Effect of Chronic Uremia on the Cell Surface Expression of B7 Family Costimulatory Molecules in an HLA-A2 Transgenic Mouse Model of Chronic Kidney Disease

Uremia due to chronic kidney disease (CKD) in humans is associated with immune dysfunction, increased susceptibility to infections, immune-activation–associated inflammation, and poor responses to vaccines. The pathophysiologic basis of these immune defects is hypothesized to be associated with a wide range of immunologic abnormalities, including an inability to sufficiently express the B7 family (B7-1, CD80; B7-2, CD86) of T-cell costimulatory molecules. However, testing the hypothesis that a state of chronic uremia contributes to attenuated expression of CD80 or CD86 has been difficult because few animal models faithfully recapitulate the immune defects observed in human CKD patients. We used a humanized mouse in a model of surgically induced renal failure and secondary chronic uremia to evaluate the effect of uremia on the expression of these markers. In a manner that resembles the changes observed in CKD patients, surgically induced CKD in mice resulted in decreased costimulatory CD86 expression compared with that in sham-operated controls. Immunodeficiency was functionally demonstrated in this mouse model by documenting an attenuated immune response to a cholera-toxin–based hepatitis B vaccine. This model will be useful for investigating the mechanisms involved in chronic uremia-associated immunodeficiency, poor response to vaccination, and problems associated with immunization of CKD patients.Abbreviations: CKD, chronic kidney disease; CT, cholera toxin; HBsAg, hepatitis B surface antigen; HLA, human leukocyte antigen; Th, T-helper cellThe B7 family of costimulatory molecules is critical in regulating adaptive immune responses and includes B7-1 (CD80) and B7-2 (CD86). These proteins are expressed on lymphoid and nonlymphoid cells and are part of a complex signaling network that includes chemokines, cytokines, adhesion molecules, and other immunoglobulin superfamily members. B7-1 and B7-2 deliver costimulatory or inhibitory signals through interaction with T-cell–associated CD28 or CTLA4 (CD152) molecules.^9,11,18,19 In particular, B7–CD28 ligation is necessary for a robust adaptive immune response to antigen that is presented in context with the MHC and T cell receptor complexes. Without B7 costimulation, T cell activation is compromised and the cell may enter an anergic state.¹³ In addition, active binding of B7, expressed by antigen-presenting cells, to CTLA4 that is expressed on responding T cells inhibits T-cell activation.^12,25 Therefore, any disease that alters the expression or function of B7 proteins has the potential to deregulate adaptive immunity.Uremia secondary to chronic kidney disease (CKD) is a disease that has been associated with reduced B7 expression and defects in innate and adaptive immunity in humans.⁹ Innate and adaptive immune dysfunction in CKD patients is associated with hyperinflammatory conditions, infections, atherosclerosis, and cardiovascular disease.^17,23 Some aspects of this dysfunction include impaired activation of T-lymphocytes, an increased T-helper cell (Th) Th1:Th2 ratio, lymphopenia, and impaired function of antigen-presenting cells.¹⁶ The function of antigen-presenting cells appears compromised in uremic patients through defective expression of B7-2 (CD86) on macrophages and dendritic cells.¹⁰ In contrast to that in human CKD patients, this pathology of reduced B7-2 expression has not previously been investigated in an animal model of uremia, and no model has been identified as having a defect in B7-2 expression.³Animal modeling of CKD is imperative to understand the pathogenesis of the disease. Furthermore, modeling will aid in the design and testing of therapeutic approaches. In the current study we examined the effect of surgically induced chronic uremia on the expression level of B7-1 and B7-2 in spleen cells of humanized B6.Cg-Tg(HLA-A/H2-D)2Enge/J transgenic mice (HLA-A2). We hypothesized that surgical induction of CKD and chronic uremia in HLA-A2 mice would result in the subsequent reduction of expression of B7-2 (that is, increased CD80:CD86 ratio) as compared with that of control mice that underwent sham surgery but did not become uremic. We show that the surgically induced chronic uremia protocol that we established results in a pathologic immune phenotype similar to that reported for immunodeficient humans with CKD, thus suggesting the utility of this approach as a new animal model of CKD.     

Characterization of 30 new microsatellite markers,developed from enriched genomic DNA library of zoysiagrass Zoysia japonica Steud.

The present study reports the development of 30 polymorphic microsatellite markers for zoysiagrass Zoysia japonica Steud. The 30 markers produced a total of 125 alleles with an average of 4.2 alleles per locus. Values for observed and expected heterozygosities ranged from 0.10 to 0.95 and from 0.15 to 0.81, respectively. At significance threshold (P < 0.05), 11 loci deviated from Hardy–Weinberg equilibrium, whereas significant linkage disequilibrium values were observed between 16 pairs of loci. These markers may provide information needed to select genetically diverse parents for developing breeding and mapping populations of zoysiagrass.     

Co-endemicity of Plasmodium falciparum and Intestinal Helminths Infection in School Age Children in Rural Communities of Kwara State Nigeria

Background

Malaria and intestinal helminths co-infection are major public health problems particularly among school age children in Nigeria. However the magnitude and possible interactions of these infections remain poorly understood. This study determined the prevalence, impact and possible interaction of Plasmodium falciparum and intestinal helminths co-infection among school children in rural communities of Kwara State, Nigeria.

Methods

Blood, urine and stool samples were collected from 1017 primary school pupils of ages 4–15 years. Stool samples were processed using both Kato-Katz and formol-ether concentration techniques and microscopically examined for intestinal helminths infection. Urine samples were analyzed using sedimentation method for Schistosoma haematobium. Plasmodium falciparum was confirmed by microscopy using thick and thin blood films methods and packed cell volume (PCV) was determined using hematocrit reader. Univariate analysis and chi-square statistical tests were used to analyze the data.

Results

Overall, 61.2% of all school children had at least an infection of either P. falciparum, S. haematobium, or intestinal helminth. S. haematobium accounted for the largest proportion (44.4%) of a single infection followed by P. falciparum (20.6%). The prevalence of malaria and helminth co-infection in the study was 14.4%. Four species of intestinal helminths were recovered from the stool samples and these were hookworm (22.5%), Hymenolepis species (9.8%), Schistosoma mansoni (2.9%) and Enterobius vermicularis (0.6%). The mean densities of P. falciparum in children co-infected with S. haematobium and hookworm were higher compared to those infected with P. falciparum only though not statistically significant (p = 0.062). The age distribution of both S. haematobium (p = 0.049) and hookworm (p = 0.034) infected children were statistically significant with the older age group (10–15 years) recording the highest prevalence of 47.2% and 25% respectively. Children who were infected with S. haematobium (RR = 1.3) and hookworm (RR = 1.4) have equal chances of being infected with P. falciparum as children with no worm infection. On the other hand children infected with Hymenolepis spp. (p<0.0001) are more likely to be infected with P. falciparum than Hymenolepis spp. uninfected children (RR = 2.0)

Conclusions

These findings suggest that multiple parasitic infections are common in school age children in rural communities of Kwara State Nigeria. The Hymenolepis spp. induced increase susceptibility to P. falciparum could have important consequences on how concurrent infections affect the expression or pathogenesis of these infections.     

Isolation and Transplantation of Different Aged Murine Thymic Grafts.

The mechanisms that regulate the efficacy of thymic selection remain ill-defined. The method presented here allows in vivo analyses of the development and selection of T cells specific for self and foreign antigens. The approach entails implantation of thymic grafts derived from various aged mice into immunodeficient scid recipients. Over a relatively short period of time the recipients are fully reconstituted with T cells derived from the implanted thymus graft. Only thymocytes seeding the thymus at the time of isolation undergo selection and develop into mature T cells. As such, changes in the nature and specificity of the engrafted T cells as a function of age-dependent thymic events can be assessed. Although technical expertise is required for successful thymic transplantation, this method provides a unique strategy to study in vivo a wide range of pathologies that are due to or a result of aberrant thymic function and/or homeostasis.     

RESCRIPt: Reproducible sequence taxonomy reference database management

Nucleotide sequence and taxonomy reference databases are critical resources for widespread applications including marker-gene and metagenome sequencing for microbiome analysis, diet metabarcoding, and environmental DNA (eDNA) surveys. Reproducibly generating, managing, using, and evaluating nucleotide sequence and taxonomy reference databases creates a significant bottleneck for researchers aiming to generate custom sequence databases. Furthermore, database composition drastically influences results, and lack of standardization limits cross-study comparisons. To address these challenges, we developed RESCRIPt, a Python 3 software package and QIIME 2 plugin for reproducible generation and management of reference sequence taxonomy databases, including dedicated functions that streamline creating databases from popular sources, and functions for evaluating, comparing, and interactively exploring qualitative and quantitative characteristics across reference databases. To highlight the breadth and capabilities of RESCRIPt, we provide several examples for working with popular databases for microbiome profiling (SILVA, Greengenes, NCBI-RefSeq, GTDB), eDNA and diet metabarcoding surveys (BOLD, GenBank), as well as for genome comparison. We show that bigger is not always better, and reference databases with standardized taxonomies and those that focus on type strains have quantitative advantages, though may not be appropriate for all use cases. Most databases appear to benefit from some curation (quality filtering), though sequence clustering appears detrimental to database quality. Finally, we demonstrate the breadth and extensibility of RESCRIPt for reproducible workflows with a comparison of global hepatitis genomes. RESCRIPt provides tools to democratize the process of reference database acquisition and management, enabling researchers to reproducibly and transparently create reference materials for diverse research applications. RESCRIPt is released under a permissive BSD-3 license at https://github.com/bokulich-lab/RESCRIPt.     

Evaluation of automated pipelines for tree and plot metric estimation from TLS data in tropical forest areas

Background and AimsTerrestrial LiDAR scanning (TLS) data are of great interest in forest ecology and management because they provide detailed 3-D information on tree structure. Automated pipelines are increasingly used to process TLS data and extract various tree- and plot-level metrics. With these developments comes the risk of unknown reliability due to an absence of systematic output control. In the present study, we evaluated the estimation errors of various metrics, such as wood volume, at tree and plot levels for four automated pipelines.MethodsWe used TLS data collected from a 1-ha plot of tropical forest, from which 391 trees >10 cm in diameter were fully processed using human assistance to obtain control data for tree- and plot-level metrics.Key ResultsOur results showed that fully automated pipelines led to median relative errors in the quantitative structural model (QSM) volume ranging from 39 to 115 % at the tree level and 10 to 134 % at the 1-ha plot level. For tree-level metrics, the median error for the crown-projected area ranged from 46 to 59 % and that for the crown-hull volume varied from 72 to 88 %. This result suggests that the tree isolation step is the weak link in automated pipeline methods. We further analysed how human assistance with automated pipelines can help reduce the error in the final QSM volume. At the tree scale, we found that isolating trees using human assistance reduced the error in wood volume by a factor of 10. At the 1-ha plot scale, locating trees with human assistance reduced the error by a factor of 3.ConclusionsOur results suggest that in complex tropical forests, fully automated pipelines may provide relatively unreliable metrics at the tree and plot levels, but limited human assistance inputs can significantly reduce errors.     

A new human prostate carcinoma cell line, 22Rv1

Summary A cell line has been derived from a human prostatic carcinoma xenograft, CWR22R. This represents one of very few available cell lines representative of this disease. The cell line is derived from a xenograft that was serially propagated in mice after castration-induced regression and relapse of the parental, androgen-dependent CWR22 xenograft. Flow cytometric and cytogenetic analysis showed that this cell line represents one hyper DNA-diploid stem line with two clonal, evolved cytogenetic sublines. The basic karyotype is close to that of the grandparent xenograft, CWR22, and is relatively simple with 50 chromosomes. In nude mice, the line forms tumors with morphology similar to that of the xenografts, and like the parental CWR22 and CWR22R xenografts, this cell line expresses prostate specific antigen. Growth is weakly stimulated by dihydroxytestosterone and lysates are immunoreactive with androgen receptor antibody by Western blot analysis. Growth is stimulated by epidermal growth factor but is not inhibited by transforming growth factor-β1.