Tissue registration and exploration user interfaces in support of a human reference atlas.

Seventeen international consortia are collaborating on a human reference atlas (HRA), a comprehensive, high-resolution, three-dimensional atlas of all the cells in the healthy human body. Laboratories around the world are collecting tissue specimens from donors varying in sex, age, ethnicity, and body mass index. However, harmonizing tissue data across 25 organs and more than 15 bulk and spatial single-cell assay types poses challenges. Here, we present software tools and user interfaces developed to spatially and semantically annotate ("register") and explore the tissue data and the evolving HRA. A key part of these tools is a common coordinate framework, providing standard terminologies and data structures for describing specimen, biological structure, and spatial data linked to existing ontologies. As of April 22, 2022, the "registration" user interface has been used to harmonize and publish data on 5,909 tissue blocks collected by the Human Biomolecular Atlas Program (HuBMAP), the Stimulating Peripheral Activity to Relieve Conditions program (SPARC), the Human Cell Atlas (HCA), the Kidney Precision Medicine Project (KPMP), and the Genotype Tissue Expression project (GTEx). Further, 5,856 tissue sections were derived from 506 HuBMAP tissue blocks. The second "exploration" user interface enables consortia to evaluate data quality, explore tissue data spatially within the context of the HRA, and guide data acquisition. A companion website is at https://cns-iu.github.io/HRA-supporting-information/ .

Combining plasma extracellular vesicle Let-7b-5p, miR-184 and circulating miR-22-3p levels for NSCLC diagnosis and drug resistance prediction.

Low-dose computed tomography (LDCT) Non-Small Cell Lung (NSCLC) screening is associated with high false-positive rates, leading to unnecessary expensive and invasive follow ups. There is a need for minimally invasive approaches to improve the accuracy of NSCLC diagnosis. In addition, NSCLC patients harboring sensitizing mutations in epidermal growth factor receptor EGFR (T790M, L578R) are treated with Osimertinib, a potent tyrosine kinase inhibitor (TKI). However, nearly all patients develop TKI resistance. The underlying mechanisms are not fully understood. Plasma extracellular vesicle (EV) and circulating microRNA (miRNA) have been proposed as biomarkers for cancer screening and to inform treatment decisions. However, the identification of highly sensitive and broadly predictive core miRNA signatures remains a challenge. Also, how these systemic and diverse miRNAs impact cancer drug response is not well understood. Using an integrative approach, we examined plasma EV and circulating miRNA isolated from NSCLC patients versus screening controls with a similar risk profile. We found that combining EV (Hsa-miR-184, Let-7b-5p) and circulating (Hsa-miR-22-3p) miRNAs abundance robustly discriminates between NSCLC patients and high-risk cancer-free controls. Further, we found that Hsa-miR-22-3p, Hsa-miR-184, and Let-7b-5p functionally converge on WNT/ßcatenin and mTOR/AKT signaling axes, known cancer therapy resistance signals. Targeting Hsa-miR-22-3p and Hsa-miR-184 desensitized EGFR-mutated (T790M, L578R) NSCLC cells to Osimertinib. These findings suggest that the expression levels of circulating hsa-miR-22-3p combined with EV hsa-miR-184 and Let-7b-5p levels potentially define a core biomarker signature for improving the accuracy of NSCLC diagnosis. Importantly, these biomarkers have the potential to enable prospective identification of patients who are at risk of responding poorly to Osimertinib alone but likely to benefit from Osimertinib/AKT blockade combination treatments.

Transcriptome dissection of candidate genes associated with lentil seed quality traits.

Lentils provide a rich plant-based protein source and staple food in many parts of the world. Despite numerous nutritional benefits, lentil seeds also possess undesirable elements, such as anti-nutritional factors. Understanding the genetic networks of seed metabolism is of great importance for improving the seed nutritional profile. We applied RNA sequencing analysis to survey the transcriptome of developing lentil seeds and compared this with that of the pod shells and leaves. In total, we identified 2622 genes differentially expressed among the tissues examined. Genes preferentially expressed in seeds were enriched in the Gene Ontology (GO) terms associated with development, nitrogen and carbon (N/C) metabolism and lipid synthesis. We further categorized seed preferentially expressed genes based on their involvement in storage protein production, starch accumulation, lipid and suberin metabolism, phytate, saponin and phenylpropanoid biosynthesis. The availability of transcript profile datasets on lentil seed metabolism and a roadmap of candidate genes presented here will be of great value for breeding strategies towards further improvement of lentil seed quality traits.

Assessing the Potential of Metal-Assisted Imaging Mass Spectrometry in Cancer Research.

In the last decade, imaging mass spectrometry (IMS) has been the primary tool for biomolecular imaging. While it is possible to map a wide range of biomolecules using matrix-assisted laser desorption/ionization IMS ranging from high-molecular-weight proteins to small metabolites, more often than not only the most abundant easily ionisable species are detected. To better understand complex diseases such as cancer more specific and sensitive methods need to be developed to enable the detection of lower abundance molecules but also molecules that have yet to be imaged by IMS. In recent years, a big shift has occurred in the imaging community from developing wide reaching methods to developing targeted ones which increases sensitivity through the use of more specific sample preparations. This has been primarily marked by the advent of solvent-free matrix deposition methods for polar lipids, chemical derivatization for hormones and metabolites, and the use of alternative ionization agents for neutral lipids. In this chapter, we discuss two of the latest sample preparations which exploit the use of alternative ionization agents to enable the detection of certain classes of neutral lipids along with free fatty acids by high-sensitivity IMS as demonstrated within our lab.

Consumer understanding of sugars claims on food and drink products.

Consumer understanding of nutrition and health claims is a key aspect of current regulations in the European Union (EU). In view of this, qualitative and quantitative research techniques were used to investigate consumer awareness and understanding of product claims in the UK, focusing particularly on nutrition claims relating to sugars. Both research methods identified a good awareness of product claims. No added sugars claims were generally preferred to reduced sugars claims, and there was a general assumption that sweeteners and other ingredients would be added in place of sugars. However, there was little awareness of the level of sugar reduction and the associated calorie reduction in products when reduced sugars claims were made on pack. In focus groups, participants felt deceived if sugar reduction claims were being made without a significant reduction in calories. This was reinforced in the quantitative research which showed that respondents expected a similar and meaningful level of calorie reduction to the level of sugar reduction. The research also identified consumer confusion around the calorie content of different nutrients, including over-estimation of the calorie content of sugars. This is crucial to consumers' expectations as they clearly link sugar to calories and therefore expect a reduction in sugar content to deliver a reduction in calorie content.

The magic nature of (132)Sn explored through the single-particle states of (133)Sn.

Atomic nuclei have a shell structure in which nuclei with 'magic numbers' of neutrons and protons are analogous to the noble gases in atomic physics. Only ten nuclei with the standard magic numbers of both neutrons and protons have so far been observed. The nuclear shell model is founded on the precept that neutrons and protons can move as independent particles in orbitals with discrete quantum numbers, subject to a mean field generated by all the other nucleons. Knowledge of the properties of single-particle states outside nuclear shell closures in exotic nuclei is important for a fundamental understanding of nuclear structure and nucleosynthesis (for example the r-process, which is responsible for the production of about half of the heavy elements). However, as a result of their short lifetimes, there is a paucity of knowledge about the nature of single-particle states outside exotic doubly magic nuclei. Here we measure the single-particle character of the levels in (133)Sn that lie outside the double shell closure present at the short-lived nucleus (132)Sn. We use an inverse kinematics technique that involves the transfer of a single nucleon to the nucleus. The purity of the measured single-particle states clearly illustrates the magic nature of (132)Sn.

Controlled fabrication of nanopores using a direct focused ion beam approach with back face particle detection.

We report a direct, ion drilling technique that enables the reproducible fabrication and placement of nanopores in membranes of different thickness. Using a 30 keV focused Ga ion beam column combined with an in situ, back face, multi-channelplate particle detector, nanopores are sputtered in Si(3)N(4) and W/Si(3)N(4) to have diameters as small as 12 nm. Transmission electron microscopy shows that focused ion beam-drilled holes are near-conical with the diameter decreasing from entry to exit side. By monitoring the detector signal during ion exposure, the drilled hole width can be minimized such that the exit-side diameter is smaller than the full width at half-maximum of the nominally Gaussian-shaped incident beam. Judicious choice of the beam defining aperture combined with back face particle detection allows for reproducible exit-side hole diameters between 18 and 100 nm. The nanopore direct drilling technique does not require potentially damaging broad area exposure to tailor hole sizes. Moreover, this technique successfully achieves breakthrough despite the effects of varying membrane thickness, redeposition, polycrystalline grain structure, and slight ion beam current fluctuations.

Importance of prudent antibiotic use on dairy farms in South Carolina: a pilot project on farmers' knowledge, attitudes and practices.

Inappropriate use of antibiotics in humans and animals contributes to decreased antimicrobial susceptibility in bacteria of medical importance. Resistant bacteria being transferred from animals to humans are causing public health concern. In-person interviews were conducted with 20 dairy farmers in rural counties of South Carolina to determine farmers' knowledge and attitudes about prudent antibiotic use among livestock. Four focus groups (n = 22) were also conducted to ascertain farmers' specific information needs about proper antibiotic use. Survey results showed that participants (100%) typically determined a need for antibiotic treatment using symptom assessment and reported following some form of operating procedures regarding administration of antibiotics. Few farmers (32%) had actual written antibiotic protocols. Preferred information sources about antibiotics were veterinarians (100%) and other dairy farmers (50%). Most farmers (86%) were not concerned that overuse of antibiotics in animals could result in antibiotic resistance among farm workers. Qualitative analysis of focus groups revealed significant barriers to following proper antibiotic procedures including limited finances and lack of time. The need for bilingual educational resources for Hispanic/Latino dairy workers was expressed. Desired formats for educational materials were posters, flowcharts, videos, and seminars. Education of South Carolina dairy farmers by veterinarians and public health professionals on the appropriate use of antibiotics in dairy cattle is needed to ensure antibiotic effectiveness in both animals and humans.

Genetic markers for ancestry are correlated with body composition traits in older African Americans.

UNLABELLED: Individual-specific percent European ancestry was assessed in 1,277 African Americans. We found significant correlations between proportion of European ancestry and several musculoskeletal traits, indicating that admixture mapping may be a useful strategy for locating genes affecting these traits. INTRODUCTION: Genotype data for admixed populations can be used to detect chromosomal regions influencing disease risk if allele frequencies at disease-related loci differ between parental populations. We assessed evidence for differentially distributed alleles affecting bone and body composition traits in African Americans. METHODS: Bone mineral density (BMD) and body composition data were collected for 1,277 African and 1,790 European Americans (aged 70-79). Maximum likelihood methods were used to estimate individual-specific percent European ancestry for African Americans genotyped at 37 ancestry-informative genetic markers. Partial correlations between body composition traits and percent European ancestry were calculated while simultaneously adjusting for the effects of covariates. RESULTS: Percent European ancestry (median = 18.7%) in African Americans was correlated with femoral neck BMD in women (r = -0.18, p < 10(-5)) and trabecular spine BMD in both sexes (r = -0.18, p < 10(-5)) independently of body size, fat, lean mass, and other covariates. Significant associations of European ancestry with appendicular lean mass (r = -0.19, p < 10(-10)), total lean mass (r = -0.12, p < 10(-4)), and total body fat (r = 0.09, p < 0.002) were also observed for both sexes. CONCLUSIONS: These results indicate that some population differences in body composition may be due to population-specific allele frequencies, suggesting the utility of admixture mapping for identifying susceptibility genes for osteoporosis, sarcopenia, and obesity.

Support for involvement of neuregulin 1 in schizophrenia pathophysiology.

Schizophrenia is a common, multigenic psychiatric disorder. Linkage studies, including a recent meta-analysis of genome scans, have repeatedly implicated chromosome 8p12-p23.1 in schizophrenia susceptibility. More recently, significant association with a candidate gene on 8p12, neuregulin 1 (NRG1), has been reported in several European and Chinese samples. We investigated NRG1 for association in schizophrenia patients of Portuguese descent to determine whether this gene is a risk factor in this population. We tested NRG1 markers and haplotypes for association in 111 parent-proband trios, 321 unrelated cases, and 242 control individuals. Associations were found with a haplotype that overlaps the risk haplotype originally reported in the Icelandic population ("Hap(ICE)"), and two haplotypes located in the 3' end of NRG1 (all P<0.05). However, association was not detected with Hap(ICE) itself. Comparison of NRG1 transcript expression in peripheral leukocytes from schizophrenia patients and unaffected siblings identified 3.8-fold higher levels of the SMDF variant in patients (P=0.039). Significant positive correlations (P<0.001) were found between SMDF and HRG-beta 2 expression and between HRG-gamma and ndf43 expression, suggesting common transcriptional regulation of NRG1 variants. In summary, our results suggest that haplotypes across NRG1 and multiple NRG1 variants are involved in schizophrenia.

Clinical characteristics of African Americans vs Caucasian Americans with multiple sclerosis.

BACKGROUND: African American (AA) individuals are thought to develop multiple sclerosis (MS) less frequently than Caucasian American (CA) individuals. OBJECTIVE: To compare the clinical characteristics of AA and CA patients with MS. METHODS: The clinical features of MS were compared in a large retrospective cohort of AA (n = 375) and CA (n = 427) subjects. RESULTS: The proportion of women to men was similar in AA and CA subjects (81% [AA] vs 77% [CA]; p = 0.122). There were no differences in the proportions of subjects with relapsing-remitting, secondary progressive, primary progressive, and progressive relapsing MS. The median time to diagnosis was 1 year after symptom onset in AA subjects and 2 years after symptom onset in CA subjects (p = 0.0013). The age at onset was approximately 2.5 years later in AA than CA subjects (33.7 vs 31.1 years; p = 0.0001). AA subjects presented with multisite signs and symptoms at disease onset more often than CA subjects (p = 0.018). Clinical involvement restricted to the optic nerves and spinal cord (opticospinal MS) occurred in 16.8% of AA patients compared with 7.9% of CA patients (p < 0.001). Transverse myelitis also occurred more frequently in AA subjects (28 vs 18%; p = 0.001). Survival analysis revealed that AA subjects were at higher risk for development of ambulatory disability than CA subjects. After adjusting for baseline variations and differences in therapeutic interventions, AAs were at 1.67-fold greater risk for requiring a cane to ambulate than CA patients (p < 0.001). There was a trend suggesting that AAs were also at greater risk for development of wheelchair dependency (p = 0.099). Adjusted Cox proportional hazard models showed that this effect was in part attributable to the older age at onset in AAs (p < 0.001). CONCLUSIONS: Compared with multiple sclerosis (MS) in Caucasian Americans, African American patients with MS have a greater likelihood of developing opticospinal MS and transverse myelitis and have a more aggressive disease course.

Burst strength of laparoscopic and open hernia repair.

BACKGROUND: There are few reports of overall strength of laparoscopic and open incisional hernia repair. METHODS: After anesthesia, a 2-inch circular defect was made in the abdominal wall of 28 female swine. Gore-Tex DualMesh Biomaterial (W. L. Gore & Associates, Flagstaff, AZ) was used for all repairs. Sixteen animals underwent open repair and 12 underwent laparoscopic repair. Burst strength was detected within 2 weeks and at 6 weeks by euthanizing the animals and insufflating the abdominal cavity with water while measuring the intraabdominal pressure until it could no longer be pressurized. RESULTS: Three events occurred after insufflation: rupture around patch (R), dissection from insufflation or pressure monitoring sites (D), or rectal prolapse (P). Failure after open early repair occurred at 289 (range 219-388) mmHg with 7-R, 1-P and late 289 (196-343) mmHg with 1-R, 6-P. Failure after laparoscopic early repair occurred at 259 (191-388) mmHg with 4-R, 1-P, 1-D and late 291 (140-330) mmHg with 2-R, 1-P, 3-D. Late groups were less likely to rupture. CONCLUSION: Both hernia repairs are durable at early and late periods. Tissue ingrowth adds to repair strength. We could not show that one repair was stronger than the other. Nonetheless, laparoscopic repair tended to degrade by dissection, which was our highest pressure event.

Induction therapy with monoclonal antibodies specific for CD80 and CD86 delays the onset of acute renal allograft rejection in non-human primates.

CD80 and CD86 (also known as B7-1 and B7-2, respectively) are both ligands for the T cell costimulatory receptors CD28 and CD152. Both CD80 and CD86 mediate T cell costimulation, and as such, have been studied for their role in promoting allograft rejection. In this study we demonstrate that administering monoclonal antibodies specific for these B7 ligands can delay the onset of acute renal allograft rejection in rhesus monkeys. The most durable effect results from simultaneous administration of both anti-B7 antibodies. The mechanism of action does not involve global depletion of T or B cells. Despite in vitro and in vivo evidence demonstrating the effectiveness of the anti-B7 antibodies in suppressing T cell responsiveness to alloantigen, their use does not result in durable tolerance. Prolonged therapy with murine anti-B7 antibodies is limited by the development of neutralizing antibodies, but that problem was avoided when humanized anti-B7 reagents are used. Most animals develop rejection and an alloantibody response although still on antibody therapy and before the development of a neutralizing antibody response. Anti-B7 antibody therapy may have use as an adjunctive agent for clinical allotransplantation, but using the dosing regimens we used, is not a tolerizing therapy in this non-human primate model.

Low dose streptozotocin-induced diabetes in rat insulin promoter-mCD80-transgenic mice is T cell autoantigen-specific and CD28 dependent.

Although transgenic mice expressing murine B7-1 (mCD80) on their pancreatic beta cells under the rat insulin-1 promoter (RIP-mCD80(+) mice) rarely develop spontaneous beta cell destruction and diabetes, we have previously reported the transgene-dependent induction of profound insulitis and lethal diabetes following multiple low dose injections of the beta cell toxin streptozotocin (MLDS) in RIP-mCD80(+) mice. Here, we have further characterized this MLDS-induced diabetes model using the RIP-mCD80(+) mice and now demonstrate that disease is critically dependent on T cell signaling via CD28. Thus, although naive RIP-mCD80(+) and nontransgenic littermates have comparable gross beta cell mass, and immediately following MLDS induction the mice display similar degrees of insulitis and decrements in the beta cell mass, only transgenic mice continued to destroy their beta cells and develop insulin-dependent diabetes mellitus. Strikingly, MLDS-induced diabetes was completely prevented in CD28-deficient mice (RIP-mCD80(+)CD28(-/-)) due to abrogation of leukocytes infiltrating their pancreatic islets. We further characterized MLDS-induced diabetes in the RIP-mCD80(+) mice by demonstrating that the MLDS-induced lymphocytic islet infiltrate contained a substantial frequency of autoantigen-specific, IFN-gamma-secreting, CD8(+) T cells. We conclude that MLDS-induced beta cell destruction and subsequent insulin-dependent diabetes mellitus in RIP-mCD80(+) mice is T cell-mediated as it involves both Ag-specific recognition of self-target molecules in the inflamed pancreatic islet (signal 1) and is CD28 costimulation dependent (signal 2).

Expression and properties of diacylglycerol acyltransferase from cell-suspension cultures of oilseed rape.

The expression of diacylglycerol acyltransferase (DGAT, EC 2.3.1.20) with predicted molecular mass of 56.9. kDa (BnDGAT1) was examined using microspore-derived cell suspension cultures of oilseed rape (Brassica napus L. cv Jet Neuf). As well, a recombinant histidine-tagged N-terminal fragment of BnDGAT1 [BnDGAT1((1-116))His(6)], which was relatively hydrophilic, was partially characterized. A temporal increase in DGAT activity occurred within a 24 h period following transfer of cells from 6% (w/v) sucrose to 14% (w/v) sucrose. Western blotting indicated that the abundance of BnDGAT1 protein was closely correlated with DGAT activity. BnDGAT1 mRNA also exhibited a temporal increase within the 24 h period following transfer of cells into higher sucrose concentrations, but the transcript level was not closely associated with DGAT activity as BnDGAT1 protein. The fragment BnDGAT1(1-116)His(6) interacted with [1-(14)C]oleoyl-CoA, suggesting that the N-terminal region of BnDGAT1 may have a role in binding cellular acyl-CoA.

Treatment with humanized monoclonal antibody against CD154 prevents acute renal allograft rejection in nonhuman primates.

CD154 is the ligand for the receptor CD40. This ligand-receptor pair mediates endothelial and antigen-presenting cell activation, and facilitates the interaction of these cells with T cells and platelets. We demonstrate here that administration of a CD154-specific monoclonal antibody (hu5C8) allows for renal allotransplantation in outbred, MHC-mismatched rhesus monkeys without acute rejection. The effect persisted for more than 10 months after therapy termination, and no additional drug was required to achieve extended graft survival. Indeed, the use of tacrolimus or chronic steroids seemed to antagonize the anti-rejection effect. Monkeys treated with antibody against CD154 remained healthy during and after therapy. The mechanism of action does not require global depletion of T or B cells. Long-term survivors lost their mixed lymphocyte reactivity in a donor-specific manner, but still formed donor-specific antibody and generated T cells that infiltrated the grafted organ without any obvious effect on graft function. Thus, therapy with antibody against CD154 is a promising agent for clinical use in human allotransplantation.

Inflammatory cytokines IFN-gamma plus TNF-alpha induce regulated expression of CD80 (B7-1) but not CD86 (B7-2) on murine fibroblasts.

Optimal T cell activation requires signals delivered via both the TCR and the costimulatory receptors. Considerable experimental data now suggest that this costimulatory signal is generated predominantly by CD28 when engaged by its ligands CD80 (B7-1) and/or CD86 (B7-2). Whether T cell activation is controlled in part by regulated CD80 and/or CD86 expression has been incompletely explored. Here, we report that CD80 can be expressed constitutively by murine fibroblasts and up-regulated after treatment with IFN-gamma plus TNF-alpha. CD80 expression and function was confirmed by 1) Northern analysis, 2) specific immunoprecipitation of a approximately 69-kDa surface protein that comigrated with CD80 precipitated from CD80-transfected CHO cells, and 3) two independent assays for costimulation of Ag-specific T cell activation. Taken together, these observations suggest that CD28/CTLA-4 ligands are expressed on a wider variety of tissues than previously suspected and that their expression is dynamically regulated. Consequently, these results might explain previous observations that inflammatory cytokines can result in autoimmune responses.