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This article examines the work behaviors related to counseling substance-abusing clients by mental health service providers in multiservice mental health agencies. The results support the need for specialized knowledge and skills in working with substance-abusing clients. (Author)

Lack of molecular targets in estrogen receptor-negative (ER-negative) breast cancer is a major therapeutic hurdle. We studied NF-[kappa]B activation in human breast tumors and in carcinoma cell lines. Activated NF-[kappa]B was detected predominantly in ER-negative vs. ER-positive breast tumors and mostly in ER-negative and ErbB2-positive tumors (86%). These in vivo results demonstrate association of activated NF-[kappa]B with a subgroup of human breast tumors and are consistent with previously reported in vitro observations using similar classes of human breast cancer cell lines. Finding such an association suggested functional and biological significance. Immunofluorescence demonstrated increased nuclear p65, a component of the active NF-[kappa]B complex, in cytokeratin 19 (CK19)-positive epithelial ceils of ER-negative/ErbB2-positive tumor samples. In contrast, nuclear NF-[kappa]B was detected mostly in stroma of ER-negative and ErbB2-negative tumors, suggesting a role of activated NF-[kappa]B in intercellular signaling between epithelial and stromal cells in this type of breast cancers. To elucidate roles of activated NF-[kappa]B, we used an ER-negative and ErbB2-positive human breast tumor cell line (SKBr3). The polypeptide heregulin [beta]1 stimulated, and herceptin, the anti-ErbB2 antibody, inhibited, NF-[kappa]B activation in SKBr3 cells. The NF-[kappa]B essential modulator (NEMO)-binding domain (NBD) peptide, an established selective inhibitor of I[kappa]B-kinase (IKK), blocked heregulin-mediated activation of NF-[kappa]B and cell proliferation, and simultaneously induced apoptosis only in proliferating and not resting cells. These results substantiate the hypothesis that certain breast cancer cells rely on NF-[kappa]B for aberrant cell proliferation and simultaneously avoid apoptosis, thus implicating activated NF-[kappa]B as a therapeutic target for distinctive subclasses of ER-negative breast cancers.

The skin of people with atopic dermatitis appears to produce smaller than normal amounts of proteins that have antibiotic properties. This would explain why people with atopic dermatitis have an increased risk of skin infections. Atopic dermatitis is a chronic inflammation of the skin.

Human liver myeloid cells are imperfectly defined, but it is broadly agreed that cells of stellate appearance in situ , expressing the markers CD11b and CD68, are the liver's resident macrophages, classically termed Kupffer cells. Recent investigations using single cell RNA sequencing and unsupervised clustering algorithms suggest there are two populations of cells with the characteristics of tissue macrophages in human liver. We therefore analyzed dissociated human liver tissue using the markers CD11b and CD68 to define macrophage-like cells and found within this population two subsets that differ in their expression of multiple surface markers. These subsets were FACS-sorted based on CD32 expression, and gene expression analysis identified them with human liver myeloid cell subsets that were previously defined by two independent single cell RNA sequencing studies. Using qRT-PCR we found that the two subsets differed in the expression of genes associated with T cell activation and immunosuppression, suggesting distinct roles in T cell tolerance. In addition, one subset expressed two markers, CD1C and CD11c, more often seen on classical dendritic cells. Criteria used to distinguish macrophages from dendritic cells in other tissues may need to be revised in the human liver. [ABSTRACT FROM AUTHOR]

High levels of protein expression are key to the successful development and manufacture of a therapeutic antibody. Here, we describe two related antibodies, Ab001 and Ab008, where Ab001 shows a markedly lower level of expression relative to Ab008 when stably expressed in Chinese hamster ovary cells. We use single-gene expression vectors and structural analysis to show that the reduced titer is associated with the VL CDR2 of Ab001. We adopted two approaches to improve the expression of Ab001. First, we used mutagenesis to change single amino-acid residues in the Ab001 VL back to the equivalent Ab008 residues but this resulted in limited improvements in expression. In contrast when we used an in silico structure-based design approach to generate a set of five individual single-point variants in a discrete region of the VL, all exhibited significantly improved expression relative to Ab001. The most successful of these, D53N, exhibited a 25-fold increase in stable transfectants relative to Ab001. The functional potency of these VL-modified antibodies was unaffected. We expect that this in silico engineering strategy can be used to improve the expression of other antibodies and proteins. [ABSTRACT FROM AUTHOR]

Phosphoinositide-dependent kinase 1 (PDK1) plays an important role in integrating the T cell antigen receptor (TCR) and CD28 signals to achieve efficient NF-κB activation. PDK1 is also an important regulator of T cell development, mediating pre-TCR induced proliferation signals. However, the role of PDK1 in B cell antigen receptor (BCR) signaling and B cell development remains largely unknown. In this study we provide genetic evidence supporting the role of PDK1 in B cell survival. We found PDK1 is required for BCR mediated survival in resting B cells, likely through regulation of Foxo activation. PDK1-dependent signaling to NF-κB is not crucial to resting B cell viability. However, PDK1 is necessary for triggering NF-κB during B cell activation and is required for activated B cell survival. Together these studies demonstrate that PDK1 is essential for BCR-induced signal transduction to Foxo and NF-κB and is indispensable for both resting and activated B cell survival. [ABSTRACT FROM AUTHOR]

Summary: Inhibition of the transcription factor nuclear factor (NF)-κB activity leads to a reduction in numbers of CD8+ single-positive (SP) thymocytes, suggesting a selective role for NF-κB in these cells. To further explore the role of NF-κB in SP thymocytes, we utilized transgenic models that allowed either inhibition or activation of NF-κB. We showed that activation of NF-κB played an important role in the selection of major histocompatibility complex (MHC) class I-restricted CD8+ T cells. Surprisingly, NF-κB was not activated in positively selected CD4+ thymocytes, and inhibition of NF-κB did not perturb positive or negative selection of CD4+ cells. However, enforced activation of NF-κB via a constitutively active inhibitor of κB (IκB) kinase transgene led to a nearly complete deletion of CD4 cells by pushing positively selecting CD4+ cells into negative selection. These studies therefore revealed a surprising difference of NF-κB activation in CD4+ and CD8+ thymocytes and suggested that NF-κB contributes to the establishment of thresholds of signaling that determine positive or negative selection of thymocytes. [Copyright &y& Elsevier]

This paper investigates the Theory of Mind ability of psychopaths. Happé's (1994) advanced test of Theory of Mind was presented to 25 psychopaths and 25 non-psychopathic incarcerated controls. The psychopaths and the non-psychopathic controls did not differ in their performance on this task. However, the psychopaths were performing significantly better than Happé's most highly able adult autistic population. It was therefore concluded that the psychopath does not have a Theory of Mind deficit. Speculations are made about the different developmental pathways of autism and psychopathy and the differences in the empathy deficit present in both these disorders. [ABSTRACT FROM PUBLISHER]

Sexual attacks on older women committed by young adult males are a cause of much concern. Despite this there is very little on the subject in the psychiatric literature. This article reviews the available literature and describes a small series of cases, highlighting not only common themes, but also the diversity of psychopathology evident in the perpetrator. [ABSTRACT FROM PUBLISHER]

Several reports have highlighted a potential role of autoreactive B-cells and autoantibodies that correlates with increased disease severity in patients with idiopathic pulmonary fibrosis (IPF). Here we show that patients with IPF have an altered B-cell phenotype and that those subjects who have autoantibodies against the intermediate filament protein periplakin (PPL) have a significantly worse outcome in terms of progression-free survival. Using a mouse model of lung fibrosis, we demonstrate that introducing antibodies targeting the endogenous protein PPL (mimicking naturally occurring autoantibodies seen in patients) directly in the lung increases lung injury, inflammation, collagen and fibronectin expression through direct activation of follicular dendritic cells, which in turn activates and drives proliferation of fibroblasts. This fibrocyte population was also observed in fibrotic foci of patients with IPF and was increased in peripheral blood of IPF patients compared to aged-matched controls. This study reiterates the complex and heterogeneous nature of IPF, identifying new pathways that may prove suitable for therapeutic intervention., Competing Interests: Conflict of interest: C. Taillé reports grants from GSK, Sanofi and AstraZeneca, advisory board participation and consulting fees from GSK, Sanofi, AstraZeneca and Novartis, lecture honoraria from GSK, Sanofi, AstraZeneca, Stallergenes and Novartis, and travel support from GSK and AstraZeneca, outside the submitted work. D. Valeyre reports travel support from ERS, ATS and CPLF, and advisory board participation with Roche and Boehringer Ingelheim, outside the submitted work. H. Nunes reports consulting fees, lecture honoraria and travel support from Boehringer Ingelheim and Roche/Genentech, and advisory board participation with Galapagos, outside the submitted work. W.K. Lim reports options and stock as part of employee compensation from Regeneron Pharmaceuticals, outside the submitted work. V. Cottin reports grants from Boehringer Ingelheim, consulting fees from Boehringer Ingelheim and Roche/Promedior, lecture honoraria from Boehringer Ingelheim, Roche/Promedior, Sanofi and AstraZeneca, travel support from Boehringer Ingelheim, Roche/Promedior and AstraZeneca, and advisory board participation with Actelion, Bayer/MSD, Roche/Promedior, Sanofi, Celgene/BMS, Galapagos, Galecto, Shionogi, Fibrogen, RedX and PureTech, outside the submitted work. D. Corkill reports support for the present manuscript from AstraZeneca/MedImmune, as a full-time employee. T. Mustelin reports grants from NIH, and consulting fees from Miro Bio, Kiniksa, Cugene, QiLu and ROME, outside the submitted work. M.A. Sleeman reports support for the present manuscript from MedImmune/AstraZeneca, and during the research was a full-time employee of MedImmune/AstraZeneca. B. Crestani reports support for the present manuscript from MedImmune/AstraZeneca; B. Crestani also reports grants from Boehringer Ingelheim, Roche, BMS and Translate Bio, consulting fees from Apellis, Boehringer Ingelheim, Roche, BMS, Translate Bio, Sanofi and Novartis, lecture honoraria from AstraZeneca, Boehringer Ingelheim, Roche, BMS and Sanofi, travel support from AstraZeneca, BMS, Boehringer Ingelheim, Roche and Novartis, and is the Vice President of the Fondation du Souffle, outside the submitted work. All other authors have nothing to disclose., (Copyright ©The authors 2023. For reproduction rights and permissions contact permissions@ersnet.org.)

High levels of protein expression are key to the successful development and manufacture of a therapeutic antibody. Here, we describe two related antibodies, Ab001 and Ab008, where Ab001 shows a markedly lower level of expression relative to Ab008 when stably expressed in Chinese hamster ovary cells. We use single-gene expression vectors and structural analysis to show that the reduced titer is associated with the VL CDR2 of Ab001. We adopted two approaches to improve the expression of Ab001. First, we used mutagenesis to change single amino-acid residues in the Ab001 VL back to the equivalent Ab008 residues but this resulted in limited improvements in expression. In contrast when we used an in silico structure-based design approach to generate a set of five individual single-point variants in a discrete region of the VL, all exhibited significantly improved expression relative to Ab001. The most successful of these, D53N, exhibited a 25-fold increase in stable transfectants relative to Ab001. The functional potency of these VL-modified antibodies was unaffected. We expect that this in silico engineering strategy can be used to improve the expression of other antibodies and proteins., (© The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

IL-13 is a major Th2 cytokine that is capable of inducing inflammation, excessive mucus production, airway hyperresponsiveness, alveolar remodeling, and fibrosis in the murine lung. Although IL-13 through its binding to IL-4Ralpha/IL-13Ralpha1 uses the canonical STAT6-signaling pathway to mediate these tissue responses, recent studies have demonstrated that other signaling pathways may also be involved. Previous studies from our laboratory demonstrated that IL-13 mediates its tissue effects by inducing a wide variety of downstream genes many of which are known to be regulated by NF-kappaB. As a result, we hypothesized that NF-kappaB activation plays a critical role in the pathogenesis of IL-13-induced tissue alterations. To test this hypothesis, we compared the effects of transgenic IL-13 in mice with normal and diminished levels of NF-kappaB activity. Three pharmacologic approaches were used to inhibit NF-kappaB including 1) PS1145, a small molecule inhibitor of IkappaBalpha kinase (IKK2), 2) antennapedia-linked NF-kappaB essential modulator-binding domain (NBD) peptide (wild-type NBD), and 3) an adenoviral construct expressing a dominant-negative version of IKK2. We also crossed IL-13-transgenic mice with mice with null mutations of p50 to generate mice that overproduced IL-13 in the presence and absence of this NF-kappaB component. These studies demonstrate that all these interventions reduced IL-13-induced tissue inflammation, fibrosis and alveolar remodeling. In addition, we show that both PS1145 and wild-type NBD inhibit lung inflammatory and structural cell apoptosis. PS1145 inhibits caspase activation and up-regulates inhibitor of apoptosis protein cellular-inhibitor of apoptosis protein 1 (c-IAP-1). Therefore, NF-kappaB is an attractive target for immunotherapy of IL-13-mediated diseases.

Lack of molecular targets in estrogen receptor-negative (ER-negative) breast cancer is a major therapeutic hurdle. We studied NF-kappa B activation in human breast tumors and in carcinoma cell lines. Activated NF-kappa B was detected predominantly in ER-negative vs. ER-positive breast tumors and mostly in ER-negative and ErbB2-positive tumors (86%). These in vivo results demonstrate association of activated NF-kappa B with a subgroup of human breast tumors and are consistent with previously reported in vitro observations using similar classes of human breast cancer cell lines. Finding such an association suggested functional and biological significance. Immunofluorescence demonstrated increased nuclear p65, a component of the active NF-kappa B complex, in cytokeratin 19 (CK19)-positive epithelial cells of ER-negative/ErbB2-positive tumor samples. In contrast, nuclear NF-kappa B was detected mostly in stroma of ER-negative and ErbB2-negative tumors, suggesting a role of activated NF-kappa B in intercellular signaling between epithelial and stromal cells in this type of breast cancers. To elucidate roles of activated NF-kappa B, we used an ER-negative and ErbB2-positive human breast tumor cell line (SKBr3). The polypeptide heregulin beta1 stimulated, and herceptin, the anti-ErbB2 antibody, inhibited, NF-kappa B activation in SKBr3 cells. The NF-kappa B essential modulator (NEMO)-binding domain (NBD) peptide, an established selective inhibitor of I kappa B-kinase (IKK), blocked heregulin-mediated activation of NF-kappa B and cell proliferation, and simultaneously induced apoptosis only in proliferating and not resting cells. These results substantiate the hypothesis that certain breast cancer cells rely on NF-kappa B for aberrant cell proliferation and simultaneously avoid apoptosis, thus implicating activated NF-kappa B as a therapeutic target for distinctive subclasses of ER-negative breast cancers.

The pathogenesis of acute rheumatic fever (ARF) is poorly understood. We identified two contiguous bacteriophage genes, designated speL and speM, encoding novel inferred superantigens in the genome sequence of an ARF strain of serotype M18 group A streptococcus (GAS). speL and speM were located at the same genomic site in 33 serotype M18 isolates, and no nucleotide sequence diversity was observed in the 33 strains analyzed. Furthermore, the genes were absent in 13 non-M18 strains tested. These data indicate a recent acquisition event by a distinct clone of serotype M18 GAS. speL and speM were transcribed in vitro and upregulated in the exponential phase of growth. Purified SpeL and SpeM were pyrogenic and mitogenic for rabbit splenocytes and human peripheral blood mononuclear cells in picogram amounts. SpeL preferentially expanded human T cells expressing T-cell receptors Vbeta1, Vbeta5.1, and Vbeta23, and SpeM had specificity for Vbeta1 and Vbeta23 subsets, indicating that both proteins had superantigen activity. SpeL was lethal in two animal models of streptococcal toxic shock, and SpeM was lethal in one model. Serologic studies indicated that ARF patients were exposed to serotype M18 GAS, SpeL, and SpeM. The data demonstrate that SpeL and SpeM are pyrogenic toxin superantigens and suggest that they may participate in the host-pathogen interactions in some ARF patients.

Glucocorticoid (GC) insensitivity is a challenging clinical problem associated with many chronic inflammatory disorders and life-threatening disease progression. The molecular basis of GC insensitivity, however, is unknown. Alternative splicing of the GC receptor (GCR) pre-mRNA generates a second GCR, termed GCRbeta, which does not bind GC but antagonizes the transactivating activity of the classic GCR, termed GCRalpha. GC-insensitive conditions have been associated with increased GCRbeta expression. Whether or not increased GCRbeta expression can contribute to GC insensitivity, however, remains controversial. To more precisely demonstrate the effect of GCRbeta on steroid responsiveness, we virally transduced GCRbeta cDNA into mouse DO-11.10 hybridoma cells, as mice are known to be deficient in the GCRbeta gene. We demonstrate that viral transduction of GCRbeta cDNA into mouse hybridoma cells to induce stable expression of GCRbeta results in GC insensitivity of these cells. Furthermore, in such cells GCRalpha is complexed with GCRbeta. Such heterodimer formation may account for the reduced effectiveness of GC action in cells overexpressing GCRbeta.

PBMC and acute skin lesions of patients with atopic dermatitis (AD) are characterized by increased IL-4 and IL-13, but decreased IFN-gamma production. This bias toward an increased Th2 cytokine profile may contribute to the elevated IgE levels and acute skin inflammation seen in AD. In this study, we examined the levels of IL-15, a Th1-like cytokine, in the PBMC and the skin lesions of AD patients. IL-15 secretion by Staphylococcal enterotoxin B-treated PBMC of AD patients was significantly lower than that of normals and psoriasis patients (p < 0.001). Membrane-bound IL-15 expression as measured by mean fluorescence intensity and percentage of IL-15-positive cells in Staphylococcal enterotoxin B-treated monocytes of AD patients (644 +/- 49% and 12.7 +/- 0.6%, respectively) were significantly lower than that of normals (869 +/- 56% and 15.8 +/- 1.2%, respectively) and psoriasis patients (1488 +/- 217% and 22.7 +/- 0.8%, respectively; p < 0.0007 and p < 0.0001, respectively). The membrane-bound IL-15 expression was also significantly lower in the control monocytes of AD patients compared with that in normals and psoriasis patients. There was no significant difference in the absolute number or percentage of monocytes between the study subjects. However, psoriasis skin lesions were found to have significantly more IL-15 mRNA-expressing cells (22.4 +/- 1.7) compared with that in acute AD (7.5 +/- 1.7) and chronic AD (13.7 +/- 1.7) skin lesions (p < 0.05). IL-15 enhanced IFN-gamma production by the PBMC of AD patients (p < 0.01), but not by that of normal individuals or psoriasis patients. In addition, IL-15 was found to suppress IgE synthesis (p < 0.01) by the PBMC of AD patients. These data support the concept that reduced IL-15 expression may contribute to the pathogenesis of AD.