bluuebunny/crossref_metadata_2025 huggingface dataset:
Optimizing approaches for targeted integration of transgenic cassettes by integrase-mediated cassette exchange in mouse and human stem cells
Authors: Rath Phalguni , Kramer Philipp , Biggs Daniel , Preece Chris , Hortin Nicole , Diaz Rebeca , Perez-Alcantara Marta , Li Xiang , Bolard Arnaud , Beer Nicola , McCarthy Mark , Davies Benjamin
Published: January 2025

To enable robust expression of transgenes in stem cells, recombinase-mediated cassette exchange at safe harbor loci is frequently adopted. The choice of recombinase enzyme is a critical parameter to ensure maximum efficiency and accuracy of the integration event. We have explored the serine recombinase family of site-specific integrases and have directly compared the efficiency of PhiC31, W-beta, and Bxb1 integrase for targeted transgene integration at the Gt(ROSA)26Sor locus in mouse embryonic stem cells. All 3 integrases were found to be suitable for efficient engineering and long-term expression of each integrase was compatible with pluripotency, as evidenced by germline transmission. Bxb1 integrase was found to be 2-3 times more efficient than PhiC31 and W-beta. The Bxb1 system was adapted for cassette exchange at the AAVS1 locus in human induced pluripotent stem (iPS) cells, and the 2 commonly used ubiquitous promoters, CAG and Ef1α (EIF1A), were tested for their suitability in driving expression of the integrated transgenic cargo. AAVS1-integrated Ef1α promoter led to a very mosaic pattern of expression in targeted hiPS cells, whereas the AAVS1-integrated CAG promoter drove consistent and stable expression. To validate the system for the integration of functional machinery, the Bxb1 integrase system was used to integrate CAG-driven CRISPR-activation and CRISPR-inhibition machinery in human iPS cells and robust sgRNA-induced up- and downregulation of target genes was demonstrated.
Application of a quantitative uncertainty assessment to develop ranges of plausible toxicity values when using observational data in risk assessment: a case study examining associations between PFOA and PFOS exposures and vaccine response
Authors: Wikoff Daniele S , Vincent Melissa J , Heintz Melissa M , Pastula Susan T , Reichert Heidi , Klaren William D , Haws Laurie C
Published: January 2025

Traditional approaches for quantitatively characterizing uncertainty in risk assessment require adaptation to accommodate increased reliance on observational (vs experimental) studies in developing toxicity values. Herein, a case study with perfluorooctanoic acid (PFOA) and PFOS and vaccine response explores approaches for qualitative and—where possible—quantitative assessments of uncertainty at each step in the toxicity value development process when using observational data, including review and appraisal of individual studies, candidate study selection, dose–response modeling, and application of uncertainty factors. Each of the 15 studies identified had uncertainties due to risk of bias in confounding, outcome, and exposure ascertainment, likely contributing to the observed inconsistencies within and across studies, and resulting in lack of candidacy for dose–response assessment. Nonetheless, 2 representative studies were selected to demonstrate possible methods to quantify uncertainty in the remaining steps. Data simulations indicated lack of a clear dose–response relationship; dose–response models fit to representative simulations indicated high uncertainty in both the magnitude and direction of effect with simulated benchmark dose and its lower limit values varying at least 66- and 86-fold for PFOA and PFOS. Uncertainty factor application added minimal uncertainty. Combined, a high level of uncertainty was observed, precluding the ability to confidently assess causal dose–response relationships with the observational data, alone. This case study highlights the need for quantitative uncertainty analysis when developing toxicity values with observational data and, importantly, emphasizes the need for application of additional techniques to directly assess causality and the specificity of dose–response when relying on studies of association in quantitative risk assessment.
The human‐specific noncoding RNA RP11‐424G14.1 functions at the intersection of sexually dimorphic pathways in inflammation, senescence, and metabolism
Authors: Kennicott Kameron , Liang Yun
Published: January 2025

Sexual dimorphism is a fundamental characteristic of various physiological and pathological processes in humans, including immune responses, senescence, and metabolism. Most studies on the sex bias have focused on sex hormones or female‐biased genes, whereas male‐biased genetic factors remain understudied. Here, we show that the Y‐linked noncoding RNA, RP11‐424G14.1, is expressed in human male keratinocytes. Microarray study suggests the NF‐κB pathway as the top biological pathway affected by RP11‐424G14.1 knockdown, consistent with known sex differences in inflammation. Additionally, IGFBP3 is identified as the top gene supported by RP11‐424G14.1 in male keratinocytes. Conversely, in female keratinocytes, IGFBP3 is the top gene repressed by the X‐linked long noncoding RNA XIST, suggesting a central role of IGFBP3 in mediating sexual dimorphism. Knockdown of RP11‐424G14.1 or IGFBP3 in male keratinocytes inhibits cellular senescence, consistent with increased longevity in females. IGFBP3 expression is dependent on insulin, and metabolomics analysis suggests that RP11‐424G14.1 and IGFBP3 regulate acrylcarnitine metabolism. Our study identifies the role of the RP11‐424G14.1‐IGFBP3 pathway in coordinating sex differences in immunity, senescence, and metabolism. With RP11‐424G14.1 being a human‐specific genetic element, our study suggests the evolving feature of sexual dimorphisms in biological processes.
Defining the Need for Causal Inference to Understand the Impact of Social Determinants of Health: A Primer on Behalf of the Consortium for the Holistic Assessment of Risk in Transplantation (CHART)
Authors: Bhavsar Nrupen A. , Patzer Rachel E. , Taber David J. , Ross-Driscoll Katie , Deierhoi Reed Rhiannon , Caicedo-Ramirez Juan C. , Gordon Elisa J. , Matsouaka Roland A. , Rogers Ursula , Webster Wendy , Adams Andrew , Kirk Allan D. , McElroy Lisa M.
Published: January 2025

Objective:

This study aims to introduce key concepts and methods that inform the design of studies that seek to quantify the causal effect of social determinants of health (SDOH) on access to and outcomes following organ transplant.

Background:

The causal pathways between SDOH and transplant outcomes are poorly understood. This is partially due to the unstandardized and incomplete capture of the complex interactions between patients, their neighborhood environments, the tertiary care system, and structural factors that impact access and outcomes. Designing studies to quantify the causal impact of these factors on transplant access and outcomes requires an understanding of the fundamental concepts of causal inference.

Methods:

We present an overview of fundamental concepts in causal inference, including the potential outcomes framework and direct acyclic graphs. We discuss how to conceptualize SDOH in a causal framework and provide applied examples to illustrate how bias is introduced.

Results:

There is a need for direct measures of SDOH, increased measurement of latent and mediating variables, and multi-level frameworks for research that examine health inequities across multiple health systems to generalize results. We illustrate that biases can arise due to socioeconomic status, race/ethnicity, and incongruencies in language between the patient and clinician.

Conclusions:

Progress towards an equitable transplant system requires establishing causal pathways between psychosocial risk factors, access, and outcomes. This is predicated on accurate and precise quantification of social risk, best facilitated by improved organization of health system data and multicenter efforts to collect and learn from it in ways relevant to specialties and service lines.
Socioeconomic Determinants and Health Outcomes of Financial Resilience Patterns
Authors: Liu Zewei , Chen Ji-Kang
Published: January 2025

Background and Objectives:

Financial resilience describes an actionable financial capability to cope with emergencies and adversities. However, little is known about whether financial resilience patterns impact holistic health outcomes.

Methods:

This study conducted a 3-step latent profile analysis using an urban community sample (n = 3710) from a nationally representative database in China. A multinomial logistic regression model and multiple linear regression models were estimated to examine the proposed argument guided by social determinants of health and resilience theory.

Results:

Three latent subgroups were identified to differentiate population heterogeneity with high, moderate, and low financial resilience. Younger generations with lower employment participation, education, income, and assets were significantly less likely to be in moderate and high financial resilience subgroups. After controlling demographics and conventional socioeconomic covariates, financial resilience patterns were significantly negatively associated with mental illness, while positively associated with physical health, subjective happiness, and life satisfaction.

Conclusion:

Financial resilience could be a strength-based and actionable socioeconomic determinant of health. Research and practical implications are provided for addressing health inequalities. A new target in community health and social welfare is to enhance financial resilience among marginalized populations through family asset-building, financial inclusion policy, financial literacy education, and social capital investment.
Nodular plantar melanoma in a black male and the importance of tumor excision and inguinal lymph node dissection: a case report
Authors: Ali Mohamed Osama Mohamed , Mohammed Osman Saadia Abdulghani Abdulrahman , Ali Khawla Alawad Elbashir , Basher Mohamed Osman Suliman , Almahdi Taiba Ahmed Mohammed , Elgmar Habab Basheir Abdelgadir
Published: January 2025

Background:

Nodular melanoma is a rare and aggressive malignant skin cancer. It is the third most common subtype of melanoma, following acral lentiginous melanoma and superficial spreading melanoma, and accounts for 5–8% of all melanoma subtypes among dark-skinned populations. This manuscript presents a case of nodular melanoma on the plantar surface of the foot with metastasis to the inguinal lymph nodes.

Case presentation:

An 82-year-old male with no significant medical history was referred by a dermatologist due to recurrent raised skin pigmentation on the plantar surface of the left foot. The lesion was black in color and associated with pain during walking. Wide-margin excision of the melanoma and inguinal lymph node dissection (ILND) were performed. The diagnosis of nodular melanoma was confirmed through clinical examination and histopathological analysis.

Clinical discussion:

This case highlights the rarity of nodular melanoma among patients of African descent. Its rarity among Africans is likely due to their darker skin and the presence of eumelanin, which filters twice as much ultraviolet radiation as lighter skin. ILND is a crucial strategy, as it can reduce recurrence and improve survival.

Conclusion:

Nodular melanoma is associated with substantially shorter disease-free survival and a higher case fatality rate compared to other melanoma subtypes. Surgical resection and lymph node dissection are effective in improving prognosis. The possibility of melanoma recurrence must always be considered.
Neurogastroenterology and motility disorders in patients with cirrhosis
Authors: Idalsoaga Francisco , Ayares Gustavo , Blaney Hanna , Cabrera Daniel , Chahuan Javier , Monrroy Hugo , Matar Ayah , Halawi Houssam , Arrese Marco , Arab Juan Pablo , Díaz Luis Antonio
Published: January 2025

Neurogastroenterology and motility disorders are complex gastrointestinal conditions that are prevalent worldwide, particularly affecting women and younger individuals. These conditions significantly impact the quality of life of people suffering from them. There is increasing evidence linking these disorders to cirrhosis, with a higher prevalence compared to the general population. However, the link between neurogastroenterology and motility disorders and cirrhosis remains unclear due to undefined mechanisms. In addition, managing these conditions in cirrhosis is often limited by the adverse effects of drugs commonly used for these disorders, presenting a significant clinical challenge in the routine management of patients with cirrhosis. This review delves into this connection, exploring potential pathophysiological links and clinical interventions between neurogastroenterology disorders and cirrhosis.
Identification of optimal portal pressure decrease to control ascites while minimizing HE after TIPS: A multicenter study
Authors: Kabelitz Martin A. , Hartl Lukas , Schaub Golda , Tiede Anja , Rieland Hannah , Kornfehl Andrea , Hübener Peter , Jachs Mathias , Hinrichs Jan , Schütte Sarah L. , Riedel Christoph , Mauz Jim B. , Tergast Tammo L. , Meyer Bernhard C. , Bannas Peter , Kappel Julia , Wedemeyer Heiner , Kluwe Johannes , Piecha Felix , Reiberger Thomas , Sandmann Lisa , Maasoumy Benjamin
Published: January 2025

Background and Aims:

Clinically significant portal hypertension in patients with liver cirrhosis can lead to refractory ascites. A TIPS treats clinically significant portal hypertension but may cause overt hepatic encephalopathy (oHE). Our aim was to determine the optimal reduction of the portal pressure gradient (PPG) through TIPS to control ascites without raising oHE risk.

Approach and Results:

This multicenter study screened 1509 patients from 3 European centers (Hannover, Vienna, and Hamburg) undergoing TIPS implantation between 2000 and 2023. Patients with TIPS indications other than refractory ascites/hepatic hydrothorax, vascular liver disease, HCC, or insufficient PPG data were excluded. PPG was measured before and after TIPS insertion. Outcome data were assessed up to 1 year after TIPS insertion. Analyses were conducted utilizing a modern machine learning model, namely a competing-risk random survival forest, partial dependence plots, and competing risk analyses with liver transplantation/death as competitors. The cohort was divided into a 60% derivation and 40% validation cohort. Overall, 729 patients (median MELD: 13 [IQR 10-16], 66% male, 23% oHE before TIPS) were analyzed. The derivation cohort comprised 438 patients, and the validation cohort comprised 291 patients. The optimal PPG reduction, determined by maximally selected Gray statistic and PDP of the random survival forest, was 60%–80%. In this range, patients showed significantly fewer hepatic decompensations due to ascites (HDA) (subdistribution hazard ratio [sHR]: 0.7 [0.52–0.96]) with similar oHE incidences (sHR: 0.92 [0.67–1.27]). The PPG range was confirmed in the validation cohort (HDA: sHR: 0.66 [0.46–0.96]; oHE: sHR: 0.89 [0.61–1.32]).

Conclusions:

A targeted PPG reduction of 60%–80% showed significantly reduced HDA without increased oHE risk. Therefore, PPG reduction within this range could be a valid reduction target.
Are we ready for the next pandemic: Disease X
Authors: Erenler Ali Kemal , Erenler Behice Hande , Baydin Ahmet
Published: January 2025

Disease X is a term that defines a disease with a potential to cause the next pandemic. Disease X may result from various agents including viruses, bacteria, fungi, parasites, and prions. Disease X is estimated to be a zoonosis, most likely an RNA virus, which may easily be transmitted among humans. Former diseases that have similarities with Disease X were HIV, severe acute respiratory syndrome (SARS), Zika, Ebola and the most recently COVID-19. Other potential agents that may cause Disease X are Ebola and Marburg virus, Crimean–Congo hemorrhagic fever, Lassa fever, Middle East respiratory syndrome (MERS), Rift valley fever, Zika fever, Nipah and Henipaviral diseases. As the number of activities in the wildlife and consumption of uncontrolled animal source foods increases, it is inevitable to experience pandemics in the future. In addition, climate change is another reason for emergence of novel viruses. Possible pathogens in exotic animals and animal products should be controlled and sampled. Measures such as scientific collaboration between clinicians and public health officials, avoidance of disinformation, travel limitations and screening of the passengers with technological devices, an active surveillance system involving widespread testing, social distancing and quarantine should be maintained.
Postintubation Sedation of Pediatric Patients in the Emergency Department
Authors: Wynia Emily , Baumgartner Kevin , Yaeger Lauren H. , Ancona Rachel , Wiltrakis Susan , Fuller Brian M.
Published: January 2025

Context

Postintubation sedation is a critical intervention for patients undergoing mechanical ventilation. Research in the intensive care unit (ICU) and adult emergency department (ED) demonstrates that appropriate postintubation sedation has a significant impact on patient outcomes. There are minimal published data regarding postintubation sedation for pediatric ED patients.

Objective

To identify, describe, and critique published literature on postintubation sedation in pediatric ED patients.

Data Sources

Embase, Ovid Medline, Scopus, Cochrane Central Register of Controlled Trials, Cochrane Database of Systematic Reviews, and ClinicalTrials.Gov.

Study Selection

Studies describing postintubation sedation in the ED for pediatric patients (<18 years of age) intubated in the ED via rapid sequence intubation (RSI) were included. Studies reporting intubation outside the ED, intubation not by RSI, or nonpediatric patients were excluded, as were studies not reporting novel human clinical research.

Data Extraction

Data were abstracted by 2 authors using a standardized worksheet. Data included study design & setting, demographics, medications for RSI and postintubation sedation, administration of any or appropriately timed postintubation sedation, and predictive factors for postintubation sedation-related outcomes.

Results

A total of 10 studies were included, all of which were nonrandomized; there was significant heterogeneity and many key variables and outcomes were not consistently reported. Meta-analysis of eligible studies demonstrated pooled estimates of 77.3% (95% confidence interval [CI]: 62.5–92.0) for proportion of patients receiving any postintubation sedation, 43.5% (95% CI: 29.3–57.6) for proportion of patients receiving appropriately timed postintubation sedation, and 18.6 minutes (95% CI: 12.5–24.7) for median time to postintubation sedation, all with significant heterogeneity.

Conclusions

Data on postintubation sedation in pediatric ED patients are limited. Administration of postintubation sedation is inconsistent and may be substantially delayed. Further high-quality research into the use of postintubation sedation in this setting is needed, and appropriate postintubation sedation should be a target for quality improvement.
Combination of Vascularized and Non-vascularized Fibula Free Flap With Dermofat Graft for Maxilla and Mandibula Reconstruction After Malignant Spindle Cell Tumor Resection
Authors: Bogari Melia , Humaira Cut Firza , Triatmoko Sara Ester , Dianti Yuanita Safitri
Published: January 2025

The spindle cell tumor is a variant of sarcomatoid carcinoma that mostly affects the oral cavity. Bone involvement in this tumor leads to a wide excision, which sometimes requires resection of both the maxilla and mandible. The maxilla and mandible are important bones that function to form the 3-dimensional dimensions of the facial bones. The fibular bone can be selected to replace the facial bone because of its strong structure with a skin paddle, long pedicle, and proper bone shape. The authors present the case of a 24-year-old female who underwent maxillectomy and total hemimandibulectomy after a spindle cell tumor resection. The free fibular flap was harvested and divided into several segments to close the mandible and maxilla. However, the vascularized bone was insufficient; thus, non-vascularized bone was used in combination to reconstruct the maxilla. Six months after bone reconstruction, a dermofat graft was placed to fill the cheek structure. During the 6 months of follow-up, the vascularized and non-vascularized fibular bones were well arranged. Intraoral placement of a skin paddle covering the maxilla and mandible was viable. Both functional and esthetic outcomes were attained in patients with high satisfaction rates. Although there are many reconstruction options for patients with post-wide tumor excision, the fibula free flap remains the main choice as a replacement for facial bones because it has a strong and firm bone structure that can resemble facial bones; thus, a non-vascularized bone graft can be used in combination to cover the defects.
‘Experimentum Crucis’: Hauksbee the Younger’s ‘decisive experiment’ for comparing the ‘Safety and Efficacy’ of new medicines (1743)
Authors: Cooper Max , Cooper Sarah
Published: January 2025

In 1743 Francis Hauksbee the Younger published a proposal for an ‘Experimentum Crucis’ (‘decisive experiment’) to compare his own medication for venereal disease with other treatments. Previously he had sought to replicate the methods of James Jurin FRS, who published outcomes from inoculation against smallpox in the 1720s. By seeking to record outcomes (‘Safety and Efficacy’) of his medicine used by local doctors, Hauksbee aimed ‘to lay the Whole of the Evidence (both for and against [his medicine]) properly attested before the Publick’. Owing to lack of engagement from doctors and bitter allegations of quackery, Hauksbee then proposed his ‘decisive experiment’ in ten practical steps. This called for twelve patients, half of whom would receive Hauksbee’s medication and the remaining half treatments of other doctors. Hauksbee insisted that patient outcomes be independently assessed and that the findings for all twelve be published, specifically long-term clinical outcomes (a ‘lasting cure’). Hauksbee’s proposal is an early model for a prospective controlled trial with patients allocated to two or more groups. The date of publication, subject (venereal disease), twelve participants, and importance of considering patients’ diet and accommodation suggest that Hauksbee’s proposal was the inspiration for James Lind’s 1747 trial of treatments for scurvy.
The importance of peripheral populations in the face of novel environmental change
Authors: Hoff Samantha , Hoyt Joseph R. , Langwig Kate E. , Johnson Luanne , Olson Elizabeth , O’Dell Danielle , Pendergast Casey J. , Herzog Carl J. , Parise Katy L. , Foster Jeffrey T. , Turner Wendy C.
Published: January 2025

Anthropogenically driven environmental change has imposed substantial threats on biodiversity, including the emergence of infectious diseases that have resulted in declines of wildlife globally. In response to pathogen invasion, maintaining diversity within host populations across heterogenous environments is essential to facilitating species persistence. White-nose syndrome is an emerging fungal pathogen that has caused mass mortalities of hibernating bats across North America. However, in the northeast, peripheral island populations of the endangered northern myotis (
Myotis septentrionalis
) appear to be persisting despite infection while mainland populations in the core of the species range have experienced sharp declines. Thus, this study investigated host and environmental factors that may contribute to divergent population responses. We compared patterns of pathogen exposure and infection intensity between populations and documented the environmental conditions and host activity patterns that may promote survival despite disease invasion. For island populations, we found lower prevalence and less severe infections, possibly due to a shorter hibernation duration compared to the mainland, which may reduce the time for disease progression. The coastal region of the northern myotis range may serve as habitat refugia that enables this species to persist despite pathogen exposure; however, conservation efforts could be critical to supporting species survival in the long term.
Impact of Donor History on the Risk of Transfusion-Related Infections
Authors: Ogolla Collince Odiwuor , Guya Benard , Maima Apollo O.
Published: January 2025

Background

Transfusion-related infections are a severe threat to the safety of transfusing blood products, internationally. Advances in screening procedures have not, nevertheless, rendered blood transfusion a risk-free procedure for transmitting infectious disease. Donor history, including previous donation history, medical conditions, and high-risk behavior, may influence transfusion-related infections.

Objective

The objective of this study was to determine donors’ histories that could influence the possibility of transfusion-related infections.

Methods

The cross-sectional study was done in January 2022 to December 2023 at KTRH. Out of 108 blood transfusions, data for 108 patients were retrieved from patient medical records and donor screening forms. Variables such as HIV and Hepatitis among donor medical conditions, previous donation history, and high-risk behaviors such as intravenous drug use were analyzed for their link with TRIs. The diagnosis of TRIs was established based on the results of a clinical examination and laboratory tests. Using descriptive statistics as well as Chi-square tests and logistic regression, data were analyzed.

Results

Of the total transfusion sample (n=108), 15 (13.9%) recipients developed infections after transfusion; in these cases, 4.6% had Hepatitis B infections, whereas 3.7% had HIV infections and 5.6% had malaria infections. Past donor experience and risky behavior, which include intravenous drug uses and other risky sexual practices, show a significant association between the increased risk of TRIs (p < 0.05). The odds of transfusion-transmitted infections among repeat donors as compared to first-time donors were marginally high (p=0.04). These independent risk factors for transfusion-related infections were hepatitis B and HIV.

Conclusion

The current study calls for a deliberate consideration of donor history, especially previous donation records, medical conditions, and high-risk behaviors, in the prevention of transfusion-related infections. Improved donor screening protocols and monitoring of high-risk behaviors can further enhance safety in blood transfusion. Future research should investigate their effectiveness in targeted intervention strategies among high-risk donor populations.
The Inversion of Naïve-to-Memory T Cell Ratios With Age: Potential Links to Autoimmunity and the Role of Lifetime Infection History
Authors: Montgomery Richard Murdoch
Published: January 2025

As individuals grow older, their T cell repertoire progressively shifts from being dominated by naïve T lymphocytes to featuring a preponderance of memory T cells. This phenomenon—well-documented in immunological literature—is frequently attributed to repeated antigenic stimulation over the course of a lifetime, coupled with thymic involution and other age-associated changes. Although memory T cells are crucial for rapid responses to previously encountered pathogens, there is increasing interest in whether this dramatically shifted balance might predispose certain individuals to a higher risk of autoimmunity, especially when compounded by chronic stress and recurrent infections. In this article, we propose a conceptual model linking the age-related inversion of the naïve-to-memory T cell ratio to an elevated likelihood of immune dysregulation. We integrate mathematical equations to capture the dynamic interactions among key variables, present simulated data in four informative graphs, and discuss the implications and limitations of our approach. Our goal is to invite a deeper consideration of how infection history, stress-induced immune breakdowns, and the intrinsic remodeling of T cell populations with age could collectively contribute to autoimmunity.
RhCMV Expands CCR5 Memory T Cells and promotes SIV reservoir genesis in the Gut Mucosa
Authors: Perdios Chrysostomos , Babu Naveen Suresh , Coleman Celeste D. , Brown Anna T. , Mostrom Matilda J. , Allers Carolina , Doyle-Meyers Lara , Fennessey Christine M. , Keele Brandon F. , Kaur Amitinder , Freeman Michael L. , Mudd Joseph C.
Published: January 2025

Cytomegalovirus (CMV) is a prevalent β-herpesvirus that persists asymptomatically in immunocompetent hosts. In people with HIV-1 (PWH), CMV is associated with persistence of the HIV-1 reservoir and particular inflammatory related co-morbidities. The true causative role of CMV in HIV-associated pathologies remains unclear given that nearly all PWH are coinfected with CMV. In this study, we examined acute phase SIV dynamics in cohorts of rhesus macaques that were seropositive or -negative for rhesus CMV (RhCMV). We observed expansion of CCR5+ target CD4+ T cells in gut and lymph nodes (LN) that existed naturally in RhCMV-seropositive animals, the majority of which did not react to RhCMV lysate. These cells expressed high levels of the chemokine receptor CXCR3 and a ligand for this receptor, CXCL9, was systemically elevated in RhCMV-seropositive animals. RhCMV+ RMs also exhibited higher peak SIV viremia. CCR5 target memory CD4 T cells in the gut of RhCMV+ RMs were maintained during acute SIV and this was associated with greater seeding of SIV DNA in the intestine. Overall, our data suggests the ability of RhCMV to regulate chemotactic axes that direct lymphocyte trafficking and promote seeding of SIV in a diverse, polyclonal pool of memory CD4+ T cells.
Optogenetic control of Protein Kinase C-epsilon activity reveals its intrinsic signaling properties with spatiotemporal resolution
Authors: Ong Qunxiang , Lim Crystal Jing Yi , Liao Yilie , Ng Justin Tze-Yang , Lim Ler Ting Rachel , Koh Shernys Xuan Yi , Chan Sher En , Lee Pheobe Yu Ying , Lim Huijun , Ye Chen Rui , Wang Loo Chien , Ler Siok Ghee , Sobota Radoslaw Mikolaj , Tan Yaw Sing , I. Shulman Gerald , Yang Xiaoyong , Han Weiping
Published: January 2025

The regulation of PKC epsilon (PKCepsilon) and its downstream effects is still not fully understood, making it challenging to develop targeted therapies or interventions. A more precise tool that enables spatiotemporal control of PKCepsilon activity is thus required. Here, we describe a photo-activatable optogenetic PKCepsilon probe (Opto-PKCepsilon) consisting of an engineered PKCepsilon catalytic domain and a blue-light inducible dimerization domain. Molecular dynamics and AlphaFold simulations enable rationalization of the dark-light activity of the optogenetic probe. We first characterize the binding partners of Opto-PKCepsilon, which are similar to those of PKCepsilon. Subsequent validation of the Opto-PKCepsilon tool is performed with phosphoproteome analysis, which reveals that only PKCepsilon substrates are phosphorylated upon light activation. Opto-PKCepsilon could be engineered for recruitment to specific subcellular locations. Activation of Opto-PKCepsilon in isolated hepatocytes reveals its sustained activation at the plasma membrane is required for its phosphorylation of the insulin receptor at Thr1160. In addition, Opto-PKCepsilon recruitment to the mitochondria results in its lowering of the spare respiratory capacity through phosphorylation of complex I NDUFS4. These results demonstrate that Opto-PKCepsilon may have broad applications for the studies of PKCepsilon signaling with high specificity and spatiotemporal resolution.
Low Magnetic Fields Stimulate Cardiac Mitochondrial Bioenergetics with a Bell-Shaped Response: Possibly Via a Radical Pair Mechanism
Authors: Beutner Gisela , Yuh Huoy-Jen , Goldenberg Ilan , Wallace Douglas C. , Porter George A. , Moss Arthur J. , Sheu Shey-Shing
Published: January 2025

Studies have shown that low magnetic fields (LMFs) of less than 1 × 10−3Tesla (T) affect numerous biological events, including bacteria and plant growth, bird migration, and human brain activity. On a cellular level, LMFs affect ion channel activities, intracellular Ca2+concentrations, and mitochondrial reactive oxygen species (mROS) generation. However, the mechanisms that could account for these effects are controversial. Here, we show that applying a static LMF, ranging from ∼2.7 × 10−4to ∼1.9 × 10−3T, to mitochondria isolated from adult rat hearts produced a bell-shaped increase in maximal respiration (Vmax) up to 40%. A similar LMF-induced increase in Vmax was also observed in mitochondria isolated from rat hearts subjected to ischemia-reperfusion (I-R) injury. We then obtained data showing that LMF- mediated bell-shaped response was also observed in the activity of several enzymes involved in oxidative phosphorylation (OXPHOS), including Complexes II, III, and V, and citrate synthase. By contrast, similar LMF caused little change in the enzymatic activity of Complex I. Interestingly, mROS generation responded to LMF with an inverted bell-shaped decrease. We propose a radical pair mechanism of magnetoreception in cytochromes, catalytical reactions, and iron-sulfur clusters within the OXPHOS enzymes to explain how an LMF can increase the likelihood of electron spin transitions from singlet to triplet state and reverse it as the magnetic field strength further increases, resulting in a bell-shaped response. Our results indicate that a narrow range of LMF can enhance mitochondrial bioenergetics and decrease mROS. This may provide a non-invasive approach to treating diseases, such as I-R injury, when energy generation is compromised and oxidative stress is magnified.
Culture-supported ecophysiology of the SAR116 clade demonstrates metabolic and spatial niche partitioning
Authors: Coelho Jordan T , Teubner Lauren , Henson Michael W , Lanclos V. Celeste , Kojima Conner Y , Thrash J. Cameron
Published: January 2025

Marine SAR116 bacterioplankton are ubiquitous in surface waters across global oceans and form their own order, Puniceispirillales, within the Alphaproteobacteria. To date no comparative physiology among diverse SAR116 isolates has been performed to capture the functional diversity within the clade, and further, diversity through the lens of metabolic potential and environmental preferences via clade-wide pangenomics remains poorly constrained. Using high-throughput dilution-to-extinction cultivation, we isolated and genome sequenced five new and diverse SAR116 isolates from the northern Gulf of Mexico. Here we present a comparative physiological analysis of these SAR116 isolates, along with a pangenomic investigation of the SAR116 clade using a combination of metagenome-assembled genomes (MAGs, n=258), single-amplified genomes (SAGs, n=84), previously existing (n=2), and new isolate genomes (n=5), totaling 349 SAR116 genomes. Phylogenomic investigation supported the division of SAR116 into three distinct subclades, each with additional structure totalling 15 monophyletic groups. Our SAR116 isolates belonged to three groups within subclade I representing distinct genera with different morphologies and varied phenotypic responses to salinity and temperature. Overall, SAR116 genomes encoded differences in vitamin and amino acid synthesis, trace metal transport, and osmolyte synthesis and transport. They also had genetic potential for diverse sulfur oxidation metabolisms, placing SAR116 at the confluence of the organic and inorganic sulfur pools. SAR116 subclades showed distinct patterns in habitat preferences across open ocean, coastal, and estuarine environments, and three of our isolates represented the most abundant coastal and estuarine subclade. This investigation provides the most comprehensive exploration of SAR116 to date anchored by new culture genomes and physiology.
Alternative splicing in 300 human cells and tissues is sparsely distributed and strongly associated with cell identity
Authors: Liang Angelita , Wilkins Marc R
Published: January 2025

In eukaryotes, alternative splicing allows RNA sequences to be changed as required in their cellular context. While the use of gene expression without considering transcript isoforms is commonly used to classify cell types, the significance of alternative splicing to cell identity is not fully understood. Using data-driven classification and clustering methods, we analysed the RNA Atlas, an ultra-deep total RNA and polyA capture sequencing dataset obtained from 300 different types of cell and tissue samples across the human body. We show that splice inclusion values can distinguish cell identity effectively, and sometimes better than gene expression. These specific differences in alternative splicing are biologically relevant, with many splice events expressed in a limited number of cell types. We show that distributions of splice inclusion levels are “sparser” than gene expression levels, being discontinuous and containing outliers – akin to a control dial with a few settings. Gene expression levels display finer variation, taking on a diverse range of values akin to a control dial with many increments. Complementary analysis of comparable RNA-Seq samples from the ENCODE dataset yielded consistent results regardless of the splice event or gene expression quantification tools used, as well as the choice of RiboMinusTMor polyA capture-derived RNA libraries. Our results highlight the utility of alternative splicing data in data-driven and functional analyses, and the unique relationship between alternative splicing and cell identity.
Simulating Longitudinal Single-cell RNA Sequencing Data with RESCUE
Authors: Wynn Elizabeth A. , Mould Kara J. , Vestal Brian E. , Moore Camille M.
Published: January 2025

As single-cell RNA-sequencing (scRNA-seq) becomes more widely used in transcriptomic research, complex experimental designs, such as longitudinal studies, become increasingly feasible. Longitudinal scRNA-seq enables the study of transcriptomic changes over time within specific cell types, yet guidance on analytical approaches and resources for study planning, such as power analysis, remains limited. Data simulation is a valuable tool for evaluating analysis method performance and informing study design decisions, including sample size selection. Currently, most scRNA-seq simulation methods simulate cells for a single sample, thus ignoring the between-sample and between-subject variability inherent to longitudinal scRNA-seq data. Here, we introduce RESCUE (REpeated measures Single Cell RNA-seqUEncing data simulation), a novel method that simulates longitudinal scRNA-seq data using a gamma-Poisson frame-work and incorporates additional variability between samples and subjects. We demonstrate our method’s ability to reproduce important data properties and demonstrate its application in study planning. RES-CUE is implemented as an R package and is available athttps://github.com/ewynn610/RESCUE.
Toxigenecity and Virulence variations ofPseudomonas aeruginosafrom out-patients hospitals in Southwest Nigeria
Authors: Akinboboye Olusola Arinola , Akingbade Olusola Abiodun , Motayo Babatunde O , Olajumoke Akinleye Eunice Folusho , Sesan Aina Emmanuel , Samuel Idowu Adeyemi , Amuche Ede Dorathy , Omotunde Nimotalahi , Amuche Ede Dorathy
Published: January 2025

Background

Multidrug resistant (MDR)Pseudomonas aeruginosaisolates harboring genes for virulence and antibiotic resistance, have grown more prevalent lately. These strains pose a major threat to the general population, especially in tertiary care settings. There is a paucity of information on toxigenic and virulence diversity of multidrug resistantP. aeruginosain Nigeria, hence, the need to characterize and determine the variations of the virulence genes.

Methods

Six hundred clinical samples from different anatomical sites were collected aseptically from Lagos University Teaching Hospital (LUTH), University of Medical Sciences, Ondo (UNIMED) and Federal Medical Centre, Abeokuta (FMC).Pseudomonas aeruginosawas isolated using cetrimide agar identified using biochemical tests. Antibiotic sensitivity was done by disc diffusion method. Protease, phospholipase C (lecithinase), caseinase and gelatinase presence were assayed for. Genomic DNA was extracted fromP. aeruginosaisolates and screened for the presence of N-Acetylneuraminate synthase (NaN), Elastase B (LasB), Exotoxin A (ExoA), Exoenzyme S (ExoS) and Exoenzyme U (ExoU) virulence genes by PCR.

Results

Three hundred and sixty bacterial isolates identified from clinical samples are as follows:Pseudomonas aeruginosa(11.3%),Escherichia coli(18.0%),Klebsiella pneumoniae(14.3%),Staphylococcus aureus(10.2%),Proteus mirabilis(3.2%),Streptococcus pnuemoniae(2.3%), Enterobacter aerogenes(0.5%) andAcinetobacter baumanni(0.1%). Enzymes detected in theP. aeruginosaisolates were Phospholipase C (77.9%), caseinase (83.9%), gelatinase (98.5%) and protease (88.2%). TheP. aeruginosaisolates were all resistant to ampicillin and cloxacillin; 26 (38.2 %) strains exhibited multidrug resistance. VirulenceLasB elastase gene was detected in all 14 multi resistantP. aeruginosa, ExoA was detected in 5, ExoS in 4, ExoU in 5 and NaN in 4 isolates: Four (28.6%)

Conclusion

The study confirmed presence and variations of toxic genes inPseudomonas aeruginosaisolated from all the three tertiary hospitals.
MULTIPLE PRIMARY CANCERS IN LUNG CANCER PATIENTS IN ATLANTIC CANADA AND IMPLICATIONS FOR SCREENING AND MANAGEMENT
Authors: Coyle Kassandra M. , Gao Yingtong , Murphy Rowan E. , Martinez Victor , Goodarzi Aaron , Dellaire Graham , Wallace Alison
Published: January 2025

Background

Lung cancer incidence rates in Atlantic Canada significantly exceed those of other Canadian provinces, with underlying causes remaining poorly understood. This regional disparity suggests potential genetic or environmental factors unique to Atlantic Canada. Here we present for the first-time data indicating that accompanying the high lung cancer rates in Atlantic Canada is an intricate landscape of multiple malignancies. This represents a phenomenon of multiple primary cancers that is unprecedented in the literature and presents unique challenges in the diagnosis and treatment of individuals diagnosed with lung cancer in Atlantic Canada.

Methods

We performed a retrospective chart review of 1,151 patients referred to the thoracic surgeons at the Queen Elizabeth II Health Sciences Centre between 2019 and 2023 with the new diagnosis of lung cancer. The primary focus was to assess the incidence of multiple primary cancers (as documented in pathology reports) and analyze demographic, clinical, and geographical factors through Fisher’s exact tests and Student’s t-tests.

Results

Forty-three precent (43.3%) of Atlantic Canadian patients presented with multiple primary cancers. Of 1,949 cancers identified, 1,440 were of primary lung origin. Sixty percent (60.0%) of our cohort was female. Thirty-one percent (31.3%) were self-reported current tobacco smokers, 50.3% were former tobacco smokers, and 18.4% reported no tobacco use. The average age and body mass index (BMI) at first diagnosis was similar between those patients with and without multiple primary cancers, averaging 67 years and 27.7 kg/m2, respectively. Thirty-nine percent (39.2%) of our study participants lived in more densely populated centers (>100,000 people), 10.2% in intermediate centers (∼98,000 people) and 50.7% in more sparsely populated centers (<55,000 people).

Conclusions

We have uncovered an unprecedented phenomenon in Atlantic Canada of a cohort of patients with lung cancer having multiple primary cancers of different tissue origin at a rate that is four times greater than previously described. The implications of this study are that Atlantic Canadians are disproportionately burdened with cancers of multiple origins that are not explained by smoking rates or BMI. Future studies will investigate the contribution of genetic predisposition, environmental exposures, and socioeconomic factors unique to Atlantic Canada underlying the high rates of multiple primary cancers documented here in a lung cancer cohort. Such studies will be essential to develop effective cancer prevention and screening strategies in Atlantic Canada.
An at-home Plant Physiology laboratory applied to dark-induced leaf senescence by college students and science teachers
Authors: Soumen Shejal , Kolstoe Alexander J. , Duncan Nora E. , Termolen Isaac P. , Solloway Amanda M. , Lu Yan
Published: January 2025

Dark-induced leaf senescence is an extreme example of leaf senescence induced by light deprivation. Prolonged dark treatments of individual leaves result in chlorophyll degradation, macromolecule catabolism, and reduction of photosynthesis. In this work, we described an at-home "Dark-induced Leaf Senescence" laboratory exercise for a junior-level undergraduate Plant Physiology course. To perform the dark-induced senescence assay on attached leaves, students may cover individual leaves of an outdoor plant with aluminum foils and record the leaf morphology with controlled vocabularies for ~9 days. To perform senescence assays on detached leaves, the students may incubate detached leaves in various aqueous solutions (e.g., tap water, sucrose solution, alkali solution, and acid solution) either in the dark or under natural light, and then record the leaf morphology with controlled vocabularies for ~9 days. This laboratory exercise provides hands-on opportunities for students to understand the relationships among sunlight, chlorophyll, and photosynthesis, in the comfort of students' own homes. Specifically, it helps students to comprehend intrinsic and dark-induced leaf senescence mechanisms, the effects of sugars on leaf senescence, and the importance of optimal pH to plant health. This laboratory exercise can be adapted to support inquiry-based learning or be implemented in a middle or high school classroom.
The mutation rate of SARS-CoV-2 is highly variable between sites and is influenced by sequence context, genomic region, and RNA structure
Authors: Haddox Hugh K. , Angehrn Georg , Sesta Luca , Jennings-Shaffer Chris , Temple Seth D. , Galloway Jared G. , DeWitt William S. , Bloom Jesse D. , Matsen Frederick A. , Neher Richard A.
Published: January 2025

RNA viruses like SARS-CoV-2 have a high mutation rate, which contributes to their rapid evolution. The rate of mutations depends on the mutation type (e.g.,A→C,A→G, etc.) and can vary between sites in the viral genome. Understanding this variation can shed light on the mutational processes at play, and is crucial for quantitative modeling of viral evolution. Using the millions of available SARS-CoV-2 full-genome sequences, we estimate rates of synonymous mutations for all 12 possible nucleotide mutation types and examine how much these rates vary between sites. We find a surprisingly high level of variability and several striking patterns: the rates of four mutation types suddenly increase at one of two gene boundaries; the rates of most mutation types strongly depend on a site’s local sequence context, with up to 56-fold differences between contexts; consistent with a previous study, the rates of some mutation types are lower at sites engaged in RNA secondary structure. A simple log-linear model of these features explains ∼15-60% of the fold-variation of mutation rates between sites, depending on mutation type; more complex models only modestly improve predictive power out of sample. We estimate the fitness effect of each mutation based on the number of times it actually occurs versus the number of times it is expected to occur based on the model. We identify several small regions of the genome where synonymous or noncoding mutations occur much less often than expected, indicative of strong purifying selection on the RNA sequence that is independent of protein sequence. Overall, this work expands our basic understanding of SARS-CoV-2’s evolution by characterizing the virus’s mutation process at the level of individual sites and uncovering several striking mutational patterns that arise from unknown mechanisms.