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Introduction: Patients with cancer have an increased risk of developing venous thromboembolism (VTE), and subsequently a higher risk of bleeding secondary to anticoagulants. Low-molecular weight heparin (LMWH) has been the standard of care for these patients, with emerging data on the use of direct oral anticoagulants (DOACs). The primary objective of the study was to determine the prevalence of major bleeding events in cancer patients taking DOACs or LMWH for VTE. Secondary objectives included the rate of first VTE recurrence and the effect of concomitant antiplatelet agents and/or significant drug interactions on major bleeding or recurrent VTE. Materials and methods: Using the electronic medical record at the University of Rochester Medical Center, we retrospectively identified adult patients with active malignancy who had a diagnosis of VTE requiring therapeutic anticoagulation within the study period of July 1st, 2015 to June 1st, 2019. Patients were excluded if they were receiving prophylactic doses of LMWH per the institution VTE guidelines. Data on anticoagulant medications were collected as well as information on major bleeding and recurrent VTE events. Results and conclusions: There is insufficient evidence of difference in risk of major bleeding among patients who received a DOAC vs LMWH (cause-specific hazard ratio (HR) = 0.77, 95% CI 0.29–2.04, P = 0.60). There was also no evidence of a difference in risk of recurrent VTE between patients who received DOAC vs. LMWH (cause-specific HR = 0.98, 95% CI 0.15–6.26, P = 0.98). These results suggest that DOACs are not significantly less safe than LMWH for patients with cancer.

Abstract The demands for dental materials continue to grow, driven by the desire to reach a better performance than currently achieved by the available materials. In the dental restorative ceramic field, the structures evolved from the metal-ceramic systems to highly translucent multilayered zirconia, aiming not only for tailored mechanical properties but also for the aesthetics to mimic natural teeth. Ceramics are widely used in prosthetic dentistry due to their attractive clinical properties, including high strength, biocompatibility, chemical stability, and a good combination of optical properties. Metal-ceramics type has always been the golden standard of dental reconstruction. However, this system lacks aesthetic aspects. For this reason, efforts are made to develop materials that met both the mechanical features necessary for the safe performance of the restoration as well as the aesthetic aspects, aiming for a beautiful smile. In this field, glass and high-strength core ceramics have been highly investigated for applications in dental restoration due to their excellent combination of mechanical properties and translucency. However, since these are recent materials when compared with the metal-ceramic system, many studies are still required to guarantee the quality and longevity of these systems. Therefore, a background on available dental materials properties is a starting point to provoke a discussion on the development of potential alternatives to rehabilitate lost hard and soft tissue structures with ceramic-based tooth and implant-supported reconstructions. This review aims to bring the most recent materials research of the two major categories of ceramic restorations: ceramic-metal system and all-ceramic restorations. The practical aspects are herein presented regarding the evolution and development of materials, technologies applications, strength, color, and aesthetics. A trend was observed to use high-strength core ceramics type due to their ability to be manufactured by CAD/CAM technology. In addition, the impacts of COVID-19 on the market of dental restorative ceramics are presented.

Cyberattacks have been rapidly increasing over the years, resulting to big financial losses to businesses for recovery, regulatory sanctions, as well as collateral damages, such as reputation and trust. In this respect, the maritime sector, which until now was considered safe due to the lack of Internet connectivity and the isolated nature of ships in the sea, is showing a 900% increase in cybersecurity breaches on operational technology as it enters the digital era. Although some research is being conducted in this area, maritime cybersecurity has not been deeply investigated. Hence, this paper provides a close investigation of the landscape of cybersecurity in the maritime sector with the aim of highlighting security problems and challenges. First, it explores the systems available on ships that could be targeted by attackers, their possible vulnerabilities that an attacker could exploit, the consequences if the system is accessed, and actual incidents. Then, it describes and analyses possible mitigation actions that can be utilised in advance to prevent such attacks. Finally, several challenges and open problems are discussed for future research.

Peripheral edema (i.e., lower limb swelling) can cause pain, weakness, and limited range of motion. However, few studies have examined its prevalence in the U.S. or its association with demographics, comorbidities, activity, or mobility. This study used data from the Health and Retirement Study, a nationally representative longitudinal survey of U.S. adults (age 51+/ N = 19,988 for 2016), to evaluate time trends and correlates of peripheral edema using weighted descriptive statistics and logistic regressions, respectively. Peripheral edema was assessed with the question "Have you had… // Persistent swelling in your feet or ankles?" The weighted prevalence of edema among older U.S. adults was 19% to 20% between 2000 and 2016. Peripheral edema was associated with older age, female sex, non-white race, low wealth, obesity, diabetes, hypertension, pain, low activity levels, and mobility limitations (odds ratios ranging from 1.2-5.6; p-values ≤0.001). This study provides the first estimates of national prevalence and correlates of peripheral edema among older Americans. Peripheral edema is common and strongly associated with comorbidities, pain, low activity levels, and mobility limitations, and disproportionately affects poorer and minority groups. Peripheral edema should be a focus of future research in order to develop novel and cost-effective interventions.

Indigestible fractions of dry matter (iDM) and neutral detergent fiber (iNDF) in the feed of ruminants are mainly estimated by in situ incubation time with regard to particle size and textile types. Samples of alfafa, ground into three particle sizes, were analyzed. Samples, processed in a Willey mill with 1.0; 2.0 and 3.0 mm sieve pores, were conditioned in F57 (Ankon®), nonwoven (100 g m-2) and polyester textile bags measuring 4 x 5 cm. Material was divided into 13 incubation periods and 8 replications, and incubated in the rumen of two multiparous cows adapted to a 70:30 diet, roughage:concentrate, respectively, for 288h. iDM and iNDF rates were evaluated sequentially for non-digested percentage and data underwent analysis of variance (ANOVA p < 0.05); means were compared by tukey’s test (p < 0.05). Particle size and material employed for incubation affected iDM and iFDN at the initial periods of incubation (p < 0.05).

Current paper assesses the relationship between in situ incubation time, particle size and types of materials on estimates of indigestible fractions of dry matter (iDM) and neutral detergent fiber (iNDF) in cattle feed and feces. Samples of soybean meal, alfalfa and feces of cattle fed on high concentrated diets were analyzed. The samples were processed in a Wiley mill, with 0.5 and 1.5 mm porosity for food and 1.5 mm for feces, which were packed in 4 × 5 cm F57 bags (Ankon®) and nonwoven fabric (100 g m-2) of two brands. The material was divided into two groups, with and without treatment with acetone, and five replications. Samples of each group were incubated in the rumen of a multiparous cow adapted to a 70:30 diet, forage:concentrate diet, for 240 hours. The iDM and iNDF levels were evaluated sequentially to interpret the percentages of undigested material. Data underwent analysis of variance (ANOVA, p < 0.05) and means were compared by Tukey’s test (p < 0.05). Material, size of particles and incubated material affected iDM and iNDF estimates.

Water deficiency during the dry seasons influences the relationship between water and gas exchange in tropical grasses, reducing their productive potential. In addition, the phosphorus (P) deficiency Brazilian soils adds to the set of factors limiting crop production. In this context, the objective of this study was to evaluate the responses of different tropical forage species to phosphorus supplementation as mitigating the damage caused by water stress. Seeds of Urochloa brizantha cv. MG-4, Urochloa decumbens cv. Basilisk, Panicum maximum cv. Áries, Panicum maximum cv. Tanzânia and Paspalum atratum cv. Pojuca were germinated in pots containing 10 liters of red-yellow podzolic type soil. Experiments were conducted by combining levels of phosphorus, 8,0 and 100,0 mg of P dm-3, with two irrigation regimes, 100% and 40% replacement of transpired water. The biometric parameters, photosynthetic capacity, leaf water potential and soil chemical characteristics were evaluated, and the data was submitted to analysis of variance (ANOVA, p < 0.05), and subsequently the means were compared using a Tukey test (p < 0.05). The results showed for tropical grasses grown under water stress, there is a clear mitigating effect of phosphorus supplementation, especially on the maintenance of biomass growth.

Este estudo teve como objetivo verificar os efeitos do fósforo sobre a fotossíntese e o crescimento de Glycine max (L.) Merr. da cultivar 'Embrapa 48' sob deficiência hídrica controlada, em condições de casa de vegetação, considerando a hipótese de que suplementações de fósforo poderiam aumentar a tolerância das plantas ao déficit hídrico. Após a formação do primeiro par de folhas totalmente expandidas, foi iniciado o processo de indução de deficiência hídrica, utilizando dois regimes de irrigação, com 100% e 25% de reposição da evapotranspiração. A deficiência hídrica causou reduções significativas no acúmulo de massa seca, no potencial de água foliar, na condutância estomática e na assimilação líquida de CO2 em todos os tratamentos. Os resultados das trocas gasosas indicaram que a suplementação de P, na adubação de G. max da cultivar 'Embrapa 48', resultou em uma redução parcial dos efeitos da deficiência hídrica como suposto inicialmente. Porém, apenas em relação à biomassa do sistema radicular detectou-se algum efeito mitigador do P nas plantas sob deficiência hídrica.The objective of this study was to investigate the effects of phosphorus supplying on growth and photosynthesis of Glycine max (L.) Merr. cv. 'Embrapa 48', cultivated under water deficit in greenhouse conditions, taking into account the hypothesis that phosphorus suply could improve plant tolerance to drough. After the development of the first pair of totally expanded leaves, it was initiate the process of induction of water deficit, using two irrigation levels, 100% and 25% of evapotranspiration replacement Water deficit reduced dry mass, leaf water potential, stomatal conduction, and net CO2 assimilation regardless P supply. The results of the photosynthetic analysis indicated that phosphorus supply was just partially effective to reduce water deficit effects on G. max cv. 'Embrapa 48'. However, additional P supply influenced positively the production of plants root biomass under water deficit.

A deficiência é um dos principais fatores limitantes da agricultura restringindo as taxas fotossintéticas das plantas e seu crescimento. Pela importância econômica das pastagens tropicais, que também sofrem com o problema de períodos de deficiência hídrica, este estudoteve como objetivo verificar os efeitos de diferentes doses de fósforo (24, 50, 100 e 150 mg P dm-3) combinadas com dois regimes de irrigação (com e sem déficit hídrico) sobre as trocas gasosasfoliares e o crescimento de B. brizantha cv. MG-5 Vitória. O experimento foi realizado em casade-vegetação, com quatro repetições por tratamento, e a deficiência hídrica foi imposta por suspensão da irrigação em plantas com 50 dias de idade após a germinação. Um tratamentocontrole foi mantido com plantas irrigadas, diariamente, com as mesmas concentrações de P. O fósforo influenciou positivamente o crescimento das plantas, sobretudo o número de novos perfilhos e área foliar. A deficiência hídrica causou reduções significativas da condutância estomática e assimilação líquida de CO2 em todos os tratamentos. Entretanto, os resultados das trocas gasosas indicaram que, efetivamente, a suplementação de P na adubação de B. brizantha cv. MG-5 pode levar à recuperação mais eficiente da fotossíntese das plantas após um período de deficiência hídrica.Water deficit is the most constraining factor on crop yield, reducing both plant photosynthesis and growth. Because of the economical importance of tropical grasses, which also are affected by water shortage, the objective of this study was to test the effects of phosphorus supply (24, 50, 100 and 150 mg P dm-3)in Brachiaria brizantha cv. MG-5 Vitória plants cultivated under water deficit. Plants were evaluated taking into account leaf gas exchanges and crop yield parameters. The study was carried out under greenhouse conditions, and the water deficit was imposed by irrigation withholding until the first sign of leaf wilting in 50-days-old plants. A control treatment was maintained, with plants under daily irrigation and the same P concentrations. Phosphorus supplying causedpositive effects on plant growth parameters, mainly in leaf area and new shoot formation. Water deficit reduced stomatal conduction and net CO2 assimilation in all treatments. However, a significant improvement was observed in gas exchange recovering after re-hydration in the plants with additional P supplying, which could support faster growth improvements after a dry season.

We investigated soil carbon characteristics of a Fluvisol as influenced by different ped sizes and crop types for a period of 10 years in Owerri, Southeastern Nigeria. The experimental design was a split-split plot arranged in a randomized complete block design, with tillage technique serving as main plot; crop regime was the split plot and NPK fertilizer as split – split plot. Tillage techniques used were conventional tillage (CT) and minimum tillage (MT) while maize and soybean were used as crop types. Soil samples were collected at 20 cm depth with soil auger while core samples with cores partitioned 0 – to 10 – and 10 – to – 20 cm depth intervals were used to obtain soil samples for bulk density determinations. Routine laboratory analysis were conducted on soil samples and obtained data on crop and soil were statistically analyzed using analysis of variance (ANOVA) and means were separated using least significant differences (LSD) at P < 0.05. Aggregate size distribution was significantly (P < 0.05) influenced by tillage and crop type at 0 – 10 cm depth and soil carbon varied significantly (P < 0.05) with aggregate–size forms, tillage technique and crop species. Prediction of carbon behaviour in floodplain soils should take into consideration variability in ped forms, tillage method and crop types. J. Appl. Sci. Environ. Manage. June, 2012, Vol. 16 (2)174 - 179

A deficiência é um dos principais fatores limitantes da agricultura restringindo as taxas fotossintéticas das plantas e seu crescimento. Pela importância econômica das pastagens tropicais, que também sofrem com o problema de períodos de deficiência hídrica, este estudo teve como objetivo verificar os efeitos de diferentes doses de fósforo (24, 50, 100 e 150 mg P DM-3) combinadas com dois regimes de irrigação (com e sem déficit hídrico) sobre as trocas gasosas foliares e o crescimento de B. brizantha cv. MG-5 Vitória. O experimento foi realizado em casa-de-vegetação, com quatro repetições por tratamento, e a deficiência hídrica foi imposta por suspensão da irrigação em plantas com 50 dias de idade após a germinação. Um tratamento-controle foi mantido com plantas irrigadas, diariamente, com as mesmas concentrações de P. O fósforo influenciou positivamente o crescimento das plantas, sobretudo o número de novos perfilhos e área foliar. A deficiência hídrica causou reduções significativas da condutância estomática e assimilação líquida de CO2 em todos os tratamentos. Entretanto, os resultados das trocas gasosas indicaram que, efetivamente, a suplementação de P na adubação de B. brizantha cv. MG-5 pode levar à recuperação mais eficiente da fotossíntese das plantas após um período de deficiência hídrica.

Background: Case tracking (following-up prior patient encounters) can help inform future clinical decisions and supplement experiential learning. Internal medicine subspecialty fellows see a high volume of patients and need to become subject matter experts within a short time frame, yet little is known about their specific needs and motivations around case tracking., Objective: The objective of this study was to explore internal medicine subspecialty fellows' motivations, preferences, and practices around case tracking., Methods: We conducted interviews with internal medicine subspecialty fellows at a single academic medical center during the 2022-2023 academic year. Interviews were analyzed using qualitative content analysis., Results: 22 fellows were interviewed for our study. We found that most fellows engaged in case tracking with lists in the electronic health record (EHR). Fellows primarily tracked cases of clinical uncertainty and patients with specific diseases or conditions of interest. Fellows sought information on patients' health outcomes, results, and clinical notes. Motivations for tracking were predominantly related to curiosity, professional growth, and development of practice patterns. Barriers to case tracking included time, challenges maintaining patient lists, losing track of patients, and lack of motivation to develop and maintain an organized system., Conclusions: Internal medicine subspecialty fellows engaged in and valued case tracking as a way to supplement their experiential learning. Our study highlighted their current practices, motivations, preferences, and challenges related to case tracking. We plan to use these findings to help develop an EHR-embedded dashboard to facilitate case tracking among subspecialty fellows.

Background and Aims: Primary sclerosing cholangitis (PSC) is a chronic inflammatory liver disorder without effective medical treatment which is characterized by inflammation and fibrotic structures around the bile ducts. Biliary epithelial cells (cholangiocytes) are the target and potential disease drivers in PSC, yet little is known if cholangiocytes from PSC patients differ from non-PSC controls. To characterize cholangiocytes at early rather than end-stage disease, cholangiocyte organoids (COs) were derived from diseased bile ducts of PSC patients and compared to organoids generated from disease controls., Methods: Cholangiocytes were obtained during endoscopic retrograde cholangiopancreatography (ERCP) brushing of diseased bile duct areas and expanded as organoids using previously established culture methods. Stable CO lines were analyzed for cell type identity, basic cholangiocyte function, and transcriptomic signature., Results: We demonstrate that cholangiocytes, derived from the damaged area within the bile ducts of PSC patients, can be expanded in culture without displaying functional or genetic disease-related features. We further show that COs from patients who later were diagnosed with dysplasia exhibit higher expression of the cancer-associated genes PGC, FXYD2, MIR4435-2HG, and HES1., Conclusions: Our results demonstrate that PSC organoids are largely similar to control organoids after culture and highlight the significance of COs as a tool for regenerative medicine approaches as well as their potential for discovering new potential biomarkers for diagnosing cholangiocarcinoma., (© 2024. The Author(s).)

The topic of this review is venous thromboembolism (VTE) during pregnancy and postpartum. The following topics will be addressed: epidemiology and pathophysiology of VTE in pregnancy and postpartum, diagnostic considerations for VTE in pregnancy, indications for prophylactic and therapeutic anticoagulation in pregnancy and postpartum, choice of anticoagulation in pregnancy and breastfeeding, anticoagulation management during labor and delivery, and anticoagulation considerations for assisted reproductive technology., (© 2024 The Author(s).)

Background: Despite similar numbers of women and men in internal medicine (IM) residency, women face unique challenges. Stereotype threat is hypothesized to contribute to underrepresentation of women in academic leadership, and exploring how it manifests in residency may provide insight into forces that perpetuate gender disparities., Objective: To quantify the prevalence of stereotype threat in IM residency and explore experiences contributing to that stereotype threat., Design: We used a mixed methods study design. First, we surveyed IM residents using the Stereotype Vulnerability Scale (SVS) to screen for stereotype threat. Second, we conducted focus groups with women who scored high on the SVS to understand experiences that led to stereotype threat., Participants: The survey was sent to all IM residents at University of California, San Francisco (UCSF), in September-November 2019. Focus groups were conducted at UCSF in Spring 2020., Approach: The survey included an adapted version of the SVS. For focus groups, we developed a focus group guide informed by literature on stereotype threat. We used a thematic approach to data analysis. The mixed methods design enabled us to draw metainferences by integrating the two data sources., Key Results: Survey response rate was 61% (110/181). Women were significantly more likely than men to have a score indicating stereotype threat vulnerability (77% vs 0%, p < 0.001). Four themes from focus groups characterized women's experiences of gender bias and stereotype threat: gender norm tension, microaggressions and sexual harassment, authority questioned, and support and allyship., Conclusions: Gender-based stereotype threat is highly prevalent among women IM residents. This phenomenon poses a threat to confidence and ability to execute patient care responsibilities, detracting from well-being and professional development. These findings indicate that, despite robust representation of women in IM training, further attention is needed to address gendered experiences and contributors to women's vulnerability to stereotype threat., (© 2023. The Author(s).)

The myeloid transcription factor CEBPA is recurrently biallelically mutated (i.e., double mutated; CEBPA DM ) in acute myeloid leukemia (AML) with a combination of hypermorphic N-terminal mutations (CEBPA NT ), promoting expression of the leukemia-associated p30 isoform, and amorphic C-terminal mutations. The most frequently co-mutated genes in CEBPA DM AML are GATA2 and TET2, however the molecular mechanisms underlying this co-mutational spectrum are incomplete. By combining transcriptomic and epigenomic analyses of CEBPA-TET2 co-mutated patients with models thereof, we identify GATA2 as a conserved target of the CEBPA-TET2 mutational axis, providing a rationale for the mutational spectra in CEBPA DM AML. Elevated CEBPA levels, driven by CEBPA NT , mediate recruitment of TET2 to the Gata2 distal hematopoietic enhancer thereby increasing Gata2 expression. Concurrent loss of TET2 in CEBPA DM AML induces a competitive advantage by increasing Gata2 promoter methylation, thereby rebalancing GATA2 levels. Of clinical relevance, demethylating treatment of Cebpa-Tet2 co-mutated AML restores Gata2 levels and prolongs disease latency., (© 2023. Springer Nature Limited.)

Background: Few studies have evaluated diagnostic yield of small volume biopsies (SVB) for the diagnosis and management of follicular lymphoma (FL)., Methods: The authors performed a multi-institutional retrospective analysis of SVBs including fine-needle aspiration (FNA) and needle core biopsy (NCB) for initial FL diagnosis and suspected recurrence or transformation of FL. A total of 676 workups beginning with SVB were assessed for the mean number of biopsies per workup, the proportion of workups requiring multiple biopsies, and the proportion with a complete diagnosis including grade, on initial biopsy., Results: Compared to workups performed for question transformation/recurrence, those done for initial FL diagnosis were significantly more likely to require multiple biopsies (p < .01), had a higher mean number of biopsies per workup (1.7 vs. 1.1, absolute standardized difference = 1.1), and a lower complete diagnosis rate at initial biopsy (39% vs. 56%). At initial FL diagnosis, NCB +/- FNA was associated with fewer biopsies per workup compared to FNA +/- CB (1.2 vs. 1.9), fewer workups requiring multiple biopsies (23% vs. 83%), and a higher complete diagnosis rate (71% vs. 18%). In contrast, during assessment for transformation/recurrence, NCB and FNA showed a similar mean number of biopsies per workup (1.2 vs. 1.2) and few workups required multiple biopsies (6% vs. 19%)., Conclusions: SVB at initial FL diagnosis often required additional biopsies to establish a complete diagnosis. In contrast, when assessing for transformed/recurrent FL, additional biopsies were generally not obtained regardless of SVB type, suggesting that in these clinical settings SVB may be sufficient for clinical decision-making., (© 2022 The Authors. Cancer Cytopathology published by Wiley Periodicals LLC on behalf of American Cancer Society.)

Small-volume biopsies (SVBs) including fine-needle aspiration (FNA), cell block, and needle core biopsies (NCB) are increasingly utilized to diagnose and guide the clinical management of lymphoma. We established a multi-institutional interdisciplinary collaboration of cytopathologists, hematopathologists, and oncologists focused on the role of SVB in the management of patients with follicular lymphoma (FL). To assess the performance characteristics of SVB in this setting, we evaluated all consecutive SVBs performed for clinical indications of initial diagnosis, recurrence, or transformation of FL over a 5-year period and focused on the 182 that had at least one subsequent biopsy within 3 months as part of the same clinical work-up. The most common outcome of a subsequent biopsy as part of the same clinical work-up was a more specific diagnosis usually assigning the pathologic grade (111/182, 61%), followed by a complete agreement with the SVB (24/182, 13%), and change from nondiagnostic on initial biopsy to diagnostic on subsequent biopsy (21/182, 12%). A minority resulted in a diagnostic change from benign to lymphoma (17/182, 9%), a change in FL grade (5/182, 3%), or change in the lymphoma diagnostic category (4/182, 2%). There were no cases where an initial diagnosis of lymphoma was overturned. The distribution of discrepancies was similar across initial SVB types (FNA, FNA + cell block, NCB with or without FNA). Tissue limitations were noted in a minority of cases (53/182, 29%) and were enriched among initially nondiagnostic biopsies (16/21, 76%). Flow cytometry immunophenotyping was performed in the majority of cases both at the first and last biopsy (147/182, 81%). SVB can be a powerful method to detect FL in various clinical indications, with discrepant cases mostly resulting from a refinement in the initial diagnosis., Competing Interests: Conflicts of Interest and Source of Funding: The authors have disclosed that they have no significant relationships with, or financial interest in, any commercial companies pertaining to this article., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

The bone marrow microenvironment provides indispensable factors to sustain blood production throughout life. It is also a hotspot for the progression of hematologic disorders and the most frequent site of solid tumor metastasis. Preclinical research relies on xenograft mouse models, but these models preclude the human-specific functional interactions of stem cells with their bone marrow microenvironment. Instead, human mesenchymal cells can be exploited for the in vivo engineering of humanized niches, which confer robust engraftment of human healthy and malignant blood samples. However, mesenchymal cells are associated with major reproducibility issues in tissue formation. Here, we report the fast and standardized generation of human mini-bones by a custom-designed human mesenchymal cell line. These resulting humanized ossicles (hOss) consist of fully mature bone and bone marrow structures hosting a human mesenchymal niche with retained stem cell properties. As compared to mouse bones, we demonstrate superior engraftment of human cord blood hematopoietic cells and primary acute myeloid leukemia samples and also validate hOss as a metastatic site for breast cancer cells. We further report the engraftment of neuroblastoma patient-derived xenograft cells in a humanized model, recapitulating clinically described osteolytic lesions. Collectively, our human mini-bones constitute a powerful preclinical platform to model bone-developing tumors using patient-derived materials.

Ribosomopathies constitute a range of disorders associated with defective protein synthesis mainly affecting hematopoietic stem cells (HSCs) and erythroid development. Here, we demonstrate that deletion of poly-pyrimidine-tract-binding protein 1 (PTBP1) in the hematopoietic compartment leads to the development of a ribosomopathy-like condition. Specifically, loss of PTBP1 is associated with decreases in HSC self-renewal, erythroid differentiation, and protein synthesis. Consistent with its function as a splicing regulator, PTBP1 deficiency results in splicing defects in hundreds of genes, and we demonstrate that the up-regulation of a specific isoform of CDC42 partly mimics the protein-synthesis defect associated with loss of PTBP1. Furthermore, PTBP1 deficiency is associated with a marked defect in ribosome biogenesis and a selective reduction in the translation of mRNAs encoding ribosomal proteins. Collectively, this work identifies PTBP1 as a key integrator of ribosomal functions and highlights the broad functional repertoire of RNA-binding proteins., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

Diagnosis of histologic transformation (HT) of follicular lymphoma (FL) requires tissue biopsy. While surgical biopsy represents the gold standard, less invasive procedures such as fine-needle aspiration biopsy (FNAB) and core needle biopsy (CNB) are frequently performed. In this retrospective multi-institutional study including 269 patients with FL and suspected HT, the median time from initial clinical suspicion to final diagnostic biopsy was similar whether the workup began with FNAB, CNB, or surgical biopsy (4, 9, and 6 days, respectively; p =.27), despite more subsequent biopsies performed following initial FNAB. Periprocedural complications were uniformly minimal. Biopsy-proven HT was more common in the initial surgery group and in workups including positron emission tomography/computed tomography (PET/CT). Our findings, derived from US academic centers with specialized procedural and pathology expertise, suggest that FNAB, CNB, and surgical biopsy are all viable initial diagnostic procedures that can inform clinical decision-making in select FL patients with suspected HT.

Background & Aims: Peroxisome proliferator-activated receptors (PPARs) are essential regulators of whole-body metabolism, but also modulate inflammation in immune cells, notably macrophages. We compared the effects of selective PPAR agonists to those of the pan-PPAR agonist lanifibranor in non-alcoholic fatty liver disease (NAFLD), and studied isoform-specific effects on hepatic macrophage biology., Methods: Lanifibranor or selective PPARα (fenofibrate), PPARγ (pioglitazone) and PPARδ (GW501516) agonists were therapeutically administered in choline-deficient, amino acid-defined high-fat diet (CDAA-HFD)- and Western diet (WD)-fed mouse models of NAFLD. Acute liver injury was induced by carbon tetrachloride (CCl 4 ). The role of PPARs on macrophage functionality was studied in isolated hepatic macrophages, bone marrow-derived macrophages stimulated with palmitic acid, and circulating monocytes from patients with NAFLD., Results: Lanifibranor improved all histological features of steatohepatitis in CDAA-HFD-fed mice, including liver fibrosis, thereby combining and exceeding specific effects of the single PPAR agonists. Its potent anti-steatotic efficacy was confirmed in a 3D liver biochip model with primary cells. Infiltrating hepatic monocyte-derived macrophages were reduced following PPAR agonist administration, especially with lanifibranor, even after short-term treatment, paralleling improved steatosis and hepatitis. Lanifibranor similarly decreased steatosis, liver injury and monocyte infiltration in the WD model. In the acute CCl 4 model, neither single nor pan-PPAR agonists directly affected monocyte recruitment. Hepatic macrophages isolated from WD-fed mice displayed a metabolically activated phenotype. Lanifibranor attenuated the accompanying inflammatory activation in both murine palmitic acid-stimulated bone marrow-derived macrophages, as well as patient-derived circulating monocytes, in a PPARδ-dependent fashion., Conclusion: Pan-PPAR agonists combine the beneficial effects of selective PPAR agonists and may counteract inflammation and disease progression more potently. PPARδ agonism and lanifibranor directly modulate macrophage activation, but not infiltration, thereby synergizing with beneficial metabolic effects of PPARα/γ agonists., Lay Summary: Peroxisome proliferated-activated receptors (PPARs) are essential regulators of metabolism and inflammation. We demonstrated that the pan-PPAR agonist lanifibranor ameliorated all aspects of non-alcoholic fatty liver disease in independent experimental mouse models. Non-alcoholic fatty liver disease and fatty acids induce a specific polarization status in macrophages, which was altered by lanifibranor to increase expression of lipid handling genes, thereby decreasing inflammation. PPAR isoforms have differential therapeutic effects on fat-laden hepatocytes, activated hepatic stellate cells and inflammatory macrophages, supporting the clinical development of pan-PPAR agonists., Competing Interests: Conflict of interest VA, ED, CE and GW are employees of Inventiva. Work in FT's laboratory has received funding by Allergan, Galapagos, Inventiva and Bristol Myers Squibb. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2020 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Pediatric liver transplantation is often required as a consequence of biliary disorders because of the lack of alternative treatments for repairing or replacing damaged bile ducts. To address the lack of availability of pediatric livers suitable for transplantation, we developed a protocol for generating bioengineered biliary tissue suitable for biliary reconstruction. Our platform allows the derivation of cholangiocyte organoids (COs) expressing key biliary markers and retaining functions of primary extra- or intrahepatic duct cholangiocytes within 2 weeks of isolation. COs are subsequently seeded on polyglycolic acid (PGA) scaffolds or densified collagen constructs for 4 weeks to generate bioengineered tissue retaining biliary characteristics. Expertise in organoid culture and tissue engineering is desirable for optimal results. COs correspond to mature functional cholangiocytes, differentiating our method from alternative organoid systems currently available that propagate adult stem cells. Consequently, COs provide a unique platform for studies in biliary physiology and pathophysiology, and the resulting bioengineered tissue has broad applications for regenerative medicine and cholangiopathies.

Background: Faculty and students debate the fairness and accuracy of medical student clerkship grades. Group decision-making is a potential strategy to improve grading., Objective: To explore how one school's grading committee members integrate assessment data to inform grade decisions and to identify the committees' benefits and challenges., Design: This qualitative study used semi-structured interviews with grading committee chairs and members conducted between November 2017 and March 2018., Participants: Participants included the eight core clerkship directors, who chaired their grading committees. We randomly selected other committee members to invite, for a maximum of three interviews per clerkship., Approach: Interviews were recorded, transcribed, and analyzed using inductive content analysis., Key Results: We interviewed 17 committee members. Within and across specialties, committee members had distinct approaches to prioritizing and synthesizing assessment data. Participants expressed concerns about the quality of assessments, necessitating careful scrutiny of language, assessor identity, and other contextual factors. Committee members were concerned about how unconscious bias might impact assessors, but they felt minimally impacted at the committee level. When committee members knew students personally, they felt tension about how to use the information appropriately. Participants described high agreement within their committees; debate was more common when site directors reviewed students' files from other sites prior to meeting. Participants reported multiple committee benefits including faculty development and fulfillment, as well as improved grading consistency, fairness, and transparency. Groupthink and a passive approach to bias emerged as the two main threats to optimal group decision-making., Conclusions: Grading committee members view their practices as advantageous over individual grading, but they feel limited in their ability to address grading fairness and accuracy. Recommendations and support may help committees broaden their scope to address these aspirations.

Polypropylene meshes that are commonly used for inguinal hernia repair may trigger granulomatous foreign body reactions. Here, we show that asymptomatic patients display mesh-associated inflammatory granulomas long after surgery, which are dominated by monocyte-derived macrophages expressing high levels of inflammatory activation markers. In mice, mesh implantation by the onlay technique induced rapid and strong myeloid cell accumulation, without substantial attenuation for up to 90 days. Myeloid cells segregated into distinct macrophage subsets with separate spatial distribution, activation profiles, and functional properties, showing a stable inflammatory phenotype in the tissue surrounding the biomaterial and a mixed, wound-healing phenotype in the surrounding stromal tissue. Protein mass spectrometry confirmed the inflammatory nature of the foreign body reaction, as characterized by cytokines, complement activation, and matrix-modulating factors. Moreover, immunoglobulin deposition increased over time around the implant, arguing for humoral immune responses in association with the cell-driven inflammation. Intravital multiphoton microscopy revealed a high motility and continuous recruitment of myeloid cells, which is partly dependent on the chemokine receptor CCR2. CCR2-dependent macrophages are particular drivers of fibroblast proliferation. Thus, our work functionally characterizes myeloid cell-dependent inflammation following mesh implantation, thereby providing insights into the dynamics and mechanisms of foreign body reactions to implanted biomaterials.

Background & Aims: We performed an integrated analysis to identify microRNAs (miRNAs) and messenger RNAs (mRNAs) with altered expression in liver tumors from 3 mouse models of hepatocellular carcinoma (HCC) and human tumor tissues., Methods: We analyzed miRNA and mRNA expression profiles of liver tissues from mice with diethylnitrosamine-induced hepatocarcinogenesis, conditional expression of lymphotoxin alpha and lymphotoxin beta, or inducible expression of a Myc transgene (Tet-O-Myc mice), as well as male C57BL/6 mice (controls). miRNA mimics were expressed and miRNAs and mRNAs were knocked down in human (Huh7, Hep3B, JHH2) hepatoma cell lines; cells were analyzed for viability, proliferation, apoptosis, migration, and invasion. Cells were grown as xenograft tumors in nude mice and analyzed. We combined in silico target gene prediction with mRNA profiles from all 3 mouse models. We quantified miRNA levels in 146 fresh-frozen tissues from patients (125 HCCs, 17 matched nontumor tissues, and 4 liver samples from patients without cancer) and published human data sets and tested correlations with patient survival times using Kaplan-Meier curves and the log-rank test. Levels of NUSAP1 mRNA were quantified in 237 HCCs and 5 nontumor liver samples using the TaqMan assay., Results: Levels of the miRNA 193a-5p (MIR193A-5p) were reduced in liver tumors from all 3 mouse tumor models and in human HCC samples, compared with nontumor liver tissues. Expression of a MIR193A-5p mimic in hepatoma cells reduced proliferation, survival, migration, and invasion and their growth as xenograft tumors in nude mice. We found nucleolar and spindle-associated protein 1 (NUSAP1) to be a target of MIR193A-5p; HCC cells and tissues with low levels of MIR193A-5p had increased expression of NUSAP1. Increased levels of NUSAP1 in HCC samples correlated with shorter survival times of patients. Knockdown of NUSAP1 in Huh7 cells reduced proliferation, survival, migration, and growth as xenograft tumors in nude mice. Hydrodynamic tail-vein injections of a small hairpin RNA against NUSAP1 reduced growth of Akt1-Myc-induced tumors in mice., Conclusions: MIR193A-5p appears to prevent liver tumorigenesis by reducing levels of NUSAP1. Levels of MIR193A-5p are reduced in mouse and human HCC cells and tissues, leading to increased levels of NUSAP1, associated with shorter survival times of patients. Integrated analyses of miRNAs and mRNAs in tumors from mouse models can lead to identification of therapeutic targets in humans. The currently reported miRNA and mRNA profiling data have been submitted to the Gene Expression Omnibus (super-series accession number GSE102418)., (Copyright © 2018 AGA Institute. Published by Elsevier Inc. All rights reserved.)

The balance between self-renewal and differentiation is crucial for the maintenance of hematopoietic stem cells (HSCs). Whereas numerous gene regulatory factors have been shown to control HSC self-renewal or drive their differentiation, we have relatively few insights into transcription factors that serve to restrict HSC differentiation. In the present work, we identify ETS (E-twenty-six)-related gene (ERG) as a critical factor protecting HSCs from differentiation. Specifically, loss of Erg accelerates HSC differentiation by >20-fold, thus leading to rapid depletion of immunophenotypic and functional HSCs. Molecularly, we could demonstrate that ERG, in addition to promoting the expression of HSC self-renewal genes, also represses a group of MYC targets, thereby explaining why Erg loss closely mimics Myc overexpression. Consistently, the BET domain inhibitor CPI-203, known to repress Myc expression, confers a partial phenotypic rescue. In summary, ERG plays a critical role in coordinating the balance between self-renewal and differentiation of HSCs., (© 2015 Knudsen et al.; Published by Cold Spring Harbor Laboratory Press.)

In-depth molecular investigation of familial leukemia has been limited by the rarity of recognized cases. This study examines the genetic events initiating leukemia and details the clinical progression of disease across multiple families harboring germ-line CEBPA mutations. Clinical data were collected from 10 CEBPA-mutated families, representing 24 members with acute myeloid leukemia (AML). Whole-exome (WES) and deep sequencing were performed to genetically profile tumors and define patterns of clonal evolution. Germline CEBPA mutations clustered within the N-terminal and were highly penetrant, with AML presenting at a median age of 24.5 years (range, 1.75-46 years). In all diagnostic tumors tested (n = 18), double CEBPA mutations (CEBPAdm) were detected, with acquired (somatic) mutations preferentially targeting the C-terminal. Somatic CEBPA mutations were unstable throughout the disease course, with different mutations identified at recurrence. Deep sequencing of diagnostic and relapse paired samples confirmed that relapse-associated CEBPA mutations were absent at diagnosis, suggesting recurrence was triggered by novel, independent clones. Integrated WES and deep sequencing subsequently revealed an entirely new complement of mutations at relapse, verifying the presentation of a de novo leukemic episode. The cumulative incidence of relapse in familial AML was 56% at 10 years (n = 11), and 3 patients experienced ≥3 disease episodes over a period of 17 to 20 years. Durable responses to secondary therapies were observed, with prolonged median survival after relapse (8 years) and long-term overall survival (10-year overall survival, 67%). Our data reveal that familial CEBPA-mutated AML exhibits a unique model of disease progression, associated with favorable long-term outcomes., (© 2015 by The American Society of Hematology.)

Dyskeratosis Congenita (DC) is a heritable multi-system disorder caused by abnormally short telomeres. Clinically diagnosed by the mucocutaneous symptoms, DC patients are at high risk for bone marrow failure, pulmonary fibrosis, and multiple types of cancers. We have recapitulated the most common DC-causing mutation in the shelterin component TIN2 by introducing a TIN2-R282H mutation into cultured telomerase-positive human cells via a knock-in approach. The resulting heterozygous TIN2-R282H mutation does not perturb occupancy of other shelterin components on telomeres, result in activation of telomeric DNA damage signaling or exhibit other characteristics indicative of a telomere deprotection defect. Using a novel assay that monitors the frequency and extension rate of telomerase activity at individual telomeres, we show instead that telomerase elongates telomeres at a reduced frequency in TIN2-R282H heterozygous cells; this recruitment defect is further corroborated by examining the effect of this mutation on telomerase-telomere co-localization. These observations suggest a direct role for TIN2 in mediating telomere length through telomerase, separable from its role in telomere protection.

Background: Chromatin-Immunoprecipitation coupled with deep sequencing (ChIP-seq) is used to map transcription factor occupancy and generate epigenetic profiles genome-wide. The requirement of nano-scale ChIP DNA for generation of sequencing libraries has impeded ChIP-seq on in vivo tissues of low cell numbers., Results: We describe a robust, simple and scalable methodology for ChIP-seq of low-abundant cell populations, verified down to 10,000 cells. By employing non-mammalian genome mapping bacterial carrier DNA during amplification, we reliably amplify down to 50 pg of ChIP DNA from transcription factor (CEBPA) and histone mark (H3K4me3) ChIP. We further demonstrate that genomic profiles are highly resilient to changes in carrier DNA to ChIP DNA ratios., Conclusions: This represents a significant advance compared to existing technologies, which involve either complex steps of pre-selection for nucleosome-containing chromatin or pre-amplification of precipitated DNA, making them prone to introduce experimental biases.

The loss of regulation of cell proliferation is a key event in leukemic transformation, and the oncogene tribbles (Trib)2 is emerging as a pivotal target of transcription factors in acute leukemias. Deregulation of the transcription factor E2F1, normally repressed by CCAAT enhancer-binding protein α (C/EBPα)-p42, occurs in acute myeloid leukemia (AML), resulting in the perturbation of cell cycle and apoptosis, emphasizing its importance in the molecular pathogenesis of AML. Here we show that E2F family members directly regulate Trib2 in leukemic cells and identify a feedback regulatory loop for E2F1, C/EBPα, and Trib2 in AML cell proliferation and survival. Further analyses revealed that E2F1-mediated Trib2 expression was repressed by C/EBPα-p42, and in normal granulocyte/macrophage progenitor cells, we detect C/EBPα bound to the Trib2 promoter. Pharmacological inhibition of the cell cycle or Trib2 knockdown resulted in a block in AML cell proliferation. Our work proposes a novel paradigm whereby E2F1 plays a key role in the regulation of Trib2 expression important for AML cell proliferation control. Importantly, we identify the contribution of dysregulated C/EBPα and E2F1 to elevated Trib2 expression and leukemic cell survival, which likely contributes to the initiation and maintenance of AML and may have significant implications for normal and malignant hematopoiesis.

Transcription factors are key regulators of hematopoietic stem cells (HSCs) and act through their ability to bind DNA and impact on gene transcription. Their functions are interpreted in the complex landscape of chromatin, but current knowledge on how this is achieved is very limited. C/EBPα is an important transcriptional regulator of hematopoiesis, but its potential functions in HSCs have remained elusive. Here we report that C/EBPα serves to protect adult HSCs from apoptosis and to maintain their quiescent state. Consequently, deletion of Cebpa is associated with loss of self-renewal and HSC exhaustion. By combining gene expression analysis with genome-wide assessment of C/EBPα binding and epigenetic configurations, we show that C/EBPα acts to modulate the epigenetic states of genes belonging to molecular pathways important for HSC function. Moreover, our data suggest that C/EBPα acts as a priming factor at the HSC level where it actively promotes myeloid differentiation and counteracts lymphoid lineage choice. Taken together, our results show that C/EBPα is a key regulator of HSC biology, which influences the epigenetic landscape of HSCs in order to balance different cell fate options.

Acute myeloid leukemia (AML) develops via a multistep process involving several genetic and epigenetic events, which ultimately leads to the formation of a heterogeneous population of malignant cells, of which only a small subpopulation termed the leukemia initiating cell (LIC) is able to sustain the leukemia. The identity of the LIC is highly diverse and ranges from populations resembling hematopoietic stem cells or multipotent progenitors (MPPs) to more committed myeloid progenitors, and the question still remains whether this is a direct consequence of which cells are targets of the final transforming events. In this study, we use premalignant cells from a Cebpa mutant AML model, in which the LIC population resembles granulocyte-macrophage progenitors (GMPs), to show that premalignant GMPs undergo spontaneous immortalization with a high clonal frequency when cultured in vitro, suggesting that these cells constitute the target of the final transforming events. Furthermore, we show that premalignant GMPs are characterized by a distinct T cell gene expression signature correlating with an increased potential for differentiation toward the T cell lineage. These findings have implications for our understanding of the transcriptional wiring in premalignant myeloid progenitors and how this contributes to the development of AML., (Copyright © 2013 ISEH - Society for Hematology and Stem Cells. Published by Elsevier Inc. All rights reserved.)

Background: Transcription factors play a key role in lineage commitment and differentiation of stem cells into distinct mature cells. In hematopoiesis, they regulate lineage-specific gene expression in a stage-specific manner through various physical and functional interactions with regulatory proteins that are simultanously recruited and activated to ensure timely gene expression. The transcription factor CCAAT/enhancer binding protein α (C/EBPα) is such a factor and is essential for the development of granulocytic/monocytic cells. The activity of C/EBPα is regulated on several levels including gene expression, alternative translation, protein interactions and posttranslational modifications, such as phosphorylation. In particular, the phosphorylation of serine 248 of the transactivation domain has been shown to be of crucial importance for granulocytic differentiation of 32Dcl3 cells in vitro., Methodology/principal Findings: Here, we use mouse genetics to investigate the significance of C/EBPα serine 248 in vivo through the construction and analysis of Cebpa(S248A/S248A) knock-in mice. Surprisingly, 8-week old Cebpa(S248A/S248A) mice display normal steady-state hematopoiesis including unaltered development of mature myeloid cells. However, over time some of the animals develop a hematopoietic disorder with accumulation of multipotent, megakaryocytic and erythroid progenitor cells and a mild impairment of differentiation along the granulocytic-monocytic lineage. Furthermore, BM cells from Cebpa(S248A/S248A) animals display a competitive advantage compared to wild type cells in a transplantation assay., Conclusions/significance: Taken together, our data shows that the substitution of C/EBPα serine 248 to alanine favors the selection of the megakaryocytic/erythroid lineage over the monocytic/granulocytic compartment in old mice and suggests that S248 phosphorylation may be required to maintain proper hematopoietic homeostasis in response to changes in the wiring of cellular signalling networks. More broadly, the marked differences between the phenotype of the S248A variant in vivo and in vitro highlight the need to exert caution when extending in vitro phenotypes to the more appropriate in vivo context.

Tumor suppressor p53 is crucial for embryonic implantation through transcriptional up-regulation of uterine leukemia inhibitory factor (LIF). This article reports that p53 and estrogen receptor α were activated in endometrial tissues during implantation to coordinately regulate LIF production. By using human p53 knockin (Hupki) mice carrying a single nucleotide polymorphism (SNP) at codon 72 (arginine/proline), the arginine allele was demonstrated to produce higher uterine LIF levels during implantation than the proline allele. In humans, the diversity of haplotypes of the p53 gene has decreased during evolution, because the arginine allele, existing in only a subset of haplotypes, is under positive selection. This observation is consistent with previous results showing that the proline allele is enriched in patients undergoing in vitro fertilization (IVF). Studies with p63- and p73-knockout mice have demonstrated the involvement of p63 and p73 in female reproduction and their roles in egg formation and apoptosis (p63) and spindle checkpoint (p73) in female mice. Here, the role of p63 and p73 in human reproduction was investigated. Selected alleles of SNPs in p63 and p73 genes were enriched in IVF patients. These findings demonstrate that the p53 family members are involved in several steps to regulate female reproduction in mice and humans.

Currently there are several dozen human polymorphisms that have been loosely associated with cancer risk. Correlating such variants with cancer risk has been challenging, primarily due to factors such as genetic heterogeneity, contributions of diet and environmental factors, and the difficulty in obtaining large sample sizes for analysis. Such difficulties can be circumvented with the establishment of mouse models for human variants. Recently, several groups have modeled human cancer susceptibility polymorphisms in the mouse. Remarkably, in each case these mouse models have accurately reflected human phenotypes, and clarified the contribution of these variants to cancer risk. We recently reported on a mouse model for the codon 72 polymorphism in p53, and found that this polymorphism regulates the ability to cooperate with NF-kB and induce apoptosis. Here-in we present evidence that this polymorphism impacts the apoptotic function of p53 in a tissue-specific manner; such tissue-specific effects of polymorphic variants represent an added challenge to human cancer risk association studies. The data presented here support the premise that modeling human polymorphisms in the mouse represents a powerful tool to assess the impact of these variants on cancer risk, progression and therapy.

Caspases play a key role in the apoptotic pathway by virtue of their ability to cleave key protein substrates within the dying cell. Caspases are produced as inactive zymogens, and need to become proteolytically processed in order to become active. A key executioner caspase, caspase-3, has previously been found to exist in both the cytosol and the mitochondria. At the mitochondria, caspase-3 is associated with both the inner and outer mitochondrial membranes, where it interacts with heat shock proteins Hsp60 and Hsp10. Like caspase-3, a small portion of the p53 tumor suppressor protein is localized to mitochondria, particularly after genotoxic stress. p53 interacts with various members of the Bcl2 family at the mitochondria, and this interaction is key to its ability to induce apoptosis. In this study, we sought to determine the identity of other mitochondrial p53-interacting proteins. Using immunoprecipitation from purified mitochondria followed by mass spectrometry we identified caspase-3 as a mitochondrial p53-interacting protein. Interestingly, we find that tumor-derived mutant forms of p53 retain the ability to interact with mitochondrial caspase-3. Further, we find evidence that these mutant forms of p53 may interfere with the ability of procaspase-3 to become proteolytically activated by caspase-9. The combined data suggest that tumor-derived mutants of p53 may be selected for in tumor cells due to their ability to bind and inhibit the activation of caspase-3.

A common polymorphism at codon 72 in the p53 tumor suppressor gene encodes either proline (P72) or arginine (R72). Several groups have reported that in cultured cells, this polymorphism influences p53's transcriptional, senescence, and apoptotic functions. However, the impact of this polymorphism within the context of a living organism is poorly understood. We generated knock-in mice with the P72 and R72 variants and analyzed the tissues of these mice for apoptosis and transcription. In the thymus, we find that the P72 variant induces increased apoptosis following ionizing radiation, along with increased transactivation of a subset of p53 target genes, which includes murine Caspase 4 (also called Caspase 11), which we show is a direct p53 target gene. Interestingly, the majority of genes in this subset have roles in inflammation, and their promoters contain NF-κB binding sites. We show that caspase 4/11 requires both p53 and NF-κB for full induction after DNA damage and that the P72 variant shows increased interaction with p65 RelA, a subunit of NF-κB. Consistent with this, we show that P72 mice have a markedly enhanced response to inflammatory challenge compared to that of R72 mice. Our data indicate that the codon 72 polymorphism impacts p53's role in inflammation.

Reconstruction of phylogenetic relationships among recently diverged species is complicated by three general problems: segregation of polymorphisms that pre-date species divergence, gene flow during and after speciation, and intra-locus recombination. In light of these difficulties, the Y chromosome offers several important advantages over other genomic regions as a source of phylogenetic information. These advantages include the absence of recombination, rapid coalescence, and reduced opportunity for interspecific introgression due to hybrid male sterility. In this report, we test the phylogenetic utility of Y-chromosomal sequences in two groups of closely related and partially inter-fertile Drosophila species. In the D. bipectinata species complex, Y-chromosomal loci unambiguously recover the phylogeny most consistent with previous multi-locus analysis and with reproductive relationships, and show no evidence of either post-speciation gene flow or persisting ancestral polymorphisms. In the D. simulans species complex, the situation is complicated by the duplication of at least one Y-linked gene region, followed by intrachromosomal recombination between the duplicate genes that scrambles their genealogy. We suggest that Y-chromosomal sequences are a useful tool for resolving phylogenetic relationships among recently diverged species, especially in male-heterogametic organisms that conform to Haldane's rule. However, duplication of Y-linked genes may not be uncommon, and special care should be taken to distinguish between orthologous and paralogous sequences.

Incipient species in the early stages of divergence can provide crucial information about the genetic basis of reproductive isolation and the evolutionary forces that promote speciation. In this report, we describe two subspecies of Drosophila bipectinata that show a continuum of reproductive isolation. Crosses between strains of the same subspecies produce fully fertile offspring. At the same time, each subspecies harbors extensive variation for the degree of reproductive isolation from the other subspecies. The percentage of fertile hybrid males varies from 0 to 90%, depending on the origin of parental strains, indicating that the genes responsible for hybrid sterility are not fixed within either subspecies, or even within local populations. Reproductive isolation is non-transitive, so that the extent of hybrid sterility depends on the particular combination of strains. The two subspecies show little or no evidence of genetic differentiation at three nuclear loci, suggesting that they diverged very recently or continue to experience significant levels of gene flow. A hybrid zone between the two subspecies may exist in New Guinea and Northeastern Australia.