Impact of HMGB1/TLR Ligand Complexes on HIV-1 Replication: Possible Role for Flagellin during HIV-1 Infection

Objective. We hypothesized that HMGB1 in complex with bacterial components, such as flagellin, CpG-ODN, and LPS, promotes HIV-1 replication. Furthermore, we studied the levels of antiflagellin antibodies during HIV-1-infection. Methods. Chronically HIV-1-infected U1 cells were stimulated with necrotic extract/recombinant HMGB1 in complex with TLR ligands or alone. HIV-1 replication was estimated by p24 antigen in culture supernatants 48–72 hours after stimulation. The presence of systemic anti-flagellin IgG was determined in 51 HIV-1-infected patients and 19 controls by immunoblotting or in-house ELISA. Results. Flagellin, LPS, and CpG-ODN induced stronger HIV-1 replication when incubated together with necrotic extract or recombinant HMGB1 than activation by any of the compounds alone. Moreover, the stimulatory effect of necrotic extract was inhibited by depletion of HMGB1. Elevated levels of anti-flagellin antibodies were present in plasma from HIV-1-infected patients and significantly decreased during 2 years of antiretroviral therapy. Conclusions. Our findings implicate a possible role of HGMB1-bacterial complexes, as a consequence of microbial translocation and cell necrosis, for immune activation in HIV-1 pathogenesis. We propose that flagellin is an important microbial product, that modulates viral replication and induces adaptive immune responses in vivo

Rebound of residual plasma viremia after initial decrease following addition of intravenous immunoglobulin to effective antiretroviral treatment of HIV

<p>Abstract</p> <p>Background</p> <p>High dosage of intravenous immunoglobulin (IVIG) has been observed as a possible activator of HIV gene expression in latently infected resting CD4⁺T-cells, leading to a substantial decrease in both the reservoir and the residual plasma viremia when added to effective ART. IVIG treatment has also been reported to expand T regulatory cells (Tregs). The aim of this study was to evaluate possible long-term effect of IVIG treatment on residual viremia and T-lymphocyte activation.</p> <p>Methods</p> <p>Nine HIV-infected subjects on effective ART included in a previously reported study on IVIG treatment were evaluated 48-104 weeks after therapy. In addition, 14 HIV-infected controls on suppressive ART were included. HIV-1 RNA was analyzed in cell-free plasma by using an ultrasensitive PCR-method with a detection limit of 2 copies/mL. T-lymphocyte activation markers and serum interleukins were measured.</p> <p>Results</p> <p>Plasma residual viremia rebounded to pre-treatment levels, 48-104 weeks after the initial decrease that was observed following treatment with high-dosage IVIG. No long-term effect was observed regarding T-lymphocyte activation markers, T-regulatory cells or serum interleukins. In a post-hoc analysis, a correlation between plasma HIV-1-RNA and CD4⁺T-cell count was found in both IVIG-treated patients and controls.</p> <p>Conclusions</p> <p>These results indicate that the decrease in the latent HIV-1 pool observed during IVIG treatment is transient. Although not our primary objective, we found a correlation between HIV-1 RNA and CD4⁺T-cell count suggesting the possibility that patients with a higher CD4⁺T-cell count might harbor a larger residual pool of latently infected CD4⁺T-cells.</p

Reduction of the HIV-1 reservoir in resting CD4+ T-lymphocytes by high dosage intravenous immunoglobulin treatment: a proof-of-concept study

<p>Abstract</p> <p>Background</p> <p>The latency of HIV-1 in resting CD4⁺T-lymphocytes constitutes a major obstacle for the eradication of virus in patients on antiretroviral therapy (ART). As yet, no approach to reduce this viral reservoir has proven effective.</p> <p>Methods</p> <p>Nine subjects on effective ART were included in the study and treated with high dosage intravenous immunoglobulin (IVIG) for five consecutive days. Seven of those had detectable levels of replication-competent virus in the latent reservoir and were thus possible to evaluate. Highly purified resting memory CD4⁺T-cells were activated and cells containing replication-competent HIV-1 were quantified. HIV-1 from plasma and activated memory CD4⁺T-cells were compared with single genome sequencing (SGS) of the <it>gag </it>region. T-lymphocyte activation markers and serum interleukins were measured.</p> <p>Results</p> <p>The latent HIV-1 pool decreased with in median 68% after IVIG was added to effective ART. The reservoir decreased in five, whereas no decrease was found in two subjects with detectable virus. Plasma HIV-1 RNA ≥ 2 copies/mL was detected in five of seven subjects at baseline, but in only one at follow-up after 8–12 weeks. The decrease of the latent HIV-1 pool and the residual plasma viremia was preceded by a transitory low-level increase in plasma HIV-1 RNA and serum interleukin 7 (IL-7) levels, and followed by an expansion of T regulatory cells. The magnitude of the viral increase in plasma correlated to the size of the latent HIV-1 pool and SGS of the <it>gag </it>region showed that viral clones from plasma clustered together with virus from activated memory T-cells, pointing to the latent reservoir as the source of HIV-1 RNA in plasma.</p> <p>Conclusion</p> <p>The findings from this uncontrolled proof-of-concept study suggest that the reservoir became accessible by IVIG treatment through activation of HIV-1 gene expression in latently-infected resting CD4⁺T-cells. We propose that IVIG should be further evaluated as an adjuvant to effective ART.</p

Aspects on latency in HIV-1 infection

The use of combination antiretroviral therapy (ART) has resulted in a substantial reduction in viremia, a rebound of CD4+ T-cells and increased survival for human immunodeficiency virus type 1 (HIV-1) infected individuals. Unfortunately, ART does not clear the infection or cure the individual from HIV-1. This is due to some unique characteristics of the virus, such as an ability to cause a latent infection in a subset of CD4+ T-cells, and a remarkable genetic variability. This allows HIV to become resistant to treatment under suboptimal conditions (i.e. adherence), and to escape the immune system.. The mechanisms of latency needs to be better understood in order to achieve eradication of HIV-1 infection, and the general aim of this thesis was to study different aspects on HIV-1 latency, with a main focus on the circulating pool of HIV-1 infected resting memory CD4+ T-cells. In paper I we set out to analyze whether resting memory CD4+ T-cells could serve as a reservoir of founder or resistant viral strains in patients with or without optimal suppressed viremia. Our results indicated a turn-over and replacement of the cell-pool in untreated and suboptimally treated patients. The memory CD4+ T-cell-pool forms an archive of the viral population, but former resistant viral variants are not always found in these circulating cells. In paper II we wanted to investigate if treatment with high dose intraveneous immunoglobulin (IVIG) in combination with ART could reduce the pool of latently infected resting memory CD4+ T-cells in vivo. Our data suggested a reduction by an average of 68% of the HIV infected latent cell pool. The findings from this proof-ofconcept study suggest that the reservoir became accessible by IVIG treatment through activation of HIV-1 in the latently-infected resting CD4+ T-cells. IVIG will be further evaluated as an adjuvant to effective ART. In paper III, we investigated the impact of DNA methylation inhibitors and NFkB activators on HIV-1 latency in resting memory CD4+ T-cells. Cytosine methylation may be an important component that controls HIV-1 latency. We found that a combination of the DNA methylation inhibitor Aza-CdR and the NFkB activator prostratin can reactivate latent HIV-1 in vitro. Our data indicate that clearance of HIV-1 from infected persons undergoing antiretroviral therapy may be enhanced by inclusion of these inhibitors. In paper IV we wanted to investigate to what extent high mobility group box protein 1, HMGB1, alone or in complex with TLR ligands activate latent HIV-1 in vitro. HMGB1 is an abundant intracellular protein found in all cells in the body. It contributes to immune activation and has been suggested to have an important impact on HIV-1 pathogenesis. We found that recombinant HMGB1, or HMGB1 as part of necrotic extract , in complex with TLR stimulating ligands LPS, CpG ODN and flagellin, increased HIV-1 replication in vitro, as compared to HMGB1 or ligands alone, in a model of latent HIV-1 infection. Our in vivo data suggest that a turn-over of the latent HIV-1 reservoir in resting memory CD4+ T-cells may appear in the natural course of HIV-1 infection in untreated and in suboptimally treated subjects. It is also possible to induce a decrease of the viral reservoirs by additional treatment, in our case IVIg, in patients with controlled viremia. Our in vitro data point at new possibilities to activate latent HIV-1 and further exploration of these mechanisms should be considered

Students’ performance of and perspective on an objective structured practical examination for the assessment of preclinical and practical skills in biomedical laboratory science students in Sweden: a 5-year longitudinal study

Purpose It aims to find students’ performance of and perspectives on an objective structured practical examination (OSPE) for assessment of laboratory and preclinical skills in biomedical laboratory science (BLS). It also aims to investigate the perception, acceptability, and usefulness of OSPE from the students’ and examiners’ point of view. Methods This was a longitudinal study to implement an OSPE in BLS. The student group consisted of 198 BLS students enrolled in semester 4, 2015–2019 at Karolinska University Hospital Huddinge, Sweden. Fourteen teachers evaluated the performance by completing a checklist and global rating scales. A student survey questionnaire was administered to the participants to evaluate the student perspective. To assess quality, 4 independent observers were included to monitor the examiners. Results Almost 50% of the students passed the initial OSPE. During the repeat OSPE, 73% of the students passed the OSPE. There was a statistically significant difference between the first and the second repeat OSPE (P<0.01) but not between the first and the third attempt (P=0.09). The student survey questionnaire was completed by 99 of the 198 students (50%) and only 63 students responded to the free-text questions (32%). According to these responses, some stations were perceived as more difficult, albeit they considered the assessment to be valid. The observers found the assessment protocols and examiner’s instructions assured the objectivity of the examination. Conclusion The introduction of an OSPE in the education of biomedical laboratory scientists was a reliable, and useful examination of practical skills

The influence of nursing home managers’ leadership on person-centred care and stress of conscience: A cross-sectional study

Background: Leadership and stress are common concepts in nursing, and this study explores empirically the connection between leadership and stress of conscience in the context of aged care practice. Previous literature has shown that when staff are unable to carry out their ethical liabilities towards the residents, feelings of guilt may occur among staff, which may be an expression of stress of conscience. Although leadership has been described as crucial for staff’s work perceptions of stress as well as for person-centred practices, the influence of nursing home managers’ leadership on stress of conscience among staff and person-centred practices is still not fully explored. This study attempts to address that knowledge gap by exploring the relationship between leadership, person-centred care, and stress of conscience. Methods: This study was based on a cross-sectional national survey of 2985 staff and their managers in 190 nursing homes throughout Sweden. Descriptive statistics and regression modelling were used to explore associations. Results: Leadership was associated with a higher degree of person-centred care and less stress of conscience. A higher degree of person-centred care was also associated with less stress of conscience. The results also showed that leadership as well as person-centred care were individually associated with lower levels of stress of conscience when adjusting for potential confounders. Conclusion: Nursing home managers’ leadership was significantly associated with less staff stress of conscience and more person-centred care. This indicates that a leadership most prominently characterised by coaching and giving feedback, relying on staff and handling conflicts constructively, experimenting with new ideas, and controlling work individually can contribute to less staff stress as well as higher degree of person-centred care provision

Exploring the impact of nursing home managers' leadership on staff job satisfaction, health and intention to leave in nursing homes

Aims and Objectives: To explore the impact of nursing home leadership and staffing characteristics on staff job satisfaction, health and intention to leave. Background: The number of older people has outpaced growth in the nursing home workforce worldwide. Identifying predictors with the potential to positively impact staff job satisfaction, health and intentions to leave are important. Leadership of the nursing home manager can be one such predictor. Design: Cross-sectional design. Methods: A sample of 2985 direct care staff in 190 nursing homes in 43 randomly selected municipalities in Sweden completed surveys on leadership, job satisfaction, self-rated health and intention to leave (response rate 52%). Descriptive statistics and Generalised Estimating Equations were conducted. The STROBE reporting checklist was applied. Results: Nursing home managers' leadership was positively related to job satisfaction, self-rated health and low intention to leave. Lower staff educational levels were related to poorer health and lower job satisfaction. Conclusions: Nursing home leadership plays a significant role in the job satisfaction, self-reported health and intention to leave of direct care staff. Low education levels among staff seem to negatively influence staff health and job satisfaction, suggesting that educational initiatives for less-educated staff could be beneficial for improving staff health and job satisfaction. Relevance to clinical practice: Managers seeking to improve staff job satisfaction can consider how they support, coach and provide feedback. Recognising staff achievement at work can contribute to high job satisfaction. One important implication for managers is to offer continuing education to staff with lower or no education, given the large amount of uneducated direct care workers in aged care and the impact this may have on staff job satisfaction and health. No patient or public contribution: No patient or public contribution was required to outcome measures in this study. Direct care staff and managers contributed with data

Longitudinal changes in nursing home leadership, direct care staff job strain and social support in Swedish nursing homes : findings from the U-AGE SWENIS study

Background: Promoting healthy work environment as a manager in nursing homes is important to safeguard staff health and well-being as well as care quality when facing increasing demands. The impact of leadership on staff work environment needs further exploration. Objectives: To describe longitudinal changes in nursing home leadership, direct care staff characteristics, job strain and social support. Methods: This study has a repeated cross-sectional design, a five-year follow-up study. Nursing home staff in 181 corresponding units (n = 1253 in 2014 and n = 1176 in 2019) completed surveys about leadership, staff job strain and social support in a five-year follow-up study. Descriptive and regression analyses were conducted. Results: A higher degree of leadership defined by coaching and providing direct feedback to care staff, handling conflicts in a constructive way and having control of the clinical work, was significantly associated with a lower degree of job strain and a higher degree of social support among staff, with stronger associations at follow-up. The proportion of enrolled nurses increased significantly at follow-up. Conclusions: Leadership is increasingly important for staff work environment, especially in times of increased workload and decreasing collegiality and deteriorating work atmosphere at work. Implications for Practice: Stakeholder and policy makers in nursing home care may reflect on how managers' leadership is prioritised in these environments because such leadership is associated with staff job strain and social support. Managers striving to improve the work situation of staff may consider their own role and allow flexibility in how and when the work can be performed

The significance of nursing home managers' leadership : longitudinal changes, characteristics and qualifications for perceived leadership, person-centredness and climate

Aims and objectives: The aim was to explore changes in nursing home managers' leadership, person-centred care and psychosocial climate comparing matched units in a five-year follow-up and to explore the significance of managers' educational qualifications and the ownership of nursing homes for perceived leadership, person-centred care and psychosocial climate in the follow-up data. Background: Leadership has been described as crucial for person-centred care and psychosocial climate even though longitudinal data are lacking. The significance of managerial leadership, its characteristics, managerial qualifications and ownership of nursing homes for perceived leadership, person-centred care and psychosocial climate also needs further exploration. Design: Repeated cross-sectional study. Methods: This study used valid and reliable measures of leadership, person-centred care, psychosocial climate and demographic variables collected from managers and staff n = 3605 in 2014 and n = 2985 in 2019. Descriptive and regression analyses were used. The STROBE checklist was used in reporting this study. Results: Leadership was still positively significantly associated to person-centred care in a five-year follow-up, but no changes in strength were seen. Leadership was still positively significantly associated with psychosocial climate, with stronger associations at follow-up. Six leadership characteristics increased over time. It was also shown that a targeted education for nursing home managers was positively associated with person-centred care. Conclusions: Leadership is still pivotal for person-centred care and psychosocial climate. Knowledge of nursing home managers' leadership, characteristics and educational qualifications of significance for person-centred delivery provides important insights when striving to improve such services. Relevance to clinical practice: The findings can be used for management and clinical practice development initiatives because it was shown that nursing home managers' leadership is vital to person-centred care practices and improves the climate for both staff and residents in these environments