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.

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.

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.

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.

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.

Laparoscopic hepaticojejunostomy after bile duct injury.

BACKGROUND: The incidence of bile duct injuries (BDI) after cholecystectomy, which is a life-threatening condition that has several medical and legal implications, currently stands at about 0.6%. The aim of this study is to describe our experience as the first center to use a laparoscopic approach for BDI repair. METHODS: A prospective study between June 2012 and September 2014 was developed. Twenty-nine consecutive patients with BDI secondary to cholecystectomy were included. Demographics, comorbidities, presenting symptoms, details of index surgery, type of lesion, preoperative and postoperative diagnostic work-up, and therapeutic interventions were registered. Videos and details of laparoscopic hepaticojejunostomy (LHJ) were recorded. Injuries were staged using Strasberg classification. A side-to-side anastomosis with Roux-en-Y reconstruction was always used. In patients with E4 and some E3 injuries, a segment 4b or 5 section was done to build a wide anastomosis. In E4 injuries, a neo-confluence was performed. Complications, mortality, and long-term evolution were recorded. RESULTS: Twenty-nine patients with BDI were operated. Women represented 82.7% of the cases. The median age was 42 years (range 21-74). Injuries at or above the confluence occurred in 62%, and primary repair at our institution was performed at 93.1% of the cases. Eight neo-confluences were performed in all E4 injuries (27.5%). The median operative time was 240 min (range 120-585) and bleeding 200 mL (range 50-1100). Oral intake was started in the first 48 h. Bile leak occurred in 5 cases (17.2%). Two patients required re-intervention (6.8%). No mortality was recorded. The maximum follow-up was 36 months (range 2-36). One patient with E4 injury developed a hepaticojejunostomy (HJ) stenosis after 15 months. This was solved with endoscopic dilatation. CONCLUSIONS: The benefits of minimally invasive approaches in BDI seem to be feasible and safe, even when this is a complex and catastrophic scenario.

Oral health-related quality of life of implant-supported overdentures versus conventional complete prostheses: Retrospective study of a cohort of edentulous patients.

BACKGROUND: This work aims to confirm if implant-supported overdentures are a good treatment option for edentulous patients and offer an improvement in quality of life compared with traditional complete prostheses (dentures). MATERIAL AND METHODS: This retrospective clinical descriptive study included three evaluation groups: validation group (n=57); control group of patients with complete removeable prostheses (n=56); study group of patients with implant-supported overdentures retained with the Locator® system (n=80). The study also validated the Oral Health Impact Profile-20 questionnaire. Individual protocols were created that included socio-demographic data, the Oral Health Impact Profile-20 (OHIP-20) questionnaire and Oral Satisfaction Scale (OSS). Descriptive and bivariate statistical analysis was carried out applying χ², Pearson, Kruskal-Wallis, and Student t tests, transferring data into SPSS-Windows® software from a Microsoft® Excel spreadsheet. RESULTS: The OHIP-20 proved to be a valid instrument and provided reliable assessment of health-related quality of life among both the Spanish general population and edentulous patients. The control and study groups proved comparable, showing socio-demographic homogeneity. For patients with overdentures retained by means of the Locator® system, these restorations had significantly lower impact on quality of life (19 vs 33), both generally and for each individual questionnaire item, and much higher satisfaction with the state of their oral cavities (8.3 vs 5.3) than patients wearing dentures; both sets of data showed a direct linear relationship, so that as the level of impact on quality of life increased, perceived oral satisfaction worsened. CONCLUSIONS: Patients rehabilitated with implant supported overdentures retained by the Locator® system, presented significantly lower levels of impact on their quality of life and significantly higher oral satisfaction than patients with conventional complete prostheses.

Dental preparation with sonic vs high-speed finishing: analysis of microleakage in bonded veneer restorations.

PURPOSE: To compare marginal microleakage in porcelain veneer restorations following dental finishing using two types of instruments to test the hypothesis that microleakage will be less when teeth are prepared with sonic oscillating burs than when prepared with high-speed rotating burs. MATERIALS AND METHODS: Fifty-six extracted human maxillary central incisors were selected and divided randomly into two groups. Group 1 samples underwent dental finishing using high-speed rotating diamond burs, while group 2 used sonic oscillating diamond burs. Buccal chamfer preparation was carried out for both groups. Forty eight of the samples (24 per group) were restored using IPS Empress ceramic veneers. 2% methylene blue was used to evaluate microleakage at the tooth/composite veneer interface. Teeth were sectioned lengthwise into three parts and microleakage was measured at two points - cervical and incisal - on each section. Before bonding, four teeth per group underwent SEM examination. RESULTS: Evaluation of microleakage at the cervical dentin margin showed a value of 10.5% in group 1 and 6.6% in group 2, which was statistically significantly different (p < 0.05). Incisal microleakage was 1.3% for group 1 and 1.2% for group 2, which was not significantly different. SEM revealed different patterns of surface texture in both areas according to the instrument used. Group 1 exhibited parallel horizontal abrasion grooves with a milled effect and thick smear layers; group 2 showed abrasive erosion, discontinuous perpendicular depressions, and thin smear layers. CONCLUSION: Tooth preparations finished with sonic burs produced significantly less microleakage in the cervical dentin area of bonded veneer restorations. No differences were found in the incisal enamel area.

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.

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.

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.

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.

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.