrients and harmful substances in food. Mass spectrometry imaging has unique advantages in food research, e. g., it is used for molecular-level detection and accurate positioning of substances, and hence, it has excellent application prospects in this field. In this paper, recent literature data about imaging technologies in the field of food research, including 72 reports published in professional local and overseas magazines, are collated. The principles of hyperspectral imaging, Raman imaging, and mass spectrometry imaging are introduced, along with the detailed applications of these methods in the quality detection, source identification, and microbial pollution of food. In addition, it also includes food physical damage, food adulteration and food chemical residues. Besides, the advantages and disadvantages of these imaging technologies are discussed. Finally, prospects for the development of imaging technologies in food research are presented. Future work related to hyperspectral imaging should focus on the development of high-sensitivity cameras and high-resolution systems. Improving the data processing efficiency and adding prediction models are also key points for the future. Future studies on Raman imaging can focus on the application of different chemometrics algorithms that would improve the evaluation of food quality and safety parameters. Expanding the scope of application of these methods in food research will also be the focus of future research. Regarding mass spectrometry imaging, attempts should be made to improve the ionization methods, detection sensitivity, spatial resolution, and data processing effectiveness. Additionally, the combination of spectral imaging and mass spectrometry imaging gives full play to their advantages, so that spectral and mass spectrometry information of the targets can be obtained. In short, the application of imaging technologies in food research is expected to be more promising.

pClean: An Algorithm To Preprocess High-Resolution Tandem Mass Spectra for Database Searching.

Database searches of MS/MS spectra are the main approach to peptide/protein identification in proteomics. Since most database search engines only utilize a small portion of the original MS/MS signals for peptide detection, how to improve the quality of MS/MS signals is a primary concern for enhancement of the peptide/protein identification rate. A fundamental issue is that some noise MS signals, informative or uninformative, have to be filtered out prior to database searching. Herein, an integrative preprocessing algorithm was designed, termed pClean, which incorporates three modules to preprocess MS/MS spectra, such as the removal of isobaric-labeling related ions, the reduction in isotopic peaks, the deconvolution of ions with higher charges, and the clearance of uninformative MS/MS signals. In contrast to the currently available approaches to MS/MS data preprocessing, pClean enables treatment of MS/MS spectra with high mass accuracy and favors filtering for the labeling or nonlabeling of peptides. Data sets at various scales gained from mass spectrometers with high resolution were used to assess the quality of peptides identified after pClean treatment and to compare the pClean improvement with those of other software programs. On the basis of the analysis of peptides identified and the Mascot ion score, pClean was proven to be effective in the removal of mass spectral noise and the reduction of random matching. Compared with other software programs, pClean appeared to be beneficial in terms of preprocessing performances for the enhancement of confidence scores and the increase in peptides identified. pClean is available at https://github.com/AimeeD90/pClean_release .

[Rapid determination of six banned fragrance components in tea by QuEChERS-gas chromatography-tandem mass spectrometry].

QuEChERS purification was established as a sample pretreatment method and combined with gas chromatography-tandem mass spectrometry (GC-MS/MS) for the simultaneous detection of six banned fragrance components in tea (coumarin, thujone, 2-exylthiophene, trans-beta-acacene, p-mentha-1,8(10)diene-9-ethyl acetate, and butyl cinnamate). Tea samples were extracted with ethyl acetate and subjected to whirlpool oscillation, following which anhydrous magnesium sulfate, primary secondary amine (PSA), and octadecylsilyl silica gel (C18) were added to the supernatant. After centrifugation, the supernatant was filtered over a membrane and separated on a DB-5MS column. The six compounds were determined by GC-MS/MS in MRM mode, and quantified by a matrix-matched external standard method. The six banned flavor compounds exhibited good linearity over the range of 1-200 µg/L, and the linear correlation coefficients (R2) were greater than 0.999. The limits of detection were 0.005-1 µg/kg and the limits of quantification were 0.02-2 µg/kg, the average recoveries were 82.3%-93.1%, and the intra-day and inter-day RSDs were each less than 10%. The method is simple, rapid, accurate, and sensitive, and can be used for the simultaneous detection of six banned fragrance components in tea.

A Mechanistic Understanding of Hydrogen Peroxide Decomposition by Vanadium Minerals for Diethyl Phthalate Degradation.

The interaction of naturally occurring minerals with H2O2 affects the remediation efficiency of polluted sites in in situ chemical oxidation (ISCO) treatments. However, interactions between vanadium(V) minerals and H2O2 have rarely been explored. In this study, H2O2 decomposition by various vanadium-containing minerals including V(III), V(IV), and V(V) oxides was examined, and the mechanism of hydroxyl radical (•OH) generation for contaminant degradation was studied. Vanadium minerals were found to catalyze H2O2 decomposition efficiently to produce •OH for diethyl phthalate (DEP) degradation in both aqueous solutions with a wide pH range and in soil slurry. Electron paramagnetic resonance (EPR), X-ray photoelectron spectroscopy (XPS), X-ray diffraction (XRD) analyses, and free radical quenching studies suggested that •OH was produced via single electron transfer from V(III)/V(IV) to H2O2 followed a Fenton-like pathway on the surface of V2O3 and VO2 particles, whereas the oxygen vacancy (OV) was mainly responsible for •OH formation on the surface of V2O5 particles. This study provides new insight into the mechanism of interactions between vanadium minerals and H2O2 during H2O2-based ISCO.

Appraisal of the Missing Proteins Based on the mRNAs Bound to Ribosomes.

Considering the technical limitations of mass spectrometry in protein identification, the mRNAs bound to ribosomes (RNC-mRNA) are assumed to reflect the mRNAs participating in the translational process. The RNC-mRNA data are reasoned to be useful for appraising the missing proteins. A set of the multiomics data including free-mRNAs, RNC-mRNAs, and proteomes was acquired from three liver cancer cell lines. On the basis of the missing proteins in neXtProt (release 2014-09-19), the bioinformatics analysis was carried out in three phases: (1) finding how many neXtProt missing proteins have or do not have RNA-seq and/or MS/MS evidence, (2) analyzing specific physicochemical and biological properties of the missing proteins that lack both RNA-seq and MS/MS evidence, and (3) analyzing the combined properties of these missing proteins. Total of 1501 missing proteins were found by neither RNC-mRNA nor MS/MS in the three liver cancer cell lines. For these missing proteins, some are expected higher hydrophobicity, unsuitable detection, or sensory functions as properties at the protein level, while some are predicted to have nonexpressing chromatin structures on the corresponding gene level. With further integrated analysis, we could attribute 93% of them (1391/1501) to these causal factors, which result in the expression products scarcely detected by RNA-seq or MS/MS.

Assessing Transcription Regulatory Elements To Evaluate the Expression Status of Missing Protein Genes on Chromosomes 11 and 19.

During an investigation of missing proteins with the RNA-seq data acquired from three liver cancer cell lines, the majority of the missing protein coding genes (MPGs) located at chromosome 11 (chr11) had no corresponding mRNAs, while a high percentage of the MPGs on chr19 were detected at the mRNA level. The phenomenon, which was also observed in more than 40 cell lines, led to an inquiry of causation of the different transcriptional statuses of the MPGs in the two chromosomes. We hypothesized that the special chromatin structure was a key element to regulate MPG transcription. Upon a systematical comparison of the effects of DNase I hypersensitive sites (DHSs), transcription factors (TFs), and histone modifications toward these genes or MPGs with/without mRNA evidence in chr11 and 19, we attributed the poor transcription of the MPGs to the weak capacity of these transcription regulatory elements, regardless of which chromosome the MPGs were located. We further analyzed the gene contents in chr11 and found a number of genes related to sensory functions in the presence of chr11. We postulate that a high number of sensory-related genes, which are located within special chromatin structure, could bring a low detection rate of MPGs in chr11.

Insights from ENCODE on Missing Proteins: Why ß-Defensin Expression Is Scarcely Detected.

ß-Defensins (DEFBs) have a variety of functions. The majority of these proteins were not identified in a recent proteome survey. Neither protein detection nor the analysis of transcriptomic data based on RNA-seq data for three liver cancer cell lines identified any expression products. Extensive investigation into DEFB transcripts in over 70 cell lines offered similar results. This fact naturally begs the question­Why are DEFB genes scarcely expressed? After examining DEFB gene annotation and the physicochemical properties of its protein products, we postulated that regulatory elements could play a key role in the resultant poor transcription of DEFB genes. Four regions containing DEFB genes and six adjacent regions on chromosomes 6, 8, and 20 were carefully investigated using The Encyclopedia of DNA Elements (ENCODE) information, such as that of DNase I hypersensitive sites (DHSs), transcription factors (TFs), and histone modifications. The results revealed that the intensities of these ENCODE features were globally weaker than those in the adjacent regions. Impressively, DEFB-related regions on chromosomes 6 and 8 containing several non-DEFB genes had lower ENCODE feature intensities, indicating that the absence of DEFB mRNAs might not depend on the gene family but may be reliant upon gene location and chromatin structure.