HostSeq: a Canadian whole genome sequencing and clinical data resource.

HostSeq was launched in April 2020 as a national initiative to integrate whole genome sequencing data from 10,000 Canadians infected with SARS-CoV-2 with clinical information related to their disease experience. The mandate of HostSeq is to support the Canadian and international research communities in their efforts to understand the risk factors for disease and associated health outcomes and support the development of interventions such as vaccines and therapeutics. HostSeq is a collaboration among 13 independent epidemiological studies of SARS-CoV-2 across five provinces in Canada. Aggregated data collected by HostSeq are made available to the public through two data portals: a phenotype portal showing summaries of major variables and their distributions, and a variant search portal enabling queries in a genomic region. Individual-level data is available to the global research community for health research through a Data Access Agreement and Data Access Compliance Office approval. Here we provide an overview of the collective project design along with summary level information for HostSeq. We highlight several statistical considerations for researchers using the HostSeq platform regarding data aggregation, sampling mechanism, covariate adjustment, and X chromosome analysis. In addition to serving as a rich data source, the diversity of study designs, sample sizes, and research objectives among the participating studies provides unique opportunities for the research community.

Images from the surface of asteroid Ryugu show rocks similar to carbonaceous chondrite meteorites.

The near-Earth asteroid (162173) Ryugu is a 900-m-diameter dark object expected to contain primordial material from the solar nebula. The Mobile Asteroid Surface Scout (MASCOT) landed on Ryugu's surface on 3 October 2018. We present images from the MASCOT camera (MASCam) taken during the descent and while on the surface. The surface is covered by decimeter- to meter-sized rocks, with no deposits of fine-grained material. Rocks appear either bright, with smooth faces and sharp edges, or dark, with a cauliflower-like, crumbly surface. Close-up images of a rock of the latter type reveal a dark matrix with small, bright, spectrally different inclusions, implying that it did not experience extensive aqueous alteration. The inclusions appear similar to those in carbonaceous chondrite meteorites.

Patterns of kidney injury in pediatric nonkidney solid organ transplant recipients.

The incidence of acute kidney injury (AKI) and its impact on chronic kidney disease (CKD) following pediatric nonkidney solid organ transplantation is unknown. We aimed to determine the incidence of AKI and CKD and examine their relationship among children who received a heart, lung, liver, or multiorgan transplant at the Hospital for Sick Children between 2002 and 2011. AKI was assessed in the first year posttransplant. Among 303 children, perioperative AKI (within the first week) occurred in 67% of children, and AKI after the first week occurred in 36%, with the highest incidence among lung and multiorgan recipients. Twenty-three children (8%) developed CKD after a median follow-up of 3.4 years. Less than 5 children developed end-stage renal disease, all within 65 days posttransplant. Those with 1 AKI episode by 3 months posttransplant had significantly greater risk for developing CKD after adjusting for age, sex, and estimated glomerular filtration rate at transplant (hazard ratio: 2.77, 95% confidence interval, 1.13-6.80, P trend = .008). AKI is common in the first year posttransplant and associated with significantly greater risk of developing CKD. Close monitoring for kidney disease may allow for earlier implementation of kidney-sparing strategies to decrease risk for progression to CKD.

Use of Everolimus After Multivisceral Transplantation: A Report of Two Cases.

Inhibitors of mechanistic target of rapamycin are used in solid organ transplant procedures to avoid calcineurin inhibitor complications, including nephrotoxicity and malignancy. We present 2 cases of multivisceral transplantation for neuroendocrine tumor (NET) for which everolimus was implemented for its potential to prevent NET recurrence as well as preserve renal function. The first case was complicated by NET recurrence in the liver before initiation of everolimus. After initiation of everolimus, the patient developed a ventral hernia and elevated aminotransferase levels with nonspecific biopsy findings. The second case was complicated by cytomegalovirus infection with elevated everolimus trough levels as well as acute cellular rejection. Everolimus was reinitiated in both cases in addition to decreasing the dosage of tacrolimus, and there were no further complications. Everolimus was beneficial in stabilizing renal function in both patients and has the theoretical potential to prevent recurrence of NET.

Intestine Transplantation Across a Positive Crossmatch With Preformed Donor-Specific Antibodies.

BACKGROUND: We describe our experience using a modified protocol for immunosuppression for intestine transplantation across a positive crossmatch. Patients who underwent transplantation in 2013 were evaluated over a 12-month period for rejection and infectious events with comparison to procedure-matched controls on our standard protocol of immunosuppression. PATIENTS AND METHODS: We used a modified protocol for intestine and multivisceral transplantation for patients with a positive flow crossmatch. In addition to our standard protocol, patients with positive crossmatch were given rituximab and intravenous immunoglobulin (IVIg) preoperatively. DSA was sent for clinical evaluation at monthly intervals. Patients were screened for rejection by endoscopic evaluation. RESULTS: Four patients underwent transplantation within a single year across a positive crossmatch. Two received isolated intestine transplants and 2 had multivisceral transplantation (MVT). During the 12-month follow-up, 1 patients had an episode of severe acute cellular rejection, which was managed with increased immunosuppression. None of the patients had episodes of cytomegalovirus infection. One patient developed major infection and 3 patients developed minor bacterial infections. Among procedure-matched controls with negative final crossmatch on standard management (no preoperative rituximab or IVIg), 2 developed mild acute cellular rejection and 2 developed minor infections. One developed cytomegalovirus viremia with invasion to the colonic mucosa. CONCLUSIONS: We report our protocol for immunosuppression for IT and MVT across a positive crossmatch. This allowed transplantation despite the presence of a positive crossmatch, with low rejection rates but potentially increased risk for major infections compared to the negative crossmatch controls on our standard protocol.

Integrin α1ß1 differentially regulates cytokine-mediated responses in chondrocytes.

OBJECTIVE: To elucidate the role of integrin α1ß1 in chondrocyte responses to inflammatory interleukin-1α (IL-1) and anabolic transforming growth factor-ß1 (TGF-ß1) in the knee. METHODS: Intracellular calcium transient responses to IL-1 and TGF-ß1 were measured in wild type and integrin α1-null chondrocytes using real time ex vivo confocal microscopy, and immunohistochemistry was performed to analyze TGF-ß1-mediated activation of Smad2/3 in tibial and femoral chondrocytes. RESULTS: Loss of integrin α1ß1 reduces intracellular calcium transient response to IL-1, while it enhances chondrocyte responses to TGF-ß1 as measured by intracellular calcium transients and activation of downstream Smad2/3. CONCLUSIONS: Integrin α1ß1 plays a vital role in mediating chondrocyte responses to two contrasting factors that are critical players in the onset and progression of osteoarthritis - inflammatory IL-1 and anabolic TGF-ß. Further investigation into the molecular mechanisms by which integrin α1ß1 mediates these responses will be an important next step in understanding the influence of increased expression of integrin α1ß1 during the early stages of osteoarthritis on disease progression.

Improved detection of semen by use of direct acid phosphatase testing.

Acid phosphatase (AP) reagent (Fast Black) is used as a presumptive test for the presence of seminal fluid on exhibits submitted in allegations of sexual assault. Research was carried out to determine whether the direct application of AP reagent to exhibits is a viable alternative to the traditional indirect (blot) testing method used routinely in the laboratory. The relative sensitivity of the indirect and direct testing methods was investigated as was the effect of AP reagent on histological staining of spermatozoa, the incidence of false positives from vaginal material and saliva, and the effect of AP reagent on subsequent DNA testing. Also included are the results of specificity studies from validations of the direct AP testing method. The results of this research show that, provided the incidence of false positives is borne in mind, direct AP testing can be especially useful when screening exhibits which are difficult to indirectly (blot) AP test or when it is problematic to relocate an AP positive stain. Direct application of AP reagent can also be beneficial for locating dilute semen stains. Three case examples are given which illustrate the use of direct AP testing in laboratory casework.

Subclinical rejection in stable positive crossmatch kidney transplant patients: incidence and correlations.

We reviewed 116 surveillance biopsies obtained approximately 1, 3, 6 and 12 months posttransplantation from 50 +XM live donor kidney transplant recipients to determine the frequency of subclinical cell-mediated rejection (CMR) and antibody-mediated rejection (AMR). Subclinical CMR was present in 39.7% of the biopsies at 1 month and >20% at all other time points. The presence of diffuse C4d on biopsies obtained at each time interval ranged from 20 to 30%. In every case, where histological and immunohistological findings were diagnostic for AMR, donor-specific antibody was found in the blood, challenging the long-held belief that low-level antibody could evade detection due to absorption on the graft. Among clinical factors, only recipient age was associated with subclinical CMR. Clinical factors associated with subclinical AMR were recipient age, positive cytotoxic crossmatch prior to desensitization and two mismatches of HLA DR 51, 52 and 53 alleles. Surveillance biopsies during the first year post-transplantation for these high-risk patients uncover clinically occult processes and phenotypes, which without intervention diminish allograft survival and function.

Undertreatment of hyperlipidemia in a cohort of United States kidney dialysis patients.

BACKGROUND: Application of national guidelines regarding cardiovascular disease risk reduction to kidney dialysis patients is complicated by the conflicting observations that dialysis patients have a high risk of atherosclerotic cardiovascular disease (ASCVD), but dialysis patients with higher serum cholesterol have lower mortality rates. Actual treatment patterns of hyperlipidemia are not well studied. METHODS: We assessed the prevalence, treatment and control of hyperlipidemia in this high-risk patient population from 1995 - 1998. We measured low-density lipoprotein cholesterol, treatment with a lipid-lowering agent, and prevalence of hyperlipidemia as defined by the National Cholesterol Education Program (NCEP), Adult Treatment Panel (ATP) II guidelines in 812 incident hemodialysis (HD), and peritoneal dialysis (PD) patients from dialysis clinics in 19 states throughout the United States. RESULTS: Hyperlipidemia was present in 40% of HD and 62% of PD patients. Among subjects with hyperlipidemia, 67% of HD and 63% of PD patients were untreated and only 22% of HD and 14% of PD patients were treated and controlled. Those who entered the study in 1997 or 1998, those with diabetes, males and Caucasians were more likely to be treated and controlled, whereas subjects on PD and those with ASCVD were less likely to be treated and controlled. CONCLUSION: These data suggest that high rates of undertreatment exist in the United States ESRD dialysis population. Whether improved rates of treatment will result in decreased cardiovascular disease events needs to be tested in randomized clinical trials.

Clinical practice guidelines for managing dyslipidemias in kidney transplant patients: a report from the Managing Dyslipidemias in Chronic Kidney Disease Work Group of the National Kidney Foundation Kidney Disease Outcomes Quality Initiative.

The incidence of cardiovascular disease (CVD) is very high in patients with chronic kidney (CKD) disease and in kidney transplant recipients. Indeed, available evidence for these patients suggests that the 10-year cumulative risk of coronary heart disease is at least 20%, or roughly equivalent to the risk seen in patients with previous CVD. Recently, the National Kidney Foundation's Kidney Disease Outcomes Quality Initiative (K/DOQI) published guidelines for the diagnosis and treatment of dyslipidemias in patients with CKD, including transplant patients. It was the conclusion of this Work Group that the National Cholesterol Education Program Guidelines are generally applicable to patients with CKD, but that there are significant differences in the approach and treatment of dyslipidemias in patients with CKD compared with the general population. In the present document we present the guidelines generated by this workgroup as they apply to kidney transplant recipients. Evidence from the general population indicates that treatment of dyslipidemias reduces CVD, and evidence in kidney transplant patients suggests that judicious treatment can be safe and effective in improving dyslipidemias. Dyslipidemias are very common in CKD and in transplant patients. However, until recently there have been no adequately powered, randomized, controlled trials examining the effects of dyslipidemia treatment on CVD in patients with CKD. Since completion of the K/DOQI guidelines on dyslipidemia in CKD, the results of the Assessment of Lescol in Renal Transplantation (ALERT) Study have been presented and published. Based on information from randomized trials conducted in the general population and the single study conducted in kidney transplant patients, these guidelines, which are a modified version of the K/DOQI dyslipidemia guidelines, were developed to aid clinicians in the management of dyslipidemias in kidney transplant patients. These guidelines are divided into four sections. The first section (Introduction) provides the rationale for the guidelines, and describes the target population, scope, intended users, and methods. The second section presents guidelines on the assessment of dyslipidemias (guidelines 1-3), while the third section offers guidelines for the treatment of dyslipidemias (guidelines 4-5). The key guideline statements are supported mainly by data from studies in the general population, but there is an urgent need for additional studies in CKD and in transplant patients. Therefore, the last section outlines recommendations for research.

hAG-2 and hAG-3, human homologues of genes involved in differentiation, are associated with oestrogen receptor-positive breast tumours and interact with metastasis gene C4.4a and dystroglycan.

hAG-2 and hAG-3 are recently discovered human homologues of the secreted Xenopus laevis proteins XAG-1/2 (AGR-1/2) that are expressed in the cement gland, an ectodermal organ in the head associated with anteroposterior fate determination during early development. Although the roles of hAG-2 and hAG-3 in mammalian cells are unknown, both proteins share a high degree of protein sequence homology and lie adjacent to one another on chromosome 7p21. hAG-2 mRNA expression has previously been demonstrated in oestrogen receptor (ER)-positive cell lines. In this study, we have used real-time quantitative RT - PCR analysis and immunohistochemistry on tissue microarrays to demonstrate concordant expression of hAG-2 and hAG-3 mRNA and protein in breast tumour tissues. Tumour expression of both genes correlated with OR (hAG2, P=0.0002; hAG-3, P=0.0012), and inversely correlated with epidermal growth factor receptor (EGFR) (P=0.003). Yeast two-hybrid cloning identified metastasis-associated GPI-anchored C4.4a protein and extracellular alpha-dystroglycan (DAG-1) as binding partners for both hAG-2 and hAG-3, which if replicated in clinical oncology would demonstrate a potential role in tumour metastasis through the regulation of receptor adhesion and functioning. hAG-2 and hAG-3 may therefore serve as useful molecular markers and/or potential therapeutic targets for hormone-responsive breast tumours.

Cardiovascular mortality in children and young adults with end-stage kidney disease.

OBJECTIVE: To analyze cardiovascular death in a national end-stage renal disease (ESRD) population. STUDY DESIGN: This retrospective, observational study with data from the US Renal Data Systems analyzed 1380 deaths from 1990 to 1996 among patients who started ESRD therapy as children and died before 30 years of age. RESULTS: Percentage of cardiac deaths (n = 311) varied by age and was higher among black patients (0-4 years, 36%; 5-9 years, 18%; 10-14 years, 35%; 15-19 years, 22%; 20-30 years, 32%) than white patients (18%, 12%, 17%, 14%, and 23%, respectively). Among black patients, cardiac deaths occurred in 11% of transplant recipients, 34% of dialysis patients, and among white patients 9% and 25%, respectively. Black patients were 1.6 times more likely to die of a cardiac death (P <.001) than white patients. Transplant recipients had 78% lower risk of cardiac death than dialysis patients (odds ratio = 0.22; P =.0001). The cardiac death rate among dialysis patients was 21.4 per 1000 patient-years in black patients compared with 20.5 in white patients. Transplantation cardiac death rates were lower in black patients, 2.1 per 1000 patient-years, and 1.3 in white patients. CONCLUSIONS: Cardiovascular death accounts for 23% of pediatric and young adult ESRD deaths. Black patients and dialysis patients are at higher risk of a cardiac death compared with white patients and transplant recipients. Further studies are needed to identify risk factors associated with cardiovascular death in patients with ESRD.

Proteomic comparison of human and great ape blood plasma reveals conserved glycosylation and differences in thyroid hormone metabolism.

Most blood plasma proteins are glycosylated. These glycoproteins typically carry sialic acid-bearing sugar chains, which can modify the observed molecular weights and isoelectric points of those proteins during electrophoretic analyses. To explore changes in protein expression and glycosylation that occurred during great ape and human evolution, we subjected multiple blood plasma samples from all these species to high-resolution proteomic analysis. We found very few species-specific differences, indicating a remarkable degree of conservation of plasma protein expression and glycosylation during approximately 12 million years of evolution. A few lineage-specific differences in protein migration were noted among the great apes. The only obvious differences between humans and all great apes were an apparent decrease in transthyretin (prealbumin) and a change in haptoglobin isoforms (the latter was predictable from prior genetic studies). Quantitative studies of transthyretin in samples of blood plasma (synthesized primarily by the liver) and of cerebrospinal fluid (synthesized locally by the choroid plexus of the brain) confirmed approximately 2-fold higher levels in chimpanzees compared to humans. Since transthyretin binds thyroid hormones, we next compared plasma thyroid hormone parameters between humans and chimpanzees. The results indicate significant differences in the status of thyroid hormone metabolism, which represent the first known endocrine difference between these species. Notably, thyroid hormones are known to play major roles in the development, differentiation, and metabolism of many organs and tissues, including the brain and the cranium. Also, transthyretin is known to be the major carrier of thyroid hormone in the cerebrospinal fluid, likely regulating delivery of this hormone to the brain. A potential secondary difference in retinoid (vitamin A) metabolism is also noted. The implications of these findings for explaining unique features of human evolution are discussed.

Alcohol: role in the development of hypertension and end-stage renal disease.

Alcohol is a common risk factor in the general population for a variety of health outcomes. In the present review, we discuss the recent literature on alcohol, hypertension, and renal disease. The regular consumption of more than two drinks per day is associated with both hypertension and renal disease. The mechanisms by which consumption of alcohol leads to hypertension and perhaps renal disease are unknown.

Constructive neural-network learning algorithms for pattern classification.

Constructive learning algorithms offer an attractive approach for the incremental construction of near-minimal neural-network architectures for pattern classification. They help overcome the need for ad hoc and often inappropriate choices of network topology in algorithms that search for suitable weights in a priori fixed network architectures. Several such algorithms are proposed in the literature and shown to converge to zero classification errors (under certain assumptions) on tasks that involve learning a binary to binary mapping (i.e., classification problems involving binary-valued input attributes and two output categories). We present two constructive learning algorithms MPyramid-real and MTiling-real that extend the pyramid and tiling algorithms, respectively, for learning real to M-ary mappings (i.e., classification problems involving real-valued input attributes and multiple output classes). We prove the convergence of these algorithms and empirically demonstrate their applicability to practical pattern classification problems. Additionally, we show how the incorporation of a local pruning step can eliminate several redundant neurons from MTiling-real networks.

Glycan composition of serum alpha-fetoprotein in patients with hepatocellular carcinoma and non-seminomatous germ cell tumour.

Although estimation of serum alpha-fetoprotein (AFP) is widely used in the diagnosis of hepatocellular carcinoma (HCC) and non-seminomatous germ cell tumours (NSGCT), the clinical usefulness of this test is limited by a low specificity. However, there exist glycoforms of AFP which may be more specific for particular tumours. Previously, detailed analysis has been prevented by the low levels of AFP in human serum. We report here the application of fluorescence labelling, sequential exoglycosidase digestion, high-performance liquid chromatography and matrix-assisted laser desorption ionization in time-of-flight mass spectrometry, to determine the glycan structures of purified serum AFP from patients with HCC and NSGCT. Eleven major glycans were found, of which seven were N-linked, and four were O-linked, to the protein backbone. The structure of the N-linked glycans (all of bi-antennary complex-type with varying degrees of sialylation, fucosylation and galactosylation) were consistent with those previously reported. The O-linked glycans (three mucin O-GalNAc type glycans with variable degrees of sialylation, one O-HexNAc monosaccharide glycan) have not previously been reported. The finding of mucin O-GalNAc type glycans was supported by the prediction of potential O-GalNAc glycosylation sites on the protein backbone by analysis of the AFP structure by molecular modelling. With knowledge of these structures it may be possible to develop more specific assays for the detection of HCC and NSGCT.

Proteomic definition of normal human luminal and myoepithelial breast cells purified from reduction mammoplasties.

Normal human luminal and myoepithelial breast cells separately purified from a set of 10 reduction mammoplasties by using a double antibody magnetic affinity cell sorting and Dynabead immunomagnetic technique were used in two-dimensional gel proteome studies. A total of 43,302 proteins were detected across the 20 samples, and a master image for each cell type comprising a total of 1,738 unique proteins was derived. Differential analysis identified 170 proteins that were elevated 2-fold or more between the two breast cell types, and 51 of these were annotated by tandem mass spectrometry. Muscle-specific enzyme isoforms and contractile intermediate filaments including tropomyosin and smooth muscle (SM22) alpha protein were detected in the myoepithelial cells, and a large number of cytokeratin subclasses and isoforms characteristic of luminal cells were detected in this cell type. A further 134 nondifferentially regulated proteins were also annotated from the two breast cell types, making this the most extensive study to date of the protein expression map of the normal human breast and the basis for future studies of purified breast cancer cells.

Proteomics: a major new technology for the drug discovery process.

Proteomics is a new enabling technology that is being integrated into the drug discovery process. This will facilitate the systematic analysis of proteins across any biological system or disease, forwarding new targets and information on mode of action, toxicology and surrogate markers. Proteomics is highly complementary to genomic approaches in the drug discovery process and, for the first time, offers scientists the ability to integrate information from the genome, expressed mRNAs, their respective proteins and subcellular localization. It is expected that this will lead to important new insights into disease mechanisms and improved drug discovery strategies to produce novel therapeutics.

Human T cell IgD receptors react with O-glycans on both human IgD and IgA1.

Previous studies on murine T cell IgD-R have shown that these receptors recognize N-glycans of murine IgD, and not of other Ig isotypes. We have now studied the specificity of IgD-R on human T cells. Human IgD digested with proteinase K to fragments of < 5 kDa inhibit the ability of T cells to form rosettes with IgD-coated ox erythrocytes. The same amount of digested IgG does not. We tested all the human Ig isotypes: IgG1, -2, -3, -4, IgA2, IgE and IgM fail to inhibit significantly at 20 microg/assay. However, IgA1 is as effective as IgD itself, showing approximately 60 % and 80 % inhibition at 5 microg and 10 microg/assay. Human IgA1 and IgD both contain Gal-1 --> 3-GalNac-rich O-linked glycans, and on this basis are both bound to ricin and jacalin. The O-linked glycans may therefore also represent the common moiety binding to IgD-R. Disaccharides Gal-1 --> 3-GalNac, and Gal-1 --> 4-Glc at 10 microg/assay blocked IgD rosetting while Gal-1 --> 6-Glc did not. We conclude that the human IgD-R is a lectin, differing from the murine IgD-R in that it has both IgA1 and IgD as ligands.