Carcinoma-associated fibroblasts (CAFs), frequently present in the stroma of human breast carcinomas, influence tumor characteristics. A forkhead box protein O1 (FOXO1) transcription factor is a critical mediator of the cellular responses to oxidative stress in various cell types. However, the roles of FOXO1 in CAFs have been poorly understood. Here, we show more intense FOXO1 staining in stromal fibroblasts in human breast cancers compared to those in non-cancerous regions. Notably, stronger stromal FOXO1 staining is significantly associated with poorer outcomes in 237 breast cancer patients. FOXO1 expression is also more abundant in cultured human breast CAFs compared to the control counterpart human mammary fibroblasts. FOXO1 expression is upregulated in control fibroblasts by malnutrition and hypoxia, suggesting such starvation to mediate increased FOXO1 expression in CAFs. Of note, treatment with AS1842856, a FOXO1 inhibitor, significantly attenuates growth and viability in CAFs relative to control fibroblasts. Suppression of FOXO1 expression by shRNA also substantially inhibits CAFs' growth in culture. When the FOXO1-knockdowned CAFs were implanted with breast cancer cells into recipient mice, the number of these fibroblasts present in developed tumors tends to decrease. Taken together, these findings indicate that FOXO1 expression in human breast CAFs is required for their growth and viability.

Initial Cell Seeding Density in Culture Influences Hematopoietic Stem and Progenitor Cell Proliferation, Expansion and Differentiation.

Multiple adsorptions shape collective T-even phage lysis dynamics: Insights from an individual-based model.

The Lyme disease spirochete Borrelia burgdorferi has a distinctive pattern of growth. Newly born cells elongate by primarily inserting peptidoglycan at mid-cell, while in longer cells, additional insertion sites form at the one-quarter and three-quarter positions along the cell length. It is not known how peptidoglycan insertion is concentrated at these locations in B. burgdorferi. In other bacteria, multi-protein complexes are known to synthesize new peptidoglycan, and are often organized by cytoskeletal proteins. We show here that B. burgdorferi's zonal concentration of peptidoglycan insertion requires BB0538 (BbbF) and BB0245 (BbbG), two members of the bactofilin class of cytoskeletal proteins. Bactofilin depletion redistributed peptidoglycan insertion along the cell length. Prolonged bactofilin depletion arrested growth in culture and induced extensive cell blebbing, indicating that B. burgdorferi bactofilins are essential for viability. Fluorescent protein fusions of BbbF and BbbG localized to new zones of growth before peptidoglycan insertion occurred at these sites, with BbbG localization dependent on BbbF. Our results show that BbbF and BbbG direct the spatial patterning of new peptidoglycan insertion in B. burgdorferi.IMPORTANCEThe spirochetal bacterium Borrelia burgdorferi causes Lyme disease, the most prevalent vector-borne infection in North America and Europe. Cellular replication, which requires growth and division of the peptidoglycan cell wall, facilitates B. burgdorferi transmission to, and dissemination within, new hosts. Cellular replication is therefore essential for pathogenesis. Bactofilins regulate peptidoglycan-related processes in several bacteria but are typically non-essential for cellular replication. Bactofilin-encoding genes can be readily deleted in multiple bacterial species. In contrast, we show that the B. burgdorferi bactofilins BbbF and BbbG are essential for cellular viability and direct zonal peptidoglycan insertion. Our findings broaden the spectrum of known bactofilin functions and advance our understanding of how peptidoglycan insertion is regulated in this unusual, medically important spirochetal bacterium.

This article examines the impact over two years of a study pilgrimage to the Holy Land by Master’s students enrolled at the University of Notre Dame Australia. The students undertook a study pilgrimage to the Holy Land in 2023 as part of the Master’s course, Scripture and Church. Through the study pilgrimage, the students showed a significant increase in skill, knowledge and understanding of their faith and the cultural, social and religious complexities within the State of Israel. The findings relating to their study pilgrimage were captured in the research paper ‘Deepening an educator’s understanding of Scripture and Church through an immersion experience in the Holy Land’ (RAiD Number 102.100.100/485606). The Master’s course is aimed at educational teachers and leaders with responsibility for formation for mission in Catholic education. Formation offers teachers of RE an opportunity to discern their place in the world and their purpose to bring Gospel values to the service of others. This research is a longitudinal investigation into the impact of the study pilgrimage to the Holy Land two years later. The impact to be explored includes the students’ professional career, faith, social and cultural engagement and growth. To measure life changes reported by students who had travelled to the Holy Land, one-on-one interviews were conducted with each participant. Open-ended questions were asked, and researchers conducted individual analyses on each. Themes were identified and compared with relevant research. Conclusions and discussions are explored in this article, giving students, teachers, leaders and Universities involved in Catholic education insights into the ongoing impact of the extraordinary human experience of study pilgrimage.

To determine the frequency of microbial DNA detection in patients with sterile pyuria and evaluate the association of leukocyte count and leukocyte esterase (LE) test results with physicians' treatment decisions. This prospective cross-sectional study was conducted over two months in the microbiology laboratories of two university hospitals. Urine samples from 201 patients with sterile pyuria (≥ 10 leukocytes/ml and/or LE positivity with negative urine culture) and 180 controls (no growth in culture and no pyuria) were analyzed. Multiplex PCR targeting 14 potential pathogens was performed. Demographic data, PCR results, and physicians' treatment decisions were recorded. Chi-square tests and logistic regression assessed associations between laboratory findings and treatment decisions. PCR detected at least one potential pathogen in 50.2% of sterile pyuria patients compared to 10% of controls (p < 0.05). Treatment initiation was more frequent in the patient group than in controls (43.3% vs. 30%, p < 0.05). Logistic regression demonstrated that leukocyte counts ≥ 10/ml were significantly associated with PCR positivity (OR = 6.84, p < 0.05) and the physician's treatment decisions (OR = 2.59, p < 0.05). The most common pathogens in patients were Ureaplasma urealyticum (21.9%), Mycoplasma hominis (12.9%), and Gardnerella vaginalis (10.9%), predominantly in adult women with both elevated leukocyte counts and positive LE tests. Although PCR positivity was significantly associated with treatment initiation, the correlation was weak (r = 0.33), indicating that treatment decisions were influenced by additional clinical factors. Molecular testing detected DNA from Ureaplasma, Mycoplasma, and G. vaginalis frequently in sterile pyuria, particularly in adult women; these findings indicate candidate microorganisms requiring clinical correlation.

A mechanistic model describing HEK293 cell growth and viral vector production was developed and calibrated for adenoviral processes and subsequently transferred to adeno‑associated and lentiviral cultures. The model represents key cellular states-viable, dead, and lysed cells-linked through substrate consumption, metabolic inhibition, and virus‑specific toxicity terms. By adjusting a limited set of parameters, the same mechanistic structure, combined with vector specific parameter sets, could accurately reproduce distinct infection and transfection dynamics, demonstrating cross‑platform applicability. As a demonstrator for real‑time deployment, the mechanistic framework was combined with an orthogonal projections to latent structures model based on dielectric spectroscopy data to form a Digital Shadow. The soft‑sensor layer provided real‑time viable cell density values that continuously updated the mechanistic simulation, yielding online predictions of culture growth and infection timing. Validation with adenoviral perfusion datasets confirmed high agreement between simulated and measured viable cell concentrations. This study demonstrates that transferable mechanistic models can serve as a robust foundation for hybrid digital systems in bioprocessing. The proposed Digital Shadow exemplifies how such models can be connected to inline monitoring tools for enhanced process understanding and decision support, paving the way toward closed‑loop Digital Twin applications in viral vector manufacturing.

Hepatitis E virus (HEV) infection is generally self-limiting in immunocompetent individuals but may progress to chronic infection in immunocompromised patients, underscoring the need for effective antiviral therapies. Although ribavirin is currently used off-label for HEV treatment, its associated adverse effects highlight the need for safer alternatives. In this study, we screened an anti-viral compound library comprising 800 compounds using three HEV reporter systems designed to target distinct stages of the viral life cycle. Candidate compounds were further evaluated in PLC/PRF/5 cells using both acute and chronic infection models with wild-type genotype 3 HEV (HEV-3). Antiviral activity was assessed by measuring HEV RNA levels in culture supernatants. Elbasvir, a known inhibitor of hepatitis C virus (HCV) non-structural protein 5A (NS5A), was identified as the most potent candidate. Although multiple compounds showed inhibitory effects in reporter assays, only elbasvir achieved sustained suppression of HEV growth in long-term culture, reducing HEV RNA levels to below the limit of detection. In a chronic infection co-culture model, elbasvir maintained antiviral activity at non-cytotoxic concentrations. Time-of-addition analysis demonstrated that elbasvir inhibits an early step in the viral life cycle, specifically viral internalization. Furthermore, combination with ribavirin enhanced antiviral efficacy, resulting in sustained viral suppression without detectable cytotoxicity and exhibiting an additive interaction. Collectively, these findings identify elbasvir as a promising candidate for repurposing as an anti-HEV drug and support a combination strategy targeting distinct steps of the viral life cycle.

Two potential sources of differentially tolerant subpopulations of Pseudomonas aeruginosa in batch cultures are aggregation and growth rates. Here, we dissected these effects by treating batch cultures with tobramycin, colistin and a combination after various time intervals. Tobramycin was significantly more effective than colistin at early time points, while both showed similar efficacy at 24 and 48 h. Combination therapy was most effective at later stages, indicating the emergence of tolerant subpopulations. Reactivating growth of stationary-phase cultures by treating in fresh media altered antibiotic tolerance, further supporting the role of growth rate in tolerance. To examine the effect of aggregation, we separated batch cultures using filtration as well as examined the WT against a hyper-aggregating ΔwspF mutant. Aggregates demonstrated lower growth rates than single cells. However, microcalorimetry and colony enumeration after treatment revealed that aggregation only partially explained the variance associated with antibiotic treatment. Given that tobramycin and colistin target metabolically distinct subpopulations, this suggests that overall metabolic rate plays a larger role than aggregation regarding antibiotic tolerance in batch cultures.

Selection and qualification of standard test organisms for BWMS type-approval testing.

Applying nephelometry for analyzing liquid yeast cultures.

Failure to rescue (FTR)-death following a potentially manageable complication-has become a key quality indicator. Although clinical and systemic contributors to FTR are well described, how surgeons make sense of these events and how this sense-making shapes communication, escalation, and learning remains underexplored. To explore surgeon interpretations of FTR events and perceptions of factors that contribute to FTR and to identify insights to inform improvements in surgical education and culture. Qualitative study using convenience sampling to recruit board-certified surgeons with 5 or more years of experience in Switzerland and Austria via email. Semistructured interviews were conducted between February and May 2023. Data analysis was performed between February 2024 and April 2025. Direct involvement in FTR events. Interviews were analyzed using a constructivist grounded theory approach to identify themes characterizing surgeons' perceptions of FTR. Fourteen surgeons were interviewed, including 12 men (85.7%) aged 33 to 67 years (mean [SD] age, 49 [9] years). Specialties included abdominal surgery (9 surgeons [64.3%]), orthopedics or traumatology (4 surgeons [29%]), and vascular surgery (1 surgeon [7.1%]). Five themes were developed: (1) being trapped in a flawed system: surgeons perceived FTR as inevitable within systemic and institutional constraints; (2) hierarchical barriers: the continued predominance of barriers hinder adequate reaction and shared decision-making; (3) being imperfect heroes: surgeons saw themselves as passionate, resilient, and driven, yet vulnerable to narcissism, peer hostility, and performative behaviors; (4) coping with failure: participants reflected that FTR experiences, though painful, contributed to improving their clinical expertise; and (5) strategies and tools for rescue: these were characterized by sharing personal strategies and proposing system-level improvements. In this qualitative study of 14 surgeons, participants described how confidence, grit, and self-reliance were essential in surgery but could also hinder timely recognition and response to complications. Fostering humility, reducing hierarchical barriers, and increasing peer support may transform FTR from a heroic surgeon's hubris into an opportunity for collective learning and cultural growth.

Sepsis in pediatric population has a high mortality rate and global health impact. Explicit diagnostic testing is critical in order to ensure timely and accurate diagnosis. Our study is aimed at comparing the efficacy of CRP and PCT, keeping blood culture as gold standard, in diagnosing sepsis in pediatric population to improve treatment and survival. This was a cross-sectional survey conducted at the Department of Pediatric Medicine at The Shaikh Zayed Hospital Lahore. Patients included were newly admitted neonates up to one month age with suspected sepsis, to the Pediatric Intensive Care Unit over a period of six months, i.e. January 3, 2025 to June 30, 2025. Data collection was done by interviewing parents using a structured proforma, CRP and PCT tests were done and their sensitivity, specificity, and diagnostic accuracy were calculated. A total of 165 neonates were enrolled in this study. Male were 46.7% and female were 53.3%. The most common age of presentation was zero day of life (Range 0-23 day, Mean one day, median zero day). C-Reactive Protein had a sensitivity and specificity of 75.5% and 67.2% respectively; while, Procalcitonin had a significantly higher sensitivity and specificity 83.6% and 86.2% respectively. The most common culture growth organism was Klebsiella (44.4%); other organisms included Pseudomonas (26.7 %), Acinetobacter (20%), and E.coli (8.9%). Procalcitonin has a better sensitivity and specificity than C reactive protein in the early diagnosis of sepsis.

Adaptive immune responses occur lymph nodes (LNs) in a microenvironment established by resident stromal cells. LNs are also a site of proliferation of chronic lymphocytic leukemia (CLL), a B cell cancer that alters LN structure in a stereotypic manner. To deeply characterize reactive and CLL LNs, we developed a single-cell RNA sequencing pipeline. We find that proliferation of CLL cells in proliferation centers (PCs), a CLL-specific niche, begins with transient upregulation of MYC, subsequent downregulation of which may limit CLL growth. PCs contain a distinct fibroblast population expressing CCL19 while CLL cells express the CCL19 receptor CCR7, providing a recruitment mechanism for CLL cells to PCs. Using informatic, spatial, and in situ analyses to identify ligand-receptor pairs involving PC CLL cells and nearby immune and stromal cells, we observe that PCs are enriched for macrophages expressing BAFF, the integrin αXβ2 heterodimer, and Galectin9, factors implicated in cell growth, adhesion, and immunosuppression. The most common predicted interactions in PCs involve CD74 and ligands such as MIF, and we find that CD74 blockade consistently inhibits CLL cell growth in culture. Our work highlights key features of the CLL proliferative niche and provides a roadmap for identifying vulnerabilities and new therapeutic strategies.

Microalgal cultivation on ricotta cheese whey: Modulating nutrient supply during cultivation to improve culture growth.

Recurrent driver mutations in genes encoding histone H3 (H3.3K27M and H3.1K27M) are observed in ∼80% of diffuse midline gliomas (DMG), which lead to aberrant gene regulation, yet the specific RNA polymerase II (Pol2) regulators that induce transcriptional dysregulation in DMG are not fully defined. We identified multiple regulators of Pol2 elongation as DMG genetic dependencies in a chromatin-focused CRISPR screen. Additional studies confirm that knockout (KO) of the Pol2 SIII complex gene elongin B (ELOB) inhibits DMG cell proliferation in tissue culture and tumor growth in xenograft models. Further genomic analyses reveal that ELOB binding sites are enriched in H3K27M oncohistones and that ELOB KO alters H3K27me3 and H3K27M incorporation at thousands of genomic regions, implicating ELOB in the maintenance of dysfunctional chromatin states in DMG. Correspondingly, PRO-seq and RNA-seq profiling reveal that ELOB loss disrupts Pol2 transcriptional activity and alters the expression of transcripts involved in metabolism, proliferation, and brain development. These findings suggest that Pol2 elongation factors like ELOB cooperate with H3K27M oncohistones to maintain the epigenetic and transcriptional landscape driving DMG malignancy.

Invasive lobular carcinoma (ILC) is the most frequently diagnosed special histological subtype of invasive breast cancer and accounts for 10 – 15% of all cases. The pathognomonic hallmark of ILC is the genetic loss of E-cadherin (CDH1) causing the disruption of adherens junctions and resulting in discohesive, linear growth. To better understand the role of E-cadherin in ILC metastasis, we generated three ILC cell lines, MDA-MB-134-VI, SUM44PE, and BCK4, with inducible E-cadherin expression, resulting in successful restoration of functional adherens junctions. E-cadherin expression reduced growth in 2D culture, and that effect was even greater in 3D ultra-low attachment (ULA) conditions where increased cell death was consistent with the previously described role of E-cadherin in anoikis. E-cadherin expression did not rescue the lack of migration and invasion of ILC cell line models; however, it decreased haptotaxis and increased adherence to Collagen I in SUM44 cells. There was no significant effect of E-cadherin expression on primary orthotopic tumor growth, but spontaneous metastasis to the reproductive tract, brain, and GI tract was reduced. Inhibition of metastasis to the reproductive tract and brain was also seen after tail vein injection of MDA-MB-134 E-cadherin-expressing cells. In summary, overexpression of functional E-cadherin in ILC models has some, but limited, effects on 2D growth in vitro and primary tumor growth in vivo, but there are pronounced effects on 3D ULA growth and metastases in vivo, with stronger effects on metastatic sites enriched in patients with ILC, especially the reproductive and GI tracts.

Evaluation of the International Consensus Meeting (ICM) diagnostic categories for shoulder periprosthetic joint infection: Cutibacterium acnes predominates even in definite infections.

Do wound protectors reduce contamination in total shoulder arthroplasty? A randomized controlled trial.

Rapid and accurate diagnosis of urinary tract infections (UTIs) is essential to guide empirical therapy and curb antimicrobial resistance. However, conventional urine culture is too slow, hindering timely diagnosis of UTI. UF‑5000, an automated flow cytometry urinalysis system equipped with the BACT-Info Gram-prediction feature, shows potential to expedite microbial screening. However, its performance under low bacterial loads re-mains unclear. We retrospectively analyzed 1,529 clinical urine specimens using urinalysis and urine culture. Any bacterial growth in urine culture was considered a culture-positive result. Receiver operating characteristic (ROC) analysis of UF‑5000 bacterial and white blood cell counts was performed to determine the optimal cutoffs. The agreement between the BACT‑Info flag feature and urine culture results was assessed using Cohen's κ value. Urine cultures identified bacteria in 700/1,529 specimens (45.8%), with 585 (38.3%) showing ≥ 105 colony-forming units/mL. ROC analysis established a bacterial count cutoff of 195.2/µL, yielding a sensitivity of 0.851, specificity of 0.885, and area under the ROC curve of 0.940 (95% confidence interval, 0.928 - 0.951). It outper-formed the cutoff value based on white blood cell count. At equivalent amounts of bacterial culture, UF‑5000 bacterial counts were significantly higher for Gram‑negative than for Gram‑positive isolates (Welch's t-test: p < 0.001). Overall concordance between BACT‑Info and urine culture results was substantial for Gram‑negative flags (κ, 0.76; accuracy, 0.904) and moderate for Gram‑positive flags (κ, 0.51; accuracy, 0.799). For Gram-negative bacteria, the sensitivity, specificity, positive predictive value, and negative predictive value were 0.739, 0.974, 0.923, and 0.898, respectively. However, 17.2% of the specimens containing only Gram-negative bacteria were flagged as "Gram Positive?". This indicates the potential for improvement. The UF‑5000 system provides a rapid, low-labor, and cost-effective method for UTI screening, even at low levels of bacteriuria. However, it is intended to complement, not replace, conventional urine culture, which remains indispensable for definitive species identification and antimicrobial susceptibility testing. Its primary value lies in accelerating the diagnostic workflow and reducing unnecessary cultures when used in conjunction with urine culture.