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Cardiac ventricular myocyte action potential dynamics are regulated by intricate and nonlinear interactions between the cell transmembrane potential and ionic currents and concentrations. Present technology limits the ability to measure transmembrane potential and multiple ionic currents simultaneously, which narrows the scope of experiments to provide a complete snapshot of the cardiac myocyte state. This limitation presents an obstacle for understanding how perturbations can trigger arrhythmias and more broadly how the myocyte responds to different conditions, such as changes in pacing rate or responses to drug treatment. In this study, we demonstrate that a data-assimilation approach can successfully reconstruct and predict the dynamics of a heterogeneous virtual cardiac ventricular myocyte population in the presence of parameter uncertainty. A population of heterogeneous cardiac ventricular myocytes is generated by varying ionic current conductance parameters, and additional observational uncertainty is mimicked by the addition of Gaussian noise to the transmembrane potential. We demonstrate that the data-assimilation approach accurately reconstructs transmembrane potential, with error less than the magnitude of the noise. Further, the data-assimilation approach successfully estimates the conductances of ionic currents generally with high accuracy and requiring low computational time. As a proof of concept, we apply the data-assimilation approach to reconstruct action potential dynamics from optical mapping experiments in an ex vivo isolated guinea pig heart. Critically, we demonstrate that the ionic conductance parameters estimated from a recording at one pacing frequency can accurately predict action potential dynamics at different rates., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Biophysical Society. Published by Elsevier Inc. All rights reserved.)

The aim of this work was to evaluate the presence of 40 pesticides in the PM 10 emitted by rural soils of the semiarid region of Argentina. Six agricultural soils for grain production under no till and with high use of pesticides (AG), 5 agricultural soils for forage and grain production under conventional tillage (AFG) and 5 unpaved rural roads (RR) were sampled. The PM 10 was generated using the Easy Dust Generator and it was collected with an electrostatic precipitator. The presence of 20 herbicides, 14 insecticides and 6 fungicides was analyzed in the soil and in the PM 10 . More than 70% of the pesticides analyzed were detected in the soil and in the PM 10 . All agricultural soils and 87% of RR soils showed at least one residue of pesticides. Multiresidues of pesticides were found in the 100% of PM 10 emitted by rural soils. The mean number of pesticides was higher in the PM 10 (7) than in the soil (5). Some pesticides were not detected in the soils but they were detected in the PM 10 (triticonazole, carbofuran, metsulfuron methyl) and vice versa. In general, the concentrations of herbicides were higher in the PM 10 than in the soil, while the concentrations of insecticides and fungicides were lower in the PM 10 than in the soil. These results suggest that the concentrations of pesticide in the PM 10 (inhalable fraction) should be used instead the concentrations of pesticide in the soil to calculate the exposure factor to pesticides by dust inhalation. This study provides the initial evidence of the presence of multiple pesticide residues in PM 10 emitted by rural soils under different land management. Also confirms that the PM 10 is a potential source of air contamination with pesticides. Future studies should be driven to measure the concentrations of pesticides and their dynamics in the PM 10 ., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

In both developed and developing countries, atmospheric pollution with particulate matter (PM) remains an important issue. Despite the health effects of poor air quality, studies on air pollution are often limited by the high costs of continuous monitoring and the need for extensive sampling. Furthermore, these particles are often enriched with potentially toxic trace elements and organic pollutants. This study evaluates both the composition of atmospheric dust accumulated during a certain timespan on Hedera helix and Senecio cineraria leaves and the potential for their use as bio-monitors. The test plants were positioned near automatic air quality monitoring stations at four different sites with respectively high, moderate and low traffic intensity. The gravimetric deposition of PM10 and PM2.5 on leaves was compared with data recorded by the monitoring stations and related to the weather conditions reported by Argentina's National Meteorological Service. To determine the presence of trace elements enriching the PM deposited on leaves, two analytical techniques were applied: XRF (not destructive) and ICP (destructive). The results indicated that only in the unpaved street location (site 2) did PM10 and PM2.5 concentrations (90 µg m -3 and 9 µg m -3 ) in the air exceed more than five times WHO guidelines (15 µg m -3 and 5 µg m -3 ). However, several trace elements were found to be enriching PM deposited on leaves from all sites. Predominantly, increased concentrations of Cd, Cu, Ti, Mn, Zn and Fe were found, which were associated with construction, traffic and unpaved street sources. Furthermore, based on its capability to sequester above 2800 µg cm -2 of PM10, 2450 µg cm -2 of PM2.5 and trace elements, Senecio cineraria can be taken into consideration for adoption as a bio-monitor or even for PM mitigation.

Background/aim: Renin-angiotensin system (RAS) is present in a diverse type of cells and plays an important role in lung physiology and pathophysiology. Angiotensin converting enzymes (ACE) are part of the RAS system. There are still controversies about the association of I/D polymorphisms of ACE1 with COVID-19 severity. The goal of the study was to determine whether there is an association of the I/D polymorphism with severity of COVID-19 in Mexican patients., Patients and Methods: The study included voluntary participants: 53 healthy individuals negative to RT-PCR COVID-19 (control), and 165 patients positive to COVID-19. Severity was defined by the need of hospitalization, invasive ventilation, shock, or multiple organ failure. The patient group consisted of 28 asymptomatic, 82 with mild, and 55 with severe COVID-19. I/D polymorphism was determined by PCR. Rutinary laboratory tests were performed in all the participants., Results: DD polymorphism was significantly associated with severe COVID-19, independently of comorbidities, or any other variable. Receiver operator characteristic curves demonstrated association of low total cholesterol, low high-density lipoproteins, and high c-reactive protein with severity of COVID-19., Conclusion: The DD polymorphism was associated with the course of the infection and severity of COVID-19 in a sample of Mexican patients., (Copyright © 2023, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

The central nervous system (CNS) can be preconditioned to resist damage by peripheral pretreatment with low-dose gram-negative bacterial endotoxin lipopolysaccharide (LPS). Underlying mechanisms associated with transient protection of the cerebral cortex against traumatic brain injury include increased neuronal production of antiapoptotic and neurotrophic molecules, microglial-mediated displacement of inhibitory presynaptic terminals innervating the soma of cortical projection neurons, and synchronized firing of cortical projection neurons. However, the cell types and signaling responsible for these neuronal and microglial changes are unknown. A fundamental question is whether LPS penetrates the CNS or acts on the luminal surface of brain endothelial cells, thereby triggering an indirect parenchymal neuroprotective response. The present study shows that a low-dose intraperitoneal LPS treatment increases brain endothelial cell activation markers CD54, but does not open the blood-brain barrier or alter brain endothelial cell tight junctions as assessed by electron microscopy. NanoString nCounter transcript analyses of CD31-positive brain endothelial cells further revealed significant upregulation of Cxcl10, C3, Ccl2, Il1β, Cxcl2, and Cxcl1 , consistent with identification of myeloid differentiation primary response 88 (MyD88) as a regulator of these transcripts by pathway analysis. Conditional genetic endothelial cell gene ablation approaches demonstrated that both MyD88-dependent Toll-like receptor 4 (TLR4) signaling and Cxcl10 expression are essential for LPS-induced neuroprotection and microglial activation. These results suggest that C-X-C motif chemokine ligand 10 (CXCL10) production by endothelial cells in response to circulating TLR ligands may directly or indirectly signal to CXCR3 on neurons and/or microglia. Targeted activation of brain endothelial receptors may thus provide an attractive approach for inducing transient neuroprotection.

Background: Cervical cancer is a public health issue of global concern. It is a preventable disease but continues to threaten the lives of women, especially in developing countries in sub-Saharan Africa., Methods: We selected two African countries in sub-Saharan Africa (the Republic of Rwanda and the Republic of Sierra Leone) to show a good example of cervical cancer prevention and constrains hindering countries from effectively implementing cervical cancer programs. Secondary data were collected from the World Health Organization (WHO), the International Agency for Research on Cancer (IARC), the Global Burden of Cancer (GLOBOCAN), the United Nations Development Programme (UNDP), and the World Bank and from official websites of the selected countries. A descriptive analysis method was used to source data and compare variables such as the associated factors, disease burden, prevention programs, health workforce, success factors, and challenges., Results: Rwanda achieved 93.3% human papillomavirus (HPV) vaccination of the three doses vaccinating girls in class 6, as a result of effective school-based platform delivery system and community partnership to identify girls who are out of school. Rwanda reduced the historical two-decade gap in vaccine introduction between high- and low-income countries. The country also introduced a nationwide cervical cancer screening and treatment program. An impressive decreased cervical cancer incidence rate in Rwanda in recent years was observed. Sierra Leone lags behind in terms of almost all cervical cancer prevention programs. Therefore, Sierra Leone needs more efforts to implement cervical cancer intervention programs at the national level, including HPV vaccination, and train and increase the number of health professionals, treatment, and palliative care services to accelerate cervical cancer activities., Conclusion: The disease burden of cervical cancer for Rwanda and Sierra Leone is heavy. There remains huge room for improvement in preventing and controlling cervical cancer in these countries. The goal of cervical cancer elimination would not be feasible in countries without the awareness and will of the policymakers and the public, the compliance to fund cervical cancer programs, the prioritization of cervical cancer activities, the availability of resources, the adequate health workforce and infrastructure, the cross-sectional collaboration and planning, inter-sectorial, national, regional, and international partnerships., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Bangura, Zhao, Gonzalez Mendez, Wang, Didier Sama, Xu, Ren, Ma and Qiao.)

Objectives: The study aimed at analyzing the prevalence of five psychological outcomes (depression, anxiety, stress, post-traumatic stress disorder (PTSD), and suicidal ideation) among Chinese healthcare workers (HCWs), and measured the total possible negative psychological impact 1 year after the COVID-19 initial outbreak. Methods: A cross-sectional nationwide multi-center study was performed between November 2020 and March 2021 in China. A self-report questionnaire was applied, and three psychological scales were used. Binary logistic regression was performed to analyze the risk factors associated with each psychological outcome. Results: The findings demonstrated that the COVID-19 pandemic had a negative psychological impact on HCWs, which was still evident 1 year after the initial outbreak. Nurses showed higher depression and anxiety than other HCWs. Female gender, passive coping, long working hours, having a chronic disease, and experiencing violence, among other factors, were all risk factors for psychological impairment. Conclusion: Developing and promoting programs to improve mental health among HCWs, and identifying those who might need psychological support is still relevant 1 year after the initial outbreak., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Gonzalez Mendez, Ma, Alvarado, Ramirez, Xu, Xu, Zhang, Bangura, Yang, Yu, Zhang, Wang, Gu, Li, Salah and Qiao.)

Epithelial-mesenchymal transition (EMT) is a biological process that plays a central role in embryonic development, tissue regeneration, and cancer metastasis. Transforming growth factor-β (TGFβ) is a potent inducer of this cellular transition, comprising transitions from an epithelial state to partial or hybrid EMT state(s), to a mesenchymal state. Recent experimental studies have shown that, within a population of epithelial cells, heterogeneous phenotypical profiles arise in response to different time- and TGFβ dose-dependent stimuli. This offers a challenge for computational models, as most model parameters are generally obtained to represent typical cell responses, not necessarily specific responses nor to capture population variability. In this study, we applied a data-assimilation approach that combines limited noisy observations with predictions from a computational model, paired with parameter estimation. Synthetic experiments mimic the biological heterogeneity in cell states that is observed in epithelial cell populations by generating a large population of model parameter sets. Analysis of the parameters for virtual epithelial cells with biologically significant characteristics (e.g., EMT prone or resistant) illustrates that these sub-populations have identifiable critical model parameters. We perform a series of in silico experiments in which a forecasting system reconstructs the EMT dynamics of each virtual cell within a heterogeneous population exposed to time-dependent exogenous TGFβ dose and either an EMT-suppressing or EMT-promoting perturbation. We find that estimating population-specific critical parameters significantly improved the prediction accuracy of cell responses. Thus, with appropriate protocol design, we demonstrate that a data-assimilation approach successfully reconstructs and predicts the dynamics of a heterogeneous virtual epithelial cell population in the presence of physiological model error and parameter uncertainty., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 Biophysical Society. Published by Elsevier Inc. All rights reserved.)

Background: Neoadjuvant immunotherapies hold promise in muscle-invasive bladder cancer (MIBC)., Objective: To report on 2-yr disease-free (DFS) and overall (OS) survival including novel tissue-based biomarkers and circulating tumor DNA (ctDNA) in the ABACUS trial., Design, Setting, and Participants: ABACUS was a multicenter, single-arm, neoadjuvant, phase 2 trial, including patients with MIBC (T2-4aN0M0) who were ineligible for or refused neoadjuvant cisplatin-based chemotherapy., Intervention: Two cycles of atezolizumab were given prior to radical cystectomy. Serial tissue and blood samples were collected., Outcome Measurements and Statistical Analysis: The primary endpoints of pathological complete response (pCR) rate and dynamic changes to T-cell biomarkers were published previously. Secondary outcomes were 2-yr DFS and OS. A biomarker analysis correlated with relapse-free survival (RFS) was performed, which includes FOXP3, major histocompatibility complex class I, CD8/CD39, and sequential ctDNA measurements., Results and Limitations: The median follow-up time was 25 mo (95% confidence interval [CI] 25-26). Ninety-five patients received at least one cycle of atezolizumab. Eight patients did not undergo cystectomy (only one due to disease progression). The pCR rate was 31% (27/88; 95% CI 21-41). Two-year DFS and OS were 68% (95% CI 58-76) and 77% (95% CI 68-85), respectively. Two-year DFS in patients achieving a pCR was 85% (95% CI 65-94). Baseline PD-L1 and tumor mutational burden did not correlate with RFS (hazard ratio [HR] 0.60 [95% CI 0.24-1.5], p = 0.26, and 0.72 [95% CI 0.31-1.7], p = 0.46, respectively). RFS correlated with high baseline stromal CD8+ (HR 0.25 [95% CI 0.09-0.68], p = 0.007) and high post-treatment fibroblast activation protein (HR 4.1 [95% CI 1.3-13], p = 0.01). Circulating tumor DNA positivity values at baseline, after neoadjuvant therapy, and after surgery were 63% (25/40), 47% (14/30), and 14% (five/36), respectively. The ctDNA status was highly prognostic at all time points. No relapses were observed in ctDNA-negative patients at baseline and after neoadjuvant therapy. The lack of randomization and exploratory nature of the biomarker analysis are limitations of this work., Conclusions: Neoadjuvant atezolizumab in MIBC is associated with clinical responses and high DFS. CD8+ expression and serial ctDNA levels correlated with outcomes, and may contribute to personalized therapy in the future., Patient Summary: We showed that bladder cancer patients receiving immunotherapy followed by cystectomy have good long-term outcomes. Furthermore, we found that certain biological features can predict patients who might have particular benefit from this therapy., (Copyright © 2022 European Association of Urology. Published by Elsevier B.V. All rights reserved.)

Aim: To design, implement and evaluate a nurse-led capacity building intervention (PromoGOB) for intersectoral action for health at local governments., Design: The programme was based on theories of the policy process and organizational change and facilitated by a nurse developing a health broker role. A complex intervention perspective was adopted in carrying out the study. The intervention was evaluated using a mixed method embedded design., Methods: Quantitative component relied on a specific questionnaire. This tool, designed and piloted ad hoc, measured the capacity in terms of knowledge, awareness, resources, skills, and commitment, both at sectoral and government levels. For the qualitative component, semi-structured interviews were conducted. These explored the perceived capacity and feasibility and acceptability issues. The programme was initiated at the end of October 2019, and it lasted a total of 5 weeks. Nineteen individuals representing various sectors at a local government in northern Spain participated in the study. The data analysis was concluded by the end of March 2020., Findings: PromoGOB positively influenced participants' capacity for addressing health promotion. Awareness component, intersectoral work and the nurse as health broker were essential in the programme. The necessity of political participation was identified as an issue to be prioritized in future studies., Conclusion: This study highlights the relevance of capacity building at local governments and the role that nurses can play in it. Further work should be undertaken to continue developing Health in All Policies approach at local level., Impact: This study offers a starting point for nurses to get involved in the policy process of health promotion, performing a specific role as health brokers, building capacity at local governments for addressing social determinants of health, and delving into theories and concepts of the Health in All Policies field., (© 2022 The Authors. Journal of Advanced Nursing published by John Wiley & Sons Ltd.)

Background: The INMUNOSUN trial had the objective of prospectively evaluating the efficacy and safety of sunitinib as a pure second-line treatment in patients with metastatic renal cell carcinoma (mRCC) who have progressed to first-line immune checkpoint inhibitor (ICI)-based therapies., Patients and Methods: A multicenter, phase II, single-arm, open-label study was carried out in patients with a histologically confirmed diagnosis of mRCC with a clear-cell component who had progressed to a first-line regimen of ICI-based therapies. All patients received sunitinib 50 mg once daily orally for 4 weeks, followed by a 2-week rest period following package insert instructions. The primary outcome was the objective response rate., Results: Twenty-one assessable patients were included in the efficacy and safety analyses. Four patients [19.0%, 95% confidence interval (CI) 2.3% to 35.8%] showed an objective response (OR), and all of them had partial responses. Additionally, 14 (67%) patients showed a stable response, leading to clinical benefit in 18 patients (85.7%, 95% CI 70.7% to 100%). Among the four assessable patients who showed an OR, the median duration of the response was 7.1 months (interquartile range 4.2-12.0 months). The median progression-free survival (PFS) was 5.6 months (95% CI 3.1-8.0 months). The median overall survival (OS) was 23.5 months (95% CI 6.3-40.7 months). Patients who had better antitumor response to first-line ICI-based treatment showed a longer PFS and OS with sunitinib. The most frequent treatment-emergent adverse events were diarrhea (n = 11, 52%), dysgeusia (n = 8, 38%), palmar-plantar erythrodysesthesia (n = 8, 38%), and hypertension (n = 8, 38%). There was 1 patient who exhibited grade 5 pancytopenia, and 11 patients experienced grade 3 adverse events. Eight (38%) patients had serious adverse events, four of which were considered to be related to sunitinib., Conclusion: Although the INMUNOSUN trial did not reach the pre-specified endpoint, it demonstrated that sunitinib is active and can be safely used as a second-line option in patients with mRCC who progress to new standard ICI-based regimens., Competing Interests: Disclosure EG has received honoraria for speaker engagements, advisory roles, or funding of continuous medical education from Adacap, AMGEN, Angelini, Astellas, Astra Zeneca, Bayer, Blueprint, Bristol Myers Squibb, Caris Life Sciences, Celgene, Clovis-Oncology, Eisai, Eusa Pharma, Genetracer, GSK, Guardant Health, HRA-Pharma, IPSEN, ITM-Radiopharma, Janssen, Lexicon, Lilly, Merck KGaA, MSD, Nanostring Technologies, Natera, Novartis, ONCODNA (Biosequence), Palex, Pharmamar, Pierre Fabre, Pfizer, Roche, Sanofi-Genzyme, Servier, Taiho, Thermo Fisher Scientific. TAG has received support for the present manuscript (protocol review and manuscript review); has received research grants from Pfizer, Roche, and Ipsen; has received consulting fees from Ipsen, Pfizer, Roche, Sanofi, Bayer, Astellas, Janssen-Cilag, BMS, and EISAI; has received payment or honoraria for lectures, presentations, speakers’ bureaus, manuscript writing, or educational events from Ipsen, Pfizer, Eisai, and Merck; has received support for attending meetings and/or travel from Pfizer, Sanofi, BMS, and IPSEN. OR has received personal payment from Eusapharma, BMS, Pfizer, and Ipsen; has received support for attending meetings and/or travel from Ipsen and Pfizer; has participated in the Advisory Board of BMS and Bayer. XGdM has received consulting fees from Roche, Pharmamar, BMS, Ipsen, Lilly, Eusapharma, Pfizer, Merck, and Astellas; has received payment or honoraria for lectures, presentations, speaker bureaus, manuscript writing, or educational events from BMS, Astellas, Pharma, Eisai, Pfizer, and EusaPharma; has received support for attending meetings and/or travel from Pfizer and Roche; and has received research funding from AstraZeneca. MJM has received payment or honoraria as invited speaker from Janssen-Cilag, Bayer Healthcare, Sanofi Aventis, Astellas Medivation, Roche, Ipsen, EISAI, Norvartis, Pfizer, and MSD; has received travel support from Roche and Ipsen; has participated on Data Safety Monitoring Board or Advisory Board of Janssen-Cilag, Bayer Healthcare, Sanofi Aventis, Astellas Medivation, Roche, Ipsen, EISAI, Novartis, Pfizer, and MSD. JGD has received grants or contracts from Eusapharma, Bristol Myers, Novartis, GSK, Merck, MSD, Sanofi, Janssen, Astellas, Astra Zeneca, and Ipsen; has received support for attending meetings and/or travel from Roche; has participated on a Data Safety Monitoring Board or Advisory Board from Pfizer and Pharmamar. MGR has received payment or honoraria for lectures, presentations, speakers’ bureaus, manuscript writing, or educational events from Roche; has received support for attending meetings and/travel from Roche, Ipsen, AstraZeneca, MSD, and Jansen. JAAA has received grants or contracts from Bristol-Myers Squibb (grant to Spanish GU Oncology Group for research in prostate cancer); has received consulting fees from Janssen, Cilag, Pfizer, MSD, and Bristol-Myers Squibb; has received payment or honoraria for lectures, presentations, speakers’ bureaus, manuscript writing, or educational events from Astellas, Pfizer, and MSD; has received support for attending meetings and/or travel from MSD, Bristol-Myers Squibb, and Astra Zeneca. PLC has received payment or honoraria for lectures, presentations, speakers’ bureaus, manuscript writing, or educational events from MSD and Bristol; has participated on a Data Safety Monitoring Board or Advisory Board from Sanofi and Roche. JMC has received support for the present manuscript (protocol review and manuscript review); has received grants or contracts from Pfizer and Ipsen; has received consulting fees from Ipsen, Pfizer, Roche, Sanofi, Astellas, Janssen Cilag, BMS, and EISAI; has received payment or honoraria for lectures, presentations, speakers’ bureaus, manuscript writing, or educational events from Ipsen, Pfizer, and BMS; and has received support for attending meetings and/or travel from Pfizer, BMS, and Ipsen. BM has received research grant/funding (self) from Jansen, Roche, Astellas, Sanofi, and Bayer; has received payment or honoraria for speaker bureau from Pfizer, Ipsen, Jansen, Roche, Astellas, Sanofi, BMS, and Bayer; has received support for travel accommodation from Pfizer, Ipsen, Janssen, and Roche; other financial or non-financial interests: advisory and speaker bureau rom Pfizer, Ipsen, Jansen, Roche, Astellas, Sanofi, BMS, and Bayer; travel accommodation from Pfizer, Ipsen, Jansen, and Roche; and research grant/funding (self) from Roche, Astellas, Sanofi Jansen, BMS, and Bayer. CAF has received consulting fees from Ipsen, Boehringer Ingelheim, and Astra Zeneca; has received support for attending meetings and/or travel from Roche, Pfizer, and Astellas. GDV has received payment or honoraria for lectures, presentations, speakers’ bureaus, manuscript writing, or educational events from BMS, Pfizer, Roche, Merk, MSD, Bayer, Eusapharma, Astellas, and Ipsen; has received support for attending meetings and/or travel from Roche; has participated on a Data Safety Monitoring Board or Advisory Board from BMS, Pfizer, Roche, Merck, MSD, Bayer, and Astellas. JFRM has received support for the present manuscript from Pfizer; has received grants or contracts from HM Hospitales; has received payment or honoraria for lectures, presentations, speakers’ bureaus, manuscript writing, or educational events from Pfizer, BMS, Novartis, and MSD; has received support for attending meetings from Pfizer, BMS, Novartis, and MSD; has participated on a Data Safety Monitoring Board or Advisory Board from BMS and Novartis; and has participated in corporate-sponsored research in his institution from AstraZeneca, BMS, Amgen, Roche, Novartis, MSD, Janssen, Pfizer, Astellas, GSK, PharmaMar, Ipsen, Tesaro, Abbvie, Aprea Therapeutics, Eisai, Bayer, and Merck. CSR has received grants or contracts from AB Science, Aragon Pharmaceuticals, Astellas Pharma, Astra Zeneca AB, Bayer, Blueprint Medicines Corporation, Boehringer Ingelheim España SA, BMS, Clovis Oncology, Exelixix Inc F, Genentech Inc, GlaxoSmithKline SA, Hoffman-La Roche LTD, Novartis Farmaceutica SA, Pfizer SLU, and Sanofi-Aventis; has received payment or honoraria for speakers’ bureau from Astellas Pharma, BMS, Hoffmann-La Roche LTD, Ipsen, Pfizer SLU; has participated on a Data Safety Monitoring Board or Advisory Board from Astellas Pharma, Bayer, BMS, Eusapharma, Hoffmann-La Roche LTD, Ipsen, MSD, Novartis, Pfizer, and Sanofi-Aventis E. All other authors have declared no conflicts of interest. Data sharing The clinical trial data are available from the corresponding author upon reasonable request., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

This article emphasizes the selection criteria for radiographic acquisition in children due to the greater sensitivity of children for radiation compared with adults. Diagnosis of common pediatric dental conditions, including dental caries, periodontitis, dental anomalies, cysts, tumors, and traumatic dental conditions, are discussed with relevant clinical scenarios., Competing Interests: Disclosure The authors have nothing to disclose., (Copyright © 2021 Elsevier Inc. All rights reserved.)

The aim of this work was to compare the concentration of glyphosate and AMPA in the PM10 and the actual PM10 emission from agricultural soils and unpaved roads, located inside and outside farm fields. To determine the actual PM10 emission by wind erosion, the actual wind erosion was estimated using the Wind Erosion Equation, and the PM10 emission efficiency was measured with the Easy Dust Generator. PM10 was collected in an electrostatic precipitator coupled to the Easy Dust Generator. Actual PM10 emission was 11.5 g ha -1  year -1 in agricultural soils and 4711.4 g ha -1  year -1 in unpaved roads. The high value of actual PM10 emission in unpaved roads was due to their high actual wind erosion and the high PM10 emission efficiency, while the low value in agricultural soils was due to their low actual wind erosion. Content of glyphosate in the PM10 ranged from 59 to 359 μg kg -1 in agricultural soils, from 382 to 454 μg kg -1 in unpaved roads inside farm fields, and from 39 to 639 μg kg -1 in unpaved roads outside farm fields. Content of AMPA in the PM10 ranged from 387 to 7228 μg kg -1 in agricultural soils, from 900 to 4138 μg kg -1 in unpaved roads inside farm fields, and 98 to 500 μg kg -1 in unpaved roads outside farm fields. AMPA concentration in PM10 was higher than that of glyphosate due to the longer persistence of AMPA than glyphosate. Glyphosate and AMPA concentrations in PM10 were higher than in soil, which is an additional risk that should be considered when the effect of PM10 emitted by agricultural soils and unpaved roads on human health are evaluated. Our results show that the amount and chemical composition of PM10 emitted by wind erosion from unpaved roads should be studied in other regions., Competing Interests: Declaration of competing interest This article presents novel results about glyphosate and AMPA concentration in PM10 emitted by rural unpaved roads and agricultural soils. The amount of glyphosate and AMPA carried in the PM10 emitted by wind erosion of unpaved roads and agricultural soils was also studied. This information is useful to understand potential contamination with glyphosate and AMPA of the respirable dust emitted by wind erosion and others process like traffic and tillage. All this information is useful to understand the environmental fate of glyphosate and their potential health effects., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Objectives: The study assessed the effectiveness of an oral health educational and behavioral intervention program in improving the knowledge, attitudes, and behaviors of refugee families., Methods: This randomized 2-arms, controlled, single site, clinical trial assessed the dental knowledge, attitudes, and behaviors related to oral health at baseline and three times over the course of the 6 months of the intervention in recent refugee families. Participating families were educated on five topics in oral health in two 1-hour sessions utilizing existing oral health education materials adapted to be linguistically and culturally appropriate for demonstration and instruction. Culturally competent techniques and motivational interviewing styles were also implemented during sessions. Pre/post surveys were used to assess changes to knowledge, attitudes, and behavior among refugee family participants., Results: Out of the 66 families enrolled in the program, 52 (72 percent) completed visits over the course of 6 months. Differences between the intervention and control groups were not significant between baseline and 3 to 6 months later (P > 0.05)., Conclusions: A short-term, culturally informed oral health educational and behavioral intervention program did not improve oral health-related knowledge, attitudes, or behaviors in a diverse group of recent refugee families., (© 2020 The Authors. Journal of Public Health Dentistry published by Wiley Periodicals LLC on behalf of American Association of Public Health Dentistry.)

Objective: The study assessed a preventive outreach educational intervention targeting improvements in dental caries and oral-health-related quality of life in the children of refugee families by comparing pre- and postintervention outcomes. Methods: This randomized controlled clinical trial assessed the outcomes at baseline and three times over six months using the WHO oral health assessment form (DMFT/dmft) and the parent version of the Michigan Oral-Health-Related Quality of Life scale. Children and at least one of their parents/caretakers were educated on oral health topics in two one-hour sessions. Results: Of the 66 enrolled families, 52 (72%) completed the six-month follow-up. DMFT/dmft scores increased significantly in both the control and intervention groups ( p < 0.05); differences in the changes in the DMFT/dmft and MOHRQoL-P scores from baseline to the three- and six-month follow-up visits between groups were not significant ( p > 0.05). Conclusions: Oral health education programs targeting a diverse group of refugee children and their parents/caregivers single-handedly did not reduce the increased number of caries lesions or improve oral-health-related quality of life.

Background: The COVID-19 pandemic has threatened the health systems of many countries worldwide. Several studies have suggested that the pandemic affects not only physical health but also all aspects of society. A lot of information has been reported about the disease since the beginning of the outbreak. For that reason, it is essential to investigate the attitudes and level of knowledge and awareness that different populations had regarding COVID-19 during the critical period of the outbreak., Objective: This study aimed to assess the knowledge and awareness of and attitudes toward the COVID-19 pandemic among different populations in Central China during the critical period of the outbreak., Methods: A cross-sectional web-based survey was conducted in Central China from February to March 2020. The study participants included three different populations: medical workers, students, and those with other occupations. In this study, a questionnaire was designed to collect information on the following four aspects: sociodemographic information, knowledge related to COVID-19, awareness of COVID-19, and attitude toward COVID-19. The chi-square test and Fisher test were used for comparison among groups. The level of significance was set at P<.05., Results: This study enrolled a total of 508 participants. Among them, there were 380 students (74.8%), 39 medical workers (7.7%), and 89 people with other occupations (17.5%). Most of the participants were female (n=272, 53.5%), lived in rural areas (n=258, 50.8%), and were single (n=423, 86.9%). The majority of the respondents had attended college (n=454, 89.4%). Most of the participants said they had heard about COVID-19 by January, and most of them looked for information on social media (Sina Weibo, 84.7%), and WeChat and QQ groups (74.2%). The participants showed an adequate level of knowledge about COVID-19 with no significant differences among the groups. However, medical workers demonstrated a slightly advanced knowledge in their responses to professional questions such as the potential susceptible population, possible host, treatment of COVID-19, and disease category. A higher proportion of medical workers (71.8%) and those in the other occupations group (52.8%) were highly concerned about the COVID-19 pandemic. More than 43% of the participants stated that the lockdown of their village/city had a significant impact on their lives. Nevertheless, the majority of respondents had an overall optimistic attitude toward the control of the disease (92.1% of students [n=350], 94.9% of medical workers [n=37], and 92.3% of those in other occupations [n=83])., Conclusions: All three groups reported an adequate background knowledge about COVID-19 but medical workers showed a slightly advanced knowledge in their responses to professional questions. Most of the participants were highly concerned about COVID-19 during the critical period of the outbreak. The majority of respondents declared that the village/city lockdown policy had a significant impact on their daily life but most of them held an optimistic attitude toward the control of COVID-19., (©Huifang Xu, Maria Jose Gonzalez Mendez, Lanwei Guo, Qiong Chen, Liyang Zheng, Peipei Chen, Xiaoqin Cao, Shuzheng Liu, Xibin Sun, Shaokai Zhang, Youlin Qiao. Originally published in the Journal of Medical Internet Research (http://www.jmir.org), 15.10.2020.)

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

The objective of this study was to analyze the healthcare encounters between nurses and parents of different cultural backgrounds in primary health care. An ethnographic study was carried out using participant observations in health centers and interviews with nurses. Data were analyzed using thematic content analysis and constant comparative method. Four main themes were identified when nurses met parents of other cultural backgrounds: lack of mutual understanding, electronic records hamper the interaction, lack of professionals' cultural awareness and skills, and nurses establish superficial or distant relationships. The concepts of ethnocentrism and cultural imposition are behind these findings, hampering the provision of culturally competent care in primary health services. There were difficulties in obtaining and registering culturally related aspects that influence children's health and development. This was due to e-records, language barriers, and the lack of cultural awareness and skills in health professionals making the encounters difficult for both nurses and parents. These findings show that there is a clear threat for health equity and safety in primary care if encounters between nurses and parents do not improve to enable nursing care to be tailored to any individual family needs., (© 2020 John Wiley & Sons Australia, Ltd.)

Background: The performance of lung cancer screening with low-dose computed tomography (CT) (LDCT) in China is uncertain. This study aimed to evaluate the performance of LDCT lung cancer screening in the Chinese setting., Methods: In 2014, a screening cohort of lung cancer with LDCT was established in Gejiu, Yunnan Province, a screening center of the Lung Cancer Screening Program in Rural China (LungSPRC). Participants received a baseline screening and four rounds of annual screening with LDCT in two local hospitals until June 2019. We analyzed the rates of participation, detection, early detection, and the clinical characteristics of lung cancer., Results: A total of 2006 participants had complete baseline screening results with a compliance rate of 98.4%. Of these, 1411 were high-risk and 558 were nonhigh-risk participants. During this period, 40 lung cancer cases were confirmed, of these, 35 were screen-detected, four were post-screening and one was an interval case. The positive rate of baseline and annual screening was 9.7% and 9.0%, while the lung cancer detection rate was 0.4% and 0.6%, respectively. The proportion of early lung cancer increased from 37.5% in T0 to 75.0% in T4. Adenocarcinoma was the most common histological subtype. Lung cancer incidence according to the criteria of LungSPRC and National Lung Cancer Screening Trial (NLST) was 513.31 and 877.41 per 100 000 person-years, respectively., Conclusions: The program of lung cancer screening with LDCT showed a successful performance in Gejiu, Yunnan. However, further studies are warranted to refine a high-risk population who will benefit most from LDCT screening and reduce the high false positive results., Key Points: This study reports the results of lung cancer screening with LDCT in Gejiu, Yunnan, a high-risk area of lung cancer, and it demonstrates that lung cancer screening with LDCT is effective in detecting early-stage lung cancer. Our program provides an opportunity to explore the performance of LDCT lung cancer screening in the Chinese context., (© 2020 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.)

Epithelial-mesenchymal transition (EMT) is a fundamental biological process that plays a central role in embryonic development, tissue regeneration, and cancer metastasis. Transforming growth factor-β (TGFβ) is a potent inducer of this cellular transition, which is composed of transitions from an epithelial state to intermediate or partial EMT state(s) to a mesenchymal state. Using computational models to predict cell state transitions in a specific experiment is inherently difficult for reasons including model parameter uncertainty and error associated with experimental observations. In this study, we demonstrate that a data-assimilation approach using an ensemble Kalman filter, which combines limited noisy observations with predictions from a computational model of TGFβ-induced EMT, can reconstruct the cell state and predict the timing of state transitions. We used our approach in proof-of-concept "synthetic" in silico experiments, in which experimental observations were produced from a known computational model with the addition of noise. We mimic parameter uncertainty in in vitro experiments by incorporating model error that shifts the TGFβ doses associated with the state transitions and reproduces experimentally observed variability in cell state by either shifting a single parameter or generating "populations" of model parameters. We performed synthetic experiments for a wide range of TGFβ doses, investigating different cell steady-state conditions, and conducted parameter studies varying properties of the data-assimilation approach including the time interval between observations and incorporating multiplicative inflation, a technique to compensate for underestimation of the model uncertainty and mitigate the influence of model error. We find that cell state can be successfully reconstructed and the future cell state predicted in synthetic experiments, even in the setting of model error, when experimental observations are performed at a sufficiently short time interval and incorporate multiplicative inflation. Our study demonstrates the feasibility and utility of a data-assimilation approach to forecasting the fate of cells undergoing EMT., (Copyright © 2020 Biophysical Society. Published by Elsevier Inc. All rights reserved.)

Antibodies targeting PD-1 or its ligand 1 PD-L1 such as atezolizumab, have great efficacy in a proportion of metastatic urothelial cancers 1,2 . Biomarkers may facilitate identification of these responding tumors 3 . Neoadjuvant use of these agents is associated with pathological complete response in a spectrum of tumors, including urothelial cancer 4-7 . Sequential tissue sampling from these studies allowed for detailed on-treatment biomarker analysis. Here, we present a single-arm phase 2 study, investigating two cycles of atezolizumab before cystectomy in 95 patients with muscle-invasive urothelial cancer (ClinicalTrials.gov identifier: NCT02662309). Pathological complete response was the primary endpoint. Secondary endpoints focused on safety, relapse-free survival and biomarker analysis. The pathological complete response rate was 31% (95% confidence interval: 21-41%), achieving the primary efficacy endpoint. Baseline biomarkers showed that the presence of preexisting activated T cells was more prominent than expected and correlated with outcome. Other established biomarkers, such as tumor mutational burden, did not predict outcome, differentiating this from the metastatic setting. Dynamic changes to gene expression signatures and protein biomarkers occurred with therapy, whereas changes in DNA alterations with treatment were uncommon. Responding tumors showed predominant expression of genes related to tissue repair after treatment, making tumor biomarker interpretation challenging in this group. Stromal factors such as transforming growth factor-β and fibroblast activation protein were linked to resistance, as was high expression of cell cycle gene signatures after treatment.

Background: Markers able to predict the response to antiangiogenics in metastatic clear cell renal cell carcinoma (ccRCC) are not available. The development of new treatment options like immunotherapy are reaching the clinic; therefore, predictors of benefit from these different available treatments are increasingly needed., Objective: In this study, we prospectively assessed the association of circulating endothelial cells (CECs) in peripheral blood with long-term benefit from first-line treatment in ccRCC., Design, Setting, and Participants: A prospective observational study was designed involving 13 institutions of the Spanish Oncology Genitourinary Group. Adult patients diagnosed with advanced ccRCC who had achieved response or disease stabilization after 3 mo on first-line therapy were eligible., Outcome Measurements and Statistical Analysis: CECs were isolated from peripheral blood, captured with ferrofluids coated with monoclonal antibodies directed against the CD146 antigen, and assessed centrally with an automated standardized system. CECs were defined as 4',6-diamidino-2-phenylindole+, CD105+, and CD45-. Blood samples were systematically taken every 6 wk for 15 mo or until tumor progression, whichever occurred first. Clinical data were externally monitored at all centers., Results and Limitations: From August 9, 2011, to January 17, 2013, 75 patients were enrolled in the study. Patients with baseline CECs above the median showed a significantly longer progression-free survival than those with low CECs (22.2 mo vs 12.2 mo) with a hazard ratio of 2.5 (95% confidence interval: 1.2-5.3, p=0.016). There was no difference between CEC levels at baseline and at tumor progression (medians of 50 CECs/4ml and 52 CECs/4ml, respectively)., Conclusions: Under antiangiogenic treatment, the detection of higher CEC levels is associated with clinical benefit in terms of progression-free survival in ccRCC., Patient Summary: Antiangiogenics are the cornerstone of treatment in kidney cancer. Since they target endothelial rather than tumor cells, we studied the correlation between levels of circulating endothelial cells in peripheral blood and long-term benefit in patients on antiangiogenic therapy. Higher levels were associated with long-term benefit, suggesting that this determination could help to separate best responders from those who could require a more intensive approach., (Copyright © 2016 European Association of Urology. Published by Elsevier B.V. All rights reserved.)

Wind erosion of freshly-deposited volcanic ash causes persistent storms, strongly affecting ecosystems and human activity. Wind erosion of the volcanic ash was measured up to 17 months after the ash deposition, at 7 sites located within the ash-deposition area. The mass flux was measured up to 1.5 m above ground level. Mass transport rates were over 125 times the soil wind-erosion rates observed before the ash deposition, reaching up to 6.3 kg m -1 day -1 . Total mass transport of ash during the 17 months ranged between 113.6 and 969.9 kg m -1 depending on topographic location and wind exposure. The vertical distribution of the mass flux at sites with higher vegetation cover was generally inverted as compared to sites with lower vegetation cover. This situation lasted 7 months and then a shift towards a more uniform vertical distribution was observed, in coincidence with the beginning of the decline of the mass transport rates. Decay rates differed between sites. Despite changes over time, an inverse linear correlation between the mass transports and the mass-flux gradients was found. Both the mass-flux gradients and the average mass-transport rates were not linked with shear-stress partition parameters, but with the ratio: ash-fall thickness to total vegetation cover.

This Point/Counterpoint discusses the long-argued debate over whether lecture attendance in dental school at the predoctoral level should be required. Current educational practice relies heavily on the delivery of content in a traditional lecture style. Viewpoint 1 asserts that attendance should be required for many reasons, including the positive impact that direct contact of students with faculty members and with each other has on learning outcomes. In lectures, students can more easily focus on subject matter that is often difficult to understand. A counter viewpoint argues that required attendance is not necessary and that student engagement is more important than physical classroom attendance. This viewpoint notes that recent technologies support active learning strategies that better engage student participation, fostering independent learning that is not supported in the traditional large lecture classroom and argues that dental education requires assimilation of complex concepts and applying them to patient care, which passing a test does not ensure. The two positions agree that attendance does not guarantee learning and that, with the surge of information technologies, it is more important than ever to teach students how to learn. At this time, research does not show conclusively if attendance in any type of setting equals improved learning or ability to apply knowledge.

Background: Skin rash is an adverse event which might be associated with longer survival in patients treated with epidermal growth factor receptor tyrosine kinase inhibitors. The aim of this nonrandomised phase II clinical trial is to prospectively evaluate the relationship between skin rash and overall survival (OS) in advanced/metastatic pancreatic cancer treated with erlotinib plus gemcitabine., Patients and Methods: Patients were given gemcitabine (1000 mg/m2/week, 3 weeks every 4 weeks) plus erlotinib (100 mg/day orally continuously) until disease progression/unacceptable toxicity. The primary end point was OS., Results: A total of 153 eligible patients were enrolled (grade≥2 rash, 25%; grade<2 rash, 75%). OS was longer in patients with grade≥2 rash versus grade<2 (11 versus 5 months; P<0.001). Progression-free survival was longer in patients with grade≥2 rash versus grade<2 (6 versus 3 months; P<0.001) and shorter in those without rash versus grade 1 (2 versus 4 months; P=0.005) or grade≥2 (2 versus 6 months; P<0.001). Patients with grade≥2 rash showed higher rates of overall response (21% versus 7%; P<0.05) and disease control (84% versus 43%; P<0.05) versus grade<2., Conclusions: This study prospectively confirms the relationship between rash and longer OS in unresectable locally advanced/metastatic pancreatic cancer treated with erlotinib plus gemcitabine.

We demonstrate a system for cloning and modifying the chloroplast genome from the green alga, Chlamydomonas reinhardtii. Through extensive use of sequence stabilization strategies, the ex vivo genome is assembled in yeast from a collection of overlapping fragments. The assembled genome is then moved into bacteria for large-scale preparations and transformed into C. reinhardtii cells. This system also allows for the generation of simultaneous, systematic and complex genetic modifications at multiple loci in vivo. We use this system to substitute genes encoding core subunits of the photosynthetic apparatus with orthologs from a related alga, Scenedesmus obliquus. Once transformed into algae, the substituted genome recombines with the endogenous genome, resulting in a hybrid plastome comprising modifications in disparate loci. The in vivo function of the genomes described herein demonstrates that simultaneous engineering of multiple sites within the chloroplast genome is now possible. This work represents the first steps toward a novel approach for creating genetic diversity in any or all regions of a chloroplast genome.

Background: In developing countries, complementary foods are often introduced earlier or later than appropriate and the quality is frequently insufficient, particularly in rural areas where complementary foods have traditionally been based on starchy gruels. Adequate intakes of a number of nutrients are recognized to be problematic in traditional complementary feeding regimens in developing societies., Aim: To determine the contribution of the complementary feeding nutrients to the estimated total nutrient intake in Guatemalan infants., Methods: Three non-consecutive 24-hr recalls were collected from a convenience sample of mothers of 64 infants, aged 6-12 month on enrolment, in the rural Guatemalan highland village of Santo Domingo Xenacoj. Additional information on early introduction of pre- and post-lacteal feeds and on first foods and beverages was included. Human milk intakes were estimated by a model based on assumptions regarding satisfaction of weight-based daily energy needs by the combined diet. The 2004 WHO/FAO recommended nutrient intakes were used as the standard for adequate nutrient consumption., Results: We observed that exclusive breastfeeding up to 6 month is rare. Mean nutrient intakes and densities were above recommended intakes for all nutrients examined, except calcium, iron and zinc. Intakes of most nutrients were greater from the complementary feeding component of the diet. Vitamin A intake was excessive due to consumption of fortified sugar., Conclusions: We conclude that intakes of most micronutrients were near recommendation levels, unusual within the complementary feeding experience in scientific literature. Calcium, iron and zinc were identified as "problem nutrients" as persistently reported in developing countries.

Ferroportin is a multipass membrane protein that serves as an iron exporter in many vertebrate cell types. Ferroportin-mediated iron export is controlled by the hormone hepcidin, which binds ferroportin, causing its internalization and degradation. Mutations in ferroportin cause a form of the iron overload hereditary disease hemochromatosis. Relatively little is known about ferroportin's properties or the mechanism by which mutations cause disease. In this study, we expressed and purified human ferroportin to characterize its biochemical/biophysical properties in solution and conducted cell biological studies in mammalian cells. We found that purified detergent-solubilized ferroportin is a well-folded monomer that binds hepcidin. In cell membranes, the N- and C-termini were both cytosolic, implying an even number of transmembrane regions, and ferroportin was mainly localized to the plasma membrane. Hepcidin addition resulted in a redistribution of ferroportin to intracellular compartments that labeled with early endosomal and lysosomal, but not Golgi, markers and that trafficked along microtubules. An analysis of 16 disease-related ferroportin mutants revealed that all were expressed and trafficked to the plasma membrane but that some were resistant to hepcidin-induced internalization. The characterizations reported here form a basis upon which models for ferroportin's role in regulating iron homeostasis in health and disease can be interpreted.

Adenoviral vectors are widely used to express transgenes in vitro and in vivo. A major obstacle to the generation of adenoviral vectors is the manipulation of the large (35 kb) adenoviral genome. We developed a hybrid yeast-bacteria cloning system for the creation of novel adenoviral vectors. The adenovirus 5 (Ad5) genome was cloned into a shuttle vector that contains both yeast and bacterial elements for replication and therefore functions as both a yeast artificial plasmid (YAP) and as a plasmid artificial chromosome (PAC). Any sequence can be introduced into any region of the adenoviral genome via the highly efficient homologous recombination in yeast and then these recombinants are rapidly amplified in bacteria. Adenoviral vectors are generated by introduction of the PAC into the appropriate complementing mammalian cell without the need for plaque purification. Vectors were constructed with deletions in the E1, E3, and/or E4 regions. We have generated more than 100 vectors with a number of different transgenes and regulatory elements. In addition, the YAP/PAC vector was used to capture a DNA fragment encompassing the human factor IX gene, demonstrating the utility of this system to clone and analyze genomic DNA. This novel cloning strategy allows the rapid and versatile construction of adenoviral vectors for gene expression and gene therapy applications.

We constructed two megabase-sized YACs containing large contiguous fragments of the human heavy and kappa (kappa) light chain immunoglobulin (Ig) loci in nearly germline configuration, including approximately 66 VH and 32 V kappa genes. We introduced these YACs into Ig-inactivated mice and observed human antibody production which closely resembled that seen in humans in all respects, including gene rearrangement, assembly, and repertoire. Diverse Ig gene usage together with somatic hypermutation enables the mice to generate high affinity fully human antibodies to multiple antigens, including human proteins. Our results underscore the importance of the large Ig fragments with multiple V genes for restoration of a normal humoral immune response. These mice are likely to be a valuable tool for the generation of therapeutic antibodies.

Objective: The effect of pulse cyclophosphamide treatment was retrospectively assessed in 25 systemic lupus erythematosus (SLE) patients with central nervous system involvement. All patients who tested positive for anti-phospholipid antibodies and/or lupus anticoagulant were excluded., Results: Low-dose intravenous cyclophosphamide pulses (500 mg) were administered weekly in all patients. Twenty-four out of 25 patients attained a good response (after a mean of 11 days). Cyclophosphamide was well tolerated in all patients with only minor side effects. None of the patients experienced ovarian failure, cystitis or herpes zoster., Conclusions: Weekly low-dose cyclophosphamide pulses appear to be safe and effective for the management of neuropsychiatric manifestations in SLE patients without antiphospholipid antibodies.

Our paper describes the introduction of large fragments of both the human heavy and light chain Ig genes into the mouse germline to create a mouse strain capable of producing a broad repertoire of antigen-specific, fully human antibodies. The human immunoglobulin gene sequences were functional in the context of the mouse machinery for antibody recombination and expression, either in the presence or absence of functional endogenous genes. This was demonstrated by their ability to undergo diverse rearrangement, to be expressed at significant levels, and to exclude expression of mouse immunoglobulins irrespective of their copy number or site of integration. The decrease in susceptibility to influence by adjacent genomic sequences may reflect the greater size, variable gene content, or structural integrity of the human Ig YACs and/or the presence of unidentified but important regulatory elements needed for optimal expression of the human immunoglobulin genes and their correct regulation. Our results show that mouse B cells coexpressing human heavy and kappa chains, upon immunization, can produce antigen-specific, fully human antibodies. Furthermore, the human heavy and kappa chain YACs induced differentiation and maturation of the growth-arrested B-cell lineage in mice with inactivated endogenous Ig genes, leading to the production of a diverse repertoire of fully human antibodies at levels approaching those in normal serum. These results suggest the potential value of these mice as a source of fully human antibodies for human therapy. Furthermore, it is expected that such mice would lack immunological tolerance to and thus readily yield antibodies to human proteins, which may constitute an important class of targets for monoclonal antibody therapy. Our findings suggest that the introduction of even larger portions of the human heavy and light chain loci, which should be achievable with the ES cell-yeast spheroplast fusion technology described, will result in strains of mice ultimately capable of recapitulating the full antibody repertoire characteristic of the human humoral response to infection and immunization. The present and future mouse strains may prove to be valuable tools for studying the molecular mechanisms and regulatory sequences influencing the programmed assembly and expression of human antibodies in the normal immune response, as well as the abnormal response characteristic of autoimmune disease and other disorders. The strategy we have described for the introduction of large segments of the human genome into mice in conjunction with the inactivation of the corresponding mouse loci may also have broad applicability to the investigation of other complex or uncharacterized loci.

With the goal of creating a strain of mice capable of producing human antibodies, we are cloning and reconstructing the human immunoglobulin germline repertoire in yeast artificial chromosomes (YACs). We describe the identification of YACs containing variable and constant region sequences from the human heavy chain (IgH) and kappa light chain (IgK) loci and the characterization of their integrity in yeast and in mouse embryonic stem (ES) cells. The IgH locus-derived YAC contains five variable (VH) genes, the major diversity (D) gene cluster, the joining (JH) genes, the intronic enhancer (EH), and the constant region genes, mu (C mu) and delta (C delta). Two IgK locus-derived YACs each contain three variable (V kappa) genes, the joining (J kappa) region, the intronic enhancer (E kappa), the constant gene (C kappa), and the kappa deleting element (kde). The IgH YAC was unstable in yeast, generating a variety of deletion derivatives, whereas both IgK YACs were stable. YACs encoding heavy chain and kappa light chain, retrofitted with the mammalian selectable marker, hypoxanthine phosphoribosyltransferase (HPRT), were each introduced into HPRT-deficient mouse ES cells. Analysis of YAC integrity in ES cell lines revealed that the majority of DNA inserts were integrated in substantially intact form.

We describe a strategy for producing human monoclonal antibodies in mice by introducing large segments of the human heavy and kappa light chain loci contained on yeast artificial chromosomes into the mouse germline. Such mice produce a diverse repertoire of human heavy and light chains, and upon immunization with tetanus toxin have been used to derive antigen-specific, fully human monoclonal antibodies. Breeding such animals with mice engineered by gene targeting to be deficient in mouse immunoglobulin (Ig) production has led to a mouse strain in which high levels of antibodies are produced, mostly comprised of both human heavy and light chains. These strains should provide insight into the adoptive human antibody response and permit the development of fully human monoclonal antibodies with therapeutic potential.

We found choroidal tubercles in two children with meningitis. This finding supposed an important clue in establishing a tuberculous etiology. Following, we discuss the evolution and fluorescein angiographic findings of choroidal tubercles. As many authors have remarked, a thorough fundus examination is of great value in cases of fever of undetermined origin and meningitis. Choroidal tubercles can also be the first sign of a common pulmonary or extrapulmonary tuberculosis.

Cytogenetic analysis of liposarcomas has demonstrated that translocation (12;16) (q13.3;p11.2) is characteristic of the myxoid subtype of this adipose tissue tumor. Our previous results suggested that the GLI gene is close to the translocation breakpoint on chromosome 12. We now describe a yeast artificial chromosome (YAC) that contains GLI and spans the chromosome 12 region involved in the t(12;16) breakpoint. This clone will permit rapid definition of the genetic region surrounding the breakpoint and allow isolation of the gene presumably affected by the translocation.

The small-cell lung carcinoma cell line U2020 contains a submicroscopic, homozygous deletion that removes a chromosomal segment within 3p13-p14, including the locus D3S3. We have sublocalized 49 additional probes to the 3p13-p14.2 region and have identified 7 new DNA markers that arise from within the U2020 deletion. The estimated size of the deletion, based on marker density, is approximately 4-5 megabases (Mb). Including D3S3, 7 of the 8 markers have been linked by pulsed-field gel (PFG) electrophoresis over an area of approximately 2 Mb. Including the one unlinked marker, PFG analysis accounts for about 3 Mb of the region. The U2020 deletion appears confined to the 3p13-p14.2 region and does not include the candidate tumor suppressor gene, protein-tyrosine phosphatase gamma (PTPG).

A new method for screening of YAC libraries is described. Individual YACs were pooled into groups of 384 clones and prepared as samples suitable for pulsed-field gel electrophoresis. A five hit human YAC library (Brownstein et al., 1989) containing approximately 60,000 clones was condensed into 150 such pools and chromosomal DNAs in each sample were separated on three pulsed field gels containing 50 samples each. Southern blots prepared from these gels were hybridized with probes of interest to identify pools containing homologous YACs. Further purification was performed using standard colony hybridization procedures. Twenty-one probes used thus far have identified 47 positive pools and corresponding YACs have been purified from 28 of these. Some significant advantages of this method include avoidance of DNA sequence analysis and primer generation prior to YAC screening and the ability to handle the entire library on three filters. The screening approach described here permits rapid isolation of YACs corresponding to unsequenced loci and will accelerate establishment of YAC contigs for large chromosomal segments.

We describe the cytogenetic findings in a recurrent neurofibrosarcoma in a patient with nonfamilial von Recklinghausen disease. The composite karyotype was: 40,Y,-X,+dic r(X;20)(:Xp22.2----q26::20p13----q13:), -1, +der(1)t(1;3) (p21;p24),-3,-4,-5,+der(5) t(5;?)(q31;?),-9,-9,+der(9)t(3;9)(q21 or q13;p24 or p22), -11,+der(11)t(11;?)(q22.2;?), -17,+der(17)t(17; 22;?)(q21;q13.1;?), -20, -21, -22, -22, +der(22)t(17; 22;?)(q21;q13.1;?),t(2;10)(q37;q22). The derivative chromosomes were demonstrated at the 500 band level. Chromosomes 17 and 22 were shown to be involved in an unbalanced three-way translocation: t(17;22;?)(q21;q13.1;?). This event was confirmed by in situ hybridization, using two probes mapped to chromosome 17. Hill H is a probe derived from the novel oncogene TRE and is located at 17q12-22. The second probe, derived from the granulocyte colony-stimulating factor (G-CSF), is located at 17q11-q21. The rearrangement between chromosomes 17 and 22 showed breakpoints similar or close to the gene loci for neurofibromatosis 1 (NF-1) and NF-2. Based on our observations we recommend that genetic studies on NF-1 tumors include both gene sites (NF-1 and NF-2) rather than focus on one gene locus.

The integration sites of human papillomavirus (HPV) DNA within the cervical carcinoma cell line C4-I and a primary cervical tumor were mapped by in situ hybridization. Cloned cellular sequences flanking the integrated viral DNA were used as probes. For the cell line, the viral integration site was mapped to chromosome region 8q21-q22.3, while in the primary tumor chromosome band 3p21 was the target for integration. The HPV DNA integration appears to occur in the vicinity of fragile sites, oncogenes, and chromosome breakpoints that are characteristic of hematologic malignancies and solid tumors. The integration of HPV may thus promote chromosome changes in cancer cells.