Cyclic immonium ion of lactyllysine reveals widespread lactylation in the human proteome.

Lactylation was initially discovered on human histones. Given its nascence, its occurrence on nonhistone proteins and downstream functional consequences remain elusive. Here we report a cyclic immonium ion of lactyllysine formed during tandem mass spectrometry that enables confident protein lactylation assignment. We validated the sensitivity and specificity of this ion for lactylation through affinity-enriched lactylproteome analysis and large-scale informatic assessment of nonlactylated spectral libraries. With this diagnostic ion-based strategy, we confidently determined new lactylation, unveiling a wide landscape beyond histones from not only the enriched lactylproteome but also existing unenriched human proteome resources. Specifically, by mining the public human Meltome Atlas, we found that lactylation is common on glycolytic enzymes and conserved on ALDOA. We also discovered prevalent lactylation on DHRS7 in the draft of the human tissue proteome. We partially demonstrated the functional importance of lactylation: site-specific engineering of lactylation into ALDOA caused enzyme inhibition, suggesting a lactylation-dependent feedback loop in glycolysis.

Chemoproteomic mapping of the glycolytic targetome in cancer cells.

Hyperactivated glycolysis is a metabolic hallmark of most cancer cells. Although sporadic information has revealed that glycolytic metabolites possess nonmetabolic functions as signaling molecules, how these metabolites interact with and functionally regulate their binding targets remains largely elusive. Here, we introduce a target-responsive accessibility profiling (TRAP) approach that measures changes in ligand binding-induced accessibility for target identification by globally labeling reactive proteinaceous lysines. With TRAP, we mapped 913 responsive target candidates and 2,487 interactions for 10 major glycolytic metabolites in a model cancer cell line. The wide targetome depicted by TRAP unveils diverse regulatory modalities of glycolytic metabolites, and these modalities involve direct perturbation of enzymes in carbohydrate metabolism, intervention of an orphan transcriptional protein's activity and modulation of targetome-level acetylation. These results further our knowledge of how glycolysis orchestrates signaling pathways in cancer cells to support their survival, and inspire exploitation of the glycolytic targetome for cancer therapy.

Detection of Interleukin-1 ß (IL-1ß) in Single Human Blastocyst-Conditioned Medium Using Ultrasensitive Bead-Based Digital Microfluidic Chip and Its Relationship with Embryonic Implantation Potential.

The implantation of human embryos is a complex process involving various cytokines and receptors expressed by both endometrium and embryos. However, the role of cytokines produced by a single embryo in successful implantation is largely unknown. This study aimed to investigate the role of IL-1ß expressed in a single-embryo-conditioned medium (ECM) in embryo implantation. Seventy samples of single ECM were analyzed by a specially designed magnetic-beads-based microfluidic chip from 15 women. We discovered that IL-1ß level increased as the embryo developed, and the difference was significant. In addition, receiver operator characteristic (ROC) curves analysis showed a higher chance of pregnancy when the IL-1ß level on day 5 ECM was below 79.37 pg/mL and the difference between day 5 and day 3 was below 24.90 pg/mL. Our study discovered a possible association between embryonic proteomic expression and successful implantation, which might facilitate single-embryo transfer in the future by helping clinicians identify the embryo with the greatest implantation potential.

Targeting YAP1 ameliorates progesterone resistance in endometriosis.

STUDY QUESTION: Does YAP1 inhibition alleviate progesterone resistance in endometriosis? SUMMARY ANSWER: YAP1 inhibition reduces progesterone resistance in vitro and in vivo. WHAT IS KNOWN ALREADY: Progesterone resistance not only causes treatment failure for endometriosis but also inhibits eutopic endometrial cell proliferation, dysregulates decidualization, and reduces the success rates of pregnancy. Hippo/yes-associated protein 1 (YAP1) signaling pathway plays an important role in the pathogenesis of endometriosis. STUDY DESIGN, SIZE, DURATION: Paraffin-embedded tissues containing paired endometriotic and endometrial specimens (n = 42) and serum samples isolated from normal controls (n = 15) or endometriotic patients with (n = 25) or without (n = 21) prior dienogest treatment were analyzed. A mouse model of endometriosis was also used to evaluate the effects of YAP1 inhibition on progesterone resistance. PARTICIPANTS/MATERIALS, SETTING, METHODS: Primary endometriotic and endometrial stromal cells treated with YAP1 inhibitor or miR-21 mimic/inhibitor were used for the in vitro studies including decidualization induction, chromatin immunoprecipitation (ChIP), and RNA immunoprecipitation. Tissue specimens and serum from human and mouse were used for immunohistochemistry staining, exosome isolation, and microRNA (miRNA) quantification, respectively. MAIN RESULTS AND THE ROLE OF CHANCE: Herein, we report, by using ChIP-PCR and RNA-IP, that YAP1 inhibits progesterone receptor (PGR) expression through upregulation of miR-21-5p. Upregulation of miR-21-5p not only reduces PGR expression but also inhibits endometrial stromal cell decidualization. Indeed, levels of YAP1 and miR-21-5p are inversely correlated with the level of PGR in human endometrial samples. In contrast, knockdown of YAP1 or treatment with verteporfin (VP), a YAP1 inhibitor, reduces miR-21-5p expression, thus leading to an increase in PGR expression in ectopic endometriotic stromal cells. In the mouse model of endometriosis, treatment with VP increases PGR expression and enhances decidualization. More importantly, VP synergistically increases the treatment effect of progestin in causing the regression of endometriotic lesions and improves the decidualization capability of the endometrium. Interestingly, treatment with dienogest, a synthetic progestin, reduces YAP1 and miR-21-5p expression in human cells and in the mouse model of endometriosis. Patients who received dienogest treatment for 6 months show a significant decrease in serum extracellular vesicle-associated miR-21-5p level. LARGE SCALE DATA: A public dataset (GSE51981) containing a large cohort of endometriotic tissues is available from the Gene Expression Omnibus (GEO). LIMITATIONS, REASONS FOR CAUTION: A large cohort of clinical samples is needed to verify the current diagnostic value of miR-21-5p in future studies. WIDER IMPLICATIONS OF THE FINDINGS: The reciprocal regulation of YAP1 and PGR suggests that combined YAP1 inhibitor and progestin may be a better therapeutic approach for treating endometriosis. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by the Ministry of Science and Technology, Taiwan (MOST-111-2636-B-006-012, MOST-111-2314-B-006-075-MY3, and MOST-106-2320-B-006-072-MY3). The authors have no conflict of interest to disclose.

From stress to responses: aluminium-induced signalling in the root apex.

Aluminium (Al) toxicity is one of the major constraints for crop growth and productivity in most of the acid soils worldwide. The primary lesion of Al toxicity is the rapid inhibition of root elongation. The root apex, especially the transition zone (TZ), has been identified as the major site of Al accumulation and injury. The signalling, in particular through phytohormones in the root apex TZ in response to Al stress, has been reported to play crucial roles in the regulation of Al-induced root growth inhibition. The binding of Al in the root apoplast is the initial event leading to inhibition of root elongation. Much progress has been made during recent years in understanding the molecular functions of cell wall modification and Al resistance-related genes in Al resistance or toxicity, and several signals including phytohormones, Ca2+, etc. have been reported to be involved in these processes. Here we summarize the recent advances in the understanding of Al-induced signalling and regulatory networks in the root apex involved in the regulation of Al-induced inhibition of root growth and Al toxicity/resistance. This knowledge provides novel insights into how Al-induced signals are recognized by root apical cells, transmitted from the apoplast to symplast, and finally initiate the defence system against Al. We conclude that the apoplast plays a decisive role in sensing and transmitting the Al-induced signals into the symplast, further stimulating a series of cellular responses (e.g. exudation of organic acid anions from roots) to adapt to the stress. We expect to stimulate new research by focusing on the signalling events in the root apex in response to Al stress, particularly taking into consideration the signal transduction between the meristem zone, TZ, and elongation zone and the apoplast and symplast.

Extracellular vesicle-associated VEGF-C promotes lymphangiogenesis and immune cells infiltration in endometriosis.

Endometriosis is a highly prevalent gynecological disease with severe negative impacts on life quality and financial burden. Unfortunately, there is no cure for this disease, which highlights the need for further investigation about the pathophysiology of this disease to provide clues for developing novel therapeutic regimens. Herein, we identified that vascular endothelial growth factor (VEGF)-C, a potent lymphangiogenic factor, is up-regulated in endometriotic cells and contributes to increased lymphangiogenesis. Bioinformatic analysis and molecular biological characterization revealed that VEGF-C is negatively regulated by an orphan nuclear receptor, chicken ovalbumin upstream promoter-transcription factor II (COUP-TFII). Further studies demonstrated that proinflammatory cytokines, via suppression of COUP-TFII level, induce VEGF-C overexpression. More importantly, we show that functional VEGF-C is transported by extracellular vesicles (EVs) to enhance the lymphangiogenic ability of lymphatic endothelial cells. Autotransplanted mouse model of endometriosis showed lenvatinib treatment abrogated the increased lymphatic vessels development in the endometriotic lesion, enlarged retroperitoneal lymph nodes, and immune cells infiltration, indicating that blocking VEGF-C signaling can reduce local chronic inflammation and concomitantly endometriosis development. Evaluation of EV-transmitted VEGF-C from patients' sera demonstrates it is a reliable noninvasive way for clinical diagnosis. Taken together, we identify the vicious cycle of inflammation, COUP-TFII, VEGF-C, and lymphangiogenesis in the endometriotic microenvironment, which opens up new horizons in understanding the pathophysiology of endometriosis. VEGF-C not only can serve as a diagnostic biomarker but also a molecular target for developing therapeutic regimens.

Assessing the effects of a mentoring program on professional identity formation.

BACKGROUND: Medical education has enjoyed mixed fortunes nurturing professional identity formation (PIF), or how medical students think, feel and act as physicians. New data suggests that structured mentoring programs like the Palliative Medicine Initiative (PMI) may offer a means of developing PIF in a consistent manner. To better understand how a well-established structured research mentoring program shapes PIF, a study of the experiences of PMI mentees is proposed. METHODOLOGY: Acknowledging PIF as a sociocultural construct, a Constructivist approach and Relativist lens were adopted for this study. In the absence of an effective tool, the Ring Theory of Personhood (RToP) and Krishna-Pisupati Model (KPM) model were used to direct this dual Systematic Evidence-Based Approach (Dual-SEBA) study in designing, employing and analysing semi-structured interviews with PMI mentees and mentoring diaries. These served to capture changes in PIF over the course of the PMI's mentoring stages. Transcripts of the interviews and mentoring diaries were concurrently analysed using content and thematic analysis. Complementary themes and categories identified from the Split Approach were combined using the Jigsaw Approach and subsequently compared with mentoring diaries in the Funnelling Process. The domains created framed the discussion. RESULTS: A total of 12 mentee interviews and 17 mentoring diaries were analysed, revealing two domains-PMI as a Community of Practice (CoP) and Identity Formation. The domains confirmed the centrality of a structured CoP capable of facilitating longitudinal mentoring support and supporting the Socialisation Process along the mentoring trajectory whilst cultivating personalised and enduring mentoring relationships. CONCLUSION: The provision of a consistent mentoring approach and personalised, longitudinal mentoring support guided along the mentoring trajectory by structured mentoring assessments lay the foundations for more effective mentoring programs. The onus must now be on developing assessment tools, such as a KPM-based tool, to guide support and oversight of mentoring relationships.

Professional identity formation amongst peer-mentors in a research-based mentoring programme.

BACKGROUND: Mentoring plays a pivotal yet poorly understood role in shaping a physician's professional identity formation (PIF) or how they see, feel and act as professionals. New theories posit that mentoring nurtures PIF by functioning as a community of practice through its structured approach and its support of a socialisation process made possible by its assessment-directed personalized support. To test this theory and reshape the design, employ and support of mentoring programs, we evaluate peer-mentor experiences within the Palliative Medicine Initiative's structured research mentoring program. METHODS: Semi-structured interviews with peer mentors under the Palliative Medicine Initiative (PMI) at National Cancer Centre Singapore were conducted and triangulated against mentoring diaries to capture longitudinal data of their PMI experiences. The Systematic Evidence-Based Approach (SEBA) was adopted to enhance the trustworthiness of the data. SEBA employed concurrent content and thematic analysis of the data to ensure a comprehensive review. The Jigsaw Perspective merged complementary themes and categories identified to create themes/categories. The themes/categories were compared with prevailing studies on mentoring in the Funnelling Process to reaffirm their accuracy. RESULTS: Twelve peer-mentors participated in the interviews and eight peer-mentors completed the mentoring diaries. The domains identified were community of practice and identity work. CONCLUSIONS: The PMI's structured mentoring program functions as a community of practice supporting the socialisation process which shapes the peer-mentor's belief system. Guided by a structured mentoring approach, stage-based assessments, and longitudinal mentoring and peer support, peer-mentors enhance their detection and evaluation of threats to their regnant belief system and adapt their self-concepts of identity and personhood to suit their context. These insights will help structure and support mentoring programs as they nurture PIF beyond Palliative Medicine.

Biomimetic Guided Bi2WO6/Bi2O3 Vertical Heterojunction with Controllable Microstructure for Efficient Photocatalysis.

To bridge the technical gap of heterojunction induction control in conventional semiconductor photocatalysts, a method of regulating the growth of heterojunctions utilizing biomimetic structures was designed to prepare a series of Bi2WO6/Bi2O3 vertical heterojunction nanocomposites for the disposal of environmentally hazardous tetracycline wastewater difficult to degrade by conventional microbial techniques. Porous Bi2O3 precursors with high-energy crystalline (110) dominant growth were produced using the sunflower straw bio-template technique (SSBT). Bi2WO6 with a (131) plane grew preferentially into 2.8 to 4 nm pieces on the (110) plane of Bi2O3, causing a significant density reduction between Bi2WO6 pieces and a dimensional decrease in the agglomerated Bi2WO6 spheres from 3 µm to 700 nm since Bi2WO6 grew on the structure of the biomimetic Bi2O3. The optimal 1:8 Bi2WO6/Bi2O3 coupling catalyst was obtained via adapting the ratio of the two semiconductors, and the coupling ratio of 1:8 minimized the adverse effects of the overgrowth of Bi2WO6 on degradation performance by securing the quantity of vertical heterojunctions. The material degradation reaction energy barrier and bandgap were significantly reduced by the presence of a large number of vertical heterojunction structures, resulting in a material with lower impedance and higher electron-hole separation efficiency; thus, the degradation efficiency of 1:8 Bi2WO6/Bi2O3 for tetracycline hydrochloride reached 99% within 60 min. In conclusion, this study not only successfully synthesized a novel photocatalyst with potential applications in water pollution remediation but also introduced a pioneering approach for semiconductor-driven synthesis.

Living Cell-Target Responsive Accessibility Profiling Reveals Silibinin Targeting ACSL4 for Combating Ferroptosis.

We report a living cell-target responsive accessibility profiling (LC-TRAP) approach to identify the targetome of silibinin (SIL), a well-established hepatoprotective natural product (NP), in HepG2 cells. Proteins showing accessibility changes, probed by covalent lysine labeling reagents and leveraged by multiplexed quantitative proteomics, following the administration of SIL to the living cells were assigned as potential targets. Among the assigned targetome, ACSL4, an enzyme essential for ferroptosis induction, might be involved in the hepatoprotective effects of SIL and hence was intensively validated. We first demonstrated that SIL protected HepG2 cells from ferroptosis dependent on ACSL4. Then, we used biophysical assays and a SIL-derivatized chemical probe to corroborate that SIL can bind to ACSL4. The ensuing enzymatic assays showed that SIL inhibited ACSL4 enzymatic activity, thereby mitigating the ACSL4-mediated ferroptosis. As such, we revealed that ACSL4 inhibition, using SIL as a model compound, represents a promising hepatoprotective strategy. Further, since TRAP probes the accessibility changes of reactive proteinaceous lysines, it can pinpoint the proximal regions where the ligand engagement may occur. Thus, the LC-TRAP analysis of SIL, the newly discovered ligand of ACSL4, and arachidonic acid (AA), the substrate, intriguingly showed that SIL and AA both affected the conformation of the K536-proximal region of ACSL4, albeit through distinct binding patterns. Collectively, we describe a straightforward LC-TRAP workflow that does not involve ligand-derived probe synthesis and is widely applicable to target discovery of NPs.

A systematic scoping review moral distress amongst medical students.

BACKGROUND: Characterised by feelings of helplessness in the face of clinical, organization and societal demands, medical students are especially prone to moral distress (MD). Despite risks of disillusionment and burnout, efforts to support them have been limited by a dearth of data and understanding of MD in medical students. Yet, new data on how healthcare professionals confront difficult care situations suggest that MD could be better understood through the lens of the Ring Theory of Personhood (RToP). A systematic scoping review (SSR) guided by the RToP is proposed to evaluate the present understanding of MD amongst medical students. METHODS: The Systematic Evidence-Based Approach (SEBA) is adopted to map prevailing accounts of MD in medical students. To enhance the transparency and reproducibility, the SEBA methodology employs a structured search approach, concurrent and independent thematic analysis and directed content analysis (Split Approach), the Jigsaw Perspective that combines complementary themes and categories, and the Funnelling Process that compares the results of the Jigsaw Perspective with tabulated summaries to ensure the accountability of these findings. The domains created guide the discussion. RESULTS: Two thousand six hundred seventy-one abstracts were identified from eight databases, 316 articles were reviewed, and 20 articles were included. The four domains identified include definitions, sources, recognition and, interventions for MD. CONCLUSIONS: MD in medical students may be explained as conflicts between the values, duties, and principles contained within the different aspects of their identity. These conflicts which are characterised as disharmony (within) and dyssynchrony (between) the rings of RToP underline the need for personalised and longitudinal evaluations and support of medical students throughout their training. This longitudinal oversight and support should be supported by the host organization that must also ensure access to trained faculty, a nurturing and safe environment for medical students to facilitate speak-up culture, anonymous reporting, feedback opportunities and supplementing positive role modelling and mentoring within the training program.

The Future of Targeted Treatment of Primary Sjögren's Syndrome: A Focus on Extra-Glandular Pathology.

Primary Sjögren's syndrome (pSS) is a chronic, systemic autoimmune disease defined by exocrine gland hypofunction resulting in dry eyes and dry mouth. Despite increasing interest in biological therapies for pSS, achieving FDA-approval has been challenging due to numerous complications in the trials. The current literature lacks insight into a molecular-target-based approach to the development of biological therapies. This review focuses on novel research in newly defined drug targets and the latest clinical trials for pSS treatment. A literature search was conducted on ClinicalTrials.gov using the search term "Primary Sjögren's syndrome". Articles published in English between 2000 and 2021 were included. Our findings revealed potential targets for pSS treatment in clinical trials and the most recent advances in understanding the molecular mechanisms underlying the pathogenesis of pSS. A prominent gap in current trials is in overlooking the treatment of extraglandular symptoms such as fatigue, depression, and anxiety, which are present in most patients with pSS. Based on dryness and these symptom-directed therapies, emerging biological agents targeting inflammatory cytokines, signal pathways, and immune reaction have been studied and their efficacy and safety have been proven. Novel therapies may complement existing non-pharmacological methods of alleviating symptoms of pSS. Better grading systems that add extraglandular symptoms to gauge disease activity and severity should be created. The future of pSS therapies may lie in gene, stem-cell, and tissue-engineering therapies.

China's energy stock market jumps: To what extent does the COVID-19 pandemic play a part?

The price jump behavior may bring tremendous challenges on risk management and asset pricing. This paper uses the BN-S test, the wavelet coherence method, and applies high-frequency data to explore whether and to what extent the COVID-19 pandemic impacts China's energy stock market jumps and its characteristics. The empirical results uncover the significant and heterogeneous interactions between the COVID-19 pandemic and China's energy stock market jumps across market specifications, investment horizons, and China/global pandemic tolls at different time scales. First, the oil stock market jumps were the most correlated with the pandemic, especially during the peak and re-deterioration phases. The pandemic played a positive and leading role in the short term (1-4 days length period) and long term (over 32 days length period). Second, the coal stock market jumps have similar characteristics to those of oil, but mainly show a negative correlation with the pandemic. Third, renewable energy stock market jumps were the least correlated, mainly showing a positive correlation in the short term and a negative correlation in the long term. In addition, the interaction characteristics of systemic co-jumps in different China's energy stock markets are also significant.

HYPOXIA AND REPRODUCTIVE HEALTH: The role of hypoxia in the development and progression of endometriosis.

Endometriosis is a benign gynecological disease that affects about 10% of women of reproductive age. Patients with endometriosis suffer from long-term coexistence with dysmenorrhea, dyspareunia, and even infertility, which severely reduces quality of life. So far, surgical removal and hormonal medication are the major treatment options; however, high recurrence and severe adverse effects hamper the therapeutic efficacy. Hypoxia is an inevitable cellular stress in many diseases that regulates the expression of a significant subset of genes involved in pathophysiological processes. A growing body of evidence demonstrates that hypoxia plays critical role in controlling the disease phenotypes of endometriosis, such as increasing adhesion ability, causing dysregulation of estrogen biosynthesis, aberrant production of proinflammatory cytokines, increasing angiogenic ability, and suppression of immune functions. In this review, we summarize the findings of the most recent studies in exploring the underlying mechanisms of hypoxia involved in endometriosis. Potential therapeutic options for targeting HIF and downstream effectors will also be discussed.

[Analysis of prescription regularity of traditional Chinese medicine for colorectal cancer based on data mining].

The tumor prescriptions contained in Dictionary of Tumor Formulas, Compendium of Good Tumor Formulas, Chinese Pharmacopoeia, Ministry of Health Drug Standards for Chinese Medicine Formulas and National Compilation of Standards for Proprietary Chinese Medicines were selected and organized to construct a database for tumor prescriptions, and the data mining techniques were applied to investigate the prescription regularity of colorectal cancer prescriptions. The formula data were extracted after screening in strict accordance with the inclusion and exclusion criteria, and were then analyzed with Microsoft Excel 2010 for frequency statistics, Apriori block provided by SPSS Clementine 12.0 software for correlation rule analysis, and arules and arulesViz packages in R 4.0.2 software for correlation rule visualization. In addition, SPSS 18.0 software was used for cluster analysis and factor analysis, in which cluster analysis was performed by Ochiai algorithm with bicategorical variables in systematic clustering method and factor analysis was performed mainly with principal component analysis. A total of 285 prescriptions were included in the statistical analysis, and the frequency statistics showed that 43 herbs had been used more than 16 times. The association rules analysis showed that 26 high-frequency me-dicine pair rules were obtained, and the association rules for those dispelling evil spirits, strengthening the body, resolving stasis, dispelling dampness, etc. were visualized. In the cluster analysis, we generated a dendrogram from which 7 groups of traditional Chinese medicines with homogeneity were extracted. 10 common factors were obtained in the factor analysis. The types of herbal medicines involved in the colorectal cancer prescription included anti-cancer antidotes, strengthening and tonifying medicines, blood-regulating medicines, and expectorant medicines, corresponding to the treatment for eliminating evil spirits, strengthening, resolving stasis, and expectorating dampness. The prescriptions for anti-cancer detoxification were normally based on the pairs composed of Scutellaria barbata-Hedyotis diffusa and Sophora flavescens, Sargentodoxa cuneata, S. barbata, often combined with stasis relieving drug and dampness eliminating drug, reflecting the characteristics of treatment for both toxicity and stasis, dampness and toxicity simultaneously. The prescriptions for strengthening the righteousness and tonifying the deficiency were composed of Astragalus membranaceus and Atractylodes macrocephala mainly, exerting the effect of benefiting Qi, strengthening the spleen and drying dampness, tonifying kidney and essence, tonifying blood and invigorating blood. Meanwhile, anti-cancer detoxification medicines shall be reduced as much as possible. The compatibility of the medicines for the intestinal tract reflected the principle of using the right medicine for the right condition and eliminating evil spirits or strengthening the body, as appropriate.

Responding to the COVID-19 Outbreak in Singapore: Staff Protection and Staff Temperature and Sickness Surveillance Systems.

BACKGROUND: Coronavirus disease 2019 (COVID-19) is an emerging infectious disease caused by a novel coronavirus (SARS-CoV-2) and first reported in Wuhan, China, in December 2019. Since the severe acute respiratory syndrome (SARS) outbreak in 2003, Tan Tock Seng Hospital (TTSH) in Singapore has routinely fit-tested staff for high-filtration N95 respirators and established Web-based staff surveillance systems. The routine systems were enhanced in response to Singapore's first imported COVID-19 case on 23 January 2020. METHODS: We conducted a cross-sectional study from 23 January to 23 February 2020 among healthcare workers to evaluate the effectiveness of the staff protection and surveillance strategy in TTSH, a 1600-bed multidisciplinary acute-care hospital colocated with the 330-bed National Centre for Infectious Diseases (NCID). As of 23 February 2020, TTSH/NCID has managed 76% of confirmed COVID-19 cases in Singapore. The hospital adopted a multipronged approach to protect and monitor staff with potential COVID-19 exposures: (1) risk-based personal protective equipment, (2) staff fever and sickness surveillance, and (3) enhanced medical surveillance of unwell staff. RESULTS: A total of 10 583 staff were placed on hospitalwide fever and sickness surveillance, with 1524 frontline staff working in COVID-19 areas under close surveillance. Among frontline staff, a median of 8 staff illness episodes was seen per day; almost 10% (n = 29) resulted in hospitalization. None of the staff was found to be infected with COVID-19. CONCLUSIONS: A robust staff protection and health surveillance system that is routinely implemented during non-outbreak periods and enhanced during the COVID-19 outbreak is effective in protecting frontline staff from the infection.

Subresidue-Resolution Footprinting of Ligand-Protein Interactions by Carbene Chemistry and Ion Mobility-Mass Spectrometry.

The knowledge of ligand-protein interactions is essential for understanding fundamental biological processes and for the rational design of drugs that target such processes. Carbene footprinting efficiently labels proteinaceous residues and has been used with mass spectrometry (MS) to map ligand-protein interactions. Nevertheless, previous footprinting studies are typically performed at the residue level, and therefore, the resolution may not be high enough to couple with conventional crystallography techniques. Herein we developed a subresidue footprinting strategy based on the discovery that carbene labeling produces subresidue peptide isomers and the intensity changes of these isomers in response to ligand binding can be exploited to delineate ligand-protein topography at the subresidue level. The established workflow combines carbene footprinting, extended liquid chromatographic separation, and ion mobility (IM)-MS for efficient separation and identification of subresidue isomers. Analysis of representative subresidue isomers located within the binding cleft of lysozyme and those produced from an amyloid-ß segment have both uncovered structural information heretofore unavailable by residue-level footprinting. Lastly, a "real-world" application shows that the reactivity changes of subresidue isomers at Phe399 can identify the interactive nuances between estrogen-related receptor α, a potential drug target for cancer and metabolic diseases, with its three ligands. These findings have significant implications for drug design. Taken together, we envision the subresidue-level resolution enabled by IM-MS-coupled carbene footprinting can bridge the gap between structural MS and the more-established biophysical tools and ultimately facilitate diverse applications for fundamental research and pharmaceutical development.

Emerging Role of TRIM Family Proteins in Cardiovascular Disease.

Ubiquitination is one of the basic mechanisms of cell protein homeostasis and degradation and is accomplished by 3 enzymes, E1, E2, and E3. Tripartite motif-containing proteins (TRIMs) constitute the largest subfamily of RING E3 ligases, with >70 current members in humans and mice. These members are involved in multiple biological processes, including growth, differentiation, and apoptosis as well as disease and tumorigenesis. Accumulating evidence has shown that many TRIM proteins are associated with various cardiac processes and pathologies, such as heart development, signal transduction, protein degradation, autophagy mediation, ion channel regulation, congenital heart disease, and cardiomyopathies. In this review, we provide an overview of the TRIM family and discuss its involvement in the regulation of cardiac proteostasis and pathophysiology and its potential therapeutic implications.

Integrative Omics Analysis Revealed that Metabolic Intervention Combined with Metronomic Chemotherapy Selectively Kills Cancer Cells.

Metronomic chemotherapy, a relatively new dosing paradigm for anticancer therapy, is an alternative to traditional chemotherapy that uses maximal tolerated dose (MTD). Although these two dosing regimens both lead to tumor cell death, how cell metabolism is differentially affected during apoptosis remains elusive. Herein, we employed metabolomics to monitor the metabolic profiles of MCF-7 cells in response to the two dosing regimens that mimic MTD and MN treatments using a model chemotherapeutic drug, doxorubicin (Dox), and correlated the changes of metabolic genes examined by PCR array to integratively describe the reprogrammed metabolic patterns. We found glycolysis, amino acid, and nucleotide synthesis-associated metabolic pathways were activated in response to the MN treatment, whereas these pathways were inhibited in a pronounced way in response to the MTD treatment. Direct supplementation of key metabolites and pharmacological modulation of targeted metabolic enzymes can both regulate cell fates. Subsequently, we tested the combined use of MN dosing with targeted metabolic intervention using a normal cell line and found the combined treatment hardly affected its apoptotic rate. Our in vitro findings using MCF-7 and MCF-10A cells thus suggest the promising perspective of combining MN dosing of chemotherapeutic agents with metabolic modulation to selectively kill cancer cells rather than normal cells.

Ligand-protein target screening from cell matrices using reactive desorption electrospray ionization-mass spectrometry via a native-denatured exchange approach.

Native mass spectrometry has been recognized as a powerful tool for probing interactions between small molecules, such as drugs and natural products, and target proteins. However, the presence of heterogeneous proteins and metabolites in real biological systems can alter the conformations of target proteins or compete with candidate ligands, thus necessitating a method for measuring binding stoichiometries in matrices aside from the extensively used pure/recombinant protein systems. Furthermore, some small molecule-protein interactions have a transient and low-affinity nature and thus can be mis-assigned as nonspecific binding complexes that are often formed during the native ESI process. A native-denatured exchange (NDX) approach was recently developed using a reactive desorption electrospray ionization-mass spectrometer (DESI-MS) setup to screen specific interacting partners. The method works by gradually increasing the composition of denaturing solvents contained in the DESI spray and thus conferring a switch from a native to denatured ionization environment. This change impairs three-dimensional structures of target proteins and disrupts specific ligand-protein interactions, leading to decreased holo/apo ratios. In contrast, ligand-protein complexes exhibiting different trends are assigned as nonspecific interactions. Herein, we applied the NDX approach to probe specific ligand-protein interactions in biological matrices. We first used mixtures of model ligands and proteins to examine the use of reactive DESI-MS in recognizing ligand-target binding in mixtures. Subsequently, we used the NDX approach to analyze binding affinity curves of ligands to target proteins spiked in cell lysates with the aid of size exclusion chromatography and demonstrated its use in probing specific ligand-protein interactions from cell matrices.