Anti-cholinergic drug burden in patients with dementia increases after hospital admission: a multicentre cross-sectional study.

BACKGROUND: Anticholinergic medications are drugs that block cholinergic transmission, either as their primary therapeutic action or as a secondary effect. Patients with dementia may be particularly sensitive to the central effects of anticholinergic drugs. Anticholinergics also antagonise the effects of the main dementia treatment, cholinesterase inhibitors. Our study aimed to investigate anticholinergic prescribing for dementia patients in UK acute hospitals before and after admission. METHODS: We included 352 patients with dementia from 17 UK hospital sites in 2019. They were all inpatients on surgical, medical or Care of the Elderly wards. Information about each patient's medications were collected using a standardised form, and the anticholinergic drug burden of each patient was calculated with an evidence-based online calculator. Wilcoxon's rank test was used to look at the correlation between two subgroups upon admission and discharge. RESULTS: On admission to hospital, 37.8% of patients had an anticholinergic burden score ≥ 1 and 5.68% ≥3. On discharge, 43.2% of patients with an anticholinergic burden score ≥ 1 and 9.1% ≥3. The increase in scores was statistically significant (p = 0.001). Psychotropics were the most common group of anticholinergic medications prescribed at discharge. Of those patients taking cholinesterase inhibitors, 44.9% were also prescribed anticholinergic medications. CONCLUSIONS: Our cross-sectional, multicentre study found that people with dementia are commonly prescribed anticholinergic medications, even if concurrently taking cholinesterase inhibitors, and are significantly more likely to be discharged from hospital with a higher anticholinergic burden than on admission.

Central sympathetic innervations to visceral and subcutaneous white adipose tissue.

There is a link between visceral white adipose tissue (WAT) and the metabolic syndrome in humans, with health improvements produced with small visceral WAT reduction. By contrast, subcutaneous WAT provides a site for lipid storage that is rather innocuous relative to ectopic lipid storage in muscle or liver. The sympathetic nervous system (SNS) is the principal initiator for lipolysis in WAT by mammals. Nothing is known, however, about the central origins of the SNS circuitry innervating the only true visceral WAT in rodents, mesenteric WAT (MWAT), which drains into the hepatic portal vein. We tested whether the central sympathetic circuits to subcutaneous [inguinal WAT (IWAT)] and visceral WAT (MWAT) are separate or shared and whether they possess differential sympathetic drives with food deprivation in Siberian hamsters. Using two isogenic strains of pseudorabies virus, a retrograde transneuronal viral tract tracer within the same hamsters, we found some overlap (∼20-55% doubly infected neurons) between the two circuitries across the neural axis with lesser overlap proximal to the depots (spinal cord and sympathetic chain) and with more neurons involved in the innervation of IWAT than MWAT in some brain regions. Food deprivation triggered a greater sympathetic drive to subcutaneous (IWAT) than visceral (MWAT) depots. Collectively, we demonstrated both shared and separate populations of brain, spinal cord, and sympathetic chain neurons ultimately project to a subcutaneous WAT depot (IWAT) and the only visceral WAT depot in rodents (MWAT). In addition, the lipolytic stimulus of food deprivation only increased SNS drive to subcutaneous fat (IWAT).

Can intense predation by bears exert a depensatory effect on recruitment in a Pacific salmon population?

It has long been recognized that, as populations increase in density, ecological processes affecting growth and survival reduce per capita recruitment in the next generation. In contrast to the evidence for such "compensatory" density dependence, the alternative "depensatory" process (reduced per capita recruitment at low density) has proven more difficult to demonstrate in the field. To test for such depensation, we measured the spawner-recruit relationship over five decades for a sockeye salmon (Oncorhynchus nerka) population in Alaska breeding in high-quality, unaltered habitat. Twenty-five years of detailed estimates of predation by brown bears, Ursus arctos, revealed strong density dependence in predation rate; the bears killed ca. 80% of the salmon in years of low salmon spawning abundance. Nevertheless, the reconstructed spawner-recruit relationship, adjusted to include salmon intercepted in the commercial fishery, provided no evidence of demographic depensation. That is, in years when few salmon returned and the great majority were killed by bears, the few that spawned were successful enough that the population remained highly productive, even when those killed by bears were included as potential spawners. We conclude that the high quality of breeding habitat at this site and the productive nature of semelparous Pacific salmon allowed this population to avoid the hypothesized depressed recruitment from depensatory processes expected at low density. The observed lack of demographic depensation is encouraging from a conservation standpoint because it implies that depleted populations may have the potential to rebound successfully given suitable spawning and rearing habitat, even in the presence of strong predation pressure.

The 'omics of obesity in B-cell acute lymphoblastic leukemia.

The obesity pandemic currently affects more than 70 million Americans and more than 650 million individuals worldwide. In addition to increasing susceptibility to pathogenic infections (eg, SARS-CoV-2), obesity promotes the development of many cancer subtypes and increases mortality rates in most cases. We and others have demonstrated that, in the context of B-cell acute lymphoblastic leukemia (B-ALL), adipocytes promote multidrug chemoresistance. Furthermore, others have demonstrated that B-ALL cells exposed to the adipocyte secretome alter their metabolic states to circumvent chemotherapy-mediated cytotoxicity. To better understand how adipocytes impact the function of human B-ALL cells, we used a multi-omic RNA-sequencing (single-cell and bulk transcriptomic) and mass spectroscopy (metabolomic and proteomic) approaches to define adipocyte-induced changes in normal and malignant B cells. These analyses revealed that the adipocyte secretome directly modulates programs in human B-ALL cells associated with metabolism, protection from oxidative stress, increased survival, B-cell development, and drivers of chemoresistance. Single-cell RNA sequencing analysis of mice on low- and high-fat diets revealed that obesity suppresses an immunologically active B-cell subpopulation and that the loss of this transcriptomic signature in patients with B-ALL is associated with poor survival outcomes. Analyses of sera and plasma samples from healthy donors and those with B-ALL revealed that obesity is associated with higher circulating levels of immunoglobulin-associated proteins, which support observations in obese mice of altered immunological homeostasis. In all, our multi-omics approach increases our understanding of pathways that may promote chemoresistance in human B-ALL and highlight a novel B-cell-specific signature in patients associated with survival outcomes.

Increased natural killer cell cytotoxicity and NKp30 expression protects against hepatitis C virus infection in high-risk individuals and inhibits replication in vitro.

UNLABELLED: CD56(pos) natural killer (NK)/natural T (NT) cells are important innate effectors providing the first line of defense against viral infection. Enhanced NK activity has been shown to protect from human immunodeficiency virus-1 infection. However, the role played by these innate effectors in protection against or development of hepatitis C virus (HCV) infection is unknown. We characterized CD56(pos) populations in 11 injection drug users (IDUs) who remained uninfected despite being repeatedly exposed to HCV. NK profiles in exposed but uninfected (EU) individuals were compared with preinfection samples (median 90 days prior to HCV seroconversion) collected from 14 IDUs who were exposed and subsequently became infected (EI) and unexposed normal control subjects (n = 8). Flow cytometric analysis of CD56(pos) populations demonstrated that EUs had a higher proportion of CD56(low) mature (P = 0.0011) NK cells compared with EI subjects. Bead-isolated NKs ( > 90% purity) from EUs had significantly higher interleukin-2 (IL-2)-induced cytolytic activity against the NK-sensitive cell line K562 at an effector-to-target ratio of 10:1 (P < 0.0001). NKp30, a natural cytotoxicity receptor involved in NK activation, is highest on NK/NT cells in EUs relative to infected subjects. Using the JFH-1 infection system, we demonstrated that NKp30(high) cells in the absence of exogenous stimulation significantly reduce infection of hepatocytes. CONCLUSION: CD56(pos) populations in EUs are enriched for effector NKs displaying enhanced IL-2-induced cytolytic activity and higher levels of the natural cytotoxicity receptor NKp30-activating receptor. In addition, NKp30(high) cells are more effective in preventing infection of Huh-7.5 cells than their NKp30(low/neg) counterparts. These data support the hypothesis that NK cells contribute to anti-HCV defense in vivo in the earliest stages of infection, providing innate protection from HCV acquisition.

Selective ablation of cochlear hair cells promotes engraftment of human embryonic stem cell-derived progenitors in the mouse organ of Corti.

BACKGROUND: Hearing loss affects 25% of the population at ages 60-69 years. Loss of the hair cells of the inner ear commonly underlies deafness and once lost this cell type cannot spontaneously regenerate in higher vertebrates. As a result, there is a need for the development of regenerative strategies to replace hair cells once lost. Stem cell-based therapies are one such strategy and offer promise for cell replacement in a variety of tissues. A number of investigators have previously demonstrated successful implantation, and certain level of regeneration of hair and supporting cells in both avian and mammalian models using rodent pluripotent stem cells. However, the ability of human stem cells to engraft and generate differentiated cell types in the inner ear is not well understood. METHODS: We differentiate human pluripotent stem cells to the pre-placodal stage in vitro then transplant them into the mouse cochlea after selective and complete lesioning of the endogenous population of hair cells. RESULTS: We demonstrate that hair cell ablation prior to transplantation leads to increased engraftment in the auditory sensory epithelium, the organ of Corti, as well as differentiation of transplanted cells into hair and supporting cell immunophenotypes. CONCLUSION: We have demonstrated the feasibility of human stem cell engraftment into an ablated mouse organ of Corti.

Comparison of platforms for testing antibodies to Chlamydia trachomatis antigens in the Democratic Republic of the Congo and Togo.

Trachoma, caused by repeated ocular infection with Chlamydia trachomatis (Ct), is targeted for elimination as a public health problem. Serological testing for antibodies is promising for surveillance; determining useful thresholds will require collection of serological data from settings with different prevalence of the indicator trachomatous inflammation-follicular (TF). Dried blood spots were collected during trachoma mapping in two districts each of Togo and Democratic Republic of the Congo. Anti-Ct antibodies were detected by multiplex bead assay (MBA) and three different lateral flow assays (LFA) and seroprevalence and seroconversion rate (SCR) were determined. By most tests, the district with > 5% TF (the elimination threshold) had five-sixfold higher seroprevalence and tenfold higher SCR than districts with < 5% TF. The agreement between LFA and MBA was improved using a black latex developing reagent. These data show optimization of antibody tests against Ct to better differentiate districts above or below trachoma elimination thresholds.

Optimization of a rapid test for antibodies to the Chlamydia trachomatis antigen Pgp3.

Serological surveillance for trachoma could allow monitoring of transmission levels in areas that have achieved elimination targets. Platforms that allow testing in basic laboratories or testing of easy-to-manage samples such as dried blood spots would contribute to the feasibility of serologic testing. Blood from 506 1-12-year-olds in 2 villages in Kongwa district, Tanzania, was tested for antibodies against the antigen Pgp3. Whole blood, plasma, and dried blood spots (DBS) were tested in lab and field settings using a cassette-enclosed Pgp3 lateral flow assay (LFA-cassette) and a pared-back "dipstick" assay (LFA-dipstick). DBS were also tested with a bead-based multiplex assay (MBA). There was no significant difference in antibody positivity between the MBA and either LFA format (ranging from 42.5% to 48.4%). Interrater agreement between an expert rater and 3 different raters in field and lab settings was uniformly good, with Cohen's kappa >0.81 in all cases.

Challenges in Cell-Based Therapies for the Treatment of Hearing Loss.

Hearing loss in mammals is an irreversible process caused by degeneration of the hair cells of the inner ear. Current therapies for hearing loss include hearing aids and cochlear implants that provide substantial benefits to most patients, but also have several shortcomings. There is great interest in the development of regenerative therapies to treat deafness in the future. Cell-based therapies, based either on adult, multipotent stem, or other types of pluripotent cells, offer promise for generating differentiated cell types to replace lost or damaged hair cells of the inner ear. In this review, we focus on the methods proposed and avenues for research that seem the most promising for stem cell-based auditory sensory cell regeneration, from work collected over the past 15 years.

Mediastinitis after pediatric cardiac surgery: a 15-year experience at a single institution.

BACKGROUND: The spectrum of sternal wound infections after cardiac surgery ranges from superficial infections to a deep sternal infection known as mediastinitis. Mediastinitis is a rare but clinically relevant source of postoperative morbidity and mortality in adult and pediatric patients after cardiac surgery. METHODS: We retrospectively identified all patients diagnosed with mediastinitis after cardiac surgery from January 1987 to December 2002 (17 patients/7,616 surgeries = 0.2%). Demographic data, cardiac diagnosis, cardiac surgery, hospital length of stay, associated medical diagnosis, and surgical treatment for mediastinitis were collected. RESULTS: Fifteen pediatric patients (age < 18 years) were diagnosed with mediastinitis (mean age at diagnosis 37.5 months, range 21 days to 17 years. The median postoperative day of diagnosis was 14 days (6 to 50 days). The most common organism was Staphylococcus species (n = 9). Six patients had an associated bacteremia. The median hospital length of stay for all patients was 42.5 days (range 16 to 163 days). The hospital mortality was 1 of 15 (6%). Each patient was treated with intravenous antibiotics; sternal debridement; and rectus abdominus flap reconstruction (n = 7), pectoralis muscle flap reconstruction (n = 3), omentum reconstruction (n = 1), or primary sternal closure (n = 4). Three patients have undergone redo-sternotomy with orthotopic heart transplantation, bidirectional cavopulmonary anastomosis, and replacement of a right ventricle to pulmonary artery homograft. CONCLUSIONS: Timely diagnosis, aggressive sternal debridement, and liberal use of rotational muscle flaps can potentially minimize the morbidity and mortality in pediatric postoperative cardiac patients. Subsequent redo-sternotomy has not been problematic.

A crucial role for Kupffer cell-derived galectin-9 in regulation of T cell immunity in hepatitis C infection.

Approximately 200 million people throughout the world are infected with hepatitis C virus (HCV). One of the most striking features of HCV infection is its high propensity to establish persistence (approximately 70-80%) and progressive liver injury. Galectins are evolutionarily conserved glycan-binding proteins with diverse roles in innate and adaptive immune responses. Here, we demonstrate that galectin-9, the natural ligand for the T cell immunoglobulin domain and mucin domain protein 3 (Tim-3), circulates at very high levels in the serum and its hepatic expression (particularly on Kupffer cells) is significantly increased in patients with chronic HCV as compared to normal controls. Galectin-9 production from monocytes and macrophages is induced by IFN-gamma, which has been shown to be elevated in chronic HCV infection. In turn, galectin-9 induces pro-inflammatory cytokines in liver-derived and peripheral mononuclear cells; galectin-9 also induces anti-inflammatory cytokines from peripheral but not hepatic mononuclear cells. Galectin-9 results in expansion of CD4(+)CD25(+)FoxP3(+)CD127(low) regulatory T cells, contraction of CD4(+) effector T cells, and apoptosis of HCV-specific CTLs. In conclusion, galectin-9 production by Kupffer cells links the innate and adaptive immune response, providing a potential novel immunotherapeutic target in this common viral infection.

Wnt signaling pathway and lung disease.

The Wnt pathway plays an important role in development and in regulating adult stem cell systems. A variety of cellular processes is mediated by Wnt signaling, which includes cellular proliferation, differentiation, survival, apoptosis, and cell motility. Loss of regulation of these pathways can lead to tumorigenesis, and the Wnt pathway has been implicated in the development of several types of cancers, including colon, lung, leukemia, breast, thyroid, and prostate. The Wnt pathway has also been associated with other lung diseases such as interstitial lung disease (ILD) and asthma. Our increasing understanding of the Wnt pathway offers great hope that new molecular-based screening tests and pharmaceutical agents that selectively target this pathway will be developed to diagnose and treat these diseases in the future.

Localization of ERK/MAP kinase is regulated by the alphaherpesvirus tegument protein Us2.

Many different viruses activate the extracellular signal-regulated kinase (ERK)/mitogen-activated protein (MAP) kinase signaling pathway during infection and require ERK activation for the efficient execution of their replication programs. Despite these findings, no virus-encoded proteins have been identified that directly modulate ERK activities. In an effort to determine the function of a conserved alphaherpesvirus structural protein called Us2, we screened a yeast two-hybrid library derived from NIH 3T3 cells and identified ERK as a Us2-interacting protein. Our studies indicate that Us2 binds to ERK in virus-infected cells, mediates the incorporation of ERK into the virion, and inhibits the activation of ERK nuclear substrates. The association of Us2 with ERK leads to the sequestration of ERK at the plasma membrane and to a perinuclear vesicular compartment, thereby keeping ERK out of the nucleus. Us2 can bind to activated ERK, and the data suggest that Us2 does not inhibit ERK enzymatic activity. The treatment of cells with U0126, a specific inhibitor of ERK activation, resulted in a substantial delay in the release of virus from infected cells that was more pronounced with a virus deleted for Us2 than with parental and repaired strains, suggesting that both ERK and Us2 activities are required for efficient virus replication. This study highlights an additional complexity to the activation of ERK by viruses, namely, that localization of active ERK can be altered by virus-encoded proteins.

Development of pseudorabies virus strains expressing red fluorescent proteins: new tools for multisynaptic labeling applications.

The transsynaptic retrograde transport of the pseudorabies virus Bartha (PRV-Bartha) strain has become an important neuroanatomical tract-tracing technique. Recently, dual viral transneuronal labeling has been introduced by employing recombinant strains of PRV-Bartha engineered to express different reporter proteins. Dual viral transsynaptic tracing has the potential of becoming an extremely powerful method for defining connections of single neurons to multiple neural circuits in the brain. However, the present use of recombinant strains of PRV expressing different reporters that are driven by different promoters, inserted in different regions of the viral genome, and detected by different methods limits the potential of these recombinant virus strains as useful reagents. We previously constructed and characterized PRV152, a PRV-Bartha derivative that expresses the enhanced green fluorescent protein. The development of a strain isogenic to PRV152 and differing only in the fluorescent reporter would have great utility for dual transsynaptic tracing. In this report, we describe the construction, characterization, and application of strain PRV614, a PRV-Bartha derivative expressing a novel monomeric red fluorescent protein, mRFP1. In contrast to viruses expressing DsRed and DsRed2, PRV614 displayed robust fluorescence both in cell culture and in vivo following transsynaptic transport through autonomic circuits afferent to the eye. Transneuronal retrograde dual PRV labeling has the potential to be a powerful addition to the neuroanatomical tools for investigation of neuronal circuits; the use of strain PRV614 in combination with strain PRV152 will eliminate many of the pitfalls associated with the presently used pairs of PRV recombinants.

The pseudorabies virus serine/threonine kinase Us3 contains mitochondrial, nuclear and membrane localization signals.

The serine/threonine kinase encoded by the Us3 gene is conserved amongst all known alphaherpesviruses. Us3 has been reported to function in a variety of aspects of the virus lifecycle including protection of cells from virus-induced apoptosis, de-envelopment of enveloped virus particles from the perinuclear space and cell-to-cell spread of virus infection. In this report, we examined the sub-cellular localization of the pseudorabies virus (PRV) Us3 homolog. The PRV Us3 gene encodes two proteins termed Us3a and Us3b. Us3a differs from Us3b in that it contains 54 additional N-terminal amino acids. In transfected cells, Us3a localized predominantly to the plasma membrane whereas the Us3b protein localized predominantly to the nucleus. To explore the differences in the localization of the Us3a and Us3b proteins, we fused the amino-terminal 54 amino acids of Us3a to the amino-terminus of the enhanced green fluorescent protein (EGFP). Surprisingly, this fusion protein localized exclusively to mitochondria in transfected cells. Analysis of mutated Us3-EGFP fusion proteins in transfected cells revealed that the carboxy-terminal 101 amino acids of Us3a and Us3b comprises a membrane/vesicular localization domain, and that the N-terminal 102 amino acids of Us3b comprises a nuclear localization domain. We provide a model to rationalize the complex localization of Us3a and Us3b in transfected cells and hypothesize that the mitochondrial, nuclear and membrane localization motifs function in the reported anti-apoptotic, egress and cell-to-cell spread functions of Us3.

The pseudorabies virus Us2 protein, a virion tegument component, is prenylated in infected cells.

The Us2 gene is conserved among alphaherpesviruses, but its function is not known. We demonstrate here that the pseudorabies virus (PRV) Us2 protein is synthesized early after infection and localizes to cytoplasmic vesicles and to the plasma membrane, despite the lack of a recognizable signal sequence or membrane-spanning domain. Us2 protein is also packaged as part of the tegument of mature virions. The Us2 carboxy-terminal four amino acids comprise a CAAX motif, a well-characterized signal for protein prenylation. Treatment of infected cells with lovastatin, a drug that disrupts protein prenylation, changed the relative electrophoretic mobility of Us2 in sodium dodecyl sulfate-polyacrylamide gels. In addition, lovastatin treatment caused a dramatic relocalization of Us2 to cytoplasmic punctate structures associated with microtubules, which appeared to concentrate over the microtubule organizing center. When the CAAX motif was changed to GAAX and the mutant protein was synthesized from an expression plasmid, it concentrated in punctate cytoplasmic structures reminiscent of Us2 localization in infected cells treated with lovastatin. We suggest that prenylation of PRV Us2 protein is required for proper membrane association. Curiously, the Us2 protein isolated from purified virions does not appear to be prenylated. This is the first report to describe the prenylation of an alphaherpesvirus protein.