Mechanisms of COVID-19-associated olfactory dysfunction.

Olfactory dysfunction is one of the most common symptoms of COVID-19. In the first 2 years of the pandemic, it was frequently reported, although its incidence has significantly decreased with the emergence of the Omicron variant, which has since become the dominant viral strain. Nevertheless, many patients continue to suffer from persistent dysosmia and dysgeusia, making COVID-19-associated olfactory dysfunction an ongoing health concern. The proposed pathogenic mechanisms of COVID-19-associated olfactory dysfunction are complex and likely multifactorial. While evidence suggests that infection of sustentacular cells and associated mucosal inflammation may be the culprit of acute, transient smell loss, alterations in other components of the olfactory system (e.g., olfactory receptor neuron dysfunction, olfactory bulb injury and alterations in the olfactory cortex) may lead to persistent, long-term olfactory dysfunction. This review aims to provide a comprehensive summary of the epidemiology, clinical manifestations and current understanding of the pathogenic mechanisms of COVID-19-associated olfactory dysfunction.

Radiologic-pathologic autopsy correlation of an internal watershed infarct, a case report.

Internal watershed infarcts (IWIs) occur at the junction of the deep and superficial perforating arterial branches of the cerebrum. Despite documentation in the radiology literature, IWIs are rarely encountered at the time of autopsy. Here, we report the case of a 59-year-old incarcerated male who was brought to the emergency department after being found unresponsive on the floor of his jail cell. Initial examination and imaging demonstrated right-sided hemiplegia, aphasia, right facial droop, and severe stenosis of the left middle cerebral artery, respectively. Repeat imaging 4 days after admission and 26 days before death demonstrated advanced stenosis of the intracranial, communicating segment of the right internal carotid artery, a large acute infarct in the right posterior cerebral artery territory, and bilateral deep white matter ischemic changes with a right-sided "rosary-like" pattern of injury that is typical of IWIs. Postmortem gross examination showed that the right deep white matter lesion had progressed to a confluent, "cigar-shaped" subacute IWI involving the right corona radiata. This is the first well-documented case of an IWI with radiologic imaging and photographic gross pathology correlation. This case uniquely highlights a rarely encountered lesion at the time of autopsy and provides an excellent visual representation of internal watershed neuroanatomy.

Neuropathological findings in COVID-19 vs. non-COVID-19 acute respiratory distress syndrome-A case-control study.

Acute brain injury (ABI) and neuroinflammation is reported in COVID-19 and acute respiratory distress syndrome (ARDS). It remains unclear if COVID-19 plays an independent role in development of ABI compared to those with non-COVID-19 ARDS. We aimed to evaluate if COVID-19 ARDS is associated with higher risk and specific patterns of ABI compared to non-COVID-19 ARDS. We conducted an age and sex matched case-control autopsy study at a tertiary academic center. Ten patients with COVID-19 ARDS were matched to 20 non-COVID-19 ARDS patients. Baseline demographics were comparable between the two groups including severity of ARDS (p = 0.3). The frequency of overall ABI (70 vs. 60%), infratentorial ABI (40 vs. 25%), ischemic infarct (40 vs. 25%), intracranial hemorrhage (30 vs. 35%), and hypoxic-ischemic brain injury (30 vs. 35%) was similar between COVID-19 and non-COVID-19 ARDS patients, respectively (p > 0.05). Intracapillary megakaryocytes were exclusively seen in 30% of COVID-19 patients. Overall, frequency and pattern of ABI in COVID-19 ARDS was comparable to non-COVID-19.

Radiologic-pathologic autopsy correlation of an internal watershed infarct, a case report

ABSTRACT Internal watershed infarcts (IWIs) occur at the junction of the deep and superficial perforating arterial branches of the cerebrum. Despite documentation in the radiology literature, IWIs are rarely encountered at the time of autopsy. Here, we report the case of a 59-year-old incarcerated male who was brought to the emergency department after being found unresponsive on the floor of his jail cell. Initial examination and imaging demonstrated right-sided hemiplegia, aphasia, right facial droop, and severe stenosis of the left middle cerebral artery, respectively. Repeat imaging 4 days after admission and 26 days before death demonstrated advanced stenosis of the intracranial, communicating segment of the right internal carotid artery, a large acute infarct in the right posterior cerebral artery territory, and bilateral deep white matter ischemic changes with a right-sided "rosary-like" pattern of injury that is typical of IWIs. Postmortem gross examination showed that the right deep white matter lesion had progressed to a confluent, "cigar-shaped" subacute IWI involving the right corona radiata. This is the first well-documented case of an IWI with radiologic imaging and photographic gross pathology correlation. This case uniquely highlights a rarely encountered lesion at the time of autopsy and provides an excellent visual representation of internal watershed neuroanatomy.

Acute appendicitis severity during the early COVID-19 pandemic period.

BACKGROUND: The early COVID-19 pandemic period significantly strained the US healthcare system. During this period, consultations and admissions for acute medical conditions decreased, which was associated with an increase in disease-specific morbidity and mortality. Therefore, we sought to determine what, if any, effect the early COVID-19 pandemic period had on the presentation, management, and histopathologic severity of acute appendicitis. METHODS: We performed a retrospective, observational study to compare the frequencies with which patients presented with acute appendicitis, the proportion of whom were managed surgically, and the distribution of histopathologic disease severity among all resected appendix specimens during the early COVID-19 pandemic period (March 6-June 30, 2020) to equivalent time periods for the 3 preceding/pre-pandemic years (2017-2019). RESULTS: Compared with equivalent pre-pandemic time periods, during the COVID-19 pandemic period there was no significant difference in the number of patients who presented for acute appendicitis, there was a decreased rate of surgical management (81% vs 94%; p=0.014), and there was an overall increase in the incidence of perforated appendicitis (31% vs 16%; p=0.004), including by histopathologic diagnosis (25% vs 11%; p=0.01). DISCUSSION: Despite potential patient hesitancy to present for care, the early COVID-19 pandemic period was associated with no significant change in the number of patients presenting with acute appendicitis; however, there was a significant increase in the incidence of perforated appendicitis. This study highlights the need to encourage patients to avoid late presentation for acute surgical conditions and for the robust planning for the medical management of otherwise surgical abnormalities during episodes of restricted or limited resources. LEVEL OF EVIDENCE: Level III.

Hematopoietic Stem and Progenitor Cells Are a Distinct HIV Reservoir that Contributes to Persistent Viremia in Suppressed Patients.

Long-lived reservoirs of persistent HIV are a major barrier to a cure. CD4+ hematopoietic stem and progenitor cells (HSPCs) have the capacity for lifelong survival, self-renewal, and the generation of daughter cells. Recent evidence shows that they are also susceptible to HIV infection in vitro and in vivo. Whether HSPCs harbor infectious virus or contribute to plasma virus (PV) is unknown. Here, we provide strong evidence that clusters of identical proviruses from HSPCs and their likely progeny often match residual PV. A higher proportion of these sequences match residual PV than proviral genomes from bone marrow and peripheral blood mononuclear cells that are observed only once. Furthermore, an analysis of near-full-length genomes isolated from HSPCs provides evidence that HSPCs harbor functional HIV proviral genomes that often match residual PV. These results support the conclusion that HIV-infected HSPCs form a distinct and functionally significant reservoir of persistent HIV in infected people.

Class 1-Selective Histone Deacetylase (HDAC) Inhibitors Enhance HIV Latency Reversal while Preserving the Activity of HDAC Isoforms Necessary for Maximal HIV Gene Expression.

Combinations of drugs that affect distinct mechanisms of HIV latency aim to induce robust latency reversal leading to cytopathicity and elimination of the persistent HIV reservoir. Thus far, attempts have focused on combinations of protein kinase C (PKC) agonists and pan-histone deacetylase inhibitors (HDIs) despite the knowledge that HIV gene expression is regulated by class 1 histone deacetylases. We hypothesized that class 1-selective HDIs would promote more robust HIV latency reversal in combination with a PKC agonist than pan-HDIs because they preserve the activity of proviral factors regulated by non-class 1 histone deacetylases. Here, we show that class 1-selective agents used alone or with the PKC agonist bryostatin-1 induced more HIV protein expression per infected cell. In addition, the combination of entinostat and bryostatin-1 induced viral outgrowth, whereas bryostatin-1 combinations with pan-HDIs did not. When class 1-selective HDIs were used in combination with pan-HDIs, the amount of viral protein expression and virus outgrowth resembled that of pan-HDIs alone, suggesting that pan-HDIs inhibit robust gene expression induced by class 1-selective HDIs. Consistent with this, pan-HDI-containing combinations reduced the activity of NF-κB and Hsp90, two cellular factors necessary for potent HIV protein expression, but did not significantly reduce overall cell viability. An assessment of viral clearance from in vitro cultures indicated that maximal protein expression induced by class 1-selective HDI treatment was crucial for reservoir clearance. These findings elucidate the limitations of current approaches and provide a path toward more effective strategies to eliminate the HIV reservoir.IMPORTANCE Despite effective antiretroviral therapy, HIV evades eradication in a latent form that is not affected by currently available drug regimens. Pharmacologic latency reversal that leads to death of cellular reservoirs has been proposed as a strategy for reservoir elimination. Because histone deacetylases (HDACs) promote HIV latency, HDAC inhibitors have been a focus of HIV cure research. However, many of these inhibitors broadly affect multiple classes of HDACs, including those that promote HIV gene expression (class 1 HDACs). Here, we demonstrate that targeted treatment with class 1-selective HDAC inhibitors induced more potent HIV latency reversal than broadly acting agents. Additionally, we provide evidence that broadly acting HDIs are limited by inhibitory effects on non-class 1 HDACs that support the activity of proviral factors. Thus, our work demonstrates that the use of targeted approaches to induce maximum latency reversal affords the greatest likelihood of reservoir elimination.

Quiescence Promotes Latent HIV Infection and Resistance to Reactivation from Latency with Histone Deacetylase Inhibitors.

Human immunodeficiency virus type 1 (HIV-1) establishes transcriptionally silent latent infections in resting memory T cells and hematopoietic stem and progenitor cells (HSPCs), which allows the virus to persist in infected individuals despite antiretroviral therapy. Developing in vitro models of HIV-1 latency that recapitulate the characteristics of latently infected cells in vivo is crucial to identifying and developing effective latency-reversing therapies. HSPCs exist in a quiescent state in vivo, and quiescence is correlated with latent infections in T cells. However, current models for culturing HSPCs and for infecting T cells in vitro require that the cells be maintained in an actively proliferating state. Here we describe a novel culture system in which primary human HSPCs cultured under hypothermic conditions are maintained in a quiescent state. We show that these quiescent HSPCs are susceptible to predominantly latent infection with HIV-1, while actively proliferating and differentiating HSPCs obtain predominantly active infections. Furthermore, we demonstrate that the most primitive quiescent HSPCs are more resistant to spontaneous reactivation from latency than more differentiated HSPCs and that quiescent HSPCs are resistant to reactivation by histone deacetylase inhibitors or P-TEFb activation but are susceptible to reactivation by protein kinase C (PKC) agonists. We also demonstrate that inhibition of HSP90, a known regulator of HIV transcription, recapitulates the quiescence and latency phenotypes of hypothermia, suggesting that hypothermia and HSP90 inhibition may regulate these processes by similar mechanisms. In summary, these studies describe a novel model for studying HIV-1 latency in human primary cells maintained in a quiescent state.IMPORTANCE Human immunodeficiency virus type 1 (HIV-1) establishes a persistent infection for which there remains no feasible cure. Current approaches are unable to clear the virus despite decades of therapy due to the existence of latent reservoirs of integrated HIV-1, which can reactivate and contribute to viral rebound following treatment interruption. Previous clinical attempts to reactivate the latent reservoirs in an individual so that they can be eliminated by the immune response or viral cytopathic effect have failed, indicating the need for a better understanding of the processes regulating HIV-1 latency. Here we characterize a novel in vitro model of HIV-1 latency in primary hematopoietic stem and progenitor cells isolated from human cord blood that may better recapitulate the behavior of latently infected cells in vivo This model can be used to study mechanisms regulating latency and potential therapeutic approaches to reactivate latent infections in quiescent cells.

CD4 is expressed on a heterogeneous subset of hematopoietic progenitors, which persistently harbor CXCR4 and CCR5-tropic HIV proviral genomes in vivo.

Latent HIV infection of long-lived cells is a barrier to viral clearance. Hematopoietic stem and progenitor cells are a heterogeneous population of cells, some of which are long-lived. CXCR4-tropic HIVs infect a broad range of HSPC subtypes, including hematopoietic stem cells, which are multi-potent and long-lived. However, CCR5-tropic HIV infection is limited to more differentiated progenitor cells with life spans that are less well understood. Consistent with emerging data that restricted progenitor cells can be long-lived, we detected persistent HIV in restricted HSPC populations from optimally treated people. Further, genotypic and phenotypic analysis of amplified env alleles from donor samples indicated that both CXCR4- and CCR5-tropic viruses persisted in HSPCs. RNA profiling confirmed expression of HIV receptor RNA in a pattern that was consistent with in vitro and in vivo results. In addition, we characterized a CD4high HSPC sub-population that was preferentially targeted by a variety of CXCR4- and CCR5-tropic HIVs in vitro. Finally, we present strong evidence that HIV proviral genomes of both tropisms can be transmitted to CD4-negative daughter cells of multiple lineages in vivo. In some cases, the transmitted proviral genomes contained signature deletions that inactivated the virus, eliminating the possibility that coincidental infection explains the results. These data support a model in which both stem and non-stem cell progenitors serve as persistent reservoirs for CXCR4- and CCR5-tropic HIV proviral genomes that can be passed to daughter cells.

Long-lived reservoirs of HIV-1.

HIV-1 persistence in long-lived cellular reservoirs remains a major barrier to a cure. In a recent Nature Medicine paper, Buzon et al. identify memory T cells with stem cell-like properties (TSCM) that harbor infectious provirus and that likely contribute to HIV-1 persistence.

Targeting IκB proteins for HIV latency activation: the role of individual IκB and NF-κB proteins.

Latently infected cell reservoirs represent the main barrier to HIV eradication. Combination antiretroviral therapy (cART) effectively blocks viral replication but cannot purge latent provirus. One approach to HIV eradication could include cART to block new infections plus an agent to activate latent provirus. NF-κB activation induces HIV expression, ending latency. Before activation, IκB proteins sequester NF-κB dimers in the cytoplasm. Three canonical IκBs, IκBα, IκBß, and IκBε, exist, but the IκB proteins' role in HIV activation regulation is not fully understood. We studied the effects on HIV activation of targeting IκBs by single and pairwise small interfering RNA (siRNA) knockdown. After determining the relative abundance of the IκBs, the relative abundance of NF-κB subunits held by the IκBs, and the kinetics of IκB degradation and resynthesis following knockdown, we studied HIV activation by IκB knockdown, in comparison with those of known HIV activators, tumor necrosis factor alpha (TNF-α), tetradecanoyl phorbol acetate (TPA), and trichostatin A (TSA), in U1 monocytic and J-Lat 10.6 lymphocytic latently infected cells. We found that IκBα knockdown activated HIV in both U1 and J-Lat 10.6 cells, IκBß knockdown did not activate HIV, and, surprisingly, IκBε knockdown produced the most HIV activation, comparable to TSA activation. Our data show that HIV reactivation can be triggered by targeting two different IκB proteins and that IκBε may be an effective target for HIV latency reactivation in T-cell and macrophage lineages. IκBε knockdown may offer attractive therapeutic advantages for HIV activation because it is not essential for mammalian growth and development and because new siRNA delivery strategies may target siRNAs to HIV latently infected cells.