Changes in Basal and Bolus Insulin Requirements with Tirzepatide as an Adjunctive Therapy in Adults with Type 1 Diabetes Using Tandem Control-IQ.

INTRODUCTION: This study was aimed at investigating changes in insulin requirements and glycemic outcomes in adults with type 1 diabetes (T1D) using Control IQ (Tandem Diabetes) automated insulin delivery system (AID) over 8 months of tirzepatide treatment. METHODS: In this single-center, observational study, we collected demographic, A1c, weight, sensor glucose, and insulin dose data for adults with T1D who were using AID and initiated tirzepatide adjunct therapy for clinical indications (n = 11, median age 37, 64% female and mean body mass index of 39.6 kg/m2). Data were compared from baseline and over 8 months. RESULTS: Within 2 months of tirzepatide treatment, there were significant reductions in total daily insulin [median (IQR) 73.9 (47.6-95.8) to 51.7 (46.7-66.8) units/day, p < 0.001], basal insulin [47 (28.2-51.8) to 32.4 (25.5-46.3) units/day, p < 0.001], and bolus insulin [31.4 (19.9-38.3) to 17.9 (14.9-22.2) units/day, p < 0.001] requirements. Insulin dose reduction from 2 to 8 months was modest. The frequency of user-initiated boluses did not differ throughout the study. Despite reductions in total insulin requirement, time in range (70-180 mg/dl) increased by 7%, A1c reduced by 0.5%, weight reduced by 9%, without increase in time below 70 mg/dl. CONCLUSIONS: This pilot study provides clinical guidance on insulin titration for adults with T1D who may initiate tirzepatide therapy. Based on the findings of this study, we recommend reducing 25% of total daily insulin dose at tirzepatide initiation in adults with T1D using AID with baseline A1c of less than 7.5%. Higher doses of tirzepatide were associated with greater weight loss, however, the reduction in insulin requirement was minimal.

Changes in the glycaemia risk index and its association with other continuous glucose monitoring metrics after initiation of an automated insulin delivery system in adults with type 1 diabetes.

AIM: To evaluate the glycaemia risk index (GRI) and its association with other continuous glucose monitoring (CGM) metrics after initiation of an automated insulin delivery (AID) system in patients with type 1 diabetes (T1D). MATERIALS AND METHODS: Up to 90 days of CGM data before and after initiation of an AID system from 185 CGM users with T1D were collected. GRI and other CGM metrics were calculated using cgmanalysis R software and were analysed for 24 hours, for both night-time and daytime. GRI values were assigned to five GRI zones: zone A (0-20), B (21-40), C (41-60), D (61-80) and E (81-100). RESULTS: Compared with baseline, GRI and its components decreased significantly after AID initiation (GRI: 48.7 ± 21.8 vs. 29 ± 13; hypoglycaemia component: 2.7 ± 2.8 vs. 1.6 ± 1.7; hyperglycaemia component: 25.3 ± 14.5 vs. 15 ± 8.5; P < .001 for all). The GRI was inversely correlated with time in range before (r = -0.962) and after (r = -0.961) AID initiation (P < .001 for both). GRI was correlated with time above range (before: r = 0.906; after = 0.910; P < .001 for both), but not with time below range (P > .05). All CGM metrics improved after AID initiation during 24 hours, for both daytime and night-time (P < .001 for all). Metrics improved significantly more during night-time than daytime (P < .01). CONCLUSIONS: GRI was highly correlated with various CGM metrics above, but not below target range, both before and after AID initiation.

Association Between Diabetes Technology Use and Glycemic Outcomes in Adults With Type 1 Diabetes Over a Decade.

OBJECTIVE: To evaluate change in mean clinic HbA1c from 2014 to 2021 with diabetes technology use in adults with type 1 diabetes. RESEARCH DESIGN AND METHODS: In this single-center study, we analyzed diabetes technology use and mean clinic HbA1c among unique adults (age ≥18 years) with type 1 diabetes (last visit of the year per patient) between 1 January 2014 and 31 December 2021 from the electronic medical record. Diabetes technology use was defined as the use of continuous glucose monitors (CGMs) without an automated insulin delivery (AID) system or an AID system. Diabetes technology use and HbA1c over time were analyzed using mixed models adjusted for age, sex, and visit year. RESULTS: A total of 15,903 clinic visits over 8 years (mean 1,988 patients per year, 4,174 unique patients, 52.7% female, 80.0% Non-Hispanic White) showed significant increases in CGM and AID use (P < 0.001 for both), resulting in an increase of diabetes technology use from 26.9% in 2014 to 82.7% in 2021. These increases were associated with a lower mean clinic HbA1c (7.7-7.5%, P < 0.001) and a higher percentage of adults achieving an HbA1c <7.0% (32.3-41.7%, P < 0.001) from 2014 to 2021. The HbA1c difference between technology users and nonusers increased over time from 0.36% (95% CI 0.26-0.47%, P < 0.001) in 2014 to 0.93% (95% CI 0.80-1.06%, P < 0.001) in 2021. CONCLUSIONS: Adopting diabetes technology in adults with type 1 diabetes decreased HbA1c and increased the number of people achieving an HbA1c <7.0%, supporting the current international recommendation to offer AID systems to most individuals with type 1 diabetes.

Assessing the relationship between low-calorie sweetener use and quality of life measures in adults with type 1 diabetes.

BACKGROUND: To evaluate use of low-calorie sweeteners (LCS) among adults with type 1 diabetes (T1D) and its impact on quality of life (QOL). METHODS: In this single center, cross-sectional survey study with 532 adults with T1D, Food related QOL (FRQOL), LCS specific questionnaire (LCSSQ), Diabetes Self-Management Questionnaire (DSMQ), Food Frequency Questionnaire (FFQ), Audit of Diabetes-Dependent QOL (AddQOL), Type 1 Diabetes and Life (T1DAL) questionnaires were administered through RedCAP, a secure, HIPAA-compliant web-based application. Demographics and scores of adults who used LCS in last month (recent users) and others (non-users) were compared. Results were adjusted for age, sex, diabetes duration and other parameters. RESULTS: Of 532 participants (mean age 36 ± 13, 69% female), 99% heard LCS before, 68% used them in the last month, 73% reported better glucose control with LCS use and 63% reported no health concerns about LCS use. Recent LCS users were older and had a longer diabetes duration and more complications (hypertension, or any complication) than non-users. However, A1c, AddQOL, T1DAL, FRQOL scores did not differ significantly between recent LCS users and non-users. DSMQ scores, DSMQ management, diet, health care scores did not differ between two groups; however, recent LCS users had lower physical activity score than non-users (p = 0.001). CONCLUSIONS: Most of the adults with T1D have used LCS and perceived that LCS use improved their QOL and glycemic control; however, these were not verified with questionnaires. There was no difference in QOL questionnaires except DSMQ physical activity between recent LCS users and not users with T1D. However, more patients in need to increase their QOL may be using LCS; therefore, associations between the exposure and outcome can be bi-directional.

Analysis of Human Leukocyte Antigen DR Alleles, Immune-Related Adverse Events, and Survival Associated With Immune Checkpoint Inhibitor Use Among Patients With Advanced Malignant Melanoma.

Importance: Treatment with immune checkpoint inhibitors (ICIs) has increased survival in patients with advanced malignant melanoma but can be associated with a wide range of immune-related adverse events (irAEs). The role of human leukocyte antigen (HLA)-DR alleles in conferring irAE risk has not been well studied. Objective: To evaluate the association between irAEs and treatment response, survival, and the presence of HLA-DR alleles after ICI therapy in advanced melanoma. Design, Setting, and Participants: This case-control study used the patient registry and biobanked samples from the tertiary referral University of Colorado Cancer Center. Specimens and clinical data were collected between January 1, 2010, and December 31, 2021. Patients with advanced (stage III unresectable and stage IV) melanoma who received ICI therapy (n = 132) were included in the analysis. Exposures: Immune checkpoint inhibitors (anti-cytotoxic T-lymphocyte antigen 4, anti-programmed cell death protein 1 or its ligand, or the combination) for the treatment of advanced melanoma. Main Outcomes and Measures: The association between irAEs and response to therapy, survival, and HLA-DR alleles. Results: Among the cohort of 132 patients with advanced melanoma (mean [SD] age, 63.4 [7.2] years; 85 men [64%] and 47 women [36%]) treated with ICIs, 73 patients had at least 1 irAE and 59 did not have an irAE. Compared with patients without an irAE, patients with an irAE had higher treatment response rates (50 of 72 [69%] vs 28 of 57 [49%]; P = .02) and increased survival (median, 4.8 [IQR, 0.2-9.6] vs 3.2 [IQR, 0.1-9.2] years; P = .02). Specific HLA-DR alleles were associated with the type of irAE that developed: 7 of 10 patients (70%) who developed type 1 diabetes had DR4; 6 of 12 (50%) who developed hypothyroidism had DR8; 5 of 8 (63%) who developed hypophysitis had DR15; 3 of 5 (60%) who developed pneumonitis had DR1; and 8 of 15 (53%) who developed hepatitis had DR4. Conclusions and Relevance: These findings suggest that IrAEs are associated with treatment response rates and increased survival after ICI therapy for advanced melanoma. Because distinct HLA-DR alleles are associated with given adverse events, HLA genotyping before ICI therapy may aid in identifying risk for specific irAEs that could develop with such treatment.

Factors affecting adherence to physical training in the outpatient phase of rehabilitation, in patients after coronary artery bypass grafting.

Aim      To evaluate the outpatient physical exercise (PE) compliance and the affecting factors in patients after coronary bypass (CB).Material and methods  The study included 67 men with ischemic heart disease younger than 75 years who had had CB. All patients were randomized to 2 groups: group 1 exercised on a bicycle ergometer at the rehabilitation center, under the monitoring of medical staff; patients of group 2 performed home-based exercise (HBE) by dosed walking. In the preoperative period, at one month after CB, and after 3 months of exercise, the following was evaluated: clinical condition of patients in different groups, plasma concentrations of lipids, body weight index, waist circumference, echocardiography and bicycle ergometry data, and questionnaire data (SF-36, Bek's Depression Inventory). At 3 months of follow-up, the outpatient exercise compliance and the affecting factors were also evaluated.Results The study demonstrated the effectiveness of the proposed alternative 3-month program of home-based PE. Both the patients exercising on a bicycle and those performing HBE had increased exercise tolerance (ET) and improved blood lipid concentrations. The number of obese patients decreased. Also, depression severity decreased, quality of life (physical and psychological components) improved, and compliance with drug therapy increased in both groups. Analysis of the training attendance in the recommended period showed that patients who had undergone CB were insufficiently adherent to physical rehabilitation programs, regardless of the program type (home-based or monitored). The highest PE adherence was observed in men with the following characteristics: married, working urban residents, with a previous history of cardiovascular diseases, who had regularly taken medications in the preoperative period, and who also had higher quality of life.Conclusion      The proposed outpatient 3-month physical rehabilitation programs increase the effectiveness of CB, which is evident as improved adherence to modifying cardiovascular risk factors, increased ET, optimization of the psychological status and quality of life, and improved compliance with drug therapy. However, despite the proposed alternative, home-based 3-month physical rehabilitation programs aimed at increasing the treatment compliance, the level of ET remained low. This requires further improvement of methods for monitoring and motivation of patients to physical rehabilitation and psychological support that would start already at the preoperative stage.

Sodium-glucose cotransporter 2 inhibitors for diabetes mellitus control after kidney transplantation: Review of the current evidence.

Sodium-glucose cotransporter type 2 inhibitors (SGLT2i) are promising drugs to treat chronic kidney disease patients with or without diabetes mellitus (DM). Besides improving glycemic control, SGLT2i are cardioprotective and kidney protective and decrease bodyweight, serum uric acid, blood pressure, albuminuria and glomerular hyperfiltration. These effects may benefit graft function and survival in kidney transplant (KT) patients. In this review, we evaluate data on the efficacy and safety of SGLT2i for KT patients with DM. Eleven studies with 214 diabetic KT patients treated with SGLT2i have been reported. SGLT2i lowered haemoglobin A1c and bodyweight. While glomerular filtration rate may be reduced in the short-term, it remained similar to baseline after 3-12 months. In two studies, blood pressure decreased and remained unchanged in the others. There were no significant changes in urine protein to creatinine ratio. Regarding safety, 23 patients had urinary tract infections, 2 patients had a genital yeast infection, one had acute kidney injury, and one had mild hypoglycaemia. No cases of ketoacidosis or acute rejection were reported. In conclusion, the limited experience so far suggests that SGLT2i are safe in KT patients with DM, decrease bodyweight and improve glycemic control. However, some of the benefits observed in larger studies in the non-KT population have yet to be demonstrated in KT recipients, including preservation of kidney function, reduction in blood pressure and decreased proteinuria.

[Prediction of Risk of Calcification of Heart Valve Bioprostheses on the Basis of Comprehensive Assessment of Recipients Clinical Factors and Their Compliance With Therapy].

PURPOSE: The development of the prediction methods of calcification risk of heart valve bioprostheses (BP) based on comprehensive assessment of the impact of clinical factors of the recipients and their adherence to medication. MATERIALS AND METHODS: We performed a retrospective study of clinical status and adherence to drug therapy of 170 recipients of heart valve BP with (n=63) and without (n=109) calcification. We used the method of comprehensive assessment of the analyzed parameters with estimation of prognostic coefficients for each of them, followed by calculation of integral indicators and construction on their basis of prognostic models for the various intervals of BP functioning. RESULTS: The most important risk factors of calcium-associated BP dysfunctions at any duration of observation were heart failure decompensation, as well as the discontinuation of ACE inhibitors and ß-blockers. At duration of BP functioning up to four years negative effect on prognosis produced the presence of concomitant multifocal atherosclerosis, during additional 4 (from 4 to 8) years - diabetes mellitus, and afterwards (over 8 years) - coronary artery disease. Lowering of probability of BP calcium degeneration was related to regular use of aldosterone antagonists during first 4 years after BP implantation and regular subsequent use of statins. CONCLUSIONS: Based on the comprehensive assessment of clinical status of patients and their compliance to medication it is possible to make adequate prediction models for assessment of the risk of structural dysfunctions due to BP calcification at various time intervals of remote postoperative period.

Drug repurposing from the perspective of pharmaceutical companies.

Drug repurposing holds the potential to bring medications with known safety profiles to new patient populations. Numerous examples exist for the identification of new indications for existing molecules, most stemming from serendipitous findings or focused recent efforts specifically limited to the mode of action of a specific drug. In recent years, the need for new approaches to drug research and development, combined with the advent of big data repositories and associated analytical methods, has generated interest in developing systematic approaches to drug repurposing. A variety of innovative computational methods to enable systematic repurposing screens, experimental as well as through in silico approaches, have emerged. An efficient drug repurposing pipeline requires the combination of access to molecular data, appropriate analytical expertise to enable robust insights, expertise and experimental set-up for validation and clinical development know-how. In this review, we describe some of the main approaches to systematic repurposing and discuss the various players in this field and the need for strategic collaborations to increase the likelihood of success in bringing existing molecules to new indications, as well as the current advantages, considerations and challenges in repurposing as a drug development strategy pursued by pharmaceutical companies. LINKED ARTICLES: This article is part of a themed section on Inventing New Therapies Without Reinventing the Wheel: The Power of Drug Repurposing. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.2/issuetoc.

[Factors for the development of persistent postoperative cognitive dysfunction in patients undergoing coronary artery bypass surgery under extracorporeal circulation]. / Faktory razvitiia stoikoi posleoperatsionnoi kognitivnoi disfunktsii u patsientov, perenesshikh koronarnoe shuntirovanie v usloviiakh iskusstvennogo krovo-obrashcheniia.

AIM: To analyze the factors contributing to the increased risk of persistent postoperative cognitive dysfunction (POCD) in patients undergoing coronary artery bypass surgery (CABS) under extracorporeal circulation (EC). SUBJECTS AND METHODS: 257 male patients aged 45 to 69 years with coronary heart disease (CHD) undergoing elective CABS under EC were examined. In addition to conventional clinical examination, all the patients underwent neuropsychological testing 3-5 days before, 7-14 days and 1 year after CABS. Persistent POCD was diagnosed if there was a 20% decline in cognitive domains at 1-year postoperatively versus preoperatively in 20% of the tests of an entire neuropsychological battery. Binary logistic regression analysis was applied to identify the factors supposedly increasing the risk of persistent POCD. RESULTS: It was found that high baseline cognitive status, early POCD after CABG under EC, low adherence to the prescribed treatment regimen, as well as progressive carotid artery (CA) stenosis could predict with a high (85%) probability that persistent POCD might develop at 1 year after surgery. CONCLUSION: The findings are suggestive of the multifactorial origin of persistent POCD, a significant role in the development of which is played by not only the preoperative cognitive status, but also by postoperative factors, such as the degree of adherence to the prescribed treatment regimen, early POCD, and progressive CA stenosis.

[PREDICTORS OF MODERATE COGNITIVE DISORDERS IN PATIENTS WITH CORONARY HEART DISEASE AND TYPE 2 DIABETES MELLITUS].

AIM: To study predictors of moderate cognitive disorders (MCD) in patients with coronary heart disease (CHD) and type 2 diabetes mellitus (DM2). MATERIALS AND METHODS: The study included 54 men with CPD andDM2 (mean age 56.8 ± 4.5 years). Standard medical examination was supplemented by the assessment of cognitive status, characteristics of lipid and carbohydrate metabolism. Factors allegedly influencing MCD development included the patients' age, education level, stenosis of carotid arteries, LV ejection fraction, arterial hypertension, insulin and HbAlc levels, HOMA and QUICKI indices, lipid metabolism, concentrations of total, HDL and LDL cholesterol, fructosamine, triglycerides, severity of coronary lesions (Syntax scale), trait and state anxiety. RESULTS: Fructosamine level and HOMA index were the most important characteristics responsible for MCD in patients with CPD and DM2. CONCLUSION: The data obtained demonstrate the significance of fructosamine level and HOMA index in the development of MCD in patients with CPD and DM2.

[Adherence to Therapy as a Factor Determining Prognosis of Coronary Artery Bypass Grafting].

AIM: to study the influence of the patients adherence to the recommended therapy after coronary artery bypass grafting (CABG) on prognosis of postoperative period. MATERIAL: We examined 197 consecutive patients with stable coronary artery disease (CAD) who had undergone CABG. Age of patients was 38-75 years. RESULTS: Assessment of modifiable cardiovascular risk factors showed that about half of patients had smoked before CABG and only a few gave up smoking after surgery. Number of patients with abdominal obesity increased by 8% after surgery. Number of patients involved in physical trainings remained unchanged. Adherence to drug therapy before CABG was low. Less than half of the patients took antiplatelet agents, beta-blockers, angiotensin-converting enzyme inhibitors, only 25% took statins. One year after CABG number of patients taking appropriate medications significantly increased. However, only half of patients managed to achieve the main objectives of secondary prevention.

[Analysis of approaches to assessment of persistent loss of capacity to work in patients who had undergone coronary bypass surgery].

Aim of the study was to assess factors influencing decisions about persistent disability of patients after coronary bypass surgery (CBS). By method of continuous sampling (registry study) we examined 427 working age patients who had undergone CBS. Although surgical treatment was effective rehabilitating factor most patients after CBS at inspection in institutions of medical social expertise were unreasonably classified as having low degree of restoration of ability to work. Possible explanations of this were incomplete volume of conducted tests, lack of relation between presence of functional class of angina and real clinical picture of this syndrome, absence of objective criteria of the presence of myocardial ischemia and tolerance to physical exercise, ill-timed referral to medical social inspection.

Bioregulators as instruments of terror.

Bioregulators are naturally occurring organic compounds that regulate diverse cellular processes. Unlike traditional disease-causing biowarfare agents that take hours or days to act, many bioregulators act within minutes of administration. If exploited for the purpose of bioterrorism, they could potentially cause profound physiologic effects. Other effects may be more subtle. The main groups of bioregulators discussed are cytokines; eicosanoids, neurotransmitters, hormones, and proteolytic enzymes. Because advances continually are being made in their development, bioregulators should be considered as weapons with increasing bioterrorism potential.

Effect of immunological adjuvant combinations on the antibody and T-cell response to vaccination with MUC1-KLH and GD3-KLH conjugates.

A year ago we described a comparison of 19 immunological adjuvants for their ability to augment antibody and T-cell responses against vaccines containing two cancer antigens, GD3 ganglioside and MUC1 peptide, covalently attached to keyhole limpet hemocyanin (KLH). As in our previous experience, the saponin fraction QS-21 was the most potent single adjuvant but several other adjuvants also had potent adjuvant activity. Induction of an immune response against cancer antigens is generally difficult because these antigens are autoantigens. To get maximal benefit from the adjuvant component of cancer vaccines we have now tested whether combinations of the optimal adjuvants induced an improved immune response compared to QS-21 alone. Since over the intervening year a new semi-synthetic saponin adjuvant (GPI-0100) containing the dodecylamide derivative of hydrolyzed naturally-occurring saponins had become available, this was tested as well. Twelve different adjuvant combinations and GPI-0100 were compared for their ability to augment (1) antibody responses against GD3 and MUC1 and (2) T-cell responses against GD3, MUC1 and KLH. GPI-0100 and five adjuvant combinations were superior to QS-21 alone for induction of IgM and IgG antibodies against MUC1 and/or GD3: QS-21 plus bacterial nucleotide CpG, QS-21 plus monophosphoryl lipid A (MPL), QS-21 plus non-ionic block copolymer CRL-1005, QS-21 plus Titermax and Titermax plus CpG. Antibody responses were documented both by ELISA against purified antigens and by FACS for cell surface reactivity. There was no evidence for T-cell immunity against GD3 or MUC1. The antibody responses against GD3 and MUC1 were, however, strongly correlated with IFN-gamma release and DTH against KLH. These results demonstrate that combinations of immunological adjuvants are able to augment antibody and T-cell responses to these conjugates beyond that attainable with QS-21 alone, and again confirm the absolute necessity of potent adjuvants or adjuvant combinations for optimal immunogenicity with conjugate vaccines.

Asbestos exposure induces MCP-1 secretion by pleural mesothelial cells.

We showed previously that both crocidolite and chrysotile asbestos inhalation induced a persistent macrophage inflammatory response within the pleural space of the rat. We postulated that the stimulus for pleural macrophage recruitment after asbestos exposure was the induction of monocyte chemoattractant protein-1 (MCP-1) synthesis by pleural mesothelial cells. To test this hypothesis, rat pleural mesothelial cells (RPMC) were cultured with or without chrysotile or crocidolite asbestos fibers (8 micrograms/cm2) in the presence (50 ng/mL) or absence of either tumor necrosis factor-alpha (TNF-alpha) or interleukin-1 beta (IL-1 beta). MCP-1 mRNA expression was assessed by RT-PCR in RPMC cultured for 2 to 24 hours, and MCP-1 protein secretion was measured by ELISA in conditioned medium from 24-hour and 48-hour cultures. Crocidolite and chrysotile fibers induced MCP-1 mRNA expression in RPMC which was maximal after 12 hours in the absence of cytokines, but which peaked after 2 hours when RPMC were challenged with asbestos + TNF-alpha or IL-1 beta. Both types of asbestos also significantly increased MCP-1 protein secretion after 24 and 48 hours (P < .0001), an effect that was potentiated by cytokine stimulation. Rats exposed by inhalation to either chrysotile or crocidolite asbestos fibers also had greater amounts of MCP-1 protein in their pleural lavage fluid than did sham-exposed rats. These findings suggest that MCP-1 secretion by RPMC may have a role in the initiation and/or potentiation of asbestos-induced pleural injury.

Peroxynitrite targets the epidermal growth factor receptor, Raf-1, and MEK independently to activate MAPK.

Activation of ERK-1 and -2 by H(2)O(2) in a variety of cell types requires epidermal growth factor receptor (EGFR) phosphorylation. In this study, we investigated the activation of ERK by ONOO(-) in cultured rat lung myofibroblasts. Western blot analysis using anti-phospho-ERK antibodies along with an ERK kinase assay using the phosphorylated heat- and acid-stable protein (PHAS-1) substrate demonstrated that ERK activation peaked within 15 min after ONOO(-) treatment and was maximally activated with 100 micrometer ONOO(-). Activation of ERK by ONOO(-) and H(2)O(2) was blocked by the antioxidant N-acetyl-l-cysteine. Catalase blocked ERK activation by H(2)O(2), but not by ONOO(-), demonstrating that the effect of ONOO(-) was not due to the generation of H(2)O(2). Both H(2)O(2) and ONOO(-) induced phosphorylation of EGFR in Western blot experiments using an anti-phospho-EGFR antibody. However, the EGFR tyrosine kinase inhibitor AG1478 abolished ERK activation by H(2)O(2), but not by ONOO(-). Both H(2)O(2) and ONOO(-) activated Raf-1. However, the Raf inhibitor forskolin blocked ERK activation by H(2)O(2), but not by ONOO(-). The MEK inhibitor PD98059 inhibited ERK activation by both H(2)O(2) and ONOO(-). Moreover, ONOO(-) or H(2)O(2) caused a cytotoxic response of myofibroblasts that was prevented by preincubation with PD98059. In a cell-free kinase assay, ONOO(-) (but not H(2)O(2)) induced autophosphorylation and nitration of a glutathione S-transferase-MEK-1 fusion protein. Collectively, these data indicate that ONOO(-) activates EGFR and Raf-1, but these signaling intermediates are not required for ONOO(-)-induced ERK activation. However, MEK-1 activation is required for ONOO(-)-induced ERK activation in myofibroblasts. In contrast, H(2)O(2)-induced ERK activation is dependent on EGFR activation, which then leads to downstream Raf-1 and MEK-1 activation.

Asbestos exposure upregulates the adhesion of pleural leukocytes to pleural mesothelial cells via VCAM-1.

This study was designed to assess the effects of in vitro and in vivo asbestos exposure on the adhesion of rat pleural leukocytes (RPLs) labeled with the fluorochrome calcein AM to rat pleural mesothelial cells (RPMCs). Exposure of RPMCs for 24 h to either crocidolite or chrysotile fibers (1.25-10 microgram/cm(2)) increased the adhesion of RPLs to RPMCs in a dose-dependent fashion, an effect that was potentiated by interleukin-1beta. These findings were not observed with nonfibrogenic carbonyl iron particles. Crocidolite and chrysotile plus interleukin-1beta also upregulated vascular cell adhesion molecule-1 mRNA and protein expression in RPMCs, and the binding of RPL to asbestos-treated RPMCs was abrogated by anti-vascular cell adhesion molecule-1 antibody. PRLs exposed by intermittent inhalation to crocidolite for 2 wk manifested significantly greater binding to RPMCs than did RPLs from sham-exposed animals. The ability of asbestos fibers to upregulate RPL adhesion to RPMCs may play a role in the induction and/or potentiation of asbestos-induced pleural injury.

Contribution of reactive oxygen and nitrogen species to particulate-induced lung injury.

Recently, a second pathway for the generation of potential oxidants with the reactivity of the hydroxyl radical without the need for metal catalysis has been described. In response to various inflammatory stimuli, lung endothelial, alveolar, and airway epithelial cells, as well as activated alveolar macrophages, produce both nitric oxide (.NO) and superoxide anion radicals (O2.-). .NO regulates pulmonary vascular and airway tone and plays an important role in lung host defense against various bacteria. However, .NO may be cytotoxic by inhibiting critical enzymes such as mitochondrial aconitase and ribonucleotide reductase, by S-nitrosolation of thiol groups, or by binding to their iron-sulfur centers. In addition, .NO reacts with O2.- at a near diffusion-limited rate to form the strong oxidant peroxynitrite (ONOO-), which can nitrate and oxidize key amino acids in various lung proteins such as surfactant protein A, and inhibit their functions. The presence of ONOO- in the lungs of patients with acute respiratory distress syndrome has been demonstrated by measuring levels of nitrotyrosine, the stable product of tyrosine nitration. Various studies have shown that inhalation or intratracheal instillation of various respirable mineral dusts or asbestos fibers increased levels of inducible nitric oxide synthase mRNA. In this presentation, we review the evidence for the upregulation of .NO in the lungs of animals exposed to mineral particulates and assess the contribution of reactive nitrogen species in the pathogenesis of the resultant lung injury.

Asbestos fibers and interleukin-1 upregulate the formation of reactive nitrogen species in rat pleural mesothelial cells.

Nitric oxide radical (.NO) and peroxynitrite anion (ONOO-) have been implicated in lung inflammation and may be important in pleural injury. The present study was undertaken to determine the effects of asbestos exposure and cytokine stimulation on .NO and ONOO- production by rat pleural mesothelial cells. Accordingly, rat parietal pleural mesothelial cells were cultured for 2 to 72 h with or without 50 ng/ml of recombinant interleukin-1beta (IL-1beta) in the presence (1.05 to 8.4 microg/cm2) or absence of crocidolite or chrysotile asbestos fibers. The effects of asbestos were compared with those of carbonyl iron, a nonfibrogenic particulate. Mesothelial cell messenger RNA (mRNA) expression of the inducible form of .NO synthase (iNOS), assessed with the reverse transcription-polymerase chain reaction (RT-PCR), increased progressively from 2 to 12 h in IL-1beta-containing cultures. Nitrite (NO2-), the stable oxidation product of .NO in mesothelial cell conditioned medium, was assayed through the Griess reaction. Both types of asbestos fibers (chrysotile > crocidolite) upregulated the formation of NO2- in mesothelial cells costimulated with IL-1beta in a concentration-dependent and time-dependent fashion. In contrast, carbonyl iron did not upregulate NO2- formation in IL-1beta-stimulated cells. Both types of asbestos fibers also induced iNOS protein expression and the formation of nitrotyrosine in mesothelial cells and greatly induced the formation of nitrate (NO3-), a surrogate marker of ONOO- formation, in IL-1beta-stimulated cells. However, the effects of chrysotile were notably greater than those of crocidolite. These findings may have significance for the induction of pleural injury by asbestos fibers.