Ptpn6 inhibits caspase-8- and Ripk3/Mlkl-dependent inflammation.

Ptpn6 is a cytoplasmic phosphatase that functions to prevent autoimmune and interleukin-1 (IL-1) receptor-dependent, caspase-1-independent inflammatory disease. Conditional deletion of Ptpn6 in neutrophils (Ptpn6∆PMN) is sufficient to initiate IL-1 receptor-dependent cutaneous inflammatory disease, but the source of IL-1 and the mechanisms behind IL-1 release remain unclear. Here, we investigate the mechanisms controlling IL-1α/ß release from neutrophils by inhibiting caspase-8-dependent apoptosis and Ripk1-Ripk3-Mlkl-regulated necroptosis. Loss of Ripk1 accelerated disease onset, whereas combined deletion of caspase-8 and either Ripk3 or Mlkl strongly protected Ptpn6∆PMN mice. Ptpn6∆PMN neutrophils displayed increased p38 mitogen-activated protein kinase-dependent Ripk1-independent IL-1 and tumor necrosis factor production, and were prone to cell death. Together, these data emphasize dual functions for Ptpn6 in the negative regulation of p38 mitogen-activated protein kinase activation to control tumor necrosis factor and IL-1α/ß expression, and in maintaining Ripk1 function to prevent caspase-8- and Ripk3-Mlkl-dependent cell death and concomitant IL-1α/ß release.

Regulation of antioxidants in cancer.

Scientists in this field often joke, "If you don't have a mechanism, say it's ROS." Seemingly connected to every biological process ever described, reactive oxygen species (ROS) have numerous pleiotropic roles in physiology and disease. In some contexts, ROS act as secondary messengers, controlling a variety of signaling cascades. In other scenarios, they initiate damage to macromolecules. Finally, in their worst form, ROS are deadly to cells and surrounding tissues. A set of molecules with detoxifying abilities, termed antioxidants, is the direct counterpart to ROS. Notably, antioxidants exist in the public domain, touted as a "cure-all" for diseases. Research has disproved many of these claims and, in some cases, shown the opposite. Of all the diseases, cancer stands out in its paradoxical relationship with antioxidants. Although the field has made numerous strides in understanding the roles of antioxidants in cancer, many questions remain.

3D Culture Models with CRISPR Screens Reveal Hyperactive NRF2 as a Prerequisite for Spheroid Formation via Regulation of Proliferation and Ferroptosis.

Cancer-associated mutations that stabilize NRF2, an oxidant defense transcription factor, are predicted to promote tumor development. Here, utilizing 3D cancer spheroid models coupled with CRISPR-Cas9 screens, we investigate the molecular pathogenesis mediated by NRF2 hyperactivation. NRF2 hyperactivation was necessary for proliferation and survival in lung tumor spheroids. Antioxidant treatment rescued survival but not proliferation, suggesting the presence of distinct mechanisms. CRISPR screens revealed that spheroids are differentially dependent on the mammalian target of rapamycin (mTOR) for proliferation and the lipid peroxidase GPX4 for protection from ferroptosis of inner, matrix-deprived cells. Ferroptosis inhibitors blocked death from NRF2 downregulation, demonstrating a critical role of NRF2 in protecting matrix-deprived cells from ferroptosis. Interestingly, proteomics analyses show global enrichment of selenoproteins, including GPX4, by NRF2 downregulation, and targeting NRF2 and GPX4 killed spheroids overall. These results illustrate the value of spheroid culture in revealing environmental or spatial differential dependencies on NRF2 and reveal exploitable vulnerabilities of NRF2-hyperactivated tumors.

Synthetic Lethal and Resistance Interactions with BET Bromodomain Inhibitors in Triple-Negative Breast Cancer.

BET bromodomain inhibitors (BBDIs) are candidate therapeutic agents for triple-negative breast cancer (TNBC) and other cancer types, but inherent and acquired resistance to BBDIs limits their potential clinical use. Using CRISPR and small-molecule inhibitor screens combined with comprehensive molecular profiling of BBDI response and resistance, we identified synthetic lethal interactions with BBDIs and genes that, when deleted, confer resistance. We observed synergy with regulators of cell cycle progression, YAP, AXL, and SRC signaling, and chemotherapeutic agents. We also uncovered functional similarities and differences among BRD2, BRD4, and BRD7. Although deletion of BRD2 enhances sensitivity to BBDIs, BRD7 loss leads to gain of TEAD-YAP chromatin binding and luminal features associated with BBDI resistance. Single-cell RNA-seq, ATAC-seq, and cellular barcoding analysis of BBDI responses in sensitive and resistant cell lines highlight significant heterogeneity among samples and demonstrate that BBDI resistance can be pre-existing or acquired.

Glutathione Primes T Cell Metabolism for Inflammation.

Activated T cells produce reactive oxygen species (ROS), which trigger the antioxidative glutathione (GSH) response necessary to buffer rising ROS and prevent cellular damage. We report that GSH is essential for T cell effector functions through its regulation of metabolic activity. Conditional gene targeting of the catalytic subunit of glutamate cysteine ligase (Gclc) blocked GSH production specifically in murine T cells. Gclc-deficient T cells initially underwent normal activation but could not meet their increased energy and biosynthetic requirements. GSH deficiency compromised the activation of mammalian target of rapamycin-1 (mTOR) and expression of NFAT and Myc transcription factors, abrogating the energy utilization and Myc-dependent metabolic reprogramming that allows activated T cells to switch to glycolysis and glutaminolysis. In vivo, T-cell-specific ablation of murine Gclc prevented autoimmune disease but blocked antiviral defense. The antioxidative GSH pathway thus plays an unexpected role in metabolic integration and reprogramming during inflammatory T cell responses.

Global proteomic assessment of the classical protein-tyrosine phosphatome and "Redoxome".

Protein-tyrosine phosphatases (PTPs), along with protein-tyrosine kinases, play key roles in cellular signaling. All Class I PTPs contain an essential active site cysteinyl residue, which executes a nucleophilic attack on substrate phosphotyrosyl residues. The high reactivity of the catalytic cysteine also predisposes PTPs to oxidation by reactive oxygen species, such as H(2)O(2). Reversible PTP oxidation is emerging as an important cellular regulatory mechanism and might contribute to diseases such as cancer. We exploited these unique features of PTP enzymology to develop proteomic methods, broadly applicable to cell and tissue samples, that enable the comprehensive identification and quantification of expressed classical PTPs (PTPome) and the oxidized subset of the PTPome (oxPTPome). We find that mouse and human cells and tissues, including cancer cells, display distinctive PTPomes and oxPTPomes, revealing additional levels of complexity in the regulation of protein-tyrosine phosphorylation in normal and malignant cells.

Visualizing DNA folding and RNA in embryos at single-cell resolution.

The establishment of cell types during development requires precise interactions between genes and distal regulatory sequences. We have a limited understanding of how these interactions look in three dimensions, vary across cell types in complex tissue, and relate to transcription. Here we describe optical reconstruction of chromatin architecture (ORCA), a method that can trace the DNA path in single cells with nanoscale accuracy and genomic resolution reaching two kilobases. We used ORCA to study a Hox gene cluster in cryosectioned Drosophila embryos and labelled around 30 RNA species in parallel. We identified cell-type-specific physical borders between active and Polycomb-repressed DNA, and unexpected Polycomb-independent borders. Deletion of Polycomb-independent borders led to ectopic enhancer-promoter contacts, aberrant gene expression, and developmental defects. Together, these results illustrate an approach for high-resolution, single-cell DNA domain analysis in vivo, identify domain structures that change with cell identity, and show that border elements contribute to the formation of physical domains in Drosophila.

Impact of simultaneous heart procurement on outcomes of donation after circulatory death lung transplantation.

Donation after circulatory death (DCD) heart procurement is done using either direct procurement (DP) or thoracoabdominal normothermic machine perfusion (TA-NRP). Both approaches could impact lung transplant outcomes with combined heart and lung procurements from the same donor. The impact of such practice on DCD lung transplant remains unstudied. We performed a retrospective analysis using the United Network for Organ Sharing (UNOS) dataset, identifying DCD lung transplants where the donor also donated the heart (cardia lung donor [CD]). A cohort of noncardiac DCD lung donors (noncardiac lung donor [NCD]) from the same era, matched for donor and recipient characteristics, was used as a comparison group. Both immediate and long-term outcomes were examined. A subanalysis was performed comparing the distinct impact of DP or TA-NRP on DCD lung transplant outcomes. Overall graft survival did not significantly differ between CD and NCD. However, recipients in the CD group trended toward a lower P/F ratio at 72 hours (CD vs NCD: 284 vs 3190; P = .054). In the subanalysis, we identified 40 DP donors and 22 TA-NRP donors. We found the both cohorts had lower P/F ratio at 72 hours than the NCD control (P = .04). Overall, 1-year graft survival was equivalent among the TA-NRP, DP, and NCD cohorts.

Evaluation of markers of immunity in different metastatic immune microenvironments suggests more suppression within breast to liver metastases in breast cancer.

PURPOSE: Programmed death receptor ligand-1 (PD-L1) expression and tumor mutational burden (TMB) are approved screening biomarkers for immune checkpoint inhibition (ICI) in advanced triple negative breast cancer. We examined these biomarkers along with characterization of the tumor microenvironment (TME) between breast tumors (BrTs), axillary metastases (AxMs), liver metastases (LvMs), non-axillary lymph node metastases, and non-liver metastases to determine differences related to site of metastatic disease. METHODS: 3076 unpaired biopsies from breast cancer patients were analyzed using whole transcriptome sequencing and NextGen DNA depicting TMB within tumor sites. The PD-L1 positivity was determined with VENTANA PD-L1 (SP142) assay. The immune cell fraction within the TME was calculated by QuantiSeq and MCP-counter. RESULTS: Compared to BrT, more LvM samples had a high TMB (≥ 10 mutations/Mb) and fewer LvM samples had PD-L1+ expression. Evaluation of the TME revealed that LvM sites harbored lower infiltration of adaptive immune cells, such as CD4+, CD8+, and regulatory T-cells compared with the BrT foci. We saw differences in innate immune cell infiltration in LvM compared to BrT, including neutrophils and NK cells. CONCLUSIONS: LvMs are less likely to express PD-L1+ tumor cells but more likely to harbor high TMB as compared to BrTs. Unlike AxMs, LvMs represent a more immunosuppressed TME and demonstrate lower gene expression associated with adaptive immunity compared to BrTs. These findings suggest biopsy site be considered when interpreting results that influence ICI use for treatment and further investigation of immune composition and biomarkers expression by metastatic site.

Reassessing the association: Evaluation of a polyalanine deletion variant of RUNX2 in non-syndromic sagittal and metopic craniosynostosis.

The RUNT-related transcription factor RUNX2 plays a critical role in osteoblast differentiation, and alterations to gene dosage cause distinct craniofacial anomalies. Uniquely amongst the RUNT-related family, vertebrate RUNX2 encodes a polyglutamine/polyalanine repeat (Gln23-Glu-Ala17 in humans), with the length of the polyalanine component completely conserved in great apes. Surprisingly, a frequent 6-amino acid deletion polymorphism, p.(Ala84_Ala89)del, occurs in humans (termed 11A allele), and a previous association study (Cuellar et al. Bone 137:115395;2020) reported that the 11A variant was significantly more frequent in non-syndromic sagittal craniosynostosis (nsSag; allele frequency [AF] = 0.156; 95% confidence interval [CI] 0.126-0.189) compared to non-syndromic metopic craniosynostosis (nsMet; AF = 0.068; 95% CI 0.045-0.098). However, the gnomAD v.2.1.1 control population used by Cuellar et al. did not display Hardy-Weinberg equilibrium, hampering interpretation. To re-examine this association, we genotyped the RUNX2 11A polymorphism in 225 individuals with sporadic nsSag as parent-child trios and 164 singletons with sporadic nsMet, restricting our analysis to individuals of European ancestry. We compared observed allele frequencies to the non-transmitted alleles in the parent-child trios, and to the genome sequencing data from gnomAD v.4, which display Hardy-Weinberg equilibrium. Observed AFs (and 95% CI) were 0.076 (0.053-0.104) in nsSag and 0.082 (0.055-0.118) in nsMet, compared with 0.062 (0.042-0.089) in non-transmitted parental alleles and 0.065 (0.063-0.067) in gnomAD v.4.0.0 non-Finnish European control genomes. In summary, we observed a non-significant excess, compared to gnomAD data, of 11A alleles in both nsSag (relative risk 1.18, 95% CI 0.83-1.67) and nsMet (relative risk 1.29, 95% CI 0.87-1.92), but we did not replicate the much higher excess of RUNX2 11A alleles in nsSag previously reported (p = 0.0001).

What Went Right? A Mixed-Methods Study of Positive Feedback Data in a Hospital-Wide Mortality Review Survey.

BACKGROUND: Toxic work culture contributes to healthcare worker burnout and attrition, but little is known about how healthcare organizations can systematically create and promote a culture of civility and collegiality. OBJECTIVE: To analyze peer-to-peer positive feedback collected as part of a systematized mortality review survey to identify themes and recognition dynamics that can inform positive organizational culture change. DESIGN: Convergent mixed-methods study design. PARTICIPANTS: A total of 388 physicians, 212 registered nurses, 64 advanced practice providers, and 1 respiratory therapist at four non-profit hospitals (2 academic and 2 community). INTERVENTION: Providing optional positive feedback in the mortality review survey. MAIN MEASURES: Key themes and subthemes that emerged from positive feedback data, associations between key themes and positive feedback respondent characteristics, and recognition dynamics between positive feedback respondents and recipients. KEY RESULTS: Approximately 20% of healthcare workers provided positive feedback. Three key themes emerged among responses with free text comments: (1) providing extraordinary patient and family-centered care; (2) demonstrating self-possession and mastery; and (3) exhibiting empathic peer support and effective team collaboration. Compared to other specialties, most positive feedback from medicine (70.2%), neurology (65.2%), hospice and palliative medicine (64.3%), and surgery (58.8%) focused on providing extraordinary patient and family-centered care (p = 0.02), whereas emergency medicine (59.1%) comments predominantly focused on demonstrating self-possession and mastery (p = 0.06). Registered nurses (40.2%) provided multidirectional positive feedback more often than other clinician types in the hospital hierarchy (p < 0.001). CONCLUSIONS: Analysis of positive feedback from a mortality review survey provided meaningful insights into a health system's culture of teamwork and values related to civility and collegiality when providing end-of-life care. Systematic collection and sharing of positive feedback is feasible and has the potential to promote positive culture change and improve healthcare worker well-being.

Subgingival biofilm microbiome in individuals with asthma and periodontitis: Metagenomic analysis.

OBJECTIVE: This observational study aimed to explore the metagenomics of subgingival biofilms in individuals with varying degrees of asthma, from severe to none, to elucidate the association between the subgingival microbiome and asthma. MATERIALS AND METHODS: Subgingival biofilm samples were collected from thirty participants at the Asthma Control Program Outpatient Clinic in Bahia (ProAR). These samples were categorized into six groups based on the severity of asthma and the presence or absence of periodontitis. We employed next-generation sequencing (Illumina MiSeq), targeting the 16S rRNA gene, to characterize the microbial communities present. Our analysis included descriptive statistics and sequencing data, evaluated using multivariate statistical methods such as the Shannon index, principal coordinate analysis, and the Bray-Curtis dissimilarity. RESULTS: Our findings indicate a higher prevalence of periodontally detrimental bacterial genera in individuals with severe asthma and periodontitis. Additionally, individuals with asthma, but without periodontitis, exhibited a tendency toward dysbiosis, particularly in cases of severe asthma. CONCLUSION: This research provides new insights into the composition of the subgingival microbiome in individuals with varying severities of asthma and periodontitis. The genera identified in this study underscore the need for further investigations to build upon these findings.

National Projections for Clinical Implications of Race-Free Creatinine-Based GFR Estimating Equations.

BACKGROUND: The National Kidney Foundation and American Society of Nephrology Task Force on Reassessing the Inclusion of Race in Diagnosing Kidney Disease recently recommended a new race-free creatinine-based equation for eGFR. The effect on recommended clinical care across race and ethnicity groups is unknown. METHODS: We analyzed nationally representative cross-sectional questionnaires and medical examinations from 44,360 participants collected between 2001 and 2018 by the National Health and Nutrition Examination Survey. We quantified the number and proportion of Black, White, Hispanic, and Asian/Other adults with guideline-recommended changes in care. RESULTS: The new equation, if applied nationally, could assign new CKD diagnoses to 434,000 (95% confidence interval [CI], 350,000 to 517,000) Black adults, reclassify 584,000 (95% CI, 508,000 to 667,000) to more advanced stages of CKD, restrict kidney donation eligibility for 246,000 (95% CI, 189,000 to 303,000), expand nephrologist referrals for 41,800 (95% CI, 19,800 to 63,800), and reduce medication dosing for 222,000 (95% CI, 169,000 to 275,000). Among non-Black adults, these changes may undo CKD diagnoses for 5.51 million (95% CI, 4.86 million to 6.16 million), reclassify 4.59 million (95% CI, 4.28 million to 4.92 million) to less advanced stages of CKD, expand kidney donation eligibility for 3.96 million (95% CI, 3.46 million to 4.46 million), reverse nephrologist referral for 75,800 (95% CI, 35,400 to 116,000), and reverse medication dose reductions for 1.47 million (95% CI, 1.22 million to 1.73 million). The racial and ethnic mix of the populations used to develop eGFR equations has a substantial effect on potential care changes. CONCLUSION: The newly recommended 2021 CKD-EPI creatinine-based eGFR equation may result in substantial changes to recommended care for US patients of all racial and ethnic groups.

Days alive and out of hospital for children born with single-ventricle heart disease.

BACKGROUND: This study describes the illness burden in the first year of life for children with single-ventricle heart disease, using the metric of days alive and out of hospital to characterize morbidity and mortality. METHODS: This is a retrospective single-centre study of single-ventricle patients born between 2005 and 2021 who had their initial operation performed at our institution. Patient demographics, anatomical details, and hospitalizations were extracted from our institutional single-ventricle database. Days alive and out of hospital were calculated by subtracting the number of days hospitalized from number of days alive during the first year of life. A multivariable linear regression with stepwise variable selection was used to determine independent risk factors associated with fewer days alive and out of hospital. RESULTS: In total, 437 patients were included. Overall median number of days alive and out of hospital in the first year of life for single-ventricle patients was 278 days (interquartile range 157-319 days). In a multivariable analysis, low birth weight (<2.5kg) (b = -37.55, p = 0.01), presence of a dominant right ventricle (b = -31.05, p = 0.01), moderate-severe dominant atrioventricular valve regurgitation at birth (b = -37.65, p < 0.05), index hybrid Norwood operation (b = -138.73, p < 0.01), or index heart transplant (b = -158.41, p < 0.01) were all independently associated with fewer days alive and out of hospital. CONCLUSIONS: Children with single-ventricle heart defects have significant illness burden in the first year of life. Identifying risk factors associated with fewer days alive and out of hospital may aid in counselling families regarding expectations and patient prognosis.

Pediatric central nervous system tumor survivor and caregiver experiences with multidisciplinary telehealth.

PURPOSE: Telehealth use to facilitate cancer survivorship care is accelerating; however, patient satisfaction and barriers to facilitation have not been studied amongst pediatric central nervous system (CNS) tumor survivors. We assessed the telehealth experiences of survivors and caregivers in the Pediatric Neuro-Oncology Outcomes Clinic at Dana-Farber/ Boston Children's Hospital. METHODS: Cross-sectional study of completed surveys among patients and caregivers with ≥ 1 telehealth multidisciplinary survivorship appointment from January 2021 through March 2022. RESULTS: Thirty-three adult survivors and 41 caregivers participated. The majority agreed or strongly agreed that telehealth visits started on time [65/67 (97%)], scheduling was convenient [59/61 (97%)], clinician's explanations were easy-to-understand [59/61 (97%)], listened carefully/addressed concerns [56/60 (93%)], and spent enough time with them [56/59 (95%)]. However, only 58% (n = 35/60) of respondents agreed or strongly agreed they would like to continue with telehealth and 48% (n = 32/67) agreed telehealth was as effective as in person office visits. Adult survivors were more likely than caregivers to prefer office visits for personal connection [23/32 (72%) vs. 18/39 (46%), p = 0.027]. CONCLUSION: Offering telehealth multi-disciplinary services may provide more efficient and accessible care for a subset of pediatric CNS tumor survivors. Despite some advantages, patients and caregivers were divided on whether they would like to continue with telehealth and whether telehealth was as effective as office visits. To improve survivor and caregiver satisfaction, initiatives to refine patient selection as well as enhance personal communication through telehealth systems should be undertaken.

Ultrasound-guided reduction of intussusception in infants in a developing world: saline hydrostatic or pneumatic technique?

Non-operative reduction has emerged as first line in the management of uncomplicated intussusception. The aim of this study was to compare the outcome of ultrasound-guided saline hydrostatic reduction and ultrasound-guided pneumatic reduction of intussusception in infants. This is a prospective study of infants with uncomplicated intussusception confirmed by ultrasound over a period of 21 months from December 2018 to August 2020. Fifty-two (69.3%) out of seventy-five infants were eligible and randomized based on simple random sampling technique into two groups: Group A included patients who had ultrasound-guided hydrostatic (saline) reduction; Group B included patients who had ultrasound-guided pneumatic (air) reduction. The success rates, time to reduction and complication rates were assessed. The success rates, between the saline hydrostatic reduction group and pneumatic reduction group, were comparable [17 (65.4%) versus 19 (73.1%); relative risk (RR) 0.8; 95% confidence interval (CI) 0.6-1.2; p = 0.54]. The mean time to reduction was higher in the saline hydrostatic reduction group (15.4 ± 5.1 min versus 10.8 ± 4.1 min; p = 0.003). There was no statistically significant difference in the perforation and recurrence rates between the two groups. CONCLUSION: Saline hydrostatic reduction and pneumatic reduction of uncomplicated intussusception under ultrasound guidance in infants might have comparable outcomes. However, pneumatic reduction may be faster. WHAT IS KNOWN: • Ileocolic intussusception is the most common cause of intestinal obstruction in infants. • Ultrasonography is useful in the diagnosis and non-operative treatment of ileocolic intussusception. WHAT IS NEW: • Ultrasound-guided hydrostatic enema and ultrasound-guided pneumatic enema are similarly effective and safe techniques in the reduction of ileocolic intussusception. • Reduction of ileocolic intussusception under ultrasound guidance is a great technique that may prove useful in the developing world due to lower cost of required equipment.

Dihydroceramides Derived from Bacteroidetes Species Sensitize TRPV1 Channels.

Bacterial colonization of open wounds is common, and patients with infected wounds often report significantly elevated pain sensitivity at the wound site. Transient Receptor Potential Vanilloid Type 1 (TRPV1) channels are known to play an important role in pain signaling and may be sensitized under pro-inflammatory conditions. Bacterial membrane components, such as phosphoethanolamine dihydroceramide (PEDHC), phosphoglycerol dihydroceramide (PGDHC), and lipopolysaccharide (LPS), are released in the environment from the Gram-negative bacteria of the Bacteroidetes species colonizing the infected wounds. Here, we used intracellular calcium imaging and patch-clamp electrophysiology approaches to determine whether bacterially derived PEDHC, PGDHC, or LPS can modulate the activity of the TRPV1 channels heterologously expressed in HEK cells. We found that PEDHC and PGDHC can sensitize TRPV1 in a concentration-dependent manner, whereas LPS treatment does not significantly affect TRPV1 activity in HEK cells. We propose that sensitization of TRPV1 channels by Bacteroidetes-derived dihydroceramides may at least in part underlie the increased pain sensitivity associated with wound infections.

Glucose Mediates Niche-Specific Repression of Staphylococcus aureus Toxic Shock Syndrome Toxin-1 through the Activity of CcpA in the Vaginal Environment.

Staphylococcus aureus chronically colonizes up to 30% of the human population on the skin or mucous membranes, including the nasal tract or vaginal canal. While colonization is often benign, this bacterium also has the capability to cause serious infections. Menstrual toxic shock syndrome (mTSS) is a serious toxinosis associated with improper use of tampons, which can induce an environment that is favorable to the production of the superantigen known as toxic shock syndrome toxin-1 (TSST-1). To better understand environmental signaling that influences TSST-1 production, we analyzed expression in the prototype mTSS strain S. aureus MN8. Using transcriptional and protein-based analysis in two niche-related media, we observed that TSST-1 expression was significantly higher in synthetic nasal medium (SNM) than in vaginally defined medium (VDM). One major divergence in medium composition was high glucose concentration in VDM. The glucose-dependent virulence regulator gene ccpA was deleted in MN8, and, compared with wild-type MN8, we observed increased TSST-1 expression in the ΔccpA mutant when grown in VDM, suggesting that TSST-1 is repressed by catabolite control protein A (CcpA) in the vaginal environment. We were able to relieve CcpA-mediated repression by modifying the glucose level in vaginal conditions, confirming that changes in nutritional conditions contribute to the overexpression of TSST-1 that can lead to mTSS. We also compared CcpA-mediated repression to other key regulators of tst, finding that CcpA regulation is dominant compared to other characterized regulatory mechanisms. This study underlines the importance of environmental signaling for S. aureus pathogenesis in the context of mTSS. IMPORTANCE Menstrual toxic shock syndrome (mTSS) is caused by strains of Staphylococcus aureus that overproduce a toxin known as toxic shock syndrome toxin-1 (TSST-1). This work studied how glucose levels in a model vaginal environment could influence the amount of TSST-1 that is produced by S. aureus. We found that high levels of glucose repress TSST-1 production, and this is done by a regulatory protein called catabolite control protein A (CcpA). The research also demonstrated that, compared with other regulatory proteins, the CcpA regulator appears to be the most important for maintaining low levels of TSST-1 in the vaginal environment, and this information helps to understand how changes in the vaginal environmental can lead to mTSS.

Effectiveness of a third dose of the BNT162b2 mRNA COVID-19 vaccine for preventing severe outcomes in Israel: an observational study.

BACKGROUND: Many countries are experiencing a resurgence of COVID-19, driven predominantly by the delta (B.1.617.2) variant of SARS-CoV-2. In response, these countries are considering the administration of a third dose of mRNA COVID-19 vaccine as a booster dose to address potential waning immunity over time and reduced effectiveness against the delta variant. We aimed to use the data repositories of Israel's largest health-care organisation to evaluate the effectiveness of a third dose of the BNT162b2 mRNA vaccine for preventing severe COVID-19 outcomes. METHODS: Using data from Clalit Health Services, which provides mandatory health-care coverage for over half of the Israeli population, individuals receiving a third vaccine dose between July 30, 2020, and Sept 23, 2021, were matched (1:1) to demographically and clinically similar controls who did not receive a third dose. Eligible participants had received the second vaccine dose at least 5 months before the recruitment date, had no previous documented SARS-CoV-2 infection, and had no contact with the health-care system in the 3 days before recruitment. Individuals who are health-care workers, live in long-term care facilities, or are medically confined to their homes were excluded. Primary outcomes were COVID-19-related admission to hospital, severe disease, and COVID-19-related death. The third dose effectiveness for each outcome was estimated as 1 - risk ratio using the Kaplan-Meier estimator. FINDINGS: 1 158 269 individuals were eligible to be included in the third dose group. Following matching, the third dose and control groups each included 728 321 individuals. Participants had a median age of 52 years (IQR 37-68) and 51% were female. The median follow-up time was 13 days (IQR 6-21) in both groups. Vaccine effectiveness evaluated at least 7 days after receipt of the third dose, compared with receiving only two doses at least 5 months ago, was estimated to be 93% (231 events for two doses vs 29 events for three doses; 95% CI 88-97) for admission to hospital, 92% (157 vs 17 events; 82-97) for severe disease, and 81% (44 vs seven events; 59-97) for COVID-19-related death. INTERPRETATION: Our findings suggest that a third dose of the BNT162b2 mRNA vaccine is effective in protecting individuals against severe COVID-19-related outcomes, compared with receiving only two doses at least 5 months ago. FUNDING: The Ivan and Francesca Berkowitz Family Living Laboratory Collaboration at Harvard Medical School and Clalit Research Institute.