Ethical leadership behaviors of senior managers perceived by the junior managers working in public hospitals in Hail, Saudi Arabia.

Objective: To examine junior managers experiences of ethical leadership behaviors exhibited by their senior managers. Methods: In this cross-sectional study, 263 junior health care managers working in public hospitals in Hail, Saudi Arabia were surveyed using a self-administered questionnaire between 20 November, 2022 and 15 February, 2023. Descriptive statistics and regression analysis were employed in the analysis. Statistical Package for Social Sciences (SPSS) version 25 (IBM, Armonk, NY, USA) was used to conduct statistical analyses. Results: The sample consisted of 118 men (44.9%) and the majority (66.6%) of the respondents were below the age of 36 years. In case of working environment, nearly 84% of the participants were satisfied with the relationships that they have had with their supervisors. Regression analysis indicate that women were more likely than men to experience healthy ethical leadership behaviors of their seniors (ß = -0.163, p < 0.05). Ethical leadership behaviors of senior health care managers would not influence by the age or work experience of their juniors. Conclusion: Ethical leadership behavior of senior health care managers was satisfactory. Longitudinal research is needed to investigate how cultural and environmental factors affect the ethical leadership behavior of healthcare managers in Saudi Arabia.

The Impact of the Working Hours Among Malaysian Nurses on Their Ill-Being, Intention to Leave, and the Perceived Quality of Care: A Cross-Sectional Study During the COVID-19 Pandemic.

Purpose: This study explores the effect of working duration on nurses and their ill-being (ie, anxiety, depression, fatigue, and sleepiness), intention to leave, and the quality of nursing care. Methods: A questionnaire survey was employed for a convenience sample of 400 nurses at Malacca General Hospital in Malaysia who voluntarily participated in this cross-sectional study. The Statistical Package for the Social Sciences (SPSS) was used to treat and analyze the data. Descriptive statistics were generated, and Post Hoc analyses and ANOVA tests were conducted. Results: Findings indicated that working hours duration was significantly associated with nurses' anxiety (F (4, 394) = 10.362, p <0.001), depression (F (4, 395) = 23.041, p< 0.001), fatigue (F (4, 395) = 24.232, p< 0.001), sleepiness (F (4, 395) = 4.324, p < 0.002), quality of nursing care (F (4, 395) = 16.21, p <0.001) and intention leave their job, (F (4, 395) = 50.29, p <0.001). The results also revealed that working more than 14 hours was negatively associated with their perceived quality of nursing care and positively associated with their perceived ill-being and intention to leave. Conclusion: Shift length is an important issue, and nursing managers must consider shift length as it can adversely correlate with the nurses' perceptions of work and life.

Hospital Work Conditions and the Mediation Role of Burnout: Residents and Practicing Physicians Reporting Adverse Events.

Background: "Patient Safety" in everyday practices is a target of healthcare leaders, and adverse events reported by healthcare providers directly reflect patient safety in the health system. Recognising how residents and practising physicians rate adverse events concerning their work conditions and burnout must be explored. Objective: This study aims to explore the mediation effect of burnout dimensions (emotional exhaustion and burnout-interpersonal disengagement) between the effects of work conditions on perceived patient safety by exploring the adverse events that residents and practising physicians reported. Methods: A quantitative and cross-sectional study collected data from 249 residents and practising physicians in a huge teaching hospital and primary health care centre (PHC) in the Eastern Province of Saudi Arabia. Hayes Macro regression analysis was employed to evaluate the multiple mediation effect of burnout dimensions, with 5000 bootstrapping and a confidence interval (CI) of 95% for statistical inference and p≤0.05 for the significance level. Results: Leadership support (B= 0.39, t= 6.24, p<0.001) and physician engagement (B=0.43, t=6.50, p<0.001) were associated with a decreased rate of adverse events to patient safety, whereas workload (B=-0.23, t=-3.73, p<0.001) was negatively associated with an increased rate of adverse events. Burnout was shown to mediate the relationship between the effects of physician's leadership support (R2=0.26, F=27.50, p<0.001), work engagement (R2=0.25, F=27.07, p<0.001) and workload (R2=0.23, F=24.23, p<0.001) on the rate of adverse events. Conclusion: This study provides insights into burnout dimensions and their consequences on patient safety indicators (ie, adverse events). Work conditions (ie, leadership support, physician engagement, and workload) directly affect the rate of adverse events and indirectly through mediators like burnout-emotional exhaustion and burnout-interpersonal disengagement.

Knowledge sharing behaviour among head nurses in online health communities: The moderating role of knowledge self-efficacy.

BACKGROUND: Head nurses are vital in understanding and encouraging knowledge sharing among their followers. However, few empirical studies have highlighted their contribution to knowledge-sharing behaviour in Online Health Communities (OHCs). In addition, scant literature has examined the moderating role of knowledge self-efficacy in this regard. PURPOSES: This study examines the moderating role of self-efficacy between the association of four selected individual factors of head nurses (i.e., Trust, Reciprocity, Reputation, and Ability to Share) and their knowledge-sharing behaviour in OHCs in Jordan. METHOD: The data were obtained by using a self-reported survey from 283 head nurses in 22 private hospitals in Jordan. A moderation regression analysis using a structural equation modelling approach (i.e. Smart PLS-SEM, Version 3) was utilised to evaluate the study's measurement and structural model. RESULTS: Knowledge self-efficacy moderates the relationship between the three individual factors (i.e., Trust, Reciprocity, and Reputation) and knowledge-sharing behaviours. However, self-efficacy did not moderate the relationship between the ability to share and knowledge-sharing behaviours. IMPLICATIONS: This study contributes to understanding the moderating role of knowledge self-efficacy among head nurses in online healthcare communities. Moreover, this study provides guidelines for head nurses to become active members in knowledge sharing in OHCs. The findings of this study offer a basis for further research on knowledge sharing in the healthcare sector.

Risk of depression, anxiety, and stress among the Saudi general population during the COVID-19 pandemic.

BACKGROUND: Promoting mental health and wellness is crucial for healthy communities. This study aims to assess the vulnerability of experiencing psychological reactions such as depression, anxiety, and stress within the general population in Saudi Arabia during the COVID-19 pandemic. METHODS: A cross-sectional online survey was completed by 754 participants recruited from thirteen regions of Saudi Arabia. The information on background variables was obtained by using a set of close-ended questions. The DASS-21, a screening tool was used to assess the risk of depression, anxiety, and stress. Pie-chart displayed the levels of risk to experience psychological reactions. The Chi-square test and Adjusted Odd Ratios (AOR) reported the risk factors associated with experiencing psychological reactions in the study population. RESULTS: The vulnerability to mild to extremely severe levels of stress, anxiety and depression were (36.5%), (34.9%), and (43.5%), respectively. Findings demonstrated that females were at higher risk to experience anxiety (AOR = 1.56; 95% CI 1.11-2.18) and stress (AOR = 1.48; 95% CI 1.10-2.10) as compared to males. Unmarried individuals had higher vulnerability for anxiety (AOR = 1.60; 95% CI 1.04-2.44) and depression (AOR = 1.55; 95% CI 1.02-2.37) as compared to married individuals. Those who lost their job during the pandemic had a higher risk to experience anxiety (AOR = 2.02; 95% CI 1.10-3.74) and depression (AOR = 2.01: 95% CI 1.09-3.87) as compared to those who remain employed. Diagnosis with COVID-19 was associated with anxiety (AOR = 2.40; 95% CI 1.27-4.52) and stress (AOR = 2. 0; 95% CI 1.10-3.69). Participants with chronic medical conditions were almost twice at risk to experience stress (AOR = 2.0; 95% CI 1.38-2.87) depression (AOR = 2.14; 95% CI 1.53-2.99) and anxiety (AOR = 2.59; 95% CI 1.78-3.78) as compared to those without such conditions. CONCLUSIONS: Findings imply the allocation of adequate psychological resources to prevent long-term psychological repercussions in at-risk populations such as females, unmarried individuals, those who lost their jobs, diagnosed with COVID-19 and those with chronic medical conditions.

Effectiveness of game-based virtual reality phone application and online education on knowledge, attitude and compliance of standard precautions among nursing students.

Game-based virtual reality phone applications can create a realistic environment to prepare for clinical applications and improve students' knowledge of and compliance with standard precautions. An experimental study was performed among 126 nursing students' from the third and fourth nursing levels to determine the effect of online education and game-based virtual reality phone applications related to standard precautions. Students were divided randomly into two groups; the experimental group used online education and game-based virtual reality phone applications, while the control group used traditional education. The study was performed between July and August 2019 to prevent clashes with lectures and midterm and final examinations. A tool package including knowledge, attitude, and compliance with standard precautions was used in pre-and post-tests among nursing students. The results showed that the knowledge of, attitudes towards, and compliance with standard precautions differed between the two groups. The performance of the experimental group of nursing students significantly improved with online instruction and game-based virtual reality phone applications. This study demonstrated the effectiveness of online education and game-based virtual reality phone application among nursing students.

Patient Safety Attitudes among Doctors and Nurses: Associations with Workload, Adverse Events, Experience.

Patient safety concept has achieved more attention from healthcare organizations to improve the safety culture. This study aimed to investigate patient safety attitudes among doctors and nurses and explore associations between workload, adverse events, and experience with patient safety attitudes. The study used a descriptive cross-sectional design and the Turkish version of the Safety Attitudes Questionnaire. Participants included 73 doctors and 246 nurses working in two private hospitals in Northern Cyprus. The participants had negative perceptions in all patient safety domains. The work conditions domain received the highest positive perception rate, and the safety climate domain received the lowest perception rate among the participants. Nurses showed a higher positive perception than doctors regarding job satisfaction, stress recognition, and perceptions of management domains. There were statistically significant differences between experiences, workloads, adverse events, and total mean scores of patient safety attitudes. Policymakers and directors can improve the quality of care of patients and patient safety by boosting the decision-making of health care providers on several domains of safety attitudes. Patient safety needs to be improved in hospitals through in-service education, management support, and institutional regulations.

Remotely Monitoring COVID-19 Patient Health Condition Using Metaheuristics Convolute Networks from IoT-Based Wearable Device Health Data.

Today, COVID-19-patient health monitoring and management are major public health challenges for technologies. This research monitored COVID-19 patients by using the Internet of Things. IoT-based collected real-time GPS helps alert the patient automatically to reduce risk factors. Wearable IoT devices are attached to the human body, interconnected with edge nodes, to investigate data for making health-condition decisions. This system uses the wearable IoT sensor, cloud, and web layers to explore the patient's health condition remotely. Every layer has specific functionality in the COVID-19 symptoms' monitoring process. The first layer collects the patient health information, which is transferred to the second layer that stores that data in the cloud. The network examines health data and alerts the patients, thus helping users take immediate actions. Finally, the web layer notifies family members to take appropriate steps. This optimized deep-learning model allows for the management and monitoring for further analysis.

Effect of Practice Environment on Nurse Reported Quality and Patient Safety: The Mediation Role of Person-Centeredness.

This study aims to explore the potential mediation role of person-centeredness between the effects of the work environment and nurse reported quality and patient safety. A quantitative cross-sectional survey collected data from 1055 nurses, working in medical and surgical units, in twelve Malaysian private hospitals. The data collection used structured questionnaires. The Hayes macro explored the mediation effect of person-centeredness between the associations of work environment dimensions and care outcomes, controlling nurses' demographics and practice characteristics. A total of 652 nurses responded completely to the survey (61.8% response rate). About 47.7% of nurses worked 7-h shifts, and 37.0% were assigned more than 15 patients. Higher workload was associated with unfavorable outcomes. Nurses working in 12-h shifts reported a lower work environment rating (3.46 ± 0.41, p < 0.01) and person-centered care (3.55 ± 0.35, p < 0.01). Nurses assigned to more than 15 patients were less likely to report a favorable practice environment (3.53 ± 0.41, p < 0.05), perceived lower person-centered care (3.61 ± 0.36, p < 0.01), and rated lower patient safety (3.54 ± 0.62, p < 0.05). Person-centeredness mediates the effect of nurse work environment dimensions on quality and patient safety. Medical and surgical nurses, working in a healthy environment, had a high level of person-centeredness, which, in turn, positively affected the reported outcomes. The function of person-centeredness was to complement the effects of the nurse work environment on care outcomes. Improving the nurse work environment (task-oriented) with a high level of person-centeredness (patient-oriented) was a mechanism through which future initiatives could improve nursing care and prevent patient harm.

Feelings, Behavioral Actions and Depressive Symptoms Related to COVID-19 among Undergraduates in Hail, Saudi Arabia.

The severe acute respiratory syndrome coronavirus 2 (COVID-19) outbreak has had a profound psychological impact on university undergraduates. Feelings, behavioral actions, and depressive symptoms related to COVID-19 in undergraduates in Hail, Saudi Arabia, were assessed using an online survey. Eighteen feelings and six behavioral acts were assessed. The Center for Epidemiological Studies Depression Scale was used to measure depressive symptomatology. Descriptive statistics and logistic regression techniques were used. The mean age of the participants (n = 418) was 20.2 years (standard deviation (SD) = 1.8 years), and 52.9% (n = 221) were males. Elevated levels of depressive symptoms were reported by 47.1% of male and 51.3% of female participants. Social isolation, loss of interest, obsessive monitoring of symptoms, concentration difficulties, recurrent negative thoughts, and worries about health services emerged as vital negative feelings related to COVID-19 that were expressed by the participants. Younger age (odds ratio (OR) = 0.636, 95% confidence interval (95% CI) = 0.428-0.946) and coming from the middle-income category (OR = 0.388, 95% CI = 0.151-0.994) were found to be protective factors against developing depressive symptoms. Frequent cleaning of hands, wearing masks when going out, and adherence to social distancing rules were practiced by 71.5% (95% CI = 67.2-75.8%), 78.7% (95% CI = (74.4-82.5%) and 66.0% (95% CI = (61.3-70.5%) of the participants, respectively. These behavioral acts were not associated with the development of depressive symptoms. Although the practice of COVID-19 precautionary measures by the participants was satisfactory, nearly half of the participants reported depressive symptoms. Innovative educational strategies are needed to curb concentration difficulties and social isolation experienced by undergraduates during outbreaks such as COVID-19.

Depressive Symptomatology and Practice of Safety Measures among Undergraduate Students during COVID-19: Impact of Gender.

The COVID-19 pandemic has greatly affected the personal and academic lives of undergraduates in Saudi Arabia. Although studies have suggested that COVID-19 increased the prevalence of psychological health problems among undergraduates, the associations between the risk of depression and safety practices and the influence of gender on these associations have not been studied in detail. A cross-sectional online survey was conducted among preparatory-year undergraduates in a large public university in Saudi Arabia during the outbreak. Depressive symptoms were assessed using the Center for Epidemiological Studies Depression (CES-D) Scale, and the practice of eight precautionary behaviors was also assessed. Data analysis was performed using the chi-square test, multiple linear regression and Spearman's correlation coefficient. In total, 3044 undergraduates were surveyed. The mean age was 18.6 years (SD = 0.84), and 61.9% (n = 1883) of the participants were female. Overall, 47.7% of the participants reported having elevated depressive symptoms. Overall mean values of CES-D scores were higher among female undergraduates than that of male undergraduates (18.08 versus 15.56, p < 0.01). There were inverse and weak but significant relationships between the CES-D score and frequent cleaning of hands (male: r = -0.116, p < 0.01; female: r = -0.098, p < 0.01), wearing a mask when going out (male: r = -0.172, p < 0.01; female: r = -0.135, p < 0.01), keeping social distance (male: r = -0.117, p < 0.01; female: r = -0.147, p < 0.01), and covering the nose when sneezing (male: r = -0.202, p < 0.01; female: r = -0.115, p < 0.01). Regression analysis indicated that adherence to precautionary measures was a strong predictor of reduction of depressive symptoms in the target population. Male gender was also found to be an independent predictor of reduction of depressive symptoms. Depressive symptoms were highly prevalent in this target group, and female undergraduates seemed to be more vulnerable to developing such symptoms. Results also indicated that female undergraduates were more likely to implement the protective measures for COVID-19. The promotion of precautionary measures seems to be effective in reducing distress in this target population, but further research is needed to confirm our assertions.

Overexpression of 15-lipoxygenase-1 in PC-3 human prostate cancer cells increases tumorigenesis.

The effect of overexpression of 15-lipoxygenase-1 (15-LO-1) was studied in the human prostate cancer cell line, PC-3. Stable PC-3 cell lines were generated by transfection with 15-LO-1-sense (15-LOS), 15-LO-1-antisense (15-LOAS) or vector (Zeo) and selection with Zeocin. After characterization by RT-PCR, western and HPLC, a PC3-15LOS clone was selected that possessed 10-fold 15-LO-1 enzyme activity compared with parental PC-3 cells. The PC3-15LOAS clone displayed little or no 15-LO-1 activity. These PC-3 cell lines were characterized for properties of tumorigenesis. The proliferation rates of the cell lines were as follows: PC3-15LOS > PC-3 = PC3-Zeo > PC3-15LOAS. Addition of a specific 15-LO-1 inhibitor, PD146176, caused a dose-dependent inhibition of proliferation in vitro. Overexpression of 15-LO-1 also caused [(3)H]thymidine incorporation to increase by 4.0-fold (P < 0.01). Compared with parental and PC-3-Zeo cells, PC3-15LOS enhanced whereas PC3-15LOAS reduced the ability of PC-3 cells to grow in an anchorage-independent manner, as assessed by colony formation in soft agar. These data suggested a pro-tumorigenic role for 15-LO-1 in PC-3 cells in vitro. Therefore, to clarify the role of 15-LO-1 in vivo, the effect of 15-LO-1 expression on the growth of tumors in nude mice was investigated. The PC-3 cell lines were inoculated subcutaneously into athymic nude mice. The frequency of tumor formation was increased and the sizes of the tumors formed were much larger in the PC3-15LOS compared with PC3-15LOAS, parental PC-3 and PC-3-Zeo cells. Immunohistochemistry for 15-LO-1 confirmed expression throughout the duration of the experiment. The expression of factor VIII, an angiogenesis marker, in tumor sections was increased in tumors derived from PC3-15LOS cells and decreased in those from PC3-15LOAS cells compared with tumors from parental or Zeo cells. These data further supported the evaluation by ELISA of vascular endothelial growth factor (VEGF) secretion by PC-3 cells in culture. Secretion of this angiogenic factor was elevated in PC3-15LOS cells compared with the other cell lines. These results support a role for 15-LO-1 in a novel growth-promoting pathway in the prostate.

F(2)-isoprostanes mediate high glucose-induced TGF-beta synthesis and glomerular proteinuria in experimental type I diabetes.

BACKGROUND: The recently discovered arachidonic acid derivatives, isoprostanes, are increased in pathological conditions associated with oxidative stress, such as diabetes. No role has yet been described for isoprostanes during the development of diabetic nephropathy. Cell culture in high ambient glucose has been used as a model in elucidating cellular mechanisms underlying diabetic nephropathy. Among the growth factors involved in the effect of high glucose, transforming growth factor-beta (TGF-beta) has been described as playing a key role in the development of nephropathy. METHODS: Streptozotocin-induced diabetic rats were supplemented in their diet with the antioxidant vitamin E (1000 U/kg diet). Blood and urine samples were taken to determine renal function and isoprostane concentration, as determined by gas chromatography/mass spectrometry. Glomerular mesangial and endothelial cells were cultured in high ambient glucose to determine the synthesis of isoprostanes and the role of isoprostanes in high glucose-induced synthesis of TGF-beta. RESULTS: Streptozotocin-induced diabetic rats had marked increases in plasma levels and urinary excretion rates of F(2)-isoprostanes. Dietary supplementation with vitamin E normalized (plasma) and reduced (urine) isoprostane levels and, surprisingly, improved proteinuria and blood urea nitrogen (BUN) levels. High ambient glucose increased F(2)-isoprostane synthesis in glomerular endothelial and mesangial cells in culture. Incubation of glomerular cells with F(2)-isoprostanes stimulated the production of TGF-beta. CONCLUSIONS: Increased F(2)-isoprostane synthesis during diabetes appears to be responsible in part for the increase in renal TGF-beta, a well-known mediator of diabetic nephropathy.

Concordant induction of 15-lipoxygenase-1 and mutant p53 expression in human prostate adenocarcinoma: correlation with Gleason staging.

We recently reported that the mutant form of the tumor-suppressor gene p53 up-regulates 15-LO-1 gene expression in a murine cell line. Here, we examine the expression of 15-lipoxygenase (LO)-1 and mutant p53 (mtp53) in human prostatic tissues and 15-LO-1 in the human prostate adenocarcinoma cell line PC-3. Reverse transcription-PCR and western analyses conclusively demonstrated expression of 15-LO-1 in PC-3 cells. Western blotting for 15-LO-1 in freshly resected 'normal' and prostate adenocarcinoma specimens showed 15-LO-1 expression in normal tissue, but significantly higher levels were detected in prostate adenocarcinomas. Prostate adenocarcinoma tissues generated chirally pure 13-S-hydroxyoctadecadienoic acid from exogenous linoleic acid, a preferred substrate of 15-LO-1. To study the correlation of 15-LO-1 expression with mtp53 in prostate cancer, we immunostained 48 prostatectomy specimens obtained by transurethral resection of the prostate and needle biopsy (median age 68 years, range 52-93) of different Gleason grades (n = 48), using antibodies specific for 15-LO-1, mtp53 and MIB-1 (a proliferation marker). We compared staining in cancerous foci with adjacent normal appearing prostate tissues. In only 5 of 48 patients did 'normal' tissue adjacent to cancerous foci display staining for 15-LO-1. However, no staining for mtp53 was observed in any of the normal tissues. In cancer foci, robust staining was observed for both 15-LO-1 (36 of 48, 75%) and mtp53 (19 of 48, 39%). Furthermore, the intensities of expression of 15-LO-1 and mtp53 correlated positively with each other (P < 0.001) and with the degree of malignancy, as assessed by Gleason grading (P < 0.01). By immunohistochemistry, 15-LO-1 was located in secretory cells of peripheral zone glands, prostatic ducts and seminal vesicles, but not in the basal cell layer or stroma. Based on these and other studies, we propose a model describing a possible role for 15-LO-1 expression in influencing the malignant potential and pathobiological behavior of adenocarcinomas.

Reciprocal regulation of LTA(4) hydrolase expression in human monocytes by gamma-interferon and interleukins 4 and 13: potential relevance to leukotriene regulation in glomerular disease.

BACKGROUND/AIMS: Leukotriene A(4) (LTA(4)) hydrolase catalyzes the final step in the synthesis of leukotriene B(4) (LTB(4)). TH-1- and TH-2-derived cytokines may regulate LTB(4) synthesis by monocytes through their actions on the expression of LTA(4) hydrolase. METHODS: Freshly isolated monocytes were incubated with pro- and anti-inflammatory cytokines for 36 h. mRNA expression was determined by Northern blot, protein expression was determined by Western blot and LTB(4) synthesis was determined by ELISA. RESULTS: Interferon-gamma (a TH-2-derived cytokine) increased significantly LTA(4) hydrolase mRNA expression, whereas interleukin (IL)-4 and IL-13 (both TH-2-derived cytokines) decreased LTA(4) hydrolase mRNA expression in these cells. The same effects were seen on the expression of immunoreactive LTA(4) hydrolase after incubating the monocytes with either TH-1- or TH-2-derived cytokines. The monocyte-derived cytokine IL-1 beta did not show any significant effect on LTA(4) hydrolase mRNA expression. When LTB(4) release was measured, both IL-1 beta and interferon-gamma significantly increased LTB(4) production by monocytes, while TH-2 cytokines (IL-4 and IL-13) decreased it. CONCLUSION: The opposing effects of TH-1- and TH-2-derived cytokines on the expression of LTA(4) hydrolase mRNA may regulate LTB(4) synthesis by monocytes during inflammation.

15-Lipoxygenase in glomerular inflammation.

Glomerulonephritis is a significant factor fueling the rapid increase in the population of patients with end-stage renal disease. Novel therapeutic strategies targeting specific mechanisms of glomerular destruction are the most reasonable approaches to arrest ongoing injury. In this review, we summarize some of our results obtained in our effort to characterize the role of 15-lipoxygenase activation as one of the mechansisms operative during the early, prefibrotic stage of glomerular immune injury. We also summarize the effects of cytokines released during these processes, as well as the activation by aspirin of the synthesis of 15-R-HETE (see text). Finally, we will propose a clinical approach to this group of disorders, based on emerging concepts of the pathophysiology of glomerulonephritis from our work and that of several other investigators.

Ku autoantigen (DNA helicase) is required for interleukins-13/-4-induction of 15-lipoxygenase-1 gene expression in human epithelial cells.

As reported previously in human monocytes, a human lung epithelial cell line, A549, showed de novo induction of 15-Lipoxygenase-1 (15-LO-1) in response to interleukins-13 (IL-13) and -4 (IL-4). In this cell line, 15-LO-1 expression, by RT-PCR and western blotting, was observed following 6 and 24 h of exposure to human IL-13 (ED50 5 ng/ml) and IL-4 (ED50 0.2 ng/ml). We have previously shown that no cis-acting regulatory elements exist within the 15-LO-1 promoter region. To define IL-13 and IL-4 responsive trans-acting elements, we identified a region (DP2: -353 to -304 bp site) within the 15-LO-1 promoter (by footprinting experiments) to which IL-13-responsive elements (or factors) bind specifically (Kelavkar et al, 1998, Mol Biol Rep 25, 173-182). To further delineate this region, we constructed (by site-directed mutagenesis) several deletion mutants in the 'LOPB5' region containing the 29 bp within the -353 to -304 bp of the DP2 core element. These were: DP3 (site totally deleted), DP4 (5 bp deleted at the center of the site), DP5 (8 bp at the 5'-end of the site) and DP6 (13 bp at the 3'-end of the site). Cotransfection of these deletion constructs (driving luciferase reporter genes) was associated with 90% (DP4, DP5 and DP6) or 100% (DP3) abrogation of promoter activity at 24 h. Purification of nuclear protein extracts from IL-13 and IL-4-stimulated A549 cells, using a DP2 core containing affinity column, identified a 150 kDa protein under non-denaturing conditions, and two, 70 and 85 kDa proteins under denaturing conditions. These were not detectable by Coomassie blue staining in control nuclear protein extracts. Matrix assisted laser desorption ionization mass spectrometry (MALDI-MS) of the tryptic digests of these proteins, identified one as the 86 kDA Lupus KU autoantigen protein P86 and the second as the 70 kDa Lupus KU autoantigen protein P70. Gel shift and supershift experiments using monoclonal antibodies toward Ku antigen and its individual subunits, and utilizing DP2 and other mutant oligonucleotides with purified nuclear protein extracts from control and cytokine-treated A549 cells, confirmed our findings. Furthermore, electroporation of neutralizing anti-Ku70, Ku 80 and Ku70/80 antibodies into A549 cells totally suppressed IL-13 and IL-4-stimulated 15-LO-1 induction in these cells. Further, immunoprecipitation experiments data suggests that IL-4 and IL-13 activate Ku antigens and 15-LO-1 expression through distinct signaling events. In summary, in A549 cells, Ku antigen is induced in response to the cytokines, IL-13 and -4, and a 29 bp region within the -353 to -304 bp region of the 15-LO-1 promoter is required for its binding and subsequent induction of 15-LO-1 gene expression. The findings may provide an important link between the established dysregulated function of Ku antigen in auto-immune diseases, such as systemic lupus erythematosus and thyroiditis, and the increasingly recognized 'anti-inflammatory' role of 15-LO-1.

Transfection of rat kidney with human 15-lipoxygenase suppresses inflammation and preserves function in experimental glomerulonephritis.

The human 15-lipoxygenase (15-LO) gene was transfected into rat kidneys in vivo via intra-renal arterial injection. Three days later, acute (passive) or accelerated forms of antiglomerular basement membrane antibody-mediated glomerulonephritis were induced in transfected and nontransfected or sham-transfected controls. Studies of glomerular functions (filtration and protein excretion) and ex vivo glomerular leukotriene B(4) biosynthesis at 3 hr, and up to 4 days, after induction of nephritis revealed preservation or normalization of these parameters in transfected kidneys that expressed human 15-LO mRNA and mature protein, but not in contralateral control kidneys or sham-transfected animals. The results provide in vivo-derived data supporting a direct anti-inflammatory role for 15-LO during immune-mediated tissue injury.

Free-radical-generated F2-isoprostane stimulates cell proliferation and endothelin-1 expression on endothelial cells.

BACKGROUND: Free-radical-generated F2-isoprostane stimulates DNA synthesis and endothelin-1 (ET-1) expression on endothelial cells. 8-Iso-prostaglandin F2alpha (8-iso-PGF2alpha) is a member of the recently discovered family of prostanoids, the F2-isoprostanes, produced in vivo by cyclooxygenase-independent, free-radical-catalyzed lipid peroxidation. The goal of our study is to establish the effect of isoprostane on ET-1 production by endothelial cells, as well to determine the receptors responsible for these effects. METHODS: The proliferative effect of isoprostanes was measured as an increase of viable cell number and [3H]-thymidine uptake. ET-1 gene expression and protein synthesis were determined by Northern blot and radioimmunoassay, respectively. We also determined inositol 1,4,5-trisphosphate synthesis. Thromboxane A2 (TXA2) receptor antagonist SQ29,548 was used to establish the role of TXA2 receptor in isoprostane effect, as well as to determine the type of receptors involved in these effects. RESULTS: Our results show that physiological concentrations of 8-iso-PGF2alpha stimulated cell proliferation, DNA synthesis, and ET-1 mRNA and protein expression in bovine aortic endothelial cells (BAECs). The proliferative effect was partially abolished by treatment with anti-endothelin antibody. 8-Iso-PGF2alpha also increased inositol 1, 4,5-trisphosphate formation in these cells. These effects were partially inhibited by SQ29,548. In competitive binding assays, two binding sites were recognized on BAECs with dissociation constants (Kd) and binding site densities at equilibrium similar to those previously described in smooth muscle cells and likely represent [3H]-8-iso-PGF2alpha binding to its own receptor (high-affinity binding site) and cross-recognition of the TXA2 receptor (low-affinity binding site). CONCLUSION: These studies expand the potential scope of the pathophysiologic significance of F2-isoprostanes, released during oxidant injury, to include alteration of endothelial cell biology.

MK-591 acutely restores glomerular size selectivity and reduces proteinuria in human glomerulonephritis.

BACKGROUND: Leukotrienes are 5-lipoxygenated (5-LO) metabolites of arachidonic acid that mediate some of the glomerular hemodynamic and structural changes in experimental and human glomerulonephritis. METHODS: We conducted an open-label, pilot study of the short-term effects of leukotriene biosynthesis inhibition using an orally active 5-LO activating protein (FLAP) antagonist (MK-591) on glomerular function in patients with glomerulonephritis. Eleven adult patients (seven women, median age 38 years) with glomerulonephritis (5 lupus nephritis, 2 IgA nephropathy, 1 membranoproliferative, 1 membranous, 1 C1q-deficiency, and 1 idiopathic crescentic) and moderate renal insufficiency [glomerular filtration rate (GFR) 62 +/- 9 ml/min/1.73 m2] were given MK-591 at a dose of 100 mg orally twice a day for four days. RESULTS: MK-591 reduced proteinuria (albumin and IgG excretion rates) from 3233 +/- 1074 to 1702 +/- 555 microg/min and from 196 +/- 78 to 148 +/- 55 microg/min for albumin and IgG, respectively (P < 0.05 for both). This was not accompanied by a reduction in systemic arterial pressure, GFR, or renal plasma flow. By analysis of the fractional clearance of polydisperse dextrans, baseline proteinuria resulted from a loss of size selectivity with enhanced passage of large (>52 A) dextrans as compared with healthy controls. Treatment with MK-591 caused a selective improvement in the enhanced passage of large (>58 A) dextrans without affecting the handling of smaller dextrans, indicating an improvement in glomerular size selectivity. MK-591 was well tolerated, and no adverse effects were observed. CONCLUSIONS: Short-term therapy with MK-591 reduces proteinuria by restoring glomerular size selectivity and thus reduces transglomerular protein trafficking. These benefits may result from glomerular leukotriene biosynthesis inhibition, but other MK-591-specific actions cannot be excluded.