Validation of the LUMIPULSE automated immunoassay for the measurement of core AD biomarkers in cerebrospinal fluid

OBJECTIVES: The core cerebrospinal fluid (CSF) biomarkers; total tau (tTau), phospho-tau (pTau), amyloid β 1-42 (Aβ 1-42), and the Aβ 1-42/Aβ 1-40 ratio have transformed Alzheimer's disease (AD) research and are today increasingly used in clinical routine laboratories as diagnostic tools. Fully automated immunoassay instruments with ready-to-use assay kits and calibrators has simplified their analysis and improved reproducibility of measurements. We evaluated the analytical performance of the fully automated immunoassay instrument LUMIPULSE G (Fujirebio) for measurement of the four core AD CSF biomarkers and determined cutpoints for AD diagnosis. METHODS: Comparison of the LUMIPULSE G assays was performed with the established INNOTEST ELISAs (Fujirebio) for hTau Ag, pTau 181, β-amyloid 1-42, and with V-PLEX Plus Aβ Peptide Panel 1 (6E10) (Meso Scale Discovery) for Aβ 1-42/Aβ 1-40, as well as with a LC-MS reference method for Aβ 1-42. Intra- and inter-laboratory reproducibility was evaluated for all assays. Clinical cutpoints for Aβ 1-42, tTau, and pTau was determined by analysis of three cohorts of clinically diagnosed patients, comprising 651 CSF samples. For the Aβ 1-42/Aβ 1-40 ratio, the cutpoint was determined by mixture model analysis of 2,782 CSF samples. RESULTS: The LUMIPULSE G assays showed strong correlation to all other immunoassays (r>0.93 for all assays). The repeatability (intra-laboratory) CVs ranged between 2.0 and 5.6%, with the highest variation observed for β-amyloid 1-40. The reproducibility (inter-laboratory) CVs ranged between 2.1 and 6.5%, with the highest variation observed for β-amyloid 1-42. The clinical cutpoints for AD were determined to be 409 ng/L for total tau, 50.2 ng/L for pTau 181, 526 ng/L for β-amyloid 1-42, and 0.072 for the Aβ 1-42/Aβ 1-40 ratio. CONCLUSIONS: Our results suggest that the LUMIPULSE G assays for the CSF AD biomarkers are fit for purpose in clinical laboratory practice. Further, they corroborate earlier presented reference limits for the biomarkers

Single-Trait and Multi-Trait Genome-Wide Association Analyses Identify Novel Loci for Blood Pressure in African-Ancestry Populations

Hypertension is a leading cause of global disease, mortality, and disability. While individuals of African descent suffer a disproportionate burden of hypertension and its complications, they have been underrepresented in genetic studies. To identify novel susceptibility loci for blood pressure and hypertension in people of African ancestry, we performed both single and multiple-trait genome-wide association analyses. We analyzed 21 genome-wide association studies comprised of 31,968 individuals of African ancestry, and validated our results with additional 54,395 individuals from multi-ethnic studies. These analyses identified nine loci with eleven independent variants which reached genome-wide significance (P \u3c 1.25×10−8) for either systolic and diastolic blood pressure, hypertension, or for combined traits. Single-trait analyses identified two loci (TARID/TCF21 and LLPH/TMBIM4) and multiple-trait analyses identified one novel locus (FRMD3) for blood pressure. At these three loci, as well as at GRP20/CDH17, associated variants had alleles common only in African-ancestry populations. Functional annotation showed enrichment for genes expressed in immune and kidney cells, as well as in heart and vascular cells/tissues. Experiments driven by these findings and using angiotensin-II induced hypertension in mice showed altered kidney mRNA expression of six genes, suggesting their potential role in hypertension. Our study provides new evidence for genes related to hypertension susceptibility, and the need to study African-ancestry populations in order to identify biologic factors contributing to hypertension

ESTABLISHMENT OF AIDS-RELATED LYMPHOMA CELL LINES FROM LYMPHOMATOUS EFFUSIONS

AIDS-related non-Hodgkin lymphomas (AIDS-NHL) are most frequently derived from B cells and include small non-cleaved cell lymphoma (SNCCL) and diffuse large cell lymphoma (DLCL) and less frequently anaplastic large cell lymphoma (ALCL) or body cavity-based lymphoma (BCBL). AIDS-NHL cell lines have proved useful to study AIDS-NHL pathogenesis. In this report, we describe the establishment and molecular characterization of two novel AIDS-NHL cell lines (HBL-4 and HBL-6) derived from lymphomatous effusions. HBL-4 was derived from a patient with SNCCL, whereas HBL-6 was derived from a patient with BCBL. The identity of the cell lines with the original tumor clone was established by immunoglobulin gene rearrangement analysis. Both HBL-4 and HBL-6 carry a monoclonal EBV infection and do not contain HIV. In addition, HBL-6 harbors DNA sequences of the recently identified Kaposi's sarcoma-associated herpesvirus (KSHV), now formally called human herpesvirus 8 (HHV8). Finally, HBL-4, but not HBL-6, harbors a rearranged c-MYC allele, while the BCL-6 gene displayed a germline configurations in both cell lines. These AIDS-NHL cell lines may prove useful in understanding the biologic events contributing to AIDS-NHL development

Negative autoregulation of c-myc gene expression is inactivated in transformed cells.

Negative feedback regulation of c-myc gene expression has been observed in some, but not all, cell types. In order to demonstrate conclusively the existence of this mechanism and gain insight into the cause of its inactivation, we have directly examined its function in B cells and then investigated its activity in a number of cell types. We demonstrate the existence of negative c-myc autoregulation by showing the rapid, dose dependent and reversible suppression of endogenous c-myc expression in EBV-immortalized B lymphoblastoid cells transfected with a c-myc gene expressed under the control of a heavy metal inducible promoter. Autoregulation occurs at the level of transcriptional initiation and is mediated by at least one stable intermediate or cofactor molecule. The c-myc autoregulatory mechanism was found operative in all (11 of 11) non-tumorigenic cells tested, including normal and immortalized lymphocytes and fibroblasts. However, this mechanism was found to be inactive in all (10 of 10) tumor cell lines derived from a variety of tissues including those carrying normal and oncogenically activated c-myc genes. These data establish the existence of an important regulatory circuit modulating c-myc expression in normal cells and suggest that its inactivation may represent a general regulatory disturbance of transformed cells

High Satellite Repeat Turnover in Great Apes Studied with Short- And Long-Read Technologies

Satellite repeats are a structural component of centromeres and telomeres, and in some instances, their divergence is known to drive speciation. Due to their highly repetitive nature, satellite sequences have been understudied and underrepresented in genome assemblies. To investigate their turnover in great apes, we studied satellite repeats of unit sizes up to 50 bp in human, chimpanzee, bonobo, gorilla, and Sumatran and Bornean orangutans, using unassembled short and long sequencing reads. The density of satellite repeats, as identified from accurate short reads (Illumina), varied greatly among great ape genomes. These were dominated by a handful of abundant repeated motifs, frequently shared among species, which formed two groups: 1) the (AATGG)n repeat (critical for heat shock response) and its derivatives; and 2) subtelomeric 32-mers involved in telomeric metabolism. Using the densities of abundant repeats, individuals could be classified into species. However, clustering did not reproduce the accepted species phylogeny, suggesting rapid repeat evolution. Several abundant repeats were enriched in males versus females; using Y chromosome assemblies or Fluorescent In Situ Hybridization, we validated their location on the Y. Finally, applying a novel computational tool, we identified many satellite repeats completely embedded within long Oxford Nanopore and Pacific Biosciences reads. Such repeats were up to 59 kb in length and consisted of perfect repeats interspersed with other similar sequences. Our results based on sequencing reads generated with three different technologies provide the first detailed characterization of great ape satellite repeats, and open new avenues for exploring their functions