Integrative temporal multi-omics reveals uncoupling of transcriptome and proteome during human T cell activation.

Engagement of the T cell receptor (TCR) triggers molecular reprogramming leading to the acquisition of specialized effector functions by CD4 helper and CD8 cytotoxic T cells. While transcription factors, chemokines, and cytokines are known drivers in this process, the temporal proteomic and transcriptomic changes that regulate different stages of human primary T cell activation remain to be elucidated. Here, we report an integrative temporal proteomic and transcriptomic analysis of primary human CD4 and CD8 T cells following ex vivo stimulation with anti-CD3/CD28 beads, which revealed major transcriptome-proteome uncoupling. The early activation phase in both CD4 and CD8 T cells was associated with transient downregulation of the mRNA transcripts and protein of the central glucose transport GLUT1. In the proliferation phase, CD4 and CD8 T cells became transcriptionally more divergent while their proteome became more similar. In addition to the kinetics of proteome-transcriptome correlation, this study unveils selective transcriptional and translational metabolic reprogramming governing CD4 and CD8 T cell responses to TCR stimulation. This temporal transcriptome/proteome map of human T cell activation provides a reference map exploitable for future discovery of biomarkers and candidates targeting T cell responses.

Ergonomics in online education of medical undergraduates: A challenge to post-COVID transformation in educational activities.

BACKGROUND: Practicing incorrect postures in online and virtual education during the COVID-19 pandemic can cause significant study-related musculoskeletal problems among students. OBJECTIVE: This study evaluated the knowledge, attitude, and practice of sitting posture and computer ergonomics and study-related musculoskeletal problems in undergraduates who followed online education during the pandemic. METHODS: A cross-sectional online survey among a cohort of Sri Lankan medical undergraduates was conducted using a structured questionnaire with 56 multiple-choice or Yes/No type questions. RESULTS: Of the 410 participants, over 85% (n = 362) knew the correct posture to sit on the chair type that they frequently used for studies. However, the majority (n = 378,92.20%) practised incorrect sitting postures in which leaning forward (n = 319,77.80%) was the most common suboptimal posture. Knowledge (n = 161,40%) and practice (n = 167,40.73%) on taking frequent breaks were poor among the majority. Their knowledge on computer ergonomics was good (>80%, n = 304) except for the recommended eye-to-screen distance (n = 129,31.46%). Importantly, ∼50% (n = 206) did not practise the recommended eye-to-screen distance. Use of non-adjustable chairs with no armrests (n = 346,84.39%) and smartphones (n = 354,86.34%) were identified as the main factors which hindered correct practices. Study-related pain/discomfort reported by the majority (n = 241,58.78%) is potentially due to suboptimal ergonomics. Their attitude toward learning and practicing correct ergonomics in home workstations was good (n = 383,93.41%). CONCLUSION: Poor practice of posture and computer ergonomics, despite the good knowledge and attitude is possibly due to the suboptimal work environments. Introducing simple practical measures to facilitate ergonomically appropriate work environments is mandatory in virtual education to prevent study-related musculoskeletal problems.

Clinico-epidemiology and aetiopathogenesis of gallstone disease in the South Asian region: a scoping review protocol.

INTRODUCTION: Pathogenesis of gallstones (GS) is multifactorial and is influenced by numerous environmental and genetic risk factors. As a result, clinico-epidemiology and aetiopathogenesis of GS vary in different populations. Understanding the aetiopathogenesis of GS for different populations is imperative in control and prevention of GS disease and its associated complications. This protocol describes the methodology of a scoping review which focuses on synthesising the most updated knowledge on GS disease in South Asia. METHODS AND ANALYSIS: The scoping review proposed in this protocol will be guided by Arksey and O'Malley's framework and the Joanna Briggs Institute Reviewers' Manual. Accordingly, population, concept and context strategy will be used to formulate the scoping review question, eligibility criteria and search strategy. In the search, electronic databases, MEDLINE/PubMed, ScienceDirect, Scopus, Cochrane library, CINAHL, Trip, and Google scholar, as well as various grey literature sources will be used in synthesising and presenting the findings on clinico-epidemiology and aetiopathogenesis of GS disease in South Asia. ETHICS AND DISSEMINATION: As secondary data will be used in the study, ethical approval will not be required. The scoping review proposed by this protocol will accurately summarise the current knowledge on GS disease in South Asia based on published and unpublished literature on the field. Thus, the evidence presented in the review will be important for healthcare providers to make decisions on the control and prevention of GS disease and as well as to identify future research priorities on GS disease in South Asia.

A high-resolution mass spectrometry based proteomic dataset of human regulatory T cells.

Regulatory T cells (Tregs) play a core role in maintaining immune tolerance, homeostasis, and host health. High-resolution analysis of the Treg proteome is required to identify enriched biological processes and pathways distinct to this important immune cell lineage. We present a comprehensive proteomic dataset of Tregs paired with conventional CD4+ (Conv CD4+) T cells in healthy individuals. Tregs and Conv CD4+ T cells were sorted to high purity using dual magnetic bead-based and flow cytometry-based methodologies. Proteins were trypsin-digested and analysed using label-free data-dependent acquisition mass spectrometry (DDA-MS) followed by label free quantitation (LFQ) proteomics analysis using MaxQuant software. Approximately 4,000 T cell proteins were identified with a 1% false discovery rate, of which approximately 2,800 proteins were consistently identified and quantified in all the samples. Finally, flow cytometry with a monoclonal antibody was used to validate the elevated abundance of the protein phosphatase CD148 in Tregs. This proteomic dataset serves as a reference point for future mechanistic and clinical T cell immunology and identifies receptors, processes, and pathways distinct to Tregs. Collectively, these data will lead to a better understanding of Treg immunophysiology and potentially reveal novel leads for therapeutics seeking Treg regulation.

Expression of CD49f defines subsets of human regulatory T cells with divergent transcriptional landscape and function that correlate with ulcerative colitis disease activity.

OBJECTIVE: Adoptive regulatory T cell (Treg) therapy is being trialled for the treatment of different autoimmune disorders, including inflammatory bowel diseases (IBD). In-depth understanding of the biological variability of Treg in the human blood may be required to improve IBD immune monitoring and treatment strategies. METHODS: Through a combination of quantitative proteomic, multiparametric flow cytometry, RNA-sequencing data analysis and functional assays on Treg enriched from the blood of ulcerative colitis (UC) patients and healthy controls, we investigated the association between CD49f expression, Treg phenotype and function, and UC disease activity. RESULTS: High-dimensional analysis and filtering defined two distinct subsets of human Treg based on the presence or absence of CD49f with divergent transcriptional landscape and functional activities. CD49f negative (CD49f-) Treg are enriched for functional Treg markers and present significantly increased suppressive capacity. In contrast, CD49fhigh Treg display a pro-inflammatory Th17-like phenotype and accumulate in the blood of patients with UC. Dysregulation on CD49f Treg subsets in patients with UC correlate with disease activity. CONCLUSION: Overall, our findings uncover the importance of CD49f expression on Treg in physiological immunity and in pathological autoimmunity.

A primary human T-cell spectral library to facilitate large scale quantitative T-cell proteomics.

Data independent analysis (DIA) exemplified by sequential window acquisition of all theoretical mass spectra (SWATH-MS) provides robust quantitative proteomics data, but the lack of a public primary human T-cell spectral library is a current resource gap. Here, we report the generation of a high-quality spectral library containing data for 4,833 distinct proteins from human T-cells across genetically unrelated donors, covering ~24% proteins of the UniProt/SwissProt reviewed human proteome. SWATH-MS analysis of 18 primary T-cell samples using the new human T-cell spectral library reliably identified and quantified 2,850 proteins at 1% false discovery rate (FDR). In comparison, the larger Pan-human spectral library identified and quantified 2,794 T-cell proteins in the same dataset. As the libraries identified an overlapping set of proteins, combining the two libraries resulted in quantification of 4,078 human T-cell proteins. Collectively, this large data archive will be a useful public resource for human T-cell proteomic studies. The human T-cell library is available at SWATHAtlas and the data are available via ProteomeXchange (PXD019446 and PXD019542) and PeptideAtlas (PASS01587).

Chemical characterization of gallstones: an approach to explore the aetiopathogenesis of gallstone disease in Sri Lanka.

INTRODUCTION: Records on gallstones and associated ailments in Sri Lankan community are scarce, despite frequent detection of gallstone disease. Identification of the chemical composition of gallstones in the local setting is important in defining aetiopathogenic factors which in turn are useful in implementing therapeutic and preventive strategies. This study aimed to describe the chemical composition of gallstones and the socio-demographic factors of a cohort of Sri Lankan patients with gallstone disease. MATERIALS AND METHODS: Data on clinical and socio-demographic factors, and gallstones removed at surgery were collected from patients with cholelithiasis admitted to Teaching Hospital, Peradeniya, Sri Lanka from May 2011 to December 2012. External and cross sectional morphological features of gallstones were recorded by naked eye observation. Compositional analysis was carried out by Fourier Transform Infrared Spectroscopy, X - ray Powder Diffraction, and Atomic Absorption Spectrophotometry. Scanning Electron Microscopy was used to identify the microstructure of gallstones. RESULTS: Data of 102 patients were analyzed. Of them majority (n = 77, 76%) were females with a female: male ratio of 3:1. Mean age of the study group was 46.1±11.6 years. All the patients had primary gallbladder stones. According to the physical and chemical analysis, majority (n = 54, 53%) were pigment gallstones followed by mixed cholesterol gallstones (n = 38, 37%). Only 10 (9%) had pure cholesterol gallstones. Calcium bilirubinate, calcium carbonate and calcium phosphate were the commonest calcium salts identified in pigment gallstones and core of mixed cholesterol gallstones. CONCLUSION: Presence of a pigment nidus in gallstones is a common feature in majority of Sri Lankan patients denoting the possible role of elevated unconjugated bilirubin in bile on the pathogenesis of GS. Hence it is imperative to explore this further to understand the aetiopathogenesis of GS among Sri Lankans.

Serum lipid concentrations in patients with cholesterol and pigment gallstones.

BACKGROUND: Gallstones (GS) are formed as a result of impaired metabolic regulation of the human body. Abnormal lipid metabolism is partly responsible for the pathogenesis of GS mainly rich in cholesterol. Thus abnormalities of serum lipids would reflect the possibilities of the formation of cholesterol GS. This study aims to identify the significance of serum lipids on the development of GS disease. METHODS: Serum lipid profiles were estimated in 73 patients with symptomatic GS, admitted to the Teaching Hospital, Peradeniya, Sri Lanka for GS removal surgeries from May 2011 to December 2012. Patients with normal serum bilirubin level and not being on lipid lowering drugs were recruited for the study. Serum lipid profile of each patient was analyzed by enzymatic kit assays and the chemical composition of GS was analyzed by Fourier Transform Infrared spectroscopy. RESULTS: Of the 73 patients, 37 (51%) had cholesterol GS while 36 (49%) had pigment GS. Serum lipid parameters of a majority of patients were within the normal range. Body mass index values of the patients with two types of GS were not significantly different (Two sample t test, p = 0.335). Out of the lipid parameters tested, only serum triglyceride concentration was significantly high in patients with cholesterol GS than that of pigment GS (Two sample t test, p = 0.038). None of the lipid parameters were significantly different between males and females (Two sample t test, p > 0.05). Compared to the patients with pigment GS who were aged below 50 years, mean total cholesterol and triglyceride concentrations were higher in the same age category patients with cholesterol GS. CONCLUSION: Abnormal serum lipid profiles doesn't seem to be an essential feature in patients with cholesterol GS. However when the two groups of patients with cholesterol and pigment GS with no significant difference of body mass indexes were compared, patients with cholesterol GS are more likely to have serum lipid parameters towards the undesirable cutoff levels of their respective normal ranges. However an effect of serum lipid concentrations on high incidence of GS among females has not been identified.

Can the type of gallstones be predicted with known possible risk factors?: A comparison between mixed cholesterol and black pigment stones.

BACKGROUND: Pathogenesis of gallstones (GS) is multifactorial and multiple genetic and environmental factors have been identified in different populations for different types of GS with varying prevalence. However the role of the each aetiological factor on the formation of mixed cholesterol and black pigment GS has not being addressed adequately. Hence in this study we attempted to compare known possible risk factors for mixed cholesterol and black pigment GS among two groups of patients with two types of GS. METHODS: The study was done on a cohort of patients with symptomatic GS admitted to the Teaching Hospital Peradeniya, Sri Lanka over a period of 18 months. Clinical and epidemiological data and physical parameters of the patients were recorded and surgically removed GS were analyzed chemically and physically to identify the type of GS. In addition lipid profile was done in all the patients with normal serum bilirubin levels. RESULTS: A total of 86 patients were included in the study. Mixed cholesterol GS was significantly common among females than males (χ2 test, p = 0.029). Mixed cholesterol GS was commonly seen among patients belonging to Moor ethnicity (χ2 test, p = 0.009). Majority of patients with mixed cholesterol GS had body mass index above 25 kg/m2 (χ2 test, p = 0.018). Black pigment GS were significantly common among patients with type II diabetes mellitus (Fisher's exact test, p = 0.035). Further all the patients with chronic haemolytic anaemia and alcoholic cirrhosis had black pigment GS. Age, family history, Fasting Blood Glucose, dyslipidaemia, lipid profile, parity and use of oral contraceptive pills in females, smoking and alcohol intake in males did not differ significantly among patients in the two groups. CONCLUSION: Gender, ethnicity and body mass index can be used to predict the formation of mixed cholesterol GS and black pigment GS.