Scientific and Regulatory Policy Committee Points to Consider: Nonclinical Research and Development of In Vivo Gene Therapy Products, Emphasizing Adeno-Associated Virus Vectors.

Sequencing of the human genome and numerous advances in molecular techniques have launched the era of genetic medicine. Increasingly precise technologies for genetic modification, manufacturing, and administration of pharmaceutical-grade biologics have proved the viability of in vivo gene therapy (GTx) as a therapeutic modality as shown in several thousand clinical trials and recent approval of several GTx products for treating rare diseases and cancers. In recognition of the rapidly advancing knowledge in this field, the regulatory landscape has evolved considerably to maintain appropriate monitoring of safety concerns associated with this modality. Nonetheless, GTx safety assessment remains complex and is designed on a case-by-case basis that is determined by the disease indication and product attributes. This article describes our current understanding of fundamental biological principles and possible procedures (emphasizing those related to toxicology and toxicologic pathology) needed to support research and development of in vivo GTx products. This article is not intended to provide comprehensive guidance on all GTx modalities but instead provides an overview relevant to in vivo GTx generally by utilizing recombinant adeno-associated virus-based GTx-the most common in vivo GTx platform-to exemplify the main points to be considered in nonclinical research and development of GTx products.

Opinion on the Use of Animal Models in Nonclinical Safety Assessment: Pros and Cons.

Nonclinical evaluation of human safety risks for new chemical entities (NCEs) is primarily conducted in conventional healthy animals (CHAs); however, in certain instances, animal models of diseases (AMDs) can play a critical role in the understanding of human health risks. Animal models of diseases may be especially important when there is a need to understand how disease conditions associated with the intended indication might impact risk assessment of NCEs or when CHAs lack the human-specific target of interest (receptor, etc). Although AMDs have potential benefits over CHAs, they also have limitations. Understanding these limitations and optimizing the AMDs of interest should be done prior to proceeding with studies that will guide development of NCE. The purpose of this manuscript is to provide an overview of the major pros and cons of utilization of AMDs in nonclinical safety assessment.

The Natural History of Pneumonic Tularemia in Female Fischer 344 Rats after Inhalational Exposure to Aerosolized Francisella tularensis Subspecies tularensis Strain SCHU S4.

The inbred Fischer 344 rat is being evaluated for testing novel vaccines and therapeutics against pneumonic tularemia. Although primary pneumonic tularemia in humans typically occurs by inhalation of aerosolized bacteria, the rat model has relied on intratracheal inoculation of organisms because of safety and equipment issues. We now report the natural history of pneumonic tularemia in female Fischer 344 rats after nose-only inhalational exposure to lethal doses of aerosolized Francisella tularensis subspecies tularensis, strain SCHU S4. Our results are consistent with initial uptake of aerosolized SCHU S4 from the nasal cavity, lungs, and possibly the gastrointestinal tract. Bacteremia with hematogenous dissemination was first detected 2 days after exposure. Shortly thereafter, the infected rats exhibited fever, tachypnea, and hypertension that persisted for 24 to 36 hours and then rapidly decreased as animals succumbed to infection between days 5 and 8 after exposure. Tachycardia was observed briefly, but only after the core body temperature and blood pressure began to decrease as the animals were near death. Initial neutrophilic and histiocytic inflammation in affected tissues became progressively more fibrinous and necrotizing over time. At death, as many as 1010 colony-forming units were found in the lungs, spleen, and liver. Death was attributed to sepsis and disseminated intravascular coagulation. Overall, the pathogenesis of pneumonic tularemia in the female F344 rat model appears to replicate the disease in humans.

Scientific and Regulatory Policy Committee Points to Consider: Data Visualization for Clinical and Anatomic Pathologists.

Assessment and communication of toxicology data are fundamental components of the work performed by veterinary anatomic and clinical pathologists involved in toxicology research. In recent years, there has been an evolution in the number and variety of software tools designed to facilitate the evaluation and presentation of toxicity study data. A working group of the Society of Toxicologic Pathology Scientific and Regulatory Policy Committee reviewed existing and emerging visualization technologies. This Points to Consider article reviews some of the currently available data visualization options, describes the utility of different types of graphical displays, and explores potential areas of controversy and ambiguity encountered with the use of these tools.

Muc1 knockout potentiates murine lung carcinogenesis involving an epiregulin-mediated EGFR activation feedback loop.

Mucin 1 (MUC1) is a tumor antigen that is aberrantly overexpressed in various cancers, including lung cancer. Our previous in vitro studies showed that MUC1 facilitates carcinogen-induced EGFR activation and transformation in human lung bronchial epithelial cells (HBECs), which along with other reports suggests an oncogenic property for MUC1 in lung cancer. However, direct evidence for the role of MUC1 in lung carcinogenesis is lacking. In this study, we used the 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)-induced A/J mouse lung tumor model to investigate the effect of whole-body Muc1 knockout (KO) on carcinogen-induced lung carcinogenesis. Surprisingly, lung tumor multiplicity was significantly increased in Muc1 KO compared to wild-type (WT) mice. The EGFR/AKT pathway was unexpectedly activated, and expression of the EGFR ligand epiregulin (EREG) was increased in the lung tissues of the Muc1 KO compared to the WT mice. EREG stimulated proliferation and protected against cigarette smoke extract (CSE)-induced cytotoxicity in in vitro cultured human bronchial epithelial cells. Additionally, we determined that MUC1 was expressed in human fibroblast cell lines where it suppressed CSE-induced EREG production. Further, suppression of MUC1 cellular activity with GO-201 enhanced EREG production in lung cancer cells, which in turn protected cancer cells from GO-201-induced cell death. Moreover, an inverse association between MUC1 and EREG was detected in human lung cancer, and EREG expression was inversely associated with patient survival. Together, these results support a promiscuous role of MUC1 in lung cancer development that may be related to cell-type specific functions of MUC1 in the tumor microenvironment, and MUC1 deficiency in fibroblasts and malignant cells results in increased EREG production that activates the EGFR pathway for lung carcinogenesis.

Lethal factor, but not edema factor, is required to cause fatal anthrax in cynomolgus macaques after pulmonary spore challenge.

Inhalational anthrax is caused by inhalation of Bacillus anthracis spores. The ability of B. anthracis to cause anthrax is attributed to the plasmid-encoded A/B-type toxins, edema toxin (edema factor and protective antigen) and lethal toxin (lethal factor and protective antigen), and a poly-d-glutamic acid capsule. To better understand the contribution of these toxins to the disease pathophysiology in vivo, we used B. anthracis Ames strain and isogenic toxin deletion mutants derived from the Ames strain to examine the role of lethal toxin and edema toxin after pulmonary spore challenge of cynomolgus macaques. Lethal toxin, but not edema toxin, was required to induce sustained bacteremia and death after pulmonary challenge with spores delivered via bronchoscopy. After intravenous challenge with bacilli to model the systemic phase of infection, lethal toxin contributed to bacterial proliferation and subsequent host death to a greater extent than edema toxin. Deletion of protective antigen resulted in greater loss of virulence after intravenous challenge with bacilli than deletion of lethal toxin or edema toxin alone. These findings are consistent with the ability of anti-protective antigen antibodies to prevent anthrax and suggest that lethal factor is the dominant toxin that contributes to the escape of significant numbers of bacilli from the thoracic cavity to cause anthrax after inhalation challenge with spores.

Autophagy protects against active tuberculosis by suppressing bacterial burden and inflammation.

Autophagy is a cell biological pathway affecting immune responses. In vitro, autophagy acts as a cell-autonomous defense against Mycobacterium tuberculosis, but its role in vivo is unknown. Here we show that autophagy plays a dual role against tuberculosis: antibacterial and anti-inflammatory. M. tuberculosis infection of Atg5(fl/fl) LysM-Cre(+) mice relative to autophagy-proficient littermates resulted in increased bacillary burden and excessive pulmonary inflammation characterized by neutrophil infiltration and IL-17 response with increased IL-1α levels. Macrophages from uninfected Atg5(fl/fl) LysM-Cre(+) mice displayed a cell-autonomous IL-1α hypersecretion phenotype, whereas T cells showed propensity toward IL-17 polarization during nonspecific activation or upon restimulation with mycobacterial antigens. Thus, autophagy acts in vivo by suppressing both M. tuberculosis growth and damaging inflammation.

Utilization of a Preemptive, Multimodal Analgesic Regimen in Adult Ambulatory Septoplasty Patients: A Quality Improvement Project.

This paper describes a quality improvement project designed to decrease postoperative pain, decrease post-operative nausea and vomiting (PONV), decrease time in the recovery room, and increase patient satisfaction in adult ambulatory septoplasty patients using a multimodal, preemptive analgesic regimen. The project was conducted in a community hospital setting with nine operating rooms, and a twenty one bed recovery room. Project participants included certified registered nurse anesthetists, anesthesiologists, operating room nurses, recovery room nurses, and otolaryngology surgeons. Following a period of departmental education, adult patients scheduled for outpatient septoplasty surgery received a preoperative regimen of medications that included gabapentin, celecoxib, and acetaminophen. Using a pre-post test design, (intervention group n = 17, non-intervention group n = 17) data was collected from patient and analyzed using SPSS version 18.0. The change in practice resulted in a significant decrease in pain scores in the recovery room and on discharge from the recovery room. In addition, patients who received the preemptive regimen also required significantly fewer opioid medications and were ready to be discharged from the recovery room in less time.

Local and systemic tolerability of a 2'O-methoxyethyl antisense oligonucleotide targeting interleukin-4 receptor-α delivery by inhalation in mouse and monkey.

Antisense oligonucleotides (ASOs) bind and facilitate degradation of RNA and inhibit protein expression in pathways not easily targeted with small molecules or antibodies. Interleukin (IL)-4 and IL-13 potentiate signaling through the shared IL-4 receptor-α (IL-4Rα) subunit of their receptors. ASO targeting of IL-4Rα mRNA in a mouse model of asthma led to attenuation of airway hyperactivity, demonstrating potential benefit in asthma patients. This study focused on tolerability of inhaled IL-4Rα-targeting ASOs. Toxicity studies were performed with mouse- (ISIS 23189) and human-specific (ISIS 369645) sequences administered by inhalation. Four week (monkey) or 13 week (mouse) repeat doses at levels of up to 15 mg/kg/exposure (exp) and 50 mg/kg/exp, respectively, demonstrated dose-dependent effects limited to increases in macrophage size and number in lung and tracheobronchial lymph nodes. The changes were largely non-specific, reflecting adaptive responses that occur during active exposure and deposition of ASO and other material in the lung. Reversibility was observed at a rate consistent with the kinetics of tissue clearance of ASO. Systemic bioavailability was minimal, and no systemic toxicity was observed at exposure levels appreciably above pharmacological doses and doses proposed for clinical trials.

Professional identity in nursing: Why it is important in graduate education.

Possessing a clear identity in nursing is a guiding principle to professional comportment. In graduate nursing education, transitioning and expanding one's professional identity requires role evolution. Nurses transitioning into the advanced professional nursing role shifts their thinking to a new level. The Conceptual Model of Professional Identity in Nursing constitutes how values and ethics, knowledge, nurse as a leader, and professional comportment are intertwined. Competency-based education requires curricular redesign. The Essentials Tool Kit aligns The Essentials with learning activities to support competency-based curriculum and assessment. The Douglass and Stager Toolkit intertwines these resources for graduate nursing educators to inform professional identity in nursing for curriculum revisions. This article aims to illustrate how faculty educate graduate nursing students in the development of professional identity using a conceptual framework to achieve competencies outlined in The Essentials (AACN, 2021).

Predictors and use of nonpharmacologic interventions for procedural pain associated with turning among hospitalized adults.

Many hospitalized adults cannot reposition themselves in their beds. Therefore, they are regularly turned by their nurses, primarily to prevent pressure ulcer formation. Earlier research indicates that turning is painful and that patients are rarely premedicated with analgesics. Nonpharmacologic interventions may be used to help with this painful procedure. However, no published research was found on the use of nonpharmacologic interventions for turning of hospitalized patients. The objectives of this study were: 1) to describe patient pain characteristics during turning and their association with patient demographic and clinical characteristics; 2) to determine the frequency of use of various nonpharmacologic interventions for hospitalized adult patients undergoing the painful procedure of turning; and 3) to identify factors that predict the use of specific nonpharmacologic interventions for pain associated with turning. Hospitalized adult patients who experienced turning, the nurses caring for them, and others who were present at the time of turning were asked if they used various nonpharmacologic interventions to manage pain during the turning. Out of 1,395 patients, 92.5% received at least one nonpharmacologic intervention. Most frequently used were calming voice (65.7%), information (60.6%), and deep breathing (37.9%). Critical-care patients were more likely to receive a calming voice (odds ratio [OR] 1.66, p < .01), receive information (OR 1.62, p < .001), and use deep breathing (OR= 1.36, p < .05) than those who were not critical-care patients. Those reporting higher pain were consistently more likely to receive each of the three interventions (OR 1.01, p < .05 for all 3). In conclusion, nonpharmacologic interventions are used frequently during a turning procedure. The specific interventions used most often are ones that can be initiated spontaneously. Our data suggest that patients, nurses, and family members respond to patients' turning-related pain by using nonpharmacologic interventions.

From provider to professor: Supporting nurse practitioner faculty through NONPF's Faculty Institute.

ABSTRACT: Over the past 15 years, the number of practicing nurse practitioners (NPs) has increased exponentially. This growth has been sustained by an increase in the number of schools offering NP preparation. Experienced NPs have assumed faculty positions in these schools. Although educational curricula prepare NPs to deliver high-quality patient care, it does not prepare graduates for academic roles in teaching, dissemination of scholarship, or service leadership to advance the profession. To address these gaps in NP faculty preparation, faculty development resources specific to NP education are needed. The National Organization of Nurse Practitioner Faculties (NONPF) has stepped forward to fill this gap. The NONPF Faculty Institute was created in 2017, and it includes the annual Fall Conference, the Faculty Leadership Mentoring Program, the Webinar Series, and, most recently, the joint NPACE/NONPF New Faculty Development Program. This comprehensive focus on faculty development aligns with NONPF's mission to promote excellence in NP education and advance the NP role in health care. Programs in the NONPF Faculty Institute are specifically designed to address the distinct learning needs of NP faculty. They use "just in time" presentations and active learning strategies to provide NP faculty with easily accessible and targeted content and opportunities for direct application of new skills. The Institute's goal is to provide NP faculty opportunities to prepare themselves to meet the teaching, scholarship and service leadership expectations of academia, the learning needs of students, and advance the impact of NP practice on patient and health care system outcomes.

Preclinical safety and biodistribution assessment of Ad-KCNH2-G628S administered via atrial painting in New Zealand white rabbits.

Post-operative atrial fibrillation (POAF) is the most common complication after cardiac surgery. Despite implementation of several pharmacological strategies, incidence of POAF remains at approximately 30%. An adenovirus vector encoding KCNH2-G628S has proven efficacious in a porcine model of AF. In this preclinical study, 1.5 × 1010 or 1.5 × 1012 Ad-KCNH2-G628S vector particles (vp) were applied to the atrial epicardium or 1.5 × 1012 vp were applied to the whole epicardial surface of New Zealand White rabbits. Saline and vector vehicle served as procedure controls. Animals were followed for up to 42 days. Vector genomes persisted in the atria up to 42 days, with no distribution to extra-thoracic organs. There were no adverse effects attributable to test article on standard toxicological endpoints or on blood pressure, left atrial or ventricular ejection fractions, electrocardiographic parameters, or serum IL-6 or troponin concentrations. Mononuclear infiltration of the myocardium of the atrial free walls of low-dose, but not high-dose animals was observed at 7 and 21 days, but these changes did not persist or affect cardiac function. After scaling for heart size, results indicate the test article is safe at doses up to 25 times the maximum proposed for the human clinical trial.

Expression of either lethal toxin or edema toxin by Bacillus anthracis is sufficient for virulence in a rabbit model of inhalational anthrax.

The development of therapeutics against biothreats requires that we understand the pathogenesis of the disease in relevant animal models. The rabbit model of inhalational anthrax is an important tool in the assessment of potential therapeutics against Bacillus anthracis. We investigated the roles of B. anthracis capsule and toxins in the pathogenesis of inhalational anthrax in rabbits by comparing infection with the Ames strain versus isogenic mutants with deletions of the genes for the capsule operon (capBCADE), lethal factor (lef), edema factor (cya), or protective antigen (pagA). The absence of capsule or protective antigen (PA) resulted in complete avirulence, while the presence of either edema toxin or lethal toxin plus capsule resulted in lethality. The absence of toxin did not influence the ability of B. anthracis to traffic to draining lymph nodes, but systemic dissemination required the presence of at least one of the toxins. Histopathology studies demonstrated minimal differences among lethal wild-type and single toxin mutant strains. When rabbits were coinfected with the Ames strain and the PA- mutant strain, the toxin produced by the Ames strain was not able to promote dissemination of the PA- mutant, suggesting that toxigenic action occurs in close proximity to secreting bacteria. Taken together, these findings suggest that a major role for toxins in the pathogenesis of anthrax is to enable the organism to overcome innate host effector mechanisms locally and that much of the damage during the later stages of infection is due to the interactions of the host with the massive bacterial burden.

Policy implications for optimizing advanced practice registered nurse use nationally.

This article examines the potential benefits of enhanced use of advanced practice registered nurses (APRNs) given health care workforce projections that predict an inadequate supply of certain types of providers. The conclusions of a systematic review comparing the effectiveness of care provided by APRNs with that of physicians alone or teams without APRNs indicate the viability of this approach. Allowing APRNs to assume roles that take full advantage of their educational preparation could mitigate the shortage of primary care physicians and improve care processes. The development of health care policy should be guided by patient-centric evidence rather than how care has been delivered in the past.

Antibodies contribute to effective vaccination against respiratory infection by type A Francisella tularensis strains.

Pneumonic tularemia is a life-threatening disease caused by inhalation of the highly infectious intracellular bacterium Francisella tularensis. The most serious form of the disease associated with the type A strains can be prevented in experimental animals through vaccination with the attenuated live vaccine strain (LVS). The protection is largely cell mediated, but the contribution of antibodies remains controversial. We addressed this issue in a series of passive immunization studies in Fischer 344 (F344) rats. Subcutaneous LVS vaccination induced a robust serum antibody response dominated by IgM, IgG2a, and IgG2b antibodies. Prophylactic administration of LVS immune serum or purified immune IgG reduced the severity and duration of disease in naïve rats challenged intratracheally with a lethal dose of the virulent type A strain SCHU S4. The level of resistance increased with the volume of immune serum given, but the maximum survivable SCHU S4 challenge dose was at least 100-fold lower than that shown for LVS-vaccinated rats. Protection correlated with reduced systemic bacterial growth, less severe histopathology in the liver and spleen during the early phase of infection, and bacterial clearance by a T cell-dependent mechanism. Our results suggest that treatment with immune serum limited the sequelae associated with infection, thereby enabling a sterilizing T cell response to develop and resolve the infection. Thus, antibodies induced by LVS vaccination may contribute to the defense of F344 rats against respiratory infection by type A strains of F. tularensis.

Advanced practice nurse outcomes 1990-2008: a systematic review.

Advanced practice registered nurses have assumed an increasing role as providers in the health care system, particularly for underserved populations. The aim of this systematic review was to answer the following question: Compared to other providers (physicians or teams without APRNs) are APRN patient outcomes of care similar? This systematic review of published literature between 1990 and 2008 on care provided by APRNs indicates patient outcomes of care provided by nurse practitioners and certified nurse midwives in collaboration with physicians are similar to and in some ways better than care provided by physicians alone for the populations and in the settings included. Use of clinical nurse specialists in acute care settings can reduce length of stay and cost of care for hospitalized patients. These results extend what is known about APRN outcomes from previous reviews by assessing all types of APRNs over a span of 18 years, using a systematic process with intentionally broad inclusion of outcomes, patient populations, and settings. The results indicate APRNs provide effective and high-quality patient care, have an important role in improving the quality of patient care in the United States, and could help to address concerns about whether care provided by APRNs can safely augment the physician supply to support reform efforts aimed at expanding access to care.

Identification of an Attenuated Substrain of Francisella tularensis SCHU S4 by Phenotypic and Genotypic Analyses.

Pneumonic tularemia is a highly debilitating and potentially fatal disease caused by inhalation of Francisella tularensis. Most of our current understanding of its pathogenesis is based on the highly virulent F. tularensis subsp. tularensis strain SCHU S4. However, multiple sources of SCHU S4 have been maintained and propagated independently over the years, potentially generating genetic variants with altered virulence. In this study, the virulence of four SCHU S4 stocks (NR-10492, NR-28534, NR-643 from BEI Resources and FTS-635 from Battelle Memorial Institute) along with another virulent subsp. tularensis strain, MA00-2987, were assessed in parallel. In the Fischer 344 rat model of pneumonic tularemia, NR-643 and FTS-635 were found to be highly attenuated compared to NR-10492, NR-28534, and MA00-2987. In the NZW rabbit model of pneumonic tularemia, NR-643 caused morbidity but not mortality even at a dose equivalent to 500x the LD50 for NR-10492. Genetic analyses revealed that NR-10492 and NR-28534 were identical to each other, and nearly identical to the reference SCHU S4 sequence. NR-643 and FTS-635 were identical to each other but were found to have nine regions of difference in the genomic sequence when compared to the published reference SCHU S4 sequence. Given the genetic differences and decreased virulence, NR-643/FTS-635 should be clearly designated as a separate SCHU S4 substrain and no longer utilized in efficacy studies to evaluate potential vaccines and therapeutics against tularemia.

Chromatin remodeling by the histone methyltransferase EZH2 drives lung pre-malignancy and is a target for cancer prevention.

BACKGROUND: Trimethylation of lysine 27 and dimethylation of lysine 9 of histone-H3 catalyzed by the histone methyltransferases EZH2 and G9a impede gene transcription in cancer. Our human bronchial epithelial (HBEC) pre-malignancy model studied the role of these histone modifications in transformation. Tobacco carcinogen transformed HBEC lines were characterized for cytosine DNA methylation, transcriptome reprogramming, and the effect of inhibiting EZH2 and G9a on the transformed phenotype. The effects of targeting EZH2 and G9a on lung cancer prevention was assessed in the A/J mouse lung tumor model. RESULTS: Carcinogen exposure induced transformation and DNA methylation of 12-96 genes in the four HBEC transformed (T) lines that was perpetuated in malignant tumors. In contrast, 506 unmethylated genes showed reduced expression in one or more HBECTs with many becoming methylated in tumors. ChIP-on-chip for HBEC2T identified 327 and 143 genes enriched for H3K27me3 and H3K9me2. Treatment of HBEC2T and HBEC13T with DZNep, a lysine methyltransferase inhibitor depleted EZH2, reversed transformation, and induced transcriptional reprogramming. The EZH2 small molecule inhibitor EPZ6438 also affected transformation and expression in HBEC2T, while a G9a inhibitor, UNC0642 was ineffective. Genetic knock down of EZH2 dramatically reduced carcinogen-induced transformation of HBEC2. Only DZNep treatment prevented progression of hyperplasia to adenomas in the NNK mouse lung tumor model through reducing EZH2 and affecting the expression of genes regulating cell growth and invasion. CONCLUSION: These studies demonstrate a critical role for EZH2 catalyzed histone modifications for premalignancy and its potential as a target for chemoprevention of lung carcinogenesis.

Granuloma formation induced by low-dose chronic silica inhalation is associated with an anti-apoptotic response in Lewis rats.

Chronic human silicosis results primarily from continued occupational exposure to silica and exhibits a long asymptomatic latency. Similarly, continued exposure of Lewis rats to low doses of silica is known to cause delayed granuloma formation with limited lung inflammation and injury. On the other hand, intratracheal exposure to large doses of silica induces acute silicosis characterized by granuloma-like formations in the lung associated with apoptosis, severe alveolitis, and alveolar lipoproteinosis. To ascertain similarities/differences between acute and chronic silicosis, in this communication, we compared cellular and molecular changes in established rat models of acute and chronic silicosis. In Lewis rats, acute silicosis was induced by intratracheal instillation of 35 mg silica, and chronic silicosis through inhalation of aerosolized silica (6.2 mg/m(3), 5 d/wk for 6 wk). Animals exposed to acute high-dose silica were sacrificed at 14 d after silica instillation while chronically silica-treated animals were sacrificed between 4 d and 28 wk after silica exposure. The lung granulomas formation in acute silicosis was associated with strong inflammation, presence of TUNEL-positive cells, and increases in caspase-3 activity and other molecular markers of apoptosis. On the other hand, lungs from chronically silica-exposed animals exhibited limited inflammation and increased expression of anti-apoptotic markers, including dramatic increases in Bcl-2 and procaspase-3, and lower caspase-3 activity. Moreover, chronic silicotic lungs were TUNEL-negative and overexpressed Bcl-3 and NF-kappaB-p50 but not NF-kappaB-p65 subunits. These results suggest that, unlike acute silicosis, chronic exposures to occupationally relevant doses of silica cause significantly lower lung inflammation and elevated expression of anti-apoptotic rather than proapoptotic markers in the lung that might result from interaction between NF-kappaB-p50 and Bcl-3.