Mechanistic Insight into the Thermal "Blueing" of Cyanine Dyes.

In recent work to develop cyanine dyes with especially large Stokes shifts, we encountered a "blueing" reaction, in which the heptamethine cyanine dye Cy7 (IUPAC: 1,3,3-trimethyl-2-((1E,3E,5E)-7-((E)-1,3,3-trimethylindolin-2-ylidene)hepta-1,3,5-trien-1-yl)-3H-indol-1-ium) undergoes shortening in two-carbon steps to form the pentamethine (Cy5) and trimethine (Cy3) analogs. Each step blue-shifts the resulting absorbance wavelength by ca. 100 nm. Though photochemical and oxidative chain-shortening reactions had been noted previously, it is simple heating alone or with amine bases that effects this unexpected net C2H2 excision. Explicit acetylene loss would be too endothermic to merit consideration. Our mechanistic studies using 2H labeling, mass spectrometric and NMR spectroscopic analyses, and quantum chemical modeling point instead to electrocyclic closure and aromatization of the heptamethine chain in Cy7 forming Fischer's base FB (1,3,3-trimethyl-2-methyleneindoline), a reactive carbon nucleophile that initiates chain shortening of the cyanine dyes by attack on their polymethine backbones. The byproduct is the cationic indolium species TMP (IUPAC: 1,3,3 trimethyl-2-phenyl indolium).

Assessing Discharge Communication and Follow-up of Acute Kidney Injury in British Columbia: A Retrospective Chart Review.

Background and objective: Acute kidney injury (AKI) affects up to 20% of hospitalizations and is associated with chronic kidney disease, cardiovascular disease, increased mortality, and increased health care costs. Proper documentation of AKI in discharge summaries is critical for optimal monitoring and treatment of these patients once discharged. Currently, there is limited literature evaluating the quality of discharge communication after AKI. This study aimed to evaluate the accuracy and quality of documentation of episodes of AKI at a tertiary care center in British Columbia, Canada. Methods design setting patients and measurements: This was a retrospective chart review study of adult patients who experienced AKI during hospital admission between January 1, 2018, and December 31, 2018. Laboratory data were used to identify all admissions to the cardiac and general medicine ward complicated by AKI defined by the Kidney Disease Improving Global Outcomes (KDIGO) criteria. A random sample of 300 AKI admissions stratified by AKI severity (eg, stages 1, 2, and 3) were identified for chart review. Patients were excluded if they required ongoing renal replacement therapy after admission, had a history of kidney transplant, died during their admission, or did not have a discharge summary available. Discharge summaries were reviewed for documentation of the following: presence of AKI, severity of AKI, AKI status at discharge, practitioner and laboratory follow-up plans, and medication changes. Results: A total of 1076 patients with 1237 AKI admissions were identified. Of the 300 patients selected for discharge summary review, 38 met exclusion criteria. In addition, AKI was documented in 140 (53%) discharge summaries and was more likely to be documented in more severe AKI: stage 1, 38%; stage 2, 51%; and stage 3, 75%. Of those with their AKI documented, 94 (67%) documented AKI severity, and 116 (83%) mentioned the AKI status or trajectory at the time of discharge. A total of 239 (91%) of discharge summaries mentioned a follow-up plan with a practitioner, but only 23 (10%) had documented follow-up with nephrology. Patients with their AKI documented were more likely to have nephrology follow-up than those without AKI documented (17% vs 1%). Regarding laboratory investigations, 92 (35%) of the summaries had documented recommendations. In summaries that included medications typically held during AKI, only about half made specific reference to those medications being held, adjusted, or documented a post-discharge plan for that medication. For those with nonsteroidal anti-inflammatory drugs (NSAIDs) listing, 64% of discharge summaries mentioned holding, and 9% mentioned a discharge plan. For those with angiotensin converting enzyme inhibitor (ACEi)/angiotensin II receptor blocker (ARB) listing, 38% mentioned holding these medications, and 46% mentioned a discharge plan. In summaries with diuretics listed, 35% mentioned holding, and 51% included a discharge plan. Conclusions and limitations: We found suboptimal quality and completeness of discharge reporting in patients hospitalized with AKI. This may contribute to inadequate follow-up and post-hospitalization care for this patient population. Strategies are required for increasing the presence and quality of AKI reporting in discharge summaries. Limitations include our definition of AKI based on lab criteria, which may have missed some of the injuries that met the criteria based on urine output. Another limitation is that our definition of AKI based on the highest and lowest creatinine during admission may have led to some overclassification. In addition, without outpatient laboratories, it is possible that we have not captured the true baseline creatinine in some patients.

Contexte et objectif: L'insuffisance rénale aiguë (IRA) complique jusqu'à 20 % des hospitalisations; elle est associée à l'insuffisance rénale chronique, aux maladies cardiovasculaires, à une mortalité accrue et à une augmentation des coûts de santé. La documentation appropriée de l'IRA dans les résumés de départ est essentielle pour optimiser la surveillance et le traitement des patients après leur sortie de l'hôpital. Il existe peu de littérature évaluant la qualité de la documentation de l'IRA dans les résumés de départ. Cette étude visait à évaluer l'exactitude et la qualité de la documentation des épisodes d'IRA dans un center de soins tertiaires de la Colombie-Britannique (Canada). Méthodologie conception et cadre de l'étude sujets et mesures: Il s'agit d'une étude rétrospective des dossiers de patients adultes ayant présenté une IRA au cours de leur admission à l'hôpital entre le 1er janvier 2018 et le 31 décembre 2018. Les données de laboratoire ont été utilisées pour répertorier toutes les admissions compliquées par une IRA (définie par les critères KDIGO) dans les services de cardiologie et de médecine générale. Un échantillon aléatoire de 300 admissions avec IRA stratifiée selon sa gravité (p. ex., stade, 1, 2 et 3) a été constitué pour l'examen des dossiers. Ont été exclus les patients qui avaient eu besoin d'une thérapie de suppléance rénale continue après leur admission, ceux qui avaient des antécédents de transplantation rénale, ceux qui étaient décédés pendant leur admission et ceux pour qui aucun résumé de départ n'était disponible. Les résumés de départ ont été examinés à la recherche d'une mention des éléments suivants : présence d'une IRA, gravité de l'IRA, statut de l'IRA à la sortie, plans de suivi pour les tests de laboratoire et suivi avec un praticien, changements dans la médication. Résultats: En tout, 1 076 patients avec un total de 1 237 admissions avec IRA ont été identifiés. Parmi les 300 patients sélectionnés pour l'examen du résumé de départ, 38 répondaient aux critères d'exclusion. L'IRA avait été documentée dans 140 (53 %) des cas et plus elle était grave, plus elle était susceptible d'être documentée (stade 1 = 38 %; stade 2 = 51 %; stade 3 = 75 %). Parmi ceux où l'IRA était documentée, 94 (67 %) mentionnaient sa gravité et 116 (83 %) mentionnaient son statut ou sa trajectoire à la sortie du patient. Un plan de suivi avec le praticien était mentionné dans 239 (91 %) des résumés de départ, mais seuls 23 (10 %) mentionnaient un suivi en néphrologie. Les patients dont l'IRA était documentée étaient plus susceptibles de faire l'objet d'un suivi en néphrologie que ceux sans mention de l'IRA (17 % contre 1 %). En ce qui concerne les plans de suivi de laboratoire, 92 (35 %) des résumés contenaient des recommandations. Dans les résumés qui mentionnaient des médicaments normalement maintenus pendant un épisode d'IRA, seule la moitié environ faisait spécifiquement référence à ces médicaments comme ayant été cessés, ajustés ou documentés dans un plan post-sortie. Dans les résumés de départ qui listaient des AINS, 64 % mentionnaient qu'ils avaient été cessés temporairement et 9 % comprenaient un plan au congé de l'hôpital. Dans les résumés de départ qui listaient des IECA/ARA, 38 % mentionnaient que ces médicaments avaient été cessés temporairement et 46 % comprenaient un plan au congé de l'hôpital. Dans les résumés qui listaient des diurétiques, 35 % mentionnaient qu'ils avaient été cessés temporairement et 51 % comprenaient un plan au congé de l'hôpital. Limites et conclusion: Nous avons constaté que la qualité et l'exhaustivité des résumés de départ étaient sous-optimales chez les patients hospitalisés ayant vécu un épisode d'IRA. Cette situation peut contribuer à l'inadéquation du suivi et des soins post-hospitalization pour cette population de patients. Des stratégies sont nécessaires pour accroître la documentation d'un épisode d'IRA dans les résumés de départ et augmenter la qualité de sa communication. Les résultats de cette étude sont notamment limités par notre définition de l'IRA fondée sur des critères de laboratoire qui pourraient avoir manqué des patients répondant aux critères fondés sur la production d'urine. Notre définition de l'IRA fondée sur le taux de créatinine le plus élevée et le plus faible pendant l'admission pourrait également avoir conduit à un surdiagnostic. En outre, sans les résultats de laboratoires externes, il est possible que nous n'ayons pas saisi la mesure initiale réelle de la créatinine chez certains patients.

Effects of COVID-19 mRNA vaccination on HIV viremia and reservoir size.

OBJECTIVE: The immunogenic nature of coronavirus disease 2019 (COVID-19) mRNA vaccines led to some initial concern that these could stimulate the HIV reservoir. We analyzed changes in plasma HIV loads (pVL) and reservoir size following COVID-19 mRNA vaccination in 62 people with HIV (PWH) receiving antiretroviral therapy (ART), and analyzed province-wide trends in pVL before and after the mass vaccination campaign. DESIGN: Longitudinal observational cohort and province-wide analysis. METHODS: Sixty-two participants were sampled prevaccination, and one month after their first and second COVID-19 immunizations. Vaccine-induced anti-SARS-CoV-2-Spike antibodies in serum were measured using the Roche Elecsys Anti-S assay. HIV reservoirs were quantified using the intact proviral DNA assay; pVL were measured using the cobas 6800 (lower limit of quantification: 20 copies/ml). The province-wide analysis included all 290 401 pVL performed in British Columbia, Canada between 2012 and 2022. RESULTS: Prevaccination, the median intact reservoir size was 77 [interquartile range (IQR): 20-204] HIV copies/million CD4 + T-cells, compared to 74 (IQR: 27-212) and 65 (IQR: 22-174) postfirst and -second dose, respectively (all comparisons P > 0.07). Prevaccination, 82% of participants had pVL <20 copies/ml (max: 110 copies/ml), compared to 79% postfirst dose (max: 183 copies/ml) and 85% postsecond dose (max: 79 copies/ml) ( P  > 0.4). There was no evidence that the magnitude of the vaccine-elicited anti-SARS-CoV-2-Spike immune response influenced pVL nor changes in reservoir size ( P  > 0.6). We found no evidence linking the COVID-19 mass vaccination campaign to population-level increases in detectable pVL frequency among all PWH in the province, nor among those who maintained pVL suppression on ART. CONCLUSION: We found no evidence that COVID-19 mRNA vaccines induced changes in HIV reservoir size nor plasma viremia.

Pediatric reference intervals for serum neurofilament light and glial fibrillary acidic protein using the Canadian Laboratory Initiative on Pediatric Reference Intervals (CALIPER) cohort.

OBJECTIVES: Blood biomarkers have the potential to transform diagnosis and prognosis for multiple neurological indications. Establishing normative data is a critical benchmark in the analytical validation process. Normative data are important in children as little is known about how brain development may impact potential biomarkers. The objective of this study is to generate pediatric reference intervals (RIs) for serum neurofilament light (NfL), an axonal marker, and glial fibrillary acidic protein (GFAP), an astrocytic marker. METHODS: Serum from healthy children and adolescents aged 1 to <19 years were obtained from the Canadian Laboratory Initiative on Pediatric Reference Intervals (CALIPER) cohort. Serum NfL (n=300) and GFAP (n=316) were quantified using Simoa technology, and discrete RI (2.5th and 97.5th percentiles) and continuous RI (5th and 95th percentiles) were generated. RESULTS: While there was no association with sex, there was a statistically significant (p<0.0001) negative association between age and serum NfL (Rho -0.400) and GFAP (Rho -0.749). Two statistically significant age partitions were generated for NfL: age 1 to <10 years (lower, upper limit; 3.13, 20.6 pg/mL) and 10 to <19 years (1.82, 7.44 pg/mL). For GFAP, three statistically significant age partitions were generated: age 1 to <3.5 years (80.4, 601 pg/mL); 3.5 to <11 years (50.7, 224 pg/mL); and 11 to <19 years (26.2, 119 pg/mL). CONCLUSIONS: Taken together with the literature on adults, NfL and GFAP display U-shaped curves with high levels in infants, decreasing levels during childhood, a plateau during adolescence and early adulthood and increasing levels in seniors. These normative data are expected to inform future pediatric studies on the importance of age on neurological blood biomarkers.

Association of admission serum sodium and outcomes following out-of-hospital cardiac arrest.

BACKGROUND: The prognostic association between dysnatremia and outcomes in out-of-hospital cardiac arrest (OHCA) is not well understood. Given hypernatremia is associated with poor outcomes in critical illness and hyponatremia may exacerbate cerebral edema, we hypothesized that dysnatremia on OHCA hospital admission would be associated with worse neurological outcomes. METHODS: We studied adults (≥19 years) with non-traumatic OHCA between 2009 and 2016 who were enrolled in the British Columbia Cardiac Arrest Registry and survived to hospital admission at 2 quaternary urban hospitals. We stratified cases by admission serum sodium into hyponatremic (<135 mmol/L), normonatremic (135-145 mmol/L), and hypernatremic (>145 mmol/L) groups. We used logistic regression models, adjusted for age, sex, shockable rhythm, admission serum lactate, and witnessed arrest, to estimate the association between admission sodium and favorable neurological outcome (cerebral performance category 1-2 or modified Rankin scale 0-3). RESULTS: Of 414 included patients, 63 were hyponatremic, 330 normonatremic, and 21 hypernatremic. In each respective group, 21 (33.3%), 159 (48.2%), and 3 (14.3%) experienced good neurological outcomes. In univariable models, hyponatremia (OR 0.53, 95% CI 0.30-0.93) and hypernatremia (OR 0.19, 95% CI 0.05-0.65) were associated with lower odds of good neurological outcomes compared to the normonatremia group. After adjustment, only hypernatremia was associated with lower odds of good neurological outcomes (OR 0.22, 95% CI 0.05-0.98). CONCLUSIONS: Hypernatremia at admission was independently associated with decreased probability of good neurological outcomes at discharge post-OHCA. Future studies should focus on elucidating the pathophysiology of dysnatremia following OHCA.

Dynamics of T-cell Responses Following COVID-19 mRNA Vaccination and Breakthrough Infection in Older Adults.

Introduction: While older adults generally mount weaker antibody responses to a primary COVID-19 vaccine series, T-cell responses remain less well characterized in this population. We compared SARS-CoV-2 spike-specific T-cell responses after 2- and 3-dose COVID-19 mRNA vaccination and subsequent breakthrough infection in older and younger adults. Methods: We quantified CD4+ and CD8+ T-cells reactive to overlapping peptides spanning the ancestral SARS-CoV-2 spike protein in 40 older adults (median age 79) and 50 younger health care workers (median age 39), all COVID-19 naive, using an activation-induced marker assay. T-cell responses were further assessed in 24 participants, including 8 older adults, who subsequently experienced their first SARS-CoV-2 breakthrough infection. Results: A third COVID-19 mRNA vaccine dose significantly boosted spike-specific CD4+ and CD8+ T-cell frequencies to above 2-dose levels in older and younger adults. T-cell frequencies did not significantly differ between older and younger adults after either dose. Multivariable analyses adjusting for sociodemographic, health, and vaccine-related variables confirmed that older age was not associated with impaired cellular responses. Instead, the strongest predictors of CD4+ and CD8+ T-cell frequencies post-third-dose were their corresponding post-second-dose frequencies. Breakthrough infection significantly increased both CD4+ and CD8+ T-cell frequencies, to comparable levels in older and younger adults. Exploratory analyses revealed an association between HLA-A*02:03 and higher post-vaccination CD8+ T-cell frequencies, which may be attributable to numerous strong-binding HLA-A*02:03-specific CD8+ T-cell epitopes in the spike protein. Conclusion: Older adults mount robust T-cell responses to 2- and 3-dose COVID-19 mRNA vaccination, which are further boosted following breakthrough infection.

Effects of COVID-19 mRNA vaccination on HIV viremia and reservoir size.

Objective: The immunogenic nature of COVID-19 mRNA vaccines led to some initial concern that these could stimulate the HIV reservoir. We analyzed changes in plasma HIV loads (pVL) and reservoir size following COVID-19 mRNA vaccination in 62 people with HIV (PWH) receiving antiretroviral therapy (ART), and analyzed province-wide trends in pVL before and after the mass vaccination campaign. Design: Longitudinal observational cohort and province-wide analysis. Methods: 62 participants were sampled pre-vaccination, and one month after their first and second COVID-19 immunizations. Vaccine-induced anti-SARS-CoV-2-Spike antibodies in serum were measured using the Roche Elecsys Anti-S assay. HIV reservoirs were quantified using the Intact Proviral DNA Assay; pVL were measured using the cobas 6800 (LLOQ:20 copies/mL). The province-wide analysis included all 290,401 pVL performed in British Columbia, Canada between 2012-2022. Results: Pre-vaccination, the median intact reservoir size was 77 (IQR:20-204) HIV copies/million CD4+ T-cells, compared to 74 (IQR:27-212) and 65 (IQR:22-174) post-first and -second dose, respectively (all comparisons p>0.07). Pre-vaccination, 82% of participants had pVL<20 copies/mL (max:110 copies/mL), compared to 79% post-first dose (max:183 copies/mL) and 85% post-second dose (max:79 copies/mL) (p>0.4). The magnitude of the vaccine-elicited anti-SARS-CoV-2-Spike antibody response did not correlate with changes in reservoir size nor detectable pVL frequency (p>0.6). We found no evidence linking the COVID-19 mass vaccination campaign to population-level increases in detectable pVL frequency among all PWH in the province, nor among those who maintained pVL suppression on ART. Conclusion: We found no evidence that COVID-19 mRNA vaccines induced changes in HIV reservoir size nor plasma viremia.

Age specific reference intervals for plasma biomarkers of neurodegeneration and neurotrauma in a Canadian population.

INTRODUCTION: In this study, we aimed to create reference intervals (RI) using a large Canadian population-based cohort, for plasma protein biomarkers with potential utility to screen, diagnosis, prognosticate and manage a variety of neurological diseases and disorders. RIs were generated for: the ratio of amyloid beta 42 over 40 (Aß42/40), phosphorylated tau-181 (p-tau-181), neurofilament light (NfL), and glial fibrillary acidic protein (GFAP). METHODS: 900 plasma specimens from male and female participants aged 3-79 years old were obtained from the Statistics Canada Biobank, which holds specimens from the Canadian Health Measures Survey. Analysis of Aß42/40, p-tau-181, NfL and GFAP was performed on the Quanterix Simoa HD-X analyzer using the Neurology 4-plex E and p-tau-181 assays. Discrete RIs were produced according to Clinical Laboratory Standards Institute guidelines (EP28-A3c). Continuous RIs were created using quantile regression. RESULTS: For discrete RIs, significant age partitions were determined for each biomarker. No significant sex partitions were found. The following ranges and age partitions were determined: Aß42/40: 3-<55y = 0.053-0.098, 55-<80y = 0.040-0.090; p-tau-181: 3-<12y = 1.4-5.6 pg/ml, 12-<60y = 0.8-3.1 pg/ml, 60-<80y = 0.9-4.0 pg/ml; NfL: 3-<40y = 2.6-11.3 pg/ml, 40-<60y = 4.6-17.7 pg/ml, 60-<80y = 8.1-47.1 pg/ml; GFAP; 3-<10y = 47.0-226 pg/ml, 10-<60y = 21.2-91.9 pg/ml, 60-<80y = 40.7-228 pg/ml. Continuous RIs produced smooth centile curves across the age range, from which point estimates for each year of age were calculated. CONCLUSIONS: Discrete and continuous RIs for neurological plasma biomarkers will help refine normative cut-offs across the lifespan and improve the precision of interpretating biomarker levels. Continuous RIs are recommended for use in age groups, such as pediatrics and older adults, that experience rapid concentration changes by age.

Is it Time to Normalize Scapular Dyskinesis? The Incidence of Scapular Dyskinesis in Those With and Without Symptoms: a Systematic Review of the Literature.

Background: Up to 67% of adults experience shoulder pain in their lifetime. Numerous factors are related to the etiology of shoulder pain, one of which is thought to be scapular dyskinesis (SD). Given the prevalence of SD among the asymptomatic population a concern is that the condition is being medicalized (clinical findings suggested to require treatment but is ultimately a normal finding). Therefore, the purpose of this systematic review was to investigate the prevalence of SD among both symptomatic and asymptomatic populations. Methods: A systematic review of the literature up to July of 2021. Relevant studies identified from PubMed, EMBASE, Cochrane and CINAHL were screened utilizing the following inclusion and exclusion criteria; inclusion: (a) individuals being assessed as having SD, including reliability and validity studies (b) subjects aged 18 or older; (c) sport and non-sport participants; (d) no date restriction; (e) symptomatic, asymptomatic, or both populations; (f) all study designs except case reports. Studies were excluded if: (a) they were not published in the English language; (b) they were a case report design; (c) the presence of SD was part of the studies inclusion criteria; (d) data were not present distinguishing the number of subjects with or without SD; (e) they did not define participants as having or not having SD. Methodological quality of the studies was assessed utilizing the Joanna Briggs Institute checklist. Results: The search resulted in 11,619 after duplicates were removed with 34 studies ultimately retained for analysis after three were removed due to low quality. A total of 2,365 individuals were studied. Within the studies for the symptomatic athletic and general orthopedic population there were 81% and 57% individuals with SD, respectively, and a total of 60% among both symptomatic groups (sport and general orthopedic population). Within the studies for the asymptomatic athletic and general population there were 42% and 59% individuals with SD, respectively, and a total of 48% among both asymptomatic groups (sport and general orthopedic population). Limitation: A strict inclusion and exclusion criteria was used to identify studies that provided the appropriate data for the purpose of this study. There was a lack of consistency for measuring SD across studies. Conclusion: A considerable number of individuals with shoulder symptoms do not present with SD. More revealing is the number of asymptomatic individuals who do present with SD, suggesting that SD may be a normal finding among nearly half of the asymptomatic population. Level of Evidence: 2a.

Lithium related thyroid and parathyroid disease: Updated clinical practice guidelines are needed.

OBJECTIVE: This study aims to estimate the prevalence of and determine physician approaches to the screening and management of lithium-associated thyroid and parathyroid disorders in British Columbia, Canada. METHODS: Serum lithium and thyroid/parathyroid laboratory data were collected retrospectively for patients with lithium levels measured at seven BC hospitals between 2012 and 2021. A mail-out survey about screening and management of thyroid/parathyroid disorders in patients on lithium was sent to the ordering physicians of patients with abnormal results. Three months after, a follow-up questionnaire was sent to respondents, and the original survey was re-sent to non-responders. RESULTS: Of 4917 patients, 1.9 % had PTH (mean 22.33 ± 23.00 pmol/L) and 77.1 % had TSH (mean 3.61 ± 6.69 pmol/L) measured. Of 222 hypercalcemic patients (defined as any serum calcium or ionized calcium above the laboratory reference), 17.6 % had a PTH level measured. From 294 surveys sent to 214 physicians, the overall response rate was 31.6 % (n = 93) with twelve fully completed surveys. All twelve respondents monitored TSH levels every 6-12 months, and eight physicians monitored PTH and/or calcium at variable intervals. Two physicians routinely ordered both thyroid and parathyroid screening laboratory tests. Of the 80 non-respondents, limited patient contact was the most common reason for opting out (n = 27). CONCLUSIONS: Our results suggest biochemical screening for lithium-associated parathyroid disorders is less common than for thyroid disorders. There is insufficient data to determine the true prevalence of lithium-associated thyroid and parathyroid disorders. This highlights the need for updated clinical guidelines for management of lithium-associated thyroid and parathyroid disorders.

A rare isocyanide derived from an unprecedented neutral yttrium(ii) bis(amide) complex.

A room temperature stable complex formulated as Y(NHAr*)2 has been prepared, where Ar* = 2,6-(2,4,6-(iPr)3C6H2)C6H3, by KC8 reduction of ClY(NHAr*)2. Based on EPR evidence, Y(NHAr*)2 is an example of a d1 Y(ii) complex with significant delocalization of the unpaired electron density from the metal to the ligand. The isolation of molecular divalent metal complexes is challenging for rare earth elements such as yttrium. In fact, stabilization of the divalent state requires judicious ligand design that allows the metal center to be coordinatively saturated. Divalent rare earth elements tend to be reactive towards various substrates. Interestingly, Y(NHAr*)2 reacts as a radical donor towards t BuNC to generate an unusual yttrium isocyanide complex, CNY(NHAr*)2, based on spectroscopic evidence and single-crystal X-ray diffraction data.

Lipid acyl chain protrusion induced by the influenza virus hemagglutinin fusion peptide detected by NMR paramagnetic relaxation enhancement.

The glycoprotein spikes of membrane-enveloped viruses include a subunit that catalyzes fusion (joining) of the viral and target cell membranes. For influenza virus, this is subunit 2 of hemagglutinin which has a âˆ¼ 20-residue N-terminal fusion peptide (Fp) region that binds target membrane. An outstanding question is whether there are associated membrane changes important for fusion. Several computational studies have found increased "protrusion" of lipid acyl chains near Fp, i.e. one or more chain carbons are closer to the aqueous region than the headgroup phosphorus. Protrusion may accelerate initial joining of outer leaflets of the two membranes into a stalk intermediate. In this study, higher protrusion probability in membrane with vs. without Fp is convincingly detected by larger Mn2+-associated increases in chain 13C NMR transverse relaxation rates (Γ2's). Data analysis provides a ratio Γ2,neighbor/Γ2,distant for lipids neighboring vs. more distant from the Fp. The calculated ratio depends on the number of Fp-neighboring lipids and the experimentally-derived range of 4 to 24 matches the range of increased protrusion probabilities from different simulations. For samples either with or without Fp, the Γ2 values are well-fitted by an exponential decay as the 13C site moves closer to the chain terminus. The decays correlate with free-energy of protrusion proportional to the number of protruded -CH2 groups, with free energy per -CH2 of ∼0.25 kBT. The NMR data support one major fusion role of the Fp to be much greater protrusion of lipid chains, with highest protrusion probability for chain regions closest to the headgroups.

Bcl-xL is required to protect endothelial cells latently infected with KSHV from virus induced intrinsic apoptosis.

Kaposi's Sarcoma herpesvirus (KSHV) is the etiologic agent of Kaposi's Sarcoma (KS), a highly vascularized tumor common in AIDS patients and many countries in Africa. KSHV is predominantly in the latent state in the main KS tumor cell, the spindle cell, a cell expressing endothelial cell markers. To identify host genes important for KSHV latent infection of endothelial cells we previously used a global CRISPR/Cas9 screen to identify genes necessary for the survival or proliferation of latently infected cells. In this study we rescreened top hits and found that the highest scoring gene necessary for infected cell survival is the anti-apoptotic Bcl-2 family member Bcl-xL. Knockout of Bcl-xL or treatment with a Bcl-xL inhibitor leads to high levels of cell death in latently infected endothelial cells but not their mock counterparts. Cell death occurs through apoptosis as shown by increased PARP cleavage and activation of caspase-3/7. Knockout of the pro-apoptotic protein, Bax, eliminates the requirement for Bcl-xL. Interestingly, neither Bcl-2 nor Mcl-1, related and often redundant anti-apoptotic proteins of the Bcl-2 protein family, are necessary for the survival of latently infected endothelial cells, likely due to their lack of expression in all the endothelial cell types we have examined. Bcl-xL is not required for the survival of latently infected primary effusion lymphoma (PEL) cells or other cell types tested. Expression of the KSHV major latent locus alone in the absence of KSHV infection led to sensitivity to the absence of Bcl-xL, indicating that viral gene expression from the latent locus induces intrinsic apoptosis leading to the requirement for Bcl-xL in endothelial cells. The critical requirement of Bcl-xL during KSHV latency makes it an intriguing therapeutic target for KS tumors.

Bone turnover markers for assessment of anti-resorptive effect in clinical practice: A good idea meets the problem of measurement uncertainty.

OBJECTIVES: Bone turnover markers (BTM) are measures for understanding the effect of anti-resorptives upon osteoclast activity. Post-hoc trial data suggests reduction in BTM of 40% may represent a target for defining appropriate response to therapy. We modeled clinical application of this target threshold in an individual patient setting where assay measurement uncertainty and biological variation are included. DESIGN: Using serum C-telo-peptide (ß-CTX), we constructed hypothetical scenarios of ß-CTX measurement pre and post bisphosphonate therapy. Using typical ß-CTX assay characteristics (analytical coefficient of variation, CV 5.0%) and published intra-individual ß-CTX data for post-menopausal women (CV 18.0%), we calculated the post-therapy ß-CTX that must be seen on single repeat measure for 95% confidence that the observed result was ≥40% below baseline. Sensitivity analyses considered greater and lesser variations in the combined sources of variation. RESULTS: The one-tailed 95% reference change value for any detectable therapeutic decrease in ß-CTX was 22%. However, to have 95% confidence of having achieved a reduction ≥40%, an observed ß-CTX decrease of ≥56% is required. Larger decreases are needed for scenarios of greater analytical or intra-individual variation. CONCLUSIONS: Although population data suggest a ß-CTX decrease of 40% is commensurate with adequate therapeutic response to anti-resorptives, application to an individual patient where measurement and natural variation are present is problematic. ß-CTX decreases much >40% are required to be confident of having achieved the optimal treatment response. It is uncertain whether this is a legitimate change to be expected in all individual patients and therefore clinical application of this threshold is uncertain.

Impact of Age and Severe Acute Respiratory Syndrome Coronavirus 2 Breakthrough Infection on Humoral Immune Responses After Three Doses of Coronavirus Disease 2019 mRNA Vaccine.

Background: Longer-term immune response data after 3 doses of coronavirus disease 2019 (COVID-19) mRNA vaccine remain limited, particularly among older adults and after Omicron breakthrough infection. Methods: We quantified wild-type- and Omicron-specific serum immunoglobulin (Ig)G levels, angiotensin-converting enzyme 2 displacement activities, and live virus neutralization up to 6 months after third dose in 116 adults aged 24-98 years who remained COVID-19 naive or experienced their first severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection during this time. Results: Among the 78 participants who remained COVID-19 naive throughout follow up, wild-type- and Omicron-BA.1-specific IgG concentrations were comparable between younger and older adults, although BA.1-specific responses were consistently significantly lower than wild-type-specific responses in both groups. Wild-type- and BA.1-specific IgG concentrations declined at similar rates in COVID-19-naive younger and older adults, with median half-lives ranging from 69 to 78 days. Antiviral antibody functions declined substantially over time in COVID-19-naive individuals, particularly in older adults: by 6 months, BA.1-specific neutralization was undetectable in 96% of older adults, versus 56% of younger adults. Severe acute respiratory syndrome coronavirus 2 infection, experienced by 38 participants, boosted IgG levels and neutralization above those induced by vaccination alone. Nevertheless, BA.1-specific neutralization remained significantly lower than wild-type, with BA.5-specific neutralization lower still. Higher Omicron BA.1-specific neutralization 1 month after third dose was an independent correlate of lower SARS-CoV-2 infection risk. Conclusions: Results underscore the immune benefits of the third COVID-19 mRNA vaccine dose in adults of all ages and identify vaccine-induced Omicron-specific neutralization as a correlate of protective immunity. Systemic antibody responses and functions however, particularly Omicron-specific neutralization, decline rapidly in COVID-19-naive individuals, particularly in older adults, supporting the need for additional booster doses.

End-to-End Data Automation for Pooled Sample SARS-CoV-2 Using R and Other Open-Source Tools.

BACKGROUND: Due to supply chain shortages of reagents for real-time (RT)-PCR for SARS-CoV-2 and increasing demand on technical staff, an end-to-end data automation strategy for SARS-CoV-2 sample pooling and singleton analysis became necessary in the summer of 2020. METHODS: Using entirely open source software tools-Linux, bash, R, RShiny, ShinyProxy, and Docker-we developed a modular software application stack to manage the preanalytical, analytical, and postanalytical processes for singleton and pooled testing in a 5-week time frame. RESULTS: Pooling was operationalized for 81 days, during which time 64 pooled runs were performed for a total of 5320 sample pools and approximately 21 280 patient samples in 4:1 format. A total of 17 580 negative pooled results were released in bulk. After pooling was discontinued, the application stack was used for singleton analysis and modified to release all viral RT-PCR results from our laboratory. To date, 236 109 samples have been processed avoiding over 610 000 transcriptions. CONCLUSIONS: We present an end-to-end data automation strategy connecting 11 devices, one network attached storage, 2 Linux servers, and the laboratory information system.

People With Human Immunodeficiency Virus Receiving Suppressive Antiretroviral Therapy Show Typical Antibody Durability After Dual Coronavirus Disease 2019 Vaccination and Strong Third Dose Responses.

BACKGROUND: Longer-term humoral responses to 2-dose coronavirus disease 2019 (COVID-19) vaccines remain incompletely characterized in people living with human immunodeficiency virus (HIV) (PLWH), as do initial responses to a third dose. METHODS: We measured antibodies against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein receptor-binding domain, angiotensin-converting enzyme 2 (ACE2) displacement, and viral neutralization against wild-type and Omicron strains up to 6 months after 2-dose vaccination, and 1 month after the third dose, in 99 PLWH receiving suppressive antiretroviral therapy and 152 controls. RESULTS: Although humoral responses naturally decline after 2-dose vaccination, we found no evidence of lower antibody concentrations or faster rates of antibody decline in PLWH compared with controls after accounting for sociodemographic, health, and vaccine-related factors. We also found no evidence of poorer viral neutralization in PLWH after 2 doses, nor evidence that a low nadir CD4+ T-cell count compromised responses. Post-third-dose humoral responses substantially exceeded post-second-dose levels, though Omicron-specific responses were consistently weaker than responses against wild-type virus. Nevertheless, post-third-dose responses in PLWH were comparable to or higher than controls. An mRNA-1273 third dose was the strongest consistent correlate of higher post-third-dose responses. CONCLUSION: PLWH receiving suppressive antiretroviral therapy mount strong antibody responses after 2- and 3-dose COVID-19 vaccination. Results underscore the immune benefits of third doses in light of Omicron.

Antibody response durability following three-dose coronavirus disease 2019 vaccination in people with HIV receiving suppressive antiretroviral therapy.

BACKGROUND: Limited data exist regarding longer term antibody responses following three-dose coronavirus disease 2019 (COVID-19) vaccination, and the impact of a first SARS-CoV-2 infection during this time, in people with HIV (PWH) receiving suppressive antiretroviral therapy (ART). We quantified wild-type-specific, Omicron BA.1-specific and Omicron BA.5-specific responses up to 6 months post-third dose in 64 PWH and 117 controls who remained COVID-19-naive or experienced their first SARS-CoV-2 infection during this time. DESIGN: Longitudinal observational cohort. METHODS: We quantified wild-type-specific and Omicron-specific anti-Spike receptor-binding domain IgG concentrations, ACE2 displacement activities and live virus neutralization at 1, 3 and 6 months post-third vaccine dose. RESULTS: Third doses boosted all antibody measures above two-dose levels, but BA.1-specific responses remained significantly lower than wild-type-specific ones, with BA.5-specific responses lower still. Serum IgG concentrations declined at similar rates in COVID-19-naive PWH and controls post-third dose (median wild-type-specific and BA.1-specific half-lives were between 66 and 74 days for both groups). Antibody function also declined significantly yet comparably between groups: 6 months post-third dose, BA.1-specific neutralization was undetectable in more than 80% of COVID-19 naive PWH and more than 90% of controls. Breakthrough SARS-CoV-2 infection boosted antibody concentrations and function significantly above vaccine-induced levels in both PWH and controls, though BA.5-specific neutralization remained significantly poorer than BA.1 even post-breakthrough. CONCLUSION: Following three-dose COVID-19 vaccination, antibody response durability in PWH receiving ART is comparable with controls. PWH also mounted strong responses to breakthrough infection. Due to temporal response declines, however, COVID-19-naive individuals, regardless of HIV status, would benefit from a fourth dose within 6 months of their third.

Corrigendum to "Migration from RIA to LC-MS/MS for aldosterone determination: Implications for clinical practice and determination of plasma and urine reference range intervals in a cohort of healthy Belgian subjects" [Clin. Mass Spectrom. 9 (2018) 7-17].

[This corrects the article DOI: 10.1016/j.clinms.2018.06.002.].

Serial infection with SARS-CoV-2 Omicron BA.1 and BA.2 following three-dose COVID-19 vaccination.

SARS-CoV-2 Omicron infections are common among individuals who are vaccinated or have recovered from prior variant infection, but few reports have immunologically assessed serial Omicron infections. We characterized SARS-CoV-2 humoral responses in an individual who acquired laboratory-confirmed Omicron BA.1.15 ten weeks after a third dose of BNT162b2, and BA.2 thirteen weeks later. Responses were compared to 124 COVID-19-naive vaccinees. One month post-second and -third vaccine doses, the participant's wild-type and BA.1-specific IgG, ACE2-displacement and virus neutralization activities were average for a COVID-19-naive triple-vaccinated individual. BA.1 infection boosted the participant's responses to the cohort ≥95th percentile, but even this strong "hybrid" immunity failed to protect against BA.2. Reinfection increased BA.1 and BA.2-specific responses only modestly. Though vaccines clearly protect against severe disease, results highlight the continued importance of maintaining additional protective measures to counteract the immune-evasive Omicron variant, particularly as vaccine-induced immune responses naturally decline over time.