Neutralization activity in chronic HIV infection is characterized by a distinct programming of follicular helper CD4 T cells.

A subset of people living with HIV (PLWH) can produce broadly neutralizing antibodies (bNAbs) against HIV, but the lymph node (LN) dynamics that promote the generation of these antibodies are poorly understood. Here, we explored LN-associated histological, immunological, and virological mechanisms of bNAb generation in a cohort of anti-retroviral therapy (ART)-naïve PLWH. We found that participants who produce bNAbs, termed neutralizers, have a superior LN-associated B cell follicle architecture compared with PLWH who do not. The latter was associated with a significantly higher in situ prevalence of Bcl-6hi follicular helper CD4 T cells (TFH), expressing a molecular program that favors their differentiation and stemness, and significantly reduced IL-10 follicular suppressor CD4 T cells. Furthermore, our data reveal possible molecular targets mediating TFH- B cell interactions in neutralizers. Together, we identify cellular and molecular mechanisms that contribute to the development of bNAbs in PLWH.

Follicular Immune Landscaping Reveals a Distinct Profile of FOXP3hiCD4hi T Cells in Treated Compared to Untreated HIV.

Follicular helper CD4hi T cells (TFH) are a major cellular pool for the maintenance of the HIV reservoir. Therefore, the delineation of the follicular (F)/germinal center (GC) immune landscape will significantly advance our understanding of HIV pathogenesis. We have applied multiplex confocal imaging, in combination with the relevant computational tools, to investigate F/GC in situ immune dynamics in viremic (vir-HIV), antiretroviral-treated (cART HIV) People Living With HIV (PLWH) and compare them to reactive, non-infected controls. Lymph nodes (LNs) from viremic and cART PLWH could be further grouped based on their TFH cell densities in high-TFH and low-TFH subgroups. These subgroups were also characterized by different in situ distributions of PD1hi TFH cells. Furthermore, a significant accumulation of follicular FOXP3hiCD4hi T cells, which were characterized by a low scattering in situ distribution profile and strongly correlated with the cell density of CD8hi T cells, was found in the cART-HIV low-TFH group. An inverse correlation between plasma viral load and LN GrzBhiCD8hi T and CD16hiCD15lo cells was found. Our data reveal the complex GC immune landscaping in HIV infection and suggest that follicular FOXP3hiCD4hi T cells could be negative regulators of TFH cell prevalence in cART-HIV.

Distinct SIV-specific CD8+ T cells in the lymph node exhibit simultaneous effector and stem-like profiles and are associated with limited SIV persistence.

Human immunodeficiency virus (HIV) cure efforts are increasingly focused on harnessing CD8+ T cell functions, which requires a deeper understanding of CD8+ T cells promoting HIV control. Here we identifiy an antigen-responsive TOXhiTCF1+CD39+CD8+ T cell population with high expression of inhibitory receptors and low expression of canonical cytolytic molecules. Transcriptional analysis of simian immunodeficiency virus (SIV)-specific CD8+ T cells and proteomic analysis of purified CD8+ T cell subsets identified TOXhiTCF1+CD39+CD8+ T cells as intermediate effectors that retained stem-like features with a lineage relationship with terminal effector T cells. TOXhiTCF1+CD39+CD8+ T cells were found at higher frequency than TCF1-CD39+CD8+ T cells in follicular microenvironments and were preferentially located in proximity of SIV-RNA+ cells. Their frequency was associated with reduced plasma viremia and lower SIV reservoir size. Highly similar TOXhiTCF1+CD39+CD8+ T cells were detected in lymph nodes from antiretroviral therapy-naive and antiretroviral therapy-suppressed people living with HIV, suggesting this population of CD8+ T cells contributes to limiting SIV and HIV persistence.

Renocardiac Effects of p-Cresyl Sulfate Administration in Acute Kidney Injury Induced by Unilateral Ischemia and Reperfusion Injury In Vivo.

The precise mechanisms underlying the cardiovascular complications due to acute kidney injury (AKI) and the retention of uremic toxins like p-cresyl sulfate (PCS) remain incompletely understood. The objective of this study was to evaluate the renocardiac effects of PCS administration in animals subjected to AKI induced by ischemia and reperfusion (IR) injury. C57BL6 mice were subjected to distinct protocols: (i) administration with PCS (20, 40, or 60 mg/L/day) for 15 days and (ii) AKI due to unilateral IR injury associated with PCS administration for 15 days. The 20 mg/L dose of PCS led to a decrease in renal mass, an increase in the gene expression of Cystatin C and kidney injury molecule 1 (KIM-1), and a decrease in the α-actin in the heart. During AKI, PCS increased the renal injury biomarkers compared to control; however, it did not exacerbate these markers. Furthermore, PCS did not enhance the cardiac hypertrophy observed after 15 days of IR. An increase, but not potentialized, in the cardiac levels of interleukin (IL)-1ß and IL-6 in the IR group treated with PCS, as well as in the injured kidney, was also noticed. In short, PCS administration did not intensify kidney injury, inflammation, and cardiac outcomes after AKI.

SARS-CoV-2 awakens ancient retroviral genes and the expression of proinflammatory HERV-W envelope protein in COVID-19 patients.

Patients with COVID-19 may develop abnormal inflammatory response, followed in some cases by severe disease and long-lasting syndromes. We show here that in vitro exposure to SARS-CoV-2 activates the expression of the human endogenous retrovirus (HERV) HERV-W proinflammatory envelope protein (ENV) in peripheral blood mononuclear cells from a subset of healthy donors, in ACE2 receptor and infection-independent manner. Plasma and/or sera of 221 COVID-19 patients from different cohorts, infected with successive SARS-CoV-2 variants including the Omicron, had detectable HERV-W ENV, which correlated with ENV expression in T lymphocytes and peaked with the disease severity. HERV-W ENV was also found in postmortem tissues of lungs, heart, gastrointestinal tract, brain olfactory bulb, and nasal mucosa from COVID-19 patients. Altogether, these results demonstrate that SARS-CoV-2 could induce HERV-W envelope protein expression and suggest its involvement in the immunopathogenesis of certain COVID-19-associated syndromes and thereby its relevance in the development of personalized treatment of patients.

Intradural extramedullary meningeal melanocytoma: a case report and literature review.

Primary meningeal melanocytomas are extremely rare, benign tumours arising from the leptomeninges. While they are considered to be benign lesions, there is potential for their growth and transformation into malignant melanomas. They are commonly found in the cervical spine, with a decreased incidence in the thoracic and lumbar regions. We present a case report of a 56-year-old man who presented to our unit with a 4-month history of lower limb weakness and a sensory level at T6. Magnetic resonance imaging shows an intradural extramedullary tumour. The patient underwent a thoracic debulking of the lesion with neurophysiological monitoring. Histopathology confirmed the diagnosis of melanocytoma of meningeal origin, with a low mitotic count. Our patient recovered well post-operatively with no complications. Surgical resection is an effective method to manage this tumour; however, adjuvant radiotherapy is advised due to the risk of recurrence and malignant transformation.

Clinical Reasoning: Progressive Hemiparesis and White Matter Abnormalities in an HIV-Negative Patient.

A 61-year-old man from India was admitted to hospital after being found unresponsive by the roadside. He was treated with dual-antiplatelet therapy for an acute coronary syndrome. Ten days into admission, he had mild left-sided face, arm, and leg weakness, which progressed significantly over the next 2 months in association with progressive white matter abnormalities on brain MRI. In this case study, we outline our clinical reasoning, which led to the detection of a rare underlying cause of a devastating neurologic disease. We also present our approach to treatment, which achieved a sustained clinical and radiologic response.

Profound phenotypic and epigenetic heterogeneity of the HIV-1-infected CD4+ T cell reservoir.

Understanding the complexity of the long-lived HIV reservoir during antiretroviral therapy (ART) remains a considerable impediment in research towards a cure for HIV. To address this, we developed a single-cell strategy to precisely define the unperturbed peripheral blood HIV-infected memory CD4+ T cell reservoir from ART-treated people living with HIV (ART-PLWH) via the presence of integrated accessible proviral DNA in concert with epigenetic and cell surface protein profiling. We identified profound reservoir heterogeneity within and between ART-PLWH, characterized by new and known surface markers within total and individual memory CD4+ T cell subsets. We further uncovered new epigenetic profiles and transcription factor motifs enriched in HIV-infected cells that suggest infected cells with accessible provirus, irrespective of reservoir distribution, are poised for reactivation during ART treatment. Together, our findings reveal the extensive inter- and intrapersonal cellular heterogeneity of the HIV reservoir, and establish an initial multiomic atlas to develop targeted reservoir elimination strategies.

The Interplay between Uremic Toxins and Albumin, Membrane Transporters and Drug Interaction.

Uremic toxins are a heterogeneous group of molecules that accumulate in the body due to the progression of chronic kidney disease (CKD). These toxins are associated with kidney dysfunction and the development of comorbidities in patients with CKD, being only partially eliminated by dialysis therapies. Importantly, drugs used in clinical treatments may affect the levels of uremic toxins, their tissue disposition, and even their elimination through the interaction of both with proteins such as albumin and cell membrane transporters. In this context, protein-bound uremic toxins (PBUTs) are highlighted for their high affinity for albumin, the most abundant serum protein with multiple binding sites and an ability to interact with drugs. Membrane transporters mediate the cellular influx and efflux of various uremic toxins, which may also compete with drugs as substrates, and both may alter transporter activity or expression. Therefore, this review explores the interaction mechanisms between uremic toxins and albumin, as well as membrane transporters, considering their potential relationship with drugs used in clinical practice.

Multiparameter immunohistochemistry analysis of HIV DNA, RNA and immune checkpoints in lymph node tissue.

The main barrier to a cure for HIV is the persistence of long-lived and proliferating latently infected CD4+ T-cells despite antiretroviral therapy (ART). Latency is well characterized in multiple CD4+ T-cell subsets, however, the contribution of regulatory T-cells (Tregs) expressing FoxP3 as well as immune checkpoints (ICs) PD-1 and CTLA-4 as targets for productive and latent HIV infection in people living with HIV on suppressive ART is less well defined. We used multiplex detection of HIV DNA and RNA with immunohistochemistry (mIHC) on formalin-fixed paraffin embedded (FFPE) cells to simultaneously detect HIV RNA and DNA and cellular markers. HIV DNA and RNA were detected by in situ hybridization (ISH) (RNA/DNAscope) and IHC was used to detect cellular markers (CD4, PD-1, FoxP3, and CTLA-4) by incorporating the tyramide system amplification (TSA) system. We evaluated latently infected cell lines, a primary cell model of HIV latency and excisional lymph node (LN) biopsies collected from people living with HIV (PLWH) on and off ART. We clearly detected infected cells that coexpressed HIV RNA and DNA (active replication) and DNA only (latently infected cells) in combination with IHC markers in the in vitro infection model as well as LN tissue from PLWH both on and off ART. Combining ISH targeting HIV RNA and DNA with IHC provides a platform to detect and quantify HIV persistence within cells identified by multiple markers in tissue samples from PLWH on ART or to study HIV latency.

In Situ Characterization of Human Lymphoid Tissue Immune Cells by Multispectral Confocal Imaging and Quantitative Image Analysis; Implications for HIV Reservoir Characterization.

CD4 T cells are key mediators of adaptive immune responses during infection and vaccination. Within secondary lymphoid organs, helper CD4 T cells, particularly those residing in germinal centers known as follicular helper T cells (Tfh), provide critical help to B-cells to promote their survival, isotype switching and selection of high affinity memory B-cells. On the other hand, the important role of Tfh cells for the maintenance of HIV reservoir is well documented. Thus, interrogating and better understanding the tissue specific micro-environment and immune subsets that contribute to optimal Tfh cell differentiation and function is important for designing successful prevention and cure strategies. Here, we describe the development and optimization of eight multispectral confocal microscopy immunofluorescence panels designed for in depth characterization and immune-profiling of relevant immune cells in formalin-fixed paraffin-embedded human lymphoid tissue samples. We provide a comprehensive library of antibodies to use for the characterization of CD4+ T-cells -including Tfh and regulatory T-cells- as well as CD8 T-cells, B-cells, macrophages and dendritic cells and discuss how the resulting multispectral confocal datasets can be quantitatively dissected using the HistoCytometry pipeline to collect information about relative frequencies and immune cell spatial distributions. Cells harboring actively transcribed virus are analyzed using an in-situ hybridization assay for the characterization of HIV mRNA positive cells in combination with additional protein markers (multispectral RNAscope). The application of this methodology to lymphoid tissues offers a means to interrogate multiple relevant immune cell targets simultaneously at increased resolution in a reproducible manner to guide CD4 T-cell studies in infection and vaccination.

Uremic Toxins: An Alarming Danger Concerning the Cardiovascular System.

The kidneys and heart share functions with the common goal of maintaining homeostasis. When kidney injury occurs, many compounds, the so-called "uremic retention solutes" or "uremic toxins," accumulate in the circulation targeting other tissues. The accumulation of uremic toxins such as p-cresyl sulfate, indoxyl sulfate and inorganic phosphate leads to a loss of a substantial number of body functions. Although the concept of uremic toxins is dated to the 1960s, the molecular mechanisms capable of leading to renal and cardiovascular injuries are not yet known. Besides, the greatest toxic effects appear to be induced by compounds that are difficult to remove by dialysis. Considering the close relationship between renal and cardiovascular functions, an understanding of the mechanisms involved in the production, clearance and overall impact of uremic toxins is extremely relevant for the understanding of pathologies of the cardiovascular system. Thus, the present study has as main focus to present an extensive review on the impact of uremic toxins in the cardiovascular system, bringing the state of the art on the subject as well as clinical implications related to patient's therapy affected by chronic kidney disease, which represents high mortality of patients with cardiac comorbidities.

Clinical behavior of posterior fixed partial dentures with a biologically oriented preparation technique: A 5-year randomized controlled clinical trial.

STATEMENT OF PROBLEM: Evidence of the behavior of the periodontal tissues around anterior teeth restored with the biologically oriented preparation technique (BOPT) is available. However, outcomes of this technique in posterior teeth restored with fixed partial dentures (FPDs) are lacking. PURPOSE: The purpose of this randomized controlled clinical trial was to evaluate the clinical, mechanical, and biological behavior of posterior 3-unit FPDs placed on teeth prepared with BOPT. MATERIAL AND METHODS: Forty participants received a 3-unit zirconia FPD in the posterior region of the mandible or maxilla. Twenty FPDs were placed on teeth prepared with BOPT (study group) and 20 on teeth with a horizontal chamfer finishing line (control group). Follow-up examinations were performed 1, 3, and 5 years after treatment to evaluate periodontal responses around the prepared teeth by means of the following parameters: plaque index, gingival index, probing depth, and marginal stability (MS). Mechanical behavior was also assessed, as were any complications. RESULTS: After the 5-year follow-up, 57.9% of the control group and 35% of the BOPT group presented a plaque index of 1. The gingival index was 1 in 68.4% of the control group and 30% of the BOPT group after the follow-up period. In the analysis of probing depth, 26.3% of teeth in the control group had pockets of more than 3 mm in depth, whereas the BOPT group had only 10%. Marginal stability appeared in 100% of the BOPT group, whereas only 10.5% of the control group exhibited gingival stability. Complications during the follow-up period were similar, 20% in the control group and 15% in the BOPT group. CONCLUSIONS: Posterior FPDs prepared by using BOPT had a good clinical response over a 5-year follow-up, with a low gingival index, a small increase in pocket depth, and a 100% marginal stability of the surrounding tissues. High survival rates after 5 years indicated that the technique produced predictable outcomes.

Assessment of HIV-1 integration in tissues and subsets across infection stages.

The integration of HIV DNA into the host genome contributes to lifelong infection in most individuals. Few studies have examined integration in lymphoid tissue, where HIV predominantly persists before and after antiretroviral treatment (ART). Of particular interest is whether integration site distributions differ between infection stages with paired blood and tissue comparisons. Here, we profiled HIV integration site distributions in sorted memory, tissue-resident, and/or follicular helper CD4+ T cell subsets from paired blood and lymphoid tissue samples from acute, chronic, and ART-treated individuals. We observed minor differences in the frequency of nonintronic and nondistal intergenic sites, varying with tissue and residency phenotypes during ART. Genomic and epigenetic annotations were generally similar. Clonal expansion of cells marked by identical integration sites was detected, with increased detection in chronic and ART-treated individuals. However, overlap between or within CD4+ T cell subsets or tissue compartments was only observed in 8 unique sites of the 3540 sites studied. Together, these findings suggest that shared integration sites between blood and tissue may, depending on the tissue site, be the exception rather than the rule and indicate that additional studies are necessary to fully understand the heterogeneity of tissue-sequestered HIV reservoirs.

Nr2f1 heterozygous knockout mice recapitulate neurological phenotypes of Bosch-Boonstra-Schaaf optic atrophy syndrome and show impaired hippocampal synaptic plasticity.

Bosch-Boonstra-Schaaf optic atrophy syndrome (BBSOAS) has been identified as an autosomal-dominant disorder characterized by a complex neurological phenotype, with high prevalence of intellectual disability and optic nerve atrophy/hypoplasia. The syndrome is caused by loss-of-function mutations in NR2F1, which encodes a highly conserved nuclear receptor that serves as a transcriptional regulator. Previous investigations to understand the protein's role in neurodevelopment have mostly used mouse models with constitutive and tissue-specific homozygous knockout of Nr2f1. In order to represent the human disease more accurately, which is caused by heterozygous NR2F1 mutations, we investigated a heterozygous knockout mouse model and found that this model recapitulates some of the neurological phenotypes of BBSOAS, including altered learning/memory, hearing defects, neonatal hypotonia and decreased hippocampal volume. The mice showed altered fear memory, and further electrophysiological investigation in hippocampal slices revealed significantly reduced long-term potentiation and long-term depression. These results suggest that a deficit or alteration in hippocampal synaptic plasticity may contribute to the intellectual disability frequently seen in BBSOAS. RNA-sequencing (RNA-Seq) analysis revealed significant differential gene expression in the adult Nr2f1+/- hippocampus, including the up-regulation of multiple matrix metalloproteases, which are known to be critical for the development and the plasticity of the nervous system. Taken together, our studies highlight the important role of Nr2f1 in neurodevelopment. The discovery of impaired hippocampal synaptic plasticity in the heterozygous mouse model sheds light on the pathophysiology of altered memory and cognitive function in BBSOAS.

Pharmacological activation of the nuclear receptor REV-ERB reverses cognitive deficits and reduces amyloid-ß burden in a mouse model of Alzheimer's disease.

Alzheimer's disease currently lacks treatment options that effectively reverse the biological/anatomical pathology and cognitive deficits associated with the disease. Loss of function of the nuclear receptor REV-ERB is associated with reduced cognitive function in mouse models. The effect of enhanced REV-ERB activity on cognitive function has not been examined. In this study, we tested the hypothesis that enhanced REV-ERB function may enhance cognitive function in a model of Alzheimer's disease. We utilized the REV-ERB agonist SR9009 to pharmacologically activate the activity of REV-ERB in the SAMP8 mouse model of Alzheimer's disease. SR9009 reversed cognitive dysfunction of an aged SAMP8 mouse in several behavioral assays including novel object recognition, T-maze foot shock avoidance, and lever press operant conditioning task assessments. SR9009 treatment reduced amyloid-ß 1-40 and 1-42 levels in the cortex, which is consistent with improved cognitive function. Furthermore, SR9009 treatment led to increased hippocampal PSD-95, cortical synaptophysin expression and the number of synapses suggesting improvement in synaptic function. We conclude that REV-ERB is a potential target for treatment of Alzheimer's disease.

Rev-erb regulation of cholesterologenesis.

REV-ERBα and REV-ERBß are heme regulated nuclear receptors that are known to regulate metabolic pathways. We previously demonstrated that treatment of mice with synthetic REV-ERB agonists suppressed plasma cholesterol levels and the hepatic levels of the rate limiting enzyme in cholesterol biosynthesis (3-hydroxy-3-methylglutaryl-CoA reductase). Here, we characterize the role of REV-ERB on the cholesterol biosynthetic pathway in greater detail. The REV-ERB agonist SR9009 reduced plasma cholesterol levels in both wild type C57Bl/6 and low density lipoprotein receptor (LDLR) null mice as well as reducing the expression of an array of genes within the cholesterol biosynthetic pathway. Consistent with these data, we observed increased expression of these genes in mice deficient in expression of Rev-erbα. Analysis of global run-on and deep sequencing (GRO-Seq) and chromatin immunoprecipitation deep sequencing (ChIP-Seq) data revealed that Rev-erb directly binds to the majority of genes involved in cholesterol biosynthesis and directly suppresses their expression. This study reveals insight into the complex mechanism by which Rev-erb directly and indirectly (via inhibition of Srebf2 expression) regulates cholesterol biosynthesis and provides information of how cholesterol levels are regulated in a circadian fashion. Additionally, these studies suggest that targeting Rev-erb may be an effective method for suppressing LDL cholesterol levels in the clinic.

Patient-centered and clinical outcomes of mandibular overdentures retained with the locator system: A prospective observational study.

STATEMENT OF PROBLEM: Whether clinical or demographic variables affect the perception of treatment in terms of quality of life and satisfaction is unknown. PURPOSE: The purpose of this prospective study was to make an evidence-based assessment of the treatment outcomes (patient- and clinically based) of locator-retained mandibular overdentures. MATERIAL AND METHODS: This prospective observational study assessed patients with edentulism who had worn mandibular overdentures supported by 2 implants and retained by the locator system for at least 1 year of functional life (N=80). Medical histories were reviewed, and patients underwent oral examinations. Prosthetic clinical outcomes and patient well-being were registered using the Oral Health Impact Profile 20 (OHIP-20) and Oral Satisfaction Scale (OSS). RESULTS: Patient well-being scored an overall OHIP-20 score of 19.0 ±14.0 of 80 (the higher the score, the greater the impact and the worse the oral health-related quality of life); overall oral satisfaction was 8.3 ±1.7 of 10. Women suffered greater social impact (0.8 ±1.0) and disability (0.4 ±0.8) than men (0.4 ±0.7 versus 0.2 ±0.4, respectively). Impact on well-being was inversely proportional to both patient age and the age of the prosthesis (r=-0.25; P<.01). Implants had been placed on average 73.6 ±39.2 months previously, showing a survival rate of 82.5%. Most of the overdentures had been functioning for over 60 months. Relining (46.3%), readjustments (82.5%), and changes of nylon retention (1.5 ±1.8 per patient over 60 months of use) devices negatively influenced well-being. CONCLUSIONS: Mandibular overdentures produced good results with regard to quality of life and oral satisfaction, but attention should be paid to factors affecting clinical outcomes and patient well-being.