Integrated Behavioral Health Prevention for Infants in Pediatric Primary Care: A Mixed-Methods Pilot Study.

OBJECTIVE: Pediatric primary care is a promising setting in which to deliver preventive behavioral health services to young children and their families. Integrated behavioral health care models typically emphasize treatment rather than prevention. This pilot study examined the efficacy of an integrated behavioral health preventive (IBH-P) intervention delivered by psychologists and focused on supporting parenting in low-income mothers of infants as part of well-child visits in the first 6 months of life. METHODS: Using a mixed-methods approach that included a pilot randomized clinical trial and post-intervention qualitative interviews, 137 mothers were randomly assigned to receive IBH-P or usual care. Self-report measures of parenting, child behavior, and stress were obtained at pre- and/or post-intervention. Direct observation of mother-infant interactions was conducted at post-intervention. RESULTS: No differences between groups were found on maternal attunement, knowledge of child development, nurturing parenting, or infant behavior. A secondary analysis on a subsample with no prior exposure to IBH-P with older siblings found that mothers in IBH-P reported increased self-efficacy relative to controls. In the qualitative interviews, mothers stated that they valued IBH-P, learning about their baby, liked the integration in primary care, and felt respected and comfortable with their provider. CONCLUSIONS: Findings are discussed in terms of the next steps in refining IBH-P approaches to prevention in primary care.

Modular Catalysis: Aptamer Enhancement of Enzyme Kinetics in a Nanoparticle Reactor.

Enzyme activity requires sequential binding and chemical transformation of substrates. While directed evolution and random mutagenesis are common methods for improving catalytic activity, these methods do not allow for independent control of KM and kcat. To achieve such control, we envisioned that the colocalization of aptamers and enzymes that act on the same molecule could increase catalytic efficiency through preconcentration of substrate. We explored this concept with cocaine esterase and anticocaine aptamers having varying KD values, both encapsulated in MS2 virus-like particles. Rate enhancements were observed with magnitudes dependent on both aptamer:enzyme stoichiometry and aptamer KD, peaking when aptamer KD and enzyme KM were roughly equivalent. This beneficial effect was lost when either aptamer binding was too tight or the aptamers were not constrained to be close to the catalyst. This work demonstrates a modular way to enhance catalysis by independently controlling substrate capture and release to the processing enzyme.

A Pilot Randomized Trial of an Obesity Prevention Program for High-Risk Infants in Primary Care.

OBJECTIVE: Rapid infant weight gain is associated with later obesity. Novel interventions to prevent rapid infant weight gain that are accessible to infants and families are needed, especially for those at the highest risk. Our aims were to examine: (a) feasibility and acceptability of a responsive parenting intervention delivered via Integrated Behavioral Health (IBH) in pediatric primary care and (b) preliminary effects on infant weight gain from birth to 6 (post-treatment) and 9 (follow-up) months. METHODS: A parallel design, proof-of-concept randomized control trial was conducted with 65 mother-infant dyads (32 randomized to intervention, 33 randomized an IBH attention control focused on promoting healthy mental health), in which the majority identify as Black (80%) and low income (91% receiving Medicaid). Participants and assessors were masked to treatment condition. Outcomes included feasibility (enrollment), acceptability (retention and adherence), and conditional weight gain (CWG), an indicator of rapid weight gain. RESULTS: The intervention was feasible (90% of eligible families enrolled) and acceptable (89% of families retained), with 81% receiving ≥3 of 4 treatment sessions. A medium effect was found on CWG (d = -0.54 post-treatment, d = -0.57 follow-up), with the infants in the treatment group showing significantly lower CWG (mean = -0.27, 95% CI, -0.63, 0.09) compared to the control group (mean = 0.29, 95% CI, -0.17, 0.76) at 9 months (p = .04). CONCLUSIONS: This study demonstrates the feasibility of implementing a responsive parenting obesity prevention intervention within primary care. Delivery in pediatric primary care is advantageous for implementation and reaching at-risk populations. The preliminary effects on CWG are promising and support testing in a larger trial.

Combating small molecule environmental contaminants: detection and sequestration using functional nucleic acids.

Small molecule contaminants pose a significant threat to the environment and human health. While regulations are in place for allowed limits in many countries, detection and remediation of contaminants in more resource-limited settings and everyday environmental sources remains a challenge. Functional nucleic acids, including aptamers and DNA enzymes, have emerged as powerful options for addressing this challenge due to their ability to non-covalently interact with small molecule targets. The goal of this perspective is to outline recent efforts toward the selection of aptamers for small molecules and describe their subsequent implementation for environmental applications. Finally, we provide an outlook that addresses barriers that hinder these technologies from being widely adopted in field friendly settings and propose a path forward toward addressing these challenges.

Systematically Modulating Aptamer Affinity and Specificity by Guanosine-to-Inosine Substitution.

Aptamers are widely used in small molecule detection applications due to their specificity, stability, and cost effectiveness. One key challenge in utilizing aptamers in sensors is matching the binding affinity of the aptamer to the desired concentration range for analyte detection. The most common methods for modulating affinity have inherent limitations, such as the likelihood of drastic changes in aptamer folding. Here, we propose that substituting guanosine for inosine at specific locations in the aptamer sequence provides a less perturbative approach to modulating affinity. Inosine is a naturally occurring nucleotide that results from hydrolytic deamination of adenosine, and like guanine, it base pairs with cytosine. Using the well-studied cocaine binding aptamer, we systematically replaced guanosine with inosine and were able to generate sequences having a range of binding affinities from 230 nM to 80 µM. Interestingly, we found that these substitutions could also modulate the specificity of the aptamers, leading to a range of binding affinities for structurally related analytes. Analysis of folding stability via melting temperature shows that, as expected, aptamer structure is impacted by guanosine-to-inosine substitutions. The ability to tune binding affinity and specificity through guanosine-to-inosine substitution provides a convenient and reliable approach for rapidly generating aptamers for diverse biosensing applications.

Thermoreversible Control of Nucleic Acid Structure and Function with Glyoxal Caging.

Controlling the structure and activity of nucleic acids dramatically expands their potential for application in therapeutics, biosensing, nanotechnology, and biocomputing. Several methods have been developed to impart responsiveness of DNA and RNA to small-molecule and light-based stimuli. However, heat-triggered control of nucleic acids has remained largely unexplored, leaving a significant gap in responsive nucleic acid technology. Moreover, current technologies have been limited to natural nucleic acids and are often incompatible with polymerase-generated sequences. Here we show that glyoxal, a well-characterized compound that covalently attaches to the Watson-Crick-Franklin face of several nucleobases, addresses these limitations by thermoreversibly modulating the structure and activity of virtually any nucleic acid scaffold. Using a variety of DNA and RNA constructs, we demonstrate that glyoxal modification is easily installed and potently disrupts nucleic acid structure and function. We also characterize the kinetics of decaging and show that activity can be restored via tunable thermal removal of glyoxal adducts under a variety of conditions. We further illustrate the versatility of this approach by reversibly caging a 2'-O-methylated RNA aptamer as well as synthetic threose nucleic acid (TNA) and peptide nucleic acid (PNA) scaffolds. Glyoxal caging can also be used to reversibly disrupt enzyme-nucleic acid interactions, and we show that caging of guide RNA allows for tunable and reversible control over CRISPR-Cas9 activity. We also demonstrate glyoxal caging as an effective method for enhancing PCR specificity, and we cage a biostable antisense oligonucleotide for time-release activation and titration of gene expression in living cells. Together, glyoxalation is a straightforward and scarless method for imparting reversible thermal responsiveness to theoretically any nucleic acid architecture, addressing a significant need in synthetic biology and offering a versatile new tool for constructing programmable nucleic acid components in medicine, nanotechnology, and biocomputing.

CD56bright NK cells exhibit potent antitumor responses following IL-15 priming.

NK cells, lymphocytes of the innate immune system, are important for defense against infectious pathogens and cancer. Classically, the CD56dim NK cell subset is thought to mediate antitumor responses, whereas the CD56bright subset is involved in immunomodulation. Here, we challenge this paradigm by demonstrating that brief priming with IL-15 markedly enhanced the antitumor response of CD56bright NK cells. Priming improved multiple CD56bright cell functions: degranulation, cytotoxicity, and cytokine production. Primed CD56bright cells from leukemia patients demonstrated enhanced responses to autologous blasts in vitro, and primed CD56bright cells controlled leukemia cells in vivo in a murine xenograft model. Primed CD56bright cells from multiple myeloma (MM) patients displayed superior responses to autologous myeloma targets, and furthermore, CD56bright NK cells from MM patients primed with the IL-15 receptor agonist ALT-803 in vivo displayed enhanced ex vivo functional responses to MM targets. Effector mechanisms contributing to IL-15-based priming included improved cytotoxic protein expression, target cell conjugation, and LFA-1-, CD2-, and NKG2D-dependent activation of NK cells. Finally, IL-15 robustly stimulated the PI3K/Akt/mTOR and MEK/ERK pathways in CD56bright compared with CD56dim NK cells, and blockade of these pathways attenuated antitumor responses. These findings identify CD56bright NK cells as potent antitumor effectors that warrant further investigation as a cancer immunotherapy.

Cytokine-Induced Memory-Like Differentiation Enhances Unlicensed Natural Killer Cell Antileukemia and FcγRIIIa-Triggered Responses.

Cytokine-induced memory-like natural killer (NK) cells differentiate after short-term preactivation with IL-12, IL-15, and IL-18 and display enhanced effector function in response to cytokines or tumor targets for weeks after the initial preactivation. Conventional NK cell function depends on a licensing signal, classically delivered by an inhibitory receptor engaging its cognate MHC class I ligand. How licensing status integrates with cytokine-induced memory-like NK cell responses is unknown. We investigated this interaction using killer cell immunoglobulin-like receptor- and HLA-genotyped primary human NK cells. Memory-like differentiation resulted in enhanced IFN-γ production triggered by leukemia targets or FcγRIIIa ligation within licensed NK cells, which exhibited the highest functionality of the NK cell subsets interrogated. IFN-γ production by unlicensed memory-like NK cells was also enhanced to a level comparable with that of licensed control NK cells. Mechanistically, differences in responses to FcγRIIIa-based triggering were not explained by alterations in key signaling intermediates, indicating that the underlying biology of memory-like NK cells is distinct from that of adaptive NK cells in human cytomegalovirus-positive individuals. Additionally, memory-like NK cells responded robustly to cytokine receptor restimulation with no impact of licensing status. These results demonstrate that both licensed and unlicensed memory-like NK cell populations have enhanced functionality, which may be translated to improve leukemia immunotherapy.

Cytokine-induced memory-like natural killer cells exhibit enhanced responses against myeloid leukemia.

Natural killer (NK) cells are an emerging cellular immunotherapy for patients with acute myeloid leukemia (AML); however, the best approach to maximize NK cell antileukemia potential is unclear. Cytokine-induced memory-like NK cells differentiate after a brief preactivation with interleukin-12 (IL-12), IL-15, and IL-18 and exhibit enhanced responses to cytokine or activating receptor restimulation for weeks to months after preactivation. We hypothesized that memory-like NK cells exhibit enhanced antileukemia functionality. We demonstrated that human memory-like NK cells have enhanced interferon-γ production and cytotoxicity against leukemia cell lines or primary human AML blasts in vitro. Using mass cytometry, we found that memory-like NK cell functional responses were triggered against primary AML blasts, regardless of killer cell immunoglobulin-like receptor (KIR) to KIR-ligand interactions. In addition, multidimensional analyses identified distinct phenotypes of control and memory-like NK cells from the same individuals. Human memory-like NK cells xenografted into mice substantially reduced AML burden in vivo and improved overall survival. In the context of a first-in-human phase 1 clinical trial, adoptively transferred memory-like NK cells proliferated and expanded in AML patients and demonstrated robust responses against leukemia targets. Clinical responses were observed in five of nine evaluable patients, including four complete remissions. Thus, harnessing cytokine-induced memory-like NK cell responses represents a promising translational immunotherapy approach for patients with AML.

The IL-15-Based ALT-803 Complex Enhances FcγRIIIa-Triggered NK Cell Responses and In Vivo Clearance of B Cell Lymphomas.

PURPOSE: Anti-CD20 monoclonal antibodies (mAb) are an important immunotherapy for B-cell lymphoma, and provide evidence that the immune system may be harnessed as an effective lymphoma treatment approach. ALT-803 is a superagonist IL-15 mutant and IL-15Rα-Fc fusion complex that activates the IL-15 receptor constitutively expressed on natural killer (NK) cells. We hypothesized that ALT-803 would enhance anti-CD20 mAb-directed NK-cell responses and antibody-dependent cellular cytotoxicity (ADCC). EXPERIMENTAL DESIGN: We tested this hypothesis by adding ALT-803 immunostimulation to anti-CD20 mAb triggering of NK cells in vitro and in vivo. Cell lines and primary human lymphoma cells were utilized as targets for primary human NK cells. Two complementary in vivo mouse models were used, which included human NK-cell xenografts in NOD/SCID-γc (-/-) mice. RESULTS: We demonstrate that short-term ALT-803 stimulation significantly increased degranulation, IFNγ production, and ADCC by human NK cells against B-cell lymphoma cell lines or primary follicular lymphoma cells. ALT-803 augmented cytotoxicity and the expression of granzyme B and perforin, providing one potential mechanism for this enhanced functionality. Moreover, in two distinct in vivo B-cell lymphoma models, the addition of ALT-803 to anti-CD20 mAb therapy resulted in significantly reduced tumor cell burden and increased survival. Long-term ALT-803 stimulation of human NK cells induced proliferation and NK-cell subset changes with preserved ADCC. CONCLUSIONS: ALT-803 represents a novel immunostimulatory drug that enhances NK-cell antilymphoma responses in vitro and in vivo, thereby supporting the clinical investigation of ALT-803 plus anti-CD20 mAbs in patients with indolent B-cell lymphoma.

MicroRNA-15/16 Antagonizes Myb To Control NK Cell Maturation.

NK cells develop in the bone marrow and complete their maturation in peripheral organs, but the molecular events controlling maturation are incompletely understood. The miR-15/16 family of microRNA regulates key cellular processes and is abundantly expressed in NK cells. In this study, we identify a critical role for miR-15/16 in the normal maturation of NK cells using a mouse model of NK-specific deletion, in which immature NK cells accumulate in the absence of miR-15/16. The transcription factor c-Myb (Myb) is expressed preferentially by immature NK cells, is a direct target of miR-15/16, and is increased in 15a/16-1 floxed knockout NK cells. Importantly, maturation of 15a/16-1 floxed knockout NK cells was rescued by Myb knockdown. Moreover, Myb overexpression in wild-type NK cells caused a defective NK cell maturation phenotype similar to deletion of miR-15/16, and Myb overexpression enforces an immature NK cell transcriptional profile. Thus, miR-15/16 regulation of Myb controls the NK cell maturation program.

PTEN regulates natural killer cell trafficking in vivo.

Phosphatase and tensin homolog (PTEN) is a critical negative regulator of the phosphoinositide-3 kinase pathway, members of which play integral roles in natural killer (NK) cell development and function. However, the functions of PTEN in NK cell biology remain unknown. Here, we used an NK cell-specific PTEN-deletion mouse model to define the ramifications of intrinsic NK cell PTEN loss in vivo. In these mice, there was a significant defect in NK cell numbers in the bone marrow and peripheral organs despite increased proliferation and intact peripheral NK cell maturation. Unexpectedly, we observed a significant expansion of peripheral blood NK cells and the premature egress of NK cells from the bone marrow. The altered trafficking of NK cells from peripheral organs into the blood was due to selective hyperresponsiveness to the blood localizing chemokine S1P. To address the importance of this trafficking defect to NK cell immune responses, we investigated the ability of PTEN-deficient NK cells to traffic to a site of tumor challenge. PTEN-deficient NK cells were defective at migrating to distal tumor sites but were more effective at clearing tumors actively introduced into the peripheral blood. Collectively, these data identify PTEN as an essential regulator of NK cell localization in vivo during both homeostasis and malignancy.

Hyperlipoproteinaemia(a) is a common cause of autosomal dominant hypercholesterolaemia.

UNLABELLED: Autosomal dominant hypercholesterolaemia (ADH) are a heterogeneous group of monogenic lipid disorders. The plasma level of lipoprotein(a) (Lp(a)) is a heritable trait associated with increased coronary heart disease (CHD) risk. OBJECTIVE: To evaluate the frequency of elevated Lp(a) as a cause of ADH and the characteristics of subjects with high Lp(a) (hyperLp(a)). MATERIAL AND METHODS: 200 healthy subjects and 933 unrelated Spanish subjects with a clinical diagnosis of ADH who were screened for low-density lipoprotein receptor (LDLR) and apolipoprotein B (APOB) gene mutations. Standard cardiovascular risk factors and blood lipid levels, including Lp(a), were evaluated. HyperLp(a) was defined as Lp(a) levels >or=95th centile of control values. RESULTS: Lp(a) was higher in 263 subjects without LDLR or APOB mutations (nonLDLR/nonAPOB group) than in 670 subjects with mutations (FH group): 40.0 mg/dl (interquartile range (IR) 15.0-89.0) versus 31.0 mg/dl (IR 11.0-73.7) respectively, p = 0.002. HyperLp(a) was present in 23% of ADH subjects (odds ratio (OR) 5.6 (95% CI, 2.9 to 10.7) versus controls) and 29% of nonLDLR/nonAPOB subjects (OR 7.7; 3.9 to 15.4). After adjusting for Lp(a), LDL cholesterol levels were <95th centile in 28 (10.6%) nonLDLR/nonAPOB subjects and in 9 (1.3%) FH subjects. Lp(a) levels were nonsignificantly higher in ADH subjects with early-onset CHD than in those without (43.5 mg/dl, (IR, 12.0-82.0) versus 31.7 mg/dl (11.8-76.5), respectively). CONCLUSIONS: HyperLp(a) is responsible for ADH in approximately 6% of nonLDLR/nonAPOB subjects. HyperLp(a) would not appear to be a risk factor for early-onset CHD in ADH, independently of whether genetic defects have or have not been demonstrated.

Tumores de la cavidad oral y huesos maxilares en pediatría. Revisión de 122 pacientes / Oral cavity and maxillary bone tumors in pediatrics. Review of 122 patients

Se presenta un estudio retrospectivo de 122 pacientes con tumores que se manifestaron en la cavidad oral, con localización inicial en maxilares o partes blandas (se excluyeron los tumores de la cara sin compromiso bucal). La edad promedio fue 9 años y 6 meses (rango de 1 día a 17 años). El 62 por ciento se presentó en varones. La localización inicial de los tumores fue en hueso en el 53 por ciento de los casos y en partes blandas en el 47 por ciento; 82 pacientes tuvieron lesiones benignas y 40 lesiones malignas. Las manifestaciones al ingreso fueron: tumor palpable o visible (39 por ciento), tumor más dolor (22 por ciento), dolor (19 por ciento) y otros como caída de dientes, parálisis, fiebre o asímetría facial (20 por ciento). La rutina de estudio comprendió radiografía panorámica de maxilar, centellografía ósea (gammacámara con Tecnesio 99), tomografía axial computada (TAC) y resonancia nuclear magnética (RNM). Los pacientes fueron tratados en forma multidisciplinaria siendo la cirugía (punción aspiración con aguja fina, biopsia y/o resección)el procedimiento inicial en la mayoría de ellos. De acuerdo al algoritmo todos los pacientes con lesión ósea fueron estudiados con Rx simple y TAC, 89 por ciento de positividad en ambas, previas a la biopsia por punción. De igual menera en los tumores de partes blandas la TAC mantuvo su utilidad, no así la Rx simple que fue reeplazada por la ecografía cuando se detectó ausencia de compromiso óseo. Las lesiones benignas predominaron (78/122) a nivel de hueso o de partes blandas. la curación en ella fue la regla. En lo que respecta a los tumores, primarios de la región (11/40) correspondieron inicialmente a partes blandas y raramente a hueso, en los que fue frecuente el compromiso metastático o multicéntrico.

Treatment and follow-up of HIV-negative multidrug-resistant tuberculosis patients in an infectious diseases reference hospital, Buenos Aires, Argentina.

SETTING: An Argentinean reference hospital specialising in infectious diseases. OBJECTIVE: To assess the outcomes of all human immunodeficiency virus (HIV) negative multidrug-resistant tuberculosis (MDR-TB) patients referred to or diagnosed at Hospital Muñiz. DESIGN: Clinical study for the period 1996-1999, with follow-up until June 2002. RESULTS: One hundred and forty-one adult patients (52.5% female) with resistance to two to seven drugs were studied. Fifty patients (35.5%) had not been treated previously. The most frequently used second-line drugs were 5-F-quinolones, cycloserine and ethionamide in susceptibility based individually tailored three- to five-drug regimens. Hospital admission was associated with treatment success. Forty-five episodes of severe toxicity occurred. Treatment was successful in 51.8% of cases, but follow-up of 73 patients yielded 11.9% relapse. The mortality rate was 19.1% and default was 19.9%. Logistic regression analysis was statistically significant for treatment success in relation to patient admission, residence and resistance pattern. CONCLUSION: The burden of MDR-TB in this setting--prolonged infection, treatment cost and difficulties, low rates of cure and treatment adherence and high rates of fatality and relapse--can be improved by strengthening TB control programme activities and fighting against poverty and HIV/AIDS.

Memories in context.

Context-dependent associative memories are models that allow the retrieval of different vectorial responses given a same vectorial stimulus, depending on the context presented to the memory. The contextualization is obtained by doing the Kronecker product between two vectorial entries to the associative memory: the key stimulus and the context. These memories are able to display a wide variety of behaviors that range from all the basic operations of the logical calculus (including fuzzy logics) to the selective extraction of features from complex vectorial patterns. In the present contribution, we show that a context-dependent memory matrix stores a large amount of possible virtual associative memories, that awaken in the presence of a context. We show how the vectorial context allows a memory matrix to be representable in terms of its singular-value decomposition. We describe a neural interpretation of the model in which the Kronecker product is performed on the same neurons that sustain the memory. We explored, with numerical experiments, the reliability of chains of contextualized associations. In some cases, random disconnection produces the emergence of oscillatory behaviors of the system. Our results show that associative chains retain their performances for relatively large dimensions. Finally, we analyze the properties of some modules of context-dependent autoassociative memories inserted in recursive nets: the perceptual autoorganization in the presence of ambiguous inputs (e.g. the disambiguation of the Necker's cube figure), the construction of intersection filters, and the feature extraction capabilities.