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Background: We report on the development and initial evaluation of a novel computerized decision support system (CDSS) that utilizes concepts from entertainment education (edutainment) to assist low-literate, multiethnic women in making initial surgical treatment decisions. Method: We randomly assigned 51 patients diagnosed with early stage breast cancer to use the decision aid. Results: Patients who viewed the CDSS improved their knowledge of breast cancer treatment; found the application easy to use and understand, informative, and enjoyable; and were less worried about treatment. Conclusion: The system clearly reached its intended objectives to create a usable decision aid for low-literate, novice computer users. [ABSTRACT FROM AUTHOR]

Background: Chronic obstructive pulmonary disease (COPD) is an inflammatory airway disease characterized by emphysema and chronic bronchitis and a leading cause of mortality worldwide. COPD is commonly associated with several comorbid diseases which contribute to exacerbated patient outcomes. Cigarette smoke (CS) is the most prominent risk factor for COPD development and progression and is known to be detrimental to numerous effector functions of lung resident immune cells, including phagocytosis and cytokine production. However, how CS mediates the various pathologies distant from the lung in COPD, and whether CS has a similar biological effect on systemic immune cells remains unknown., Methods: C57BL/6 mice were exposed to 8 weeks of CS as an experimental model of COPD. Bone marrow cells were isolated from both CS-exposed and room air (RA) control mice and differentiated to bone marrow-derived macrophages (BMDMs). Airspace macrophages (AMs) were isolated from the same CS-exposed and RA mice and bulk RNA-Seq performed. The functional role of differentially expressed genes was assessed through gene ontology analyses. Ingenuity Pathway Analysis was used to determine the activation states of canonical pathways and upstream regulators enriched in differentially expressed genes in both cell types, and to compare the differences between the two cell types., Results: CS induced transcriptomic changes in BMDMs, including an upregulation of genes in sirtuin signalling and oxidative phosphorylation pathways and a downregulation of genes involved in histone and lysine methylation. In contrast, CS induced decreased expression of genes involved in pathogen response, phagosome formation, and immune cell trafficking in AMs. Little overlap was observed in differentially expressed protein-coding genes in BMDMs compared to AMs and their associated pathways, highlighting the distinct effects of CS on immune cells in different compartments., Conclusions: CS exposure can induce transcriptomic remodelling in BMDMs which is distinct to that of AMs. Our study highlights the ability of CS exposure to affect immune cell populations distal to the lung and warrants further investigation into the functional effects of these changes and the ensuing role in driving multimorbid disease., (© 2024. The Author(s).)

Cigarette smoke (CS) exposure is a risk factor for many chronic diseases, including chronic obstructive pulmonary disease, but the mechanism by which smoke exposure can alter homeostasis and bring about chronic inflammation is poorly understood. Here, we showcase a novel role for smoke in regulating long noncoding RNAs, showing that it activates lincRNA-Cox2 , which we previously characterized as functional in inflammatory regulation. Exposing lincRNA-Cox2 murine models to smoke in vivo confirmed lincRNA-Cox2 as a regulator of inflammatory gene expression in response to smoke both systemically and within the lung. We also report that lincRNA-Cox2 negatively regulates genes in smoked bone marrow-derived macrophages exposed to LPS stimulation. In addition to the effects on long noncoding RNAs, we also report dysregulated transcription and splicing of inflammatory protein-coding genes in the bone marrow niche after CS exposure in vivo . Collectively, this work provides insights into how innate immune signaling from gene expression to splicing is altered after in vivo exposure to CS and highlights an important new role for lincRNA-Cox2 in regulating immune genes after smoke exposure.

Helicobacter pylori colonizes half of the world's population and is responsible for a significant disease burden by causing gastritis, peptic ulcers, and gastric cancer. The development of host inflammation drives these diseases, but there are still open questions in the field about how H. pylori controls this process. We characterized H. pylori inflammation using an 8-month mouse infection time course and comparison of the wild type (WT) and a previously identified mutant lacking the TlpA chemoreceptor that causes elevated inflammation. Our work shows that H. pylori chronic-stage corpus inflammation undergoes surprising fluctuations, with changes in Th17 and eosinophil numbers. The H. pylori tlpA mutant changed the inflammation temporal characteristics, resulting in different inflammation from the wild type at some time points. tlpA mutants have equivalent total and gland colonization in late-stage infections. During early infection, in contrast, they show elevated gland and total colonization compared to those by WT. Our results suggest the chronic inflammation setting is dynamic and may be influenced by colonization properties of early infection.

Our respiratory system is vital to protect us from the surrounding nonsterile environment; therefore, it is critical for a state of homeostasis to be maintained through a balance of inflammatory cues. Recent studies have shown that actively transcribed noncoding regions of the genome are emerging as key regulators of biological processes, including inflammation. lincRNA-Cox2 is one such example of an inflammatory inducible long intergenic noncoding RNA functioning to fine-tune immune gene expression. Using bulk and single-cell RNA sequencing, in addition to FACS, we find that lincRNA-Cox2 is most highly expressed in the lung and is most upregulated after LPS-induced lung injury (acute lung injury [ALI]) within alveolar macrophages, where it functions to regulate inflammation. We previously reported that lincRNA-Cox2 functions to regulate its neighboring protein Ptgs2 in cis , and in this study, we use genetic mouse models to confirm its role in regulating gene expression more broadly in trans during ALI. Il6, Ccl3, and Ccl5 are dysregulated in the lincRNA-Cox2 -deficient mice and can be rescued to wild type levels by crossing the deficient mice with our newly generated lincRNA-Cox2 transgenic mice, confirming that this gene functions in trans. Many genes are specifically regulated by lincRNA-Cox2 within alveolar macrophages originating from the bone marrow because the phenotype can be reversed by transplantation of wild type bone marrow into the lincRNA-Cox2 -deficient mice. In conclusion, we show that lincRNA-Cox2 is a trans -acting long noncoding RNA that functions to regulate immune responses and maintain homeostasis within the lung at baseline and on LPS-induced ALI., (Copyright © 2022 by The American Association of Immunologists, Inc.)

Chlorpyrifos, an organophosphate (OP) insecticide, is prevalent in aquatic systems globally and is often implicated in aquatic toxicity during storm events. Chlorpyrifos induces toxicity by inhibition of acetylcholinesterase (AChE) activity, which has been related to alterations to fish swimming performance. Resistance to organophosphate insecticides, including chlorpyrifos, is prevalent in populations of the epibenthic amphipod Hyalella azteca in areas with known OP exposure. Previous studies have demonstrated an elevated bioaccumulation potential of insecticide-resistant prey items, however the potential for trophic transfer of chlorpyrifos from OP-resistant prey items and associated neurotoxic effects in fish predators has not been studied. Consequently, the present study aimed to determine the potential for trophic transfer of chlorpyrifos from OP-resistant H. azteca to a known predator, the inland silverside, Menidia beryllina at two temperatures (18 and 23 °C) to simulate temperature changes associated with global climate change (GCC). Fish were fed either 14 C-chlorpyrifos-dosed H. azteca or control animals for 7 d, after which total bioaccumulation, percent parent chlorpyrifos, brain AChE activity and swimming performance (ramp-U crit ) were determined. Fish fed chlorpyrifos-dosed H. azteca bioaccumulated chlorpyrifos ranging from 29.9 to 1250 ng/g lipid, demonstrating the potential for trophic transfer. Lower bioaccumulation and greater biotransformation were observed in M. beryllina at 23 °C as compared to 18 °C, though this was not statistically significant. A significant 36.5% reduction in brain AChE activity was observed in fish fed chlorpyrifos-dosed H. azteca at 23 °C only, which may be attributed to increased biotransformation of parent chlorpyrifos to more potent AChE-inhibiting metabolites. Dietary chlorpyrifos exposure had no significant effect on swimming performance in M. beryllina, though ramp-U crit was significantly increased by 25% at 23 as compared to 18 °C. These findings confirm the potential for trophic transfer of chlorpyrifos from OP-resistant prey to fish predators and the potential for elevated temperatures to exacerbate the neurotoxic effects of chlorpyrifos., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The burden of asthma disproportionately affects minority and low-income communities, resulting in racial and socioeconomic disparities in asthma prevalence, asthma exacerbations, and asthma-related death. Social determinants of health are increasingly implicated as root causes of disparities, and healthy housing is perhaps the most critical social determinant in asthma health disparities. In many minority communities, poor housing conditions and value are a legacy of historical policies and practices imbued with structural racism, including redlining, displacement, and exclusionary zoning. As a result, poor-quality, substandard housing is a characteristic feature of many underrepresented minority communities. Consequently, structurally deficient housing stock cultivates home environments rife with indoor asthma triggers. In this review we consider the historical context of urban housing policies and practices and how these policies and practices have contributed to the substandard housing conditions for many minoritized children in the present day. We describe the impact of poor housing quality on asthma and interventions that have attempted to mitigate its influence on asthma symptoms and health care utilization. We discuss the need to promote asthma health equity by reinvesting in these neighborhoods and communities to provide healthy housing., (Copyright © 2021 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Determining the layers of gene regulation within the innate immune response is critical to our understanding of the cellular responses to infection and dysregulation in disease. We identified a conserved mechanism of gene regulation in human and mouse via changes in alternative first exon (AFE) usage following inflammation, resulting in changes to the isoforms produced. Of these AFE events, we identified 95 unannotated transcription start sites in mice using a de novo transcriptome generated by long-read native RNA-sequencing, one of which is in the cytosolic receptor for dsDNA and known inflammatory inducible gene, Aim2 . We show that this unannotated AFE isoform of Aim2 is the predominant isoform expressed during inflammation and contains an iron-responsive element in its 5'UTR enabling mRNA translation to be regulated by iron levels. This work highlights the importance of examining alternative isoform changes and translational regulation in the innate immune response and uncovers novel regulatory mechanisms of Aim2 ., Competing Interests: ER, PJ, SC, MC, VS, RS, BS, RA, MJ, SC, SC No competing interests declared, MA holds options in Oxford Nanopore Technologies (ONT), is a paid consultant to ONT, received reimbursement for travel, accommodation and conference fees to speak at events organized by ONT,received research funding from ONT and is an inventor on 11 UC patents licensed to ONT (6,267,872, 6,465,193, 6,746,594, 6,936,433, 7,060,50, 8,500,982, 8,679,747, 9,481,908, 9,797,013, 10,059,988, and 10,081,835), AB received reimbursement for travel, accommodation and conference fees to speak at events organized by Oxford Nanopore Technologies (ONT), (© 2021, Robinson et al.)

Macrophages and dendritic cells (DCs) are innate immune cells that play a key role in defense against pathogens. In vitro cultures of bone marrow-derived macrophages (BMDMs) and dendritic cells (BMDCs) are well-established and valuable methods for immunological studies. Typically, commercially available recombinant GM-CSF is utilized to generate BMDCs and is also used to culture alveolar macrophages. We have generated a new HEK-293T cell line expressing murine GM-CSF that secretes high levels of GM-CSF (~180 ng/ml) into complete media as an alternative to commercial GM-CSF. Differentiation of dendritic cells and expression of various markers were kinetically assessed using the GM-CSF HEK293T cell line, termed supGM-CSF and compared directly to purified commercial GMCSF. After 7-9 days of cell culture the supGM-CSF yielded twice as many viable cells compared to the commercial purified GM-CSF. In addition to differentiating BMDCs, the supGM-CSF can be utilized to culture functionally active alveolar macrophages. Collectively, our results show that supernatant from our GM-CSF HEK293T cell line supports the differentiation of mouse BMDCs or alveolar macrophage culturing, providing an economical alternative to purified GM-CSF., Competing Interests: The authors have declared that no competing interests exist.

Purpose: The aim of this study was to determine the upgrade rate of image-guided core needle biopsy (CNB)-proven benign breast intraductal papillomas (IDPs) without atypia to high-risk benign lesions or malignancy after surgical excision., Methods: A retrospective database search at a single institution identified 102 adult female patients with benign breast IDPs without atypia diagnosed on imaging-guided CNBs who subsequently had surgical excisions between 2011 and 2016. Patient characteristics, imaging features, biopsy techniques, and the pathology reports from imaging-guided CNBs and subsequent surgical excisions were reviewed. The upgrade rate to malignancies or high-risk benign lesions was determined at the patient level., Results: The upgrade rate to malignancy was 2.9% (3/102), including two cases of ductal carcinoma in situ (DCIS) and one case of microinvasive (< 1 mm) ductal carcinoma arising from DCIS. The upgrade rate to high-risk benign lesions was 7.8% (8/102), with seven cases of atypical ductal hyperplasia and one case of atypical lobular hyperplasia. A personal history of breast cancer and a larger mean lesion size were significantly associated with an upgrade to malignancy (p < 0.05)., Conclusions: The management of benign breast IDPs without atypia detected on imaging-guided CNBs is controversial. Our results suggest risk stratification is important in approaching these patients. Although surgical excision should be considered for all benign breast IDPs without atypia, observation with serial imaging may be appropriate in selected low-risk patients. This approach will save many women from surgeries and decrease the cost of medical care.

Macrophages are critical effector cells of the immune system, and understanding genes involved in their viability and function is essential for gaining insights into immune system dysregulation during disease. We use a high-throughput, pooled-based CRISPR-Cas screening approach to identify essential genes required for macrophage viability. In addition, we target 3' UTRs to gain insights into previously unidentified cis-regulatory regions that control these essential genes. Next, using our recently generated nuclear factor κB (NF-κB) reporter line, we perform a fluorescence-activated cell sorting (FACS)-based high-throughput genetic screen and discover a number of previously unidentified positive and negative regulators of the NF-κB pathway. We unravel complexities of the TNF signaling cascade, showing that it can function in an autocrine manner in macrophages to negatively regulate the pathway. Utilizing a single complex library design, we are capable of interrogating various aspects of macrophage biology, thus generating a resource for future studies., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Axonal dystrophy, indicative of perturbed axonal transport, occurs early during Alzheimer's disease (AD) pathogenesis. Little is known about the mechanisms underlying this initial sign of the pathology. This study proves that Golgi-localized γ-ear-containing ARF binding protein 3 (GGA3) loss of function, due to Gga3 genetic deletion or a GGA3 rare variant that cosegregates with late-onset AD, disrupts the axonal trafficking of the β-site APP-cleaving enzyme 1 (BACE1) resulting in its accumulation in axonal swellings in cultured neurons and in vivo. We show that BACE pharmacological inhibition ameliorates BACE1 axonal trafficking and diminishes axonal dystrophies in Gga3 null neurons in vitro and in vivo. These data indicate that axonal accumulation of BACE1 engendered by GGA3 loss of function results in local toxicity leading to axonopathy. Gga3 deletion exacerbates axonal dystrophies in a mouse model of AD before β-amyloid (Aβ) deposition. Our study strongly supports a role for GGA3 in AD pathogenesis, where GGA3 loss of function triggers BACE1 axonal accumulation independently of extracellular Aβ, and initiates a cascade of events leading to the axonal damage distinctive of the early stage of AD., (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

This study aims at prospectively evaluating the difference in the effect of cholecystokinin (CCK) and half-and-half milk (HHM) administered in the same patient on gallbladder contractility and correlation with clinical outcomes. Upon gallbladder visualization during standard hepatobiliary imaging, 0.02 μg/kg of CCK was injected over 3 min, and additional 30 min of dynamic imaging was obtained. Patients with gallbladder ejection fraction (GBEF) <35% after CCK were administered 8 oz of HHM followed by 30 min of imaging. The GBEF was recalculated. The number of patients whom GBEF changed from below 35% (abnormal) after CCK to above 35% (normal) after HHM was recorded. Follow-up of the clinical outcome at 6 months was performed. Fifty patients with abnormal GBEF were prospectively included. The average GBEF after CCK was 14.7% ± 8.5% and after HHM was 30.7% ± 20.8%. The average increase in GBEF with HHM was 16.0% ± 22.2%. The GBEF changed from abnormal to normal in 17 patients (34%). The remaining 33 patients remained abnormal. Clinical outcomes at 6 months were available in 47 patients. Cholecystectomy was performed in 60% of patients with abnormal GBEF with CCK and HHM with resolution or improvement of pain. Two of 16 patients (12%) with abnormal GBEF after CCK but normal after HHM had cholecystectomies with pain improvement, while 8 out of these patients (50%) were diagnosed and treated with other disorders and improved. Hepatobiliary imaging with HHM stimulation is a superior physiologic test which can lower the number of unnecessary cholecystectomies and misdiagnoses as functional cholecystitis., Competing Interests: There are no conflicts of interest., (Copyright: © 2020 World Journal of Nuclear Medicine.)

Next-generation sequencing has provided a more complete picture of the composition of the human transcriptome indicating that much of the "blueprint" is a vastness of poorly understood non-protein-coding transcripts. This includes a newly identified class of genes called long noncoding RNAs (lncRNAs). The lack of sequence conservation for lncRNAs across species meant that their biological importance was initially met with some skepticism. LncRNAs mediate their functions through interactions with proteins, RNA, DNA, or a combination of these. Their functions can often be dictated by their localization, sequence, and/or secondary structure. Here we provide a review of the approaches typically adopted to study the complexity of these genes with an emphasis on recent discoveries within the innate immune field. Finally, we discuss the challenges, as well as the emergence of new technologies that will continue to move this field forward and provide greater insight into the biological importance of this class of genes. This article is part of a Special Issue entitled: ncRNA in control of gene expression edited by Kotb Abdelmohsen., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Macrophages undergo metabolic changes during activation that are coupled to functional responses. The gram negative bacterial product lipopolysaccharide (LPS) is especially potent at driving metabolic reprogramming, enhancing glycolysis and altering the Krebs cycle. Here we describe a role for the citrate-derived metabolite malonyl-CoA in the effect of LPS in macrophages. Malonylation of a wide variety of proteins occurs in response to LPS. We focused on one of these, glyceraldehyde-3-phosphate dehydrogenase (GAPDH). In resting macrophages, GAPDH binds to and suppresses translation of several inflammatory mRNAs, including that encoding TNFα. Upon LPS stimulation, GAPDH undergoes malonylation on lysine 213, leading to its dissociation from TNFα mRNA, promoting translation. We therefore identify for the first time malonylation as a signal, regulating GAPDH mRNA binding to promote inflammation.

An inducible gene expression program is a hallmark of the host inflammatory response. Recently, long intergenic non-coding RNAs (lincRNAs) have been shown to regulate the magnitude, duration, and resolution of these responses. Among these is lincRNA-Cox2, a dynamically regulated gene that broadly controls immune gene expression. To evaluate the in vivo functions of this lincRNA, we characterized multiple models of lincRNA-Cox2-deficient mice. LincRNA-Cox2-deficient macrophages and murine tissues had altered expression of inflammatory genes. Transcriptomic studies from various tissues revealed that deletion of the lincRNA-Cox2 locus also strongly impaired the basal and inducible expression of the neighboring gene prostaglandin-endoperoxide synthase (Ptgs2), encoding cyclooxygenase-2, a key enzyme in the prostaglandin biosynthesis pathway. By utilizing different genetic manipulations in vitro and in vivo, we found that lincRNA-Cox2 functions through an enhancer RNA mechanism to regulate Ptgs2. More importantly, lincRNA-Cox2 also functions in trans, independently of Ptgs2, to regulate critical innate immune genes in vivo., (Published by Elsevier Inc.)

The innate immune system protects against infections by initiating an inducible inflammatory response. NF-κB is one of the critical transcription factors controlling this complex response, but some aspects of its regulation remain unclear. For example, although long non-coding RNAs (lncRNAs) have been shown to critically regulate gene expression, only a fraction of these have been functionally characterized, and the extent to which lncRNAs control NF-κB expression is unknown. Here, we describe the generation of a GFP-based NF-κB reporter system in immortalized murine bone marrow-derived macrophages (iBMDM). Activation of this reporter, using Toll-like receptor ligands, resulted in GFP expression, which could be monitored by flow cytometry. We also established a CRISPR/Cas9 gene deletion system in this NF-κB reporter line, enabling us to screen for genes that regulate NF-κB signaling. Our deletion-based approach identified two long intergenic non-coding(linc)RNAs, lincRNA-Cox2 and lincRNA-AK170409, that control NF-κB signaling. We demonstrate a potential novel role for lincRNA-Cox2 in promoting IκBα degradation in the cytoplasm. For lincRNA-AK170409, we provide evidence that this nuclearly-localized lincRNA regulates a number of inflammation-related genes. In conclusion, we have established an NF-κB-GFP iBMDM reporter cell line and a line that stably expresses Cas9. Our approach enabled the identification of lincRNA-Cox2 and lincRNA-AK170409 as NF-κB regulators, and this tool will be useful for identifying additional genes involved in regulating this transcription factor critical for immune function., (© 2017 by The American Society for Biochemistry and Molecular Biology, Inc.)

Background: Breast cancer survival disparities by race are likely multifactorial. In a small pilot cohort, we demonstrated a statistical interaction between age and race. The purpose of this study was to validate earlier findings in a larger, more diverse cohort and to test the hypothesis that breast cancer survival is influenced by the dependent relationship of age and race., Methods: We conducted a retrospective analysis of a multi-institutional breast cancer database for patients treated between 1999 and 2009. Study variables included age and disease stage at diagnosis, race, treatment (surgery, chemotherapy, radiotherapy, hormone therapy) and overall survival. Statistical analysis and regression models were performed by Stata software., Results: A total of 9,249 patients were included in this study. African American, Hispanic, and Asian patients were more likely to present at a younger age with metastases. African American and Hispanic race were associated with increased mortality after adjusting for stage, age, and treatment. A 2-way interaction between age and race was identified in the Cox regression model (p < 0.001). To further define this interaction, a postestimation analysis was performed to determine the predicted relative hazard for each race with age fixed at 40, 50, 60, 70, and 80 years. At younger ages, the predicted relative hazard was significantly higher for both African American and Hispanic race., Conclusions: Despite adjusting for stage and treatment differences, African American and Hispanic race predicted poor survival. The effect of age and treatment on breast cancer survival differs across races. Additional research is needed to accurately determine the reasons for worsened survival.

Background: Undocumented immigrants have been shown to be predisposed to worse clinical outcomes than the general population. This study examines survival in socioeconomically disadvantaged Hispanic documented and undocumented breast cancer patients., Methods: Analysis of a prospective breast cancer database of patients treated in a safety-net hospital system. Overall survival was the primary outcome, and advanced stage at diagnosis (regional and metastatic) was a secondary outcome. Survival analysis and multivariate regression modeling were performed., Results: Seven hundred fifty-one breast cancer patients were identified. Undocumented patients presented at an earlier age and were likely to present with advanced stage. After adjusting for covariates, undocumented status was not associated with increased mortality. The diagnosis-to-treatment interval was significantly longer in undocumented patients., Conclusions: Despite undocumented patients presenting at a younger age, they have similar mortality compared with documented patients. This finding is partly explained by the local treatment afforded by undocumented patients, further studies are necessary to detail the reasons for these differences in presentation and outcome., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Objective: To evaluate an entertainment-based patient decision aid for early stage breast cancer surgery in low health literacy patients., Methods: Newly diagnosed female patients with early stage breast cancer from two public hospitals were randomized to receive an entertainment-based decision aid for breast cancer treatment along with usual care (intervention arm) or to receive usual care only (control arm). Pre-decision (baseline), pre-surgery, and 1-year follow-up assessments were conducted., Results: Patients assigned to the intervention arm of the study were more likely than the controls to choose mastectomy rather than breast-conserving surgery; however, they appeared better informed and clearer about their surgical options than women assigned to the control group. No differences in satisfaction with the surgical decision or the decision-making process were observed between the patients who viewed the intervention and those assigned to the control group., Conclusions: Entertainment education may be a desirable strategy for informing lower health literate women about breast cancer surgery options., Practice Implications: Incorporating patient decision aids, particularly computer-based decision aids, into standard clinical practice remains a challenge; however, patients may be directed to view programs at home or at public locations (e.g., libraries, community centers)., (Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.)

Background: Matrix metalloproteinases (MMPs) are a group of endopeptidases induced under inflammatory conditions in the intestine which possess the capacity to degrade components of the extracellular matrix. We have previously demonstrated that MMP-2 expression correlates with increased inducible nitric oxide synthase (iNOS) production in the stomach and that iNOS is upregulated in the postischemic gut by the luminal nutrient arginine and repressed by luminal glutamine. We therefore hypothesized that arginine would enhance expression of MMP-2 in the postischemic gut., Methods: Jejunal sacs were created in rats at laparotomy and filled with either 60 mM glutamine, arginine, or magnesium sulfate (osmotic control) followed by 60 minutes of superior mesenteric artery occlusion (SMAO) and 6 hours of reperfusion and compared with shams. Jejunum was harvested, and membrane type-1 matrix metalloproteinase (MT1-MMP), MMP-2, and iNOS protein expression was determined by Western analysis and MMP-9 production by gelatin zymography., Results: MMP-2, MT1-MMP, MMP-9, and iNOS were all increased after SMAO compared with shams. Arginine maintained while glutamine inhibited the increase in iNOS, MT1-MMP, and MMP-2 expression in the postischemic gut. Pretreatment of the arginine group with a selective iNOS inhibitor blunted the induction of MMP-2 in the postischemic gut. There was no differential modulation of MMP-9 by the luminal nutrients., Conclusions: The arginine-induced upregulation of iNOS may contribute to increased activity of MT1-MMP and MMP-2. The mechanism for this differential regulation by arginine warrants further investigation.

Objective: To report on the initial testing of a values clarification exercise utilizing a jewellery box within a computerized patient decision aid (CPtDA) designed to assist women in making a surgical breast cancer treatment decision., Design: Pre-post design, with patients interviewed after diagnosis, and then after completing the CPtDA sometime later at their preoperative visit., Sample: Fifty-one female patients, who are low literate and naïve computer users, newly diagnosed with early stage breast cancer from two urban public hospitals., Intervention: A computerized decision aid that combines entertainment-education (edutainment) with enhanced (factual) content. An interactive jewellery box is featured to assist women in: (1) recording and reflecting over issues of concern with possible treatments, (2) deliberating over surgery decision, and (3) communicating with physician and significant others., Outcomes: Patients' use of the jewellery box to store issues during completion of the CPtDA, and perceived clarity of values in making a treatment decision, as measured by a low literacy version of the Decisional Conflict Scale (DCS)., Results: Over half of the participants utilized the jewellery box to store issues they found concerning about the treatments. On average, users flagged over 13 issues of concern with the treatments. Scores on the DCS Uncertainty and Feeling Unclear about Values subscales were lower after the intervention compared to before the decision was made., Conclusions: A values clarification exercise using an interactive jewellery box may be a promising method for promoting informed treatment decision making by low literacy breast cancer patients.

Background: Increased matrix metalloproteinase (MMP) activity is associated with tissue injury in some organs. Their role in gut injury remains to be fully elucidated. We recently demonstrated that increased MMP-2 activity participated in lipopolysaccharide (LPS)-induced gastric injury. Thus we hypothesized that MMPs may play a role in other models of gastric injury., Materials and Methods: The effect of L-NAME (10 mg/kg IP) or salicylate (100 mg/kg IP) on gastric injury from 20% ethanol was evaluated in an anesthetized model of gastric injury. In a separate experiment, gastric metalloproteinase activity was assessed after salicylate or L-NAME administration. Rats were given either L-NAME (10 mg/kg), salicylate (100 mg/kg), or saline IP and sacrificed after 6 hours. Gastric mucosa was harvested and portions of the glandular stomach snap frozen for gelatin and in situ zymography as indices of MMP activity. Subsequently the effect of MMP inhibition on macroscopic gastric injury from salicylate and a dilute luminal irritant was determined., Results: Both L-NAME and salicylate significantly increased gastric injury from 20% ethanol versus saline controls. Salicylate treatment significantly increased gelatinase activity as determined by in situ zymography and gelatin zymography while L-NAME did not. MMP inhibition ameliorated macroscopic gastric injury secondary to salicylate and a dilute luminal irritant., Conclusions: This is the first study to report that MMP activity increases in the stomach following salicylate treatment. These data suggest that MMPs may play a role in the ability of salicylate to exacerbate gastric injury from irritants, but likely do not play a role in mediating the deleterious effects of L-NAME.

Unlabelled: Matrix metalloproteinases (MMPs) degrade the extracellular matrix and contribute to LPS-induced gastric injury. MMPs are closely modulated by their activators, membrane type-MMP (MT-MMPs) and their endogenous inhibitors, the tissue inhibitors of metalloproteinases (TIMPs). As LPS-induced gastric injury is mediated in part by iNOS, and NO modulates MMP production in vitro, we hypothesized that NOS inhibition would similarly modulate LPS-induced gastric MMP production. Therefore, the purpose of these studies was to compare the effects of selective and nonselective NOS inhibition on LPS-induced gastric MMP production., Methods: Sprague-Dawley rats were given either the nonselective NOS inhibitor NG-nitro-L-arginine methyl ester (L-NAME; 5 mg/kg, s.c.), a selective iNOS inhibitor, aminoguanidine (45 mg/kg, i.p.) or L-N-iminoethyl-lysine (L-NIL; 10 mg/kg, i.p.), or vehicle 15 min before saline or LPS (20 mg/kg, i.p.) and killed 24 h after LPS administration. Stomachs were assessed for macroscopic injury (computed planimetry), and gastric mucosal MMP production was assessed by gelatin zymography, in situ zymography, and Western analysis for MMP-2, MT1-MMP, and TIMP-2. (n > or = 4/group; ANOVA)., Results: Aminoguanidine treatment decreased LPS-induced macroscopic gastric injury as well as MMP-2 and MT1-MMP protein production while having no effect on TIMP-2 protein levels. L-NIL similarly attenuated the induction of MMP-2 and MT1-MMP by LPS. L-NAME failed to attenuate LPS induced gastric injury or MT1-MMP protein induction and increased MMP-2 levels. L-NAME similarly had no effect on gastric TIMP-2 production., Conclusions: Selective iNOS inhibition decreases gastric MMP-2 activity after LPS administration, whereas nonselective inhibition increases MMP-2 levels. The ability of selective iNOS inhibition to ameliorate LPS-induced gastric injury may be due in part to its inhibition of active MMP-2 production, whereas nonselective NOS inhibitors increase MMP-2 levels and maintain gastric injury after LPS administration.

Despite continued investigation, the pathogenesis of tissue injury secondary to sepsis remains elusive. Further evaluation of the mechanisms by which endotoxemia and sepsis induce tissue injury is necessary to formulate rational and effective treatment strategies. The purpose of these studies was to evaluate the role of the matrix metalloproteinases MMP-2 and MMP-9 in gastric injury during lipopolysaccharide induced endotoxemia. Lipopolysaccharide increased gastric gelatinase activity as determined by in situ and gelatin zymography. Specifically, lipopolysaccharide induced MMP-2, MMP-9, and tissue inhibitor of metalloproteinase-1 (TIMP-1) transcription, with subsequent increases in MMP-2 and TIMP-1 protein expression. Furthermore, selective metalloproteinase inhibition ameliorated gastric injury in this model. These data suggest that lipopolysaccharide-induced gastric injury is mediated, at least in part, by increased MMP-2 production.

Background: This study was conducted to test the hypothesis that nonselective nitric oxide synthase (NOS) inhibitors have different effects on lipopolysaccharide (LPS)-induced gastric injury depending upon whether they are given concurrently with LPS or after LPS at a time point that inducible NOS is up-regulated., Materials and Methods: Female Sprague-Dawley rats received intraperitoneal (IP) LPS (20 mg/kg) for 3 h. Western immunoblot was used to determine iNOS, eNOS, and nNOS immunoreactivity after 3 h. In an additional set of experiments, we assessed the time dependent effects of nitric oxide synthase inhibition by giving rats LPS (20 mg/kg, IP) concurrently with Nitro-l-arginine methyl ester (l-NAME; 2-5 mg/kg, SC) or l-N(G)-(1-iminoethyl) lysine (l-NIL; 10 mg/kg, IP) for 5 h or LPS and delayed administration of l-NAME or l-NIL 3 h following LPS injection in identical doses. For these NOS inhibition experiments microscopic and macroscopic injury was assessed by a blinded observer using previously published scoring systems. Injury studies were conducted by exposing the stomach to 3 ml of 5 mM acidified taurocholate for 5 minutes in an anesthetized prep., Results: A 3-h treatment with LPS (20 mg/kg IP) significantly increased iNOS protein immunoreactivity (Western immunoblot) but not eNOS or nNOS. N(G)-l-NAME (2-5 mg/kg SC) dose dependently aggravated macroscopic (computerized planimetry) and morphological gastric injury caused by the intraluminal bile irritant 5 mm acidified taurocholate for 10 min when given concurrently with LPS, an effect reversed by l- but not D-arginine (300 mg/kg). In contrast, delayed administration of l-NAME (3 h after LPS) dose dependently attenuated the ability of LPS to exacerbate gastric injury from bile. Both concurrent and delayed administration of the selective iNOS inhibitor, l-NIL (10 mg/kg IP) attenuated the effects of LPS., Conclusions: These data indicate that during endotoxemia, the stomach is rendered more susceptible to damage from luminal irritants such as bile, a frequent occurrence in septic patients with a gastrointestinal ileus. In this setting, iNOS has a pathologic role while the constitutive NOS isoforms play gastroprotective roles.

Background: Short-term treatment with lipopolysaccharide (LPS) causes morphologic, but not macroscopic, gastric injury and decreases gastric injury caused by a subsequent challenge with a luminal irritant. This effect is abrogated by inducible nitric oxide synthase (iNOS) inhibition. The effects of long-term treatment with LPS on gastric injury are unknown as is the role of iNOS. We hypothesized that LPS would cause macroscopic gastric injury at later time points through an iNOS-dependent pathway., Methods: Conscious rats were given saline or LPS (1 or 20 mg/kg intraperitoneal) as a single intraperitoneal injection and killed 24 to 72 hours after injection. Macroscopic gastric injury (computerized planimetry), gastric luminal fluid volume and pH, and iNOS protein levels were assessed., Results: When compared with saline, high-dose but not low-dose LPS caused macroscopic gastric injury, increased gastric luminal fluid and pH, and up-regulated iNOS at 24 and 48 hours. All assessments returned to baseline by 72 hours. Inhibition of iNOS with 1400W (1 mg/kg intraperitoneal) given 15 minutes before saline or LPS (20 mg/kg) attenuated the deleterious effects of LPS on gastric injury and pH, but not fluid accumulation., Conclusions: These data suggest that prolonged treatment with high-dose LPS causes gastric injury through an iNOS-mediated pathway.

Background: Endotoxic shock can cause end-organ dysfunction and liver injury. Critically ill patients frequently require surgical intervention under general anesthesia for source control. However, the effects of anesthetics on organ function during sepsis and their influence on inflammatory mediators such as cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) remain to be fully elucidated. Because ketamine anesthesia has anti-inflammatory effects in some tissues, we hypothesized that it would attenuate lipopolysaccharide (LPS)-induced liver injury., Methods: Adult rats were given no anesthesia (saline), continuous isoflurane inhalation, or intraperitoneal (i.p.) injection of ketamine 70 mg/kg. One hour later, the rats received saline or LPS (20 mg/kg i.p.) for 5 hours. The rats were killed, and serum hepatocellular enzymes, liver COX-2, iNOS protein (Western immunoblot), and nuclear factor kappa B (NF-kappaB)-binding activity (electrophoretic mobility shift assay) determined. In a separate study, the role of COX-2 in LPS-induced liver injury was examined by pretreating rats with the selective COX-2 inhibitor NS-398 (3 mg/kg, i.p.) and the role of iNOS examined with the use of the selective inhibitor aminoguanidine (45 mg/kg, i.p.) 1 hour before LPS., Results: LPS increased serum aspartate aminotransferase and alanine aminotransferase levels, hepatic iNOS and COX-2 protein, and nuclear factor NF-kappaB. Ketamine, but not isoflurane, attenuated these effects caused by LPS. COX-2 inhibition with NS-398 as well as iNOS inhibition with aminoguanidine diminished LPS-induced changes in aspartate aminotransferase and alanine aminotransferase levels., Conclusions: These data indicate that anesthetics differ in their effects on liver injury caused by LPS. Ketamine has hepatoprotective effects, while isoflurane does not. Moreover, the protective effects of ketamine are mediated, at least in part, through a reduction in COX-2 and iNOS protein that could be regulated via changes in NF-kappaB-binding activity.

Background: The purpose of this study was to examine the presentation of diverticulitis at an urban county hospital serving predominantly indigent patients and to analyze the differences, if any, in presentation and treatment in younger patients., Methods: A retrospective review of medical records from 1995 to 2001 was performed at a single institution to identify patients admitted to the surgical service with the diagnosis of diverticular disease. Inclusion criteria were either diverticulitis confirmed at operation or radiographic findings consistent with the disease. Patient demographics, history, pertinent physical findings, and treatment were recorded. The data were analyzed after dividing the patients into two populations: a younger population 50 years of age or less, and a second population of patients older than 50., Results: During the interval, a total of 64 patients were admitted to the surgical service with the diagnosis of diverticulitis. The mean age of this population was 45.5 years (range 21 to 86). Forty-six patients were under 50 years of age (72%). Analysis of sex differences, type and timing of surgical procedure, and complication rate with respect to age showed no significant difference between the two age groups., Conclusions: We are clearly treating a younger patient population than previous reports on patients with diverticulitis. Although there was a trend toward increased surgical intervention in the younger population, this number did not reach statistical significance. Diverticulitis in young patients at our institution does not appear to take a more aggressive course than the same disease in older patients.

Lipopolysaccharide (LPS) and gut ischemia/reperfusion (I/R) injury cause reversible liver injury. Because nitric oxide (NO) can have both beneficial and deleterious effects in the gastrointestinal tract, and because the role of NO in gut I/R-induced hepatic injury is unknown, this study examined its role in LPS and gut I/R-induced hepatic injury in the rat. Both LPS and gut I/R caused a similar increase in serum hepatocellular enzymes. LPS but not gut I/R caused a significant increase in upregulation of hepatic inducible NO synthase (iNOS) according to quantitative real-time RT-PCR and Western immunoblot analysis. Aminoguanidine, a selective iNOS inhibitor, attenuated LPS-induced hepatic injury and hypotension, but did not prevent gut I/R-induced hepatic injury. In contrast, the non-selective NOS inhibitor N(G)-nitro-L-arginine methyl ester aggravated liver damage from both LPS and gut I/R. These data indicate that iNOS plays a role in mediating LPS-induced hepatic injury, but not gut I/R-induced hepatic injury. The data also suggest that the constitutive isoforms of NOS play a hepatoprotective role in both models of hepatic injury.

Background: Surgical wound infection and intra-abdominal abscess remain common infectious complications after appendectomy, especially in the setting of a perforated or gangrenous appendix. We therefore developed a clinical protocol for the management of appendicitis to decrease postoperative infectious complications., Methods: Between January 1, 1999, and December 31, 1999, 206 patients with appendicitis were treated on protocol. Retrospectively, the charts were reviewed for all protocol patients as well as for 232 patients with appendicitis treated in the year prior to protocol initiation. Data were collected on surgical wound infections and intra-abdominal abscesses., Results: There were significantly fewer infectious complications in the protocol group than in the nonprotocol group (20 [9%] versus 8 [4%]; P <0.05). In patients with a perforated or gangrenous appendix, the infectious complication rate was reduced from 33% to 13% (P <0.05)., Conclusions: The incidence of infectious complications after appendectomy can be significantly reduced with a standardized approach to antibiotic therapy and wound management.

Our previous studies have characterized Dexamethasone (Dex)-induced apoptotic signaling pathways in multiple myeloma (MM) cells; however, related transcriptional events are not fully defined. In the present study, gene expression profiles of Dex-treated MM cells were determined using oligonucleotide arrays. Dex triggers early transient induction of many genes involved in cell defense/repair-machinery. This is followed by induction of genes known to mediate cell death and repression of growth/survival-related genes. The molecular and genetic alterations associated with Dex resistance in MM cells are also unknown. We compared the gene expression profiles of Dex-sensitive and Dex-resistant MM cells and identified a number of genes which may confer Dex-resistance. Finally, gene profiling of freshly isolated MM patient cells validates our in vitro MM cell line data, confirming an in vivo relevance of these studies. Collectively, these findings provide insights into the basic mechanisms of Dex activity against MM, as well as mechanisms of Dex-resistance in MM cells. These studies may therefore allow improved therapeutic uses of Dex, based upon targeting genes that regulate MM cell growth and survival.

The authors hypothesized that IL-6 production by breast tumour tissues would correlate with OR-positivity, as only those tumours that contain oestrogen receptors (OR) and use oestrogen as a mitogen would benefit from locally increased oestrogen. IL-6 increases the activity of the 17beta-oxidoreductase, which converts oestrone to oestradiol, a process that may contribute to the increased concentration of oestrogen around breast tumours. IL-1alpha upregulates IL-6 production; therefore, the correlation between IL-1alpha and IL-6 immunoreactivity and OR-positivity in paraffin-embedded human breast tumours was further investigated.The results indicate IL-6 immunoreactivity in 40 of 66 paraffin embedded breast tumour specimens, a finding which did not correlate with the clinical evaluation of oestrogen receptor positivity (P=0.32 by Fisher's exact test). However, there was a correlation between IL-6 and IL-1alpha immunoreactivity (P<0.05). To study an in vitro model for this phenomenon, the IL-6 immunoreactivity in available cell lines was tested. Surprisingly, no production of IL-6 protein or mRNA could be detected in any of the cell lines, and this did not change with IL-1alpha stimulation. Therefore, none of the cell lines apparently reflected the immunological potential observed in the majority of surgical specimens., (Copyright 1998 Academic Press.)

Background: Clinical trials have suggested a survival advantage for selected patients with metastatic pancreatic cancer treated with tamoxifen. We sought to identify the molecular mechanism by which tamoxifen inhibits human pancreatic cancer cell (HPCC) growth., Methods: HPCCs were grown in tamoxifen and growth inhibition was determined by 3H-thymidine uptake and by the MTT assay; changes in cell viability were determined by cell counts. Cell cycle alterations were evaluated by FACS, and the induction of apoptosis was evaluated using the TUNEL assay. Total cellular RNA was isolated after tamoxifen treatment, and Northern blot analysis was performed for p21waf1., Results: Tamoxifen inhibited HPCC growth as measured by inhibition of 3H-thymidine incorporation and by the MTT assay. However, there was no decrease in the total number of viable cells after 6 days of treatment with 10 microM of tamoxifen and no evident apoptosis, confirming the absence of a cytotoxic effect. Cell cycle analysis revealed cellular arrest in the G0/G1 phase, which correlated with p21waf1 mRNA upregulation in response to tamoxifen treatment., Conclusions: Tamoxifen inhibits HPCC growth by inducing G0/G1 arrest with an associated increase in p21waf1 mRNA expression. Tamoxifen is an effective inhibitor of HPCC growth in vitro and warrants further in vivo study.

Pancreaticoduodenectomy is performed on carefully selected patients as part of a protocol-based clinical research program emphasizing the importance of multimodality management for patients with potentially resectable adenocarcinoma of the pancreatic head. Treatment schemas emphasize the importance of minimizing toxicity and treatment duration, while attempting to improve therapeutic efficacy. Cytotoxicity is enhanced by combining radiation therapy with more potent radiation-sensitizing agents. Because of the high incidence of liver metastases, systemic therapy is continued after chemoradiation and surgery with systemic agents of low toxicity directed at specific molecular events involved in pancreatic tumorigenesis such as inhibition of angiogenesis, induction of apoptosis, or arrest of the cell cycle.

Background: We have noted a continued increase in the number of patients referred to our institution for presumed or biopsy-proven periampullary carcinoma following an "exploratory" laparotomy during which tumor resection was not performed. Although previous work has demonstrated the safety of reoperative pancreaticoduodenectomy (PD), the need to avoid nontherapeutic laparotomy in these patients is obvious. In the current study, we sought to determine why PD was not performed at the initial operation., Methods: Using the prospective pancreatic cancer database, we identified all patients who underwent reoperative PD at our institution between June 1990 and October 1995. Radiologic imaging prior to reoperation was standardized and based on thin-section, contrast-enhanced computed tomography (CT); helical CT was used in more recent cases. Pathologic data were obtained, and initial outside operative reports were reviewed to determine why a PD was not performed at the initial procedure., Results: Twenty-nine patients underwent reoperative PD. Resection was not performed at the initial laparotomy because of the surgeon's assessment of local unresectability (17 patients), lack of a tissue diagnosis of malignancy (9), misdiagnoses (2), and error in intraoperative management (1). In the 17 patients deemed to have unresectable disease, successful reoperative PD required vascular resection in 10. All 10 of these patients had resection with negative microscopic margins of excision. Of the 9 patients who did not have resection owing to diagnostic uncertainty, all 9 had undergone multiple intraoperative biopsies interpreted as negative for malignancy; 6 of 9 had carcinoma confirmed on permanent-section analysis of the biopsy specimens. Four patients suffered major complications from intraoperative large-needle biopsy., Conclusions: Detailed preoperative imaging and a clearly defined operative plan would have allowed successful resection at the initial operation in 27 of 29 patients who underwent reoperative PD. Avoidable patient morbidity and the cost of unnecessary surgery argue strongly against "exploratory" surgery in patients with presumed periampullary neoplasms.

Developmentally controlled genomic deletion-ligations occur during ciliate macronuclear differentiation. We have identified a novel activity in Tetrahymena cell-free extracts that efficiently catalyzes a specific set of intramolecular DNA deletion-ligation reactions. When synthetic DNA oligonucleotide substrates were used, all the deletion-ligation products resembled those formed in vivo in that they resulted from deletions between pairs of short direct repeats. The reaction is ATP-dependent, salt-sensitive, and strongly influenced by the oligonucleotide substrate sequence. The deletion-ligation activity has an apparent size of 200-500 kd, no nuclease-sensitive component, and is highly enriched in cells developing new macronuclei. The temperature inactivation profile of the activity parallels the temperature lethality profile specific for Tetrahymena cells developing new macronuclei. We suggest that this deletion-ligation activity carries out the genomic deletions in developing macronuclei in vivo.