Characterisation of transgenic pigs expressing a human T cell-depleting anti-CD2 monoclonal antibody.

BACKGROUND: Pig islet xenotransplantation is a potential treatment for type 1 diabetes. We have shown that maintenance immunosuppression is required to protect genetically modified (GM) porcine islet xenografts from T cell-mediated rejection in baboons. Local expression of a depleting anti-CD2 monoclonal antibody (mAb) by the xenograft may provide an alternative solution. We have previously reported the generation of GGTA1 knock-in transgenic pigs expressing the chimeric anti-CD2 mAb diliximab under an MHC class I promoter (MHCIP). In this study, we generated GGTA1 knock-in pigs in which MHCIP was replaced by the ß-cell-specific porcine insulin promoter (PIP), and compared the pattern of diliximab expression in the two lines. METHODS: A PIP-diliximab knock-in construct was prepared and validated by transfection of NIT-1 mouse insulinoma cells. The construct was knocked into GGTA1 in wild type (WT) porcine fetal fibroblasts using CRISPR, and knock-in cells were used to generate pigs by somatic cell nuclear transfer (SCNT). Expression of the transgene in MHCIP-diliximab and PIP-diliximab knock-in pigs was characterised at the mRNA and protein levels using RT-qPCR, flow cytometry, ELISA and immunohistochemistry. Islets from MHCIP-diliximab and control GGTA1 KO neonatal pigs were transplanted under the kidney capsule of streptozotocin-diabetic SCID mice. RESULTS: NIT-1 cells stably transfected with the PIP-diliximab knock-in construct secreted diliximab into the culture supernatant, confirming correct expression and processing of the mAb in ß cells. PIP-diliximab knock-in pigs showed a precise integration of the transgene within GGTA1. Diliximab mRNA was detected in all tissues tested (spleen, kidney, heart, liver, lung, pancreas) in MHCIP-diliximab pigs, but was not detectable in PIP-diliximab pigs. Likewise, diliximab was present in the serum of MHCIP-diliximab pigs, at a mean concentration of 1.8 µg/mL, but was not detected in PIP-diliximab pig serum. An immunohistochemical survey revealed staining for diliximab in all organs of MHCIP-diliximab pigs but not of PIP-diliximab pigs. Whole genome sequencing (WGS) of a PIP-diliximab pig identified a missense mutation in the coding region for the dixilimab light chain. This mutation was also found to be present in the fibroblast knock-in clone used to generate the PIP-diliximab pigs. Islet xenografts from neonatal MHCIP-diliximab pigs restored normoglycemia in diabetic immunodeficient mice, indicating no overt effect of the transgene on islet function, and demonstrated expression of diliximab in situ. CONCLUSION: Diliximab was widely expressed in MHCIP-diliximab pigs, including in islets, consistent with the endogenous expression pattern of MHC class I. Further investigation is required to determine whether the level of expression in islets from the MHCIP-diliximab pigs is sufficient to prevent T cell-mediated islet xenograft rejection. The unexpected absence of diliximab expression in the islets of PIP-diliximab pigs was probably due to a mutation in the transgene arising during the generation of the knock-in cells used for SCNT.

Suptavumab for the Prevention of Medically Attended Respiratory Syncytial Virus Infection in Preterm Infants.

BACKGROUND: Respiratory syncytial virus (RSV) is a major cause of childhood medically attended respiratory infection (MARI). METHODS: We conducted a randomized, double-blind, placebo-controlled phase 3 trial in 1154 preterm infants of 1 or 2 doses of suptavumab, a human monoclonal antibody that can bind and block a conserved epitope on RSV A and B subtypes, for the prevention of RSV MARI. The primary endpoint was proportion of subjects with RSV-confirmed hospitalizations or outpatient lower respiratory tract infection (LRTI). RESULTS: There were no significant differences between primary endpoint rates (8.1%, placebo; 7.7%, 1-dose; 9.3%, 2-dose). Suptavumab prevented RSV A infections (relative risks, .38; 95% confidence interval [CI], .14-1.05 in the 1-dose group and .39 [95% CI, .14-1.07] in the 2-dose group; nominal significance of combined suptavumab group vs placebo; P = .0499), while increasing the rate of RSV B infections (relative risk 1.36 [95% CI, .73-2.56] in the 1-dose group and 1.69 [95% CI, .92-3.08] in the 2-dose group; nominal significance of combined suptavumab group vs placebo; P = .12). Sequenced RSV isolates demonstrated no suptavumab epitope changes in RSV A isolates, while all RSV B isolates had 2-amino acid substitution in the suptavumab epitope that led to loss of neutralization activity. Treatment emergent adverse events were balanced across treatment groups. CONCLUSIONS: Suptavumab did not reduce overall RSV hospitalizations or outpatient LRTI because of a newly circulating mutant strain of RSV B. Genetic variation in circulating RSV strains will continue to challenge prevention efforts. CLINICAL TRIALS REGISTRATION: NCT02325791.

A Protein-Truncating HSD17B13 Variant and Protection from Chronic Liver Disease.

BACKGROUND: Elucidation of the genetic factors underlying chronic liver disease may reveal new therapeutic targets. METHODS: We used exome sequence data and electronic health records from 46,544 participants in the DiscovEHR human genetics study to identify genetic variants associated with serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST). Variants that were replicated in three additional cohorts (12,527 persons) were evaluated for association with clinical diagnoses of chronic liver disease in DiscovEHR study participants and two independent cohorts (total of 37,173 persons) and with histopathological severity of liver disease in 2391 human liver samples. RESULTS: A splice variant (rs72613567:TA) in HSD17B13, encoding the hepatic lipid droplet protein hydroxysteroid 17-beta dehydrogenase 13, was associated with reduced levels of ALT (P=4.2×10-12) and AST (P=6.2×10-10). Among DiscovEHR study participants, this variant was associated with a reduced risk of alcoholic liver disease (by 42% [95% confidence interval {CI}, 20 to 58] among heterozygotes and by 53% [95% CI, 3 to 77] among homozygotes), nonalcoholic liver disease (by 17% [95% CI, 8 to 25] among heterozygotes and by 30% [95% CI, 13 to 43] among homozygotes), alcoholic cirrhosis (by 42% [95% CI, 14 to 61] among heterozygotes and by 73% [95% CI, 15 to 91] among homozygotes), and nonalcoholic cirrhosis (by 26% [95% CI, 7 to 40] among heterozygotes and by 49% [95% CI, 15 to 69] among homozygotes). Associations were confirmed in two independent cohorts. The rs72613567:TA variant was associated with a reduced risk of nonalcoholic steatohepatitis, but not steatosis, in human liver samples. The rs72613567:TA variant mitigated liver injury associated with the risk-increasing PNPLA3 p.I148M allele and resulted in an unstable and truncated protein with reduced enzymatic activity. CONCLUSIONS: A loss-of-function variant in HSD17B13 was associated with a reduced risk of chronic liver disease and of progression from steatosis to steatohepatitis. (Funded by Regeneron Pharmaceuticals and others.).

Neonatal immune activation depletes the ovarian follicle reserve and alters ovarian acute inflammatory mediators in neonatal rats.

Normal ovarian development is crucial for female reproductive success and longevity. Interruptions to the delicate process of initial folliculogenesis may lead to ovarian dysfunction. We have previously demonstrated that an early life immune challenge in the rat, induced by administration of lipopolysaccharide (LPS) on postnatal day (PND) 3 and 5, depletes ovarian follicle reserve long term. Here, we hypothesized that this neonatal immune challenge leads to an increase in peripheral and ovarian inflammatory signaling, contributing to an acute depletion of ovarian follicles. Morphological analysis of neonatal ovaries indicated that LPS administration significantly depleted PND 5 primordial follicle populations and accelerated follicle maturation. LPS exposure upregulated circulating interleukin 6, tumor necrosis factor alpha (TNFa), and C-reactive protein on PND 5, and upregulated ovarian mRNA expression of Tnfa, mitogen-activated protein kinase 8 (Mapk8/Jnk1), and growth differentiation factor 9 (Gdf9) (P < 0.05). Mass spectrometry and cell signaling pathway analysis indicated upregulation of cellular pathways associated with acute phase signaling, and cellular survival and assembly. Apoptosis assessed by terminal deoxynucleotidyl transferase dUTP nick end labeling indicated significantly increased positive staining in the ovaries of LPS-treated neonates. These findings suggest that increased proinflammatory signaling within the neonatal ovary may be responsible for the LPS-induced depletion of the primordial follicle pool. These findings also have implications for female reproductive health, as the ovarian reserve is a major determinate of female reproductive longevity.

Replicating Evidence-Based Practices with Flexibility for Perinatal Home Visiting by Paraprofessionals.

Introduction Strategies are needed to improve the efficacy of paraprofessional home visitors for pregnant women in the United States. This study evaluates the maternal and child outcomes when evidence-based practices (EBP) are replicated with flexibility, rather than fidelity to a manualized intervention. Methods Pregnant mothers (N = 203) in five clinics were recruited in the waiting rooms and randomized to standard clinic care as the control condition (n = 104) or standard care plus home visiting (n = 99). Home visitors (n = 9) were selected, trained in foundational skills common to EBP and four problem domains (weight control, breastfeeding, daily habits, and depression). Independent interviewers assessed targeted outcomes at birth (82%) and 6 months later (83%). RESULTS: Home visitors, called Mentor Mothers [MM], made an average of 14.9 home visits or telephone contacts (SD = 9; total contacts = 1491) addressing maternal daily habits, breastfeeding, and depression. Intervention and control mothers were similar in weight, Body Mass Index (BMI), depression and social support at baseline and 6 months later. The percentage of low birth weight babies was similar; intervention infants' growth (weight/height Z score) tended to be significantly better compared to the control condition. DISCUSSION: There are many explanations for the failure to find significant benefits: insufficient statistical power; the benefits of repeated assessments by warm, supportive peers to improve outcomes; or the failure of EBP and the need to maintain replication with fidelity. All study mothers had better outcomes than documented among comparable published samples of low-income, Latina and Korean-American mothers in Los Angeles, CA. ClinicalTrials.gov registration NCT01687634.

Adapting Current Strategies to Implement Evidence-Based Prevention Programs for Paraprofessional Home Visiting.

This paper describes a strategy for using evidence-based interventions (EBI) that does not require replication and fidelity. Eight parents, identified as positive role models, conducted home visits for 101 low-income Latina and Korean pregnant women. The home visitors, called mentor mothers (MM), were trained in 10 of the practice elements common to 80% of child-focused EBI and how to apply these skills to support mothers in obesity prevention, to increase the duration of breastfeeding, and to reduce depression. MM reported the content and skills utilized on each home visit on mobile phones. Each MM made an average of 153 home visits (SD = 173.3), with 28 of these visits being phone contacts. Body mass index (BMI) at 6 months was significantly associated with the frequency of MM focused on coping with depression (r = .24), but was not related to practice elements used by MM. The duration of breastfeeding was significantly related to the frequency with which MM focused on the topic of breastfeeding (r = .28) and parenting (r = .3), and MM use of attending (r = .24) and relaxation (r = .27). Depression was significantly correlated with the frequency of addressing depression (r = .27), but not to practice elements. MM did use different strategies in the first 150 visits compared to their last 150 visits, reflecting data-informed supervision. Evidence synthesized from EBI was used as a novel training method, with real-time monitoring and data-informed supervision providing evidence of iterative quality improvements in MM behaviors over time, as well as a way for linking implementation processes to outcomes.

Factors in Early-Life Programming of Reproductive Fitness.

Fertility rates have been declining worldwide, with a growing number of young women suffering from infertility. Infectious and inflammatory diseases are important causes of infertility, and recent evidence points to the critical role of the early-life microbial environment in developmental programming of adult reproductive fitness. Our laboratory and others have demonstrated that acute exposure to an immunological challenge early in life has a profound and prolonged impact on male and female reproductive development. This review presents evidence that perinatal exposure to immunological challenge by a bacterial endotoxin, lipopolysaccharide, acts at all levels of the hypothalamic-pituitary-gonadal axis, resulting in long-lasting changes in reproductive function, suggesting that disposition to infertility may begin early in life.

Optical design considerations for efficient light collection from liquid scintillation counters.

Liquid scintillation counters measure charged particle-emitting radioactive isotopes and are used for environmental studies, nuclear chemistry, and life science. Alpha and beta emissions arising from the material under study interact with the scintillation cocktail to produce light. The prototypical liquid scintillation counter employs low-level photon-counting detectors to measure the arrival of the scintillation. For reliable operation, the counting instrument must convey the scintillation light to the detectors efficiently and predictably. Current best practices employ the use of two or more detectors for coincidence processing to discriminate true scintillation events from background events due to instrumental effects such as photomultiplier tube dark rates, tube flashing, or other light emission not generated in the scintillation cocktail vial. In low-background liquid scintillation counters, additional attention is paid to shielding the scintillation cocktail from naturally occurring radioactive material present in the laboratory and within the instrument's construction materials. Low-background design is generally at odds with optimal light collection. This study presents the evolution of a light collection design for liquid scintillation counting (LSC) in a low-background shield. The basic approach to achieve both good light collection and a low-background measurement is described. The baseline signals arising from the scintillation vial are modeled and methods to efficiently collect scintillation light are presented as part of the development of a customized low-background, high-sensitivity LSC system.

Concurrent, Convergent, and Discriminant Validity of the DSM-5 Section III Psychopathy Specifier.

Section III of the fifth iteration of the Diagnostic and Statistical Manual of Mental Disorders (DSM-5) includes an alternative model of personality disorder diagnosis that conceptualizes antisocial personality disorder as an interpersonal, rather than behavioral, construct. However, the diagnostic specifier for psychopathy has been met with recent controversy due to its conceptual and empirical overlap with triarchic boldness, which has been debated as a necessary and sufficient domain of psychopathy. This study examined the concurrent, convergent, and discriminant validity of the specifier using canonical correlation analysis in samples of undergraduate students (N = 224) and community adults with prior criminal involvement (N = 306). Findings highlight the specifier as a multidimensional construct with divergent associations across its three facets. There was limited validity evidence for two of the three facets, raising concerns regarding the clinical utility of the psychopathy specifier.

Making the connection: randomized controlled trial of social skills at school for children with autism spectrum disorders.

BACKGROUND: This study compared two interventions for improving the social skills of high functioning children with autism spectrum disorders in general education classrooms. One intervention involved a peer-mediated approach (PEER) and the other involved a child-assisted approach (CHILD). METHOD: The two interventions were crossed in a 2 × 2 factorial design yielding control, PEER, CHILD, and both PEER and CHILD conditions. Sixty children participated from 56 classrooms in 30 schools. Interventions involved 12 sessions over 6 weeks, with a 3-month follow-up. Outcome measures included self, peer and teacher reports of social skills and independent weekly observations of children on their school playground over the course of the intervention. RESULTS: Significant improvements were found in social network salience, number of friendship nominations, teacher report of social skills in the classroom, and decreased isolation on the playground for children who received PEER interventions. Changes obtained at the end of the treatment persisted to the 3-month follow-up. CONCLUSIONS: These data suggest that significant improvements can be made in peer social connections for children with autism spectrum disorders in general education classrooms with a brief intervention, and that these gains persist over time.

Increasing the Utility of the Comprehensive Assessment of Psychopathic Personality-Lexical Rating Scale (CAPP-LRS): Instrument Adaptation and Simplification.

The Comprehensive Assessment of Psychopathic Personality-Lexical Rating Scale (CAPP-LRS) is a self-report instrument designed to index psychopathy according to the CAPP psychopathy framework. Developed with the expressed goal of advancing the state of knowledge regarding the specific features of psychopathy, the CAPP model and associated instruments have garnered increasing attention and support in the field. Despite the conceptual strength of the CAPP model, the advanced lexical structure of its primary research tool (the CAPP-LRS) has led researchers to question the utility of the instrument for use with some populations of interest (e.g., forensic/correctional and adolescent/young adult samples). The aim of the present work was to address this issue by creating a lexically simplified, though functionally equivalent, version of the CAPP-LRS to increase accessibility to critically relevant populations. A set of two studies (N = 602) describes the adaptation protocol and the initial validation of the modified instrument.

A tale of two plaques: convergent mechanisms of T-cell-mediated inflammation in psoriasis and atherosclerosis.

Psoriasis and atherosclerosis are diseases in which effector T lymphocytes such as Helper T cells type 1 (Th1) and 17 (Th17) play integral roles in disease pathogenesis and progression. Regulatory T cells (Treg) also exert clinically important anti-inflammatory effects that are pathologically altered in psoriasis and atherosclerosis. We review the immunological pathways involving Th1, Th17 and Treg cells that are common to psoriasis and atherosclerosis. These shared pathways provide the basis for mechanisms that may explain the epidemiologic observation that patients with psoriasis have an increased risk of heart disease. Improved understanding of these pathways will guide future experiments and may lead to the development of therapeutics that prevent or treat cardiovascular complications in patients with psoriasis.

Content Validation of the Comprehensive Assessment of Psychopathic Personality-Basic Version (CAPP-Basic) Using Prototypical Analysis.

The Comprehensive Assessment of Psychopathic Personality (CAPP) is a lexically based conceptual model of psychopathy. Despite widespread use in research settings, the Lexical Rating Scale for the CAPP (CAPP-LRS) requires an advanced reading level for completion. The present study is the first to evaluate the content validity of the CAPP-Basic, a lexically simplified version of the CAPP-LRS designed for use with individuals possessing lower verbal skills. Symptoms were rated by mental health professionals (N = 121). Findings were largely in line with results of previous studies, indicating good content validity of the CAPP-Basic at the domain and symptom level.

Construct Validity of the Comprehensive Assessment of Psychopathic Personality (CAPP): Examining the Internal Structure and Generalizability of CAPP Self-Ratings Across Gender and Ethnicity.

The Comprehensive Assessment of Psychopathic Personality (CAPP) is a recently developed conceptual model of psychopathy designed to index the disorder across 33 personality traits. Although recent research has evidenced support for the CAPP model with respect to the convergent, criterion, and predictive validity of CAPP instruments, little work has examined the optimal internal structure and generalizability of the model and associated measures. The present study sought to elaborate on the construct validity and psychometric properties of the CAPP Lexical Self Rating Scale and determine the utility of the instrument across men and women, and individuals of Caucasian and East Asian descent. Within a large sample of self-identified offenders (N = 1,414), we found strong support for a three-factor model comprising domains reflecting interpersonal dominance, behavioral disinhibition, and deficient emotional attachment. Analyses examining the generalizability of the model provide further insight into its applicability for diverse populations.

Do Community Structural Characteristics Moderate the Association Between Mental Health and the Frequency and Severity of Violent-Behavioral Outcomes in Community Respondents?

In this prospective study, we examined the association between three types of mental health symptom clusters (i.e., psychotic, internalizing, and externalizing) and the frequency and severity of violent-behavioral outcomes, and whether community disadvantage, residential instability, and criminogenic facility density moderated these associations. Study data were derived from 258 community-dwelling adults nested in 60 postal forward sortation areas (FSAs) in a large metropolitan area in Western Canada who were assessed twice over a 6-month period. In addition, census and administrative data were obtained on the same areas. Controlling for sociodemographic characteristics (i.e., age, gender, ethnicity, relationship status, and employment status), lifetime history of violent-behavioral outcomes, and community structural characteristics, internalizing and externalizing mental health symptoms were significantly positively associated with the frequency and severity of subsequent violence perpetration and with the severity of subsequent violent victimization. Several significant interactions were observed: internalizing symptoms increased the risk of frequent and severe violence perpetration in FSAs with high but not low disadvantage, and externalizing symptoms increased the risk of frequent violent victimization in FSAs with a high but not low criminogenic facility density. Only the interactive association of internalizing symptoms and community disadvantage with the severity of violence perpetration, however, remained significant after Bonferroni correction was applied. These findings provide tentative support that associations between mental health and violent-behavioral outcomes can vary with community context. The implication of these findings for assessing and managing violent-behavioral outcomes in the community is discussed.

Maternal depressed mood and child development over the first five years of life in South Africa.

BACKGROUND: While the negative impact of peri-natal depression is well-documented in high-income countries, the long-term effects across the life course in low and middle-income countries is less clear. Children's adjustment over the first five years is examined as a function of patterns of maternal depressed mood. METHODS: Pregnant women in 24 peri-urban townships (N = 1,238) were randomized to a home-visiting intervention or standard care and reassessed five times, with high retention. There were no intervention effects on children past 18 months. Multilevel regression models examined the impact of depressed mood on child outcomes. Using the Edinburgh Postnatal Depression Scale, four patterns of maternal depressed mood were identified: never (40.6%); antenatal (13.0%); early childhood (26.1%); and recurrent episodes of depressed mood (20.3 %). FINDINGS: Mothers' patterns of depressive symptoms and child outcomes were similar, regardless of intervention. Never depressed mothers were significantly younger, had higher income, less food insecurity, were more likely to have electricity, be living with HIV or have an HIV positive partner, and had fewer problems with alcohol than depressed mothers. Children of mothers who experienced depressed mood weighed less, were more aggressive, and were hospitalized more often than children of never depressed mothers, but were similar in cognitive and social development. INTERPRETATIONS: Depressed mood, has significant negative impacts on South African children's growth and aggressive behavior. The timing of maternal depressed mood was less important than never having a depressed mood or a recurrent depressed mood. FUNDING: There were no funding conflicts in executing this trial.

Social involvement of children with autism spectrum disorders in elementary school classrooms.

BACKGROUND: Children with autism spectrum disorders (ASD) are increasingly included in general education classrooms in an effort to improve their social involvement. METHODS: Seventy-nine children with ASD and 79 randomly selected, gender-matched peers (88.6% male) in 75 early (K-1), middle (2nd-3rd), and late (4th-5th) elementary classrooms across 30 schools completed social network surveys examining each child's reciprocal friendships, peer rejection, acceptance, and social involvement. RESULTS: Across grade levels, peers less frequently reciprocated friendships with children with ASD than students in the matched sample. While children with ASD were not more likely to be rejected by peers, they were less accepted and had fewer reciprocal friendships than matched peers at each grade level. Although 48.1% of children with ASD were involved in the social networks of their classrooms, children with ASD were more likely to be isolated or peripheral to social relationships within the classroom across all grade levels, and this difference is even more dramatic in later elementary grades. CONCLUSIONS: In inclusive classrooms, children with ASD are only involved in peers' social relationships about half of the time, and appear to be even less connected with increasing grade level. Promoting children with ASD's skills in popular activities to share with peers in early childhood may be a key preventive intervention to protect social relationships in late elementary school grades.