Miracle-dz/UniFAR-FTUs huggingface dataset:
Genomic Determinants of Gene Regulation by 1,25-Dihydroxyvitamin D3 during Osteoblast-lineage Cell Differentiation*♦

Background: The biological activities of 1,25(OH)2D3 in osteoblasts are dependent upon their differentiation state. Results: Genome-wide analyses reveal transcriptomic responses to 1,25(OH)2D3, and VDR, RUNX2, and C/EBPβ cistromes are modified during differentiation. Conclusion: Differentiation-induced changes in expression and transcription factor genome occupancy underlie the response to 1,25(OH)2D3. Significance: Transcription factor occupancy at genome sites is dynamic and significantly influenced by the state of cellular differentiation. The biological effects of 1α,25-dihydroxyvitamin D3 (1,25 (OH)2D3) on osteoblast differentiation and function differ significantly depending upon the cellular state of maturation. To explore this phenomenon mechanistically, we examined the impact of 1,25(OH)2D3 on the transcriptomes of both pre-osteoblastic (POBs) and differentiated osteoblastic (OBs) MC3T3-E1 cells, and assessed localization of the vitamin D receptor (VDR) at sites of action on a genome-scale using ChIP sequence analysis. We observed that the 1,25(OH)2D3-induced transcriptomes of POBs and OBs were quantitatively and qualitatively different, supporting not only the altered biology observed but the potential for a change in VDR interaction at the genome as well. This idea was confirmed through discovery that VDR cistromes in POBs and OBs were also strikingly different. Depletion of VDR-binding sites in OBs, due in part to reduced VDR expression, was the likely cause of the loss of VDR-target gene interaction. Continued novel regulation by 1,25(OH)2D3, however, suggested that factors in addition to the VDR might also be involved. Accordingly, we show that transcriptomic modifications are also accompanied by changes in genome binding of the master osteoblast regulator RUNX2 and the chromatin remodeler CCAAT/enhancer-binding protein β. Importantly, genome occupancy was also highlighted by the presence of epigenetic enhancer signatures that were selectively changed in response to both differentiation and 1,25(OH)2D3. The impact of VDR, RUNX2, and C/EBPβ on osteoblast differentiation is exemplified by their actions at the Runx2 and Sp7 gene loci. We conclude that each of these mechanisms may contribute to the diverse actions of 1,25(OH)2D3 on differentiating osteoblasts.
Antigen‐specific adaptive immune responses in fingolimod‐treated multiple sclerosis patients

T cells exit secondary lymphoid organs along a sphingosine1‐phosphate (S1P) gradient and, accordingly, are reduced in blood upon fingolimod‐mediated S1P‐receptor (S1PR)‐blockade. Serving as a model of adaptive immunity, we characterized cellular and humoral immune responses to influenza vaccine in fingolimod‐treated patients with multiple sclerosis (MS) and in untreated healthy controls. Although the mode of action of fingolimod might predict reduced immunity, vaccine‐triggered T cells accumulated normally in blood despite efficient S1PR‐blockade. Concentrations of anti–influenza A/B immunoglobulin (Ig)M and IgG also increased similarly in both groups. These results indicate that fingolimod‐treated individuals can mount vaccine‐specific adaptive immune responses comparable to healthy controls. Ann Neurol 2011
Hyaluronan Synthase 2 Protects Skin Fibroblasts against Apoptosis Induced by Environmental Stress*

Background: Hyaluronan (HA), an extracellular glycosaminoglycan, is normally produced by three HA synthase (Has) enzymes. Results: Skin fibroblasts from Has1/Has3 double knock-out mice have higher Has2 expression and HA levels and are resistant to cell death after UVB exposure or serum starvation. Conclusion: HA modulates injury-induced apoptotic responses in fibroblasts. Significance: HA has an important role in cell death responses. A balanced turnover of dermal fibroblasts is crucial for structural integrity and normal function of the skin. During recovery from environmental injury (such as UV exposure and physical wounding), apoptosis is an important mechanism regulating fibroblast turnover. We are interested in the role that hyaluronan (HA), an extracellular matrix molecule synthesized by HA synthase enzymes (Has), plays in regulating apoptosis in fibroblasts. We previously reported that Has1 and Has3 double knock-out (Has1/3 null) mice show accelerated wound closure and increased numbers of fibroblasts in the dermis. In the present study, we report that HA levels and Has2 mRNA expression are higher in cultured Has1/3 null primary skin fibroblasts than in wild type (WT) cells. Apoptosis induced by two different environmental stressors, UV exposure and serum starvation (SS), was reduced in the Has1/3 null cells. Hyaluronidase, added to cultures to remove extracellular HA, surprisingly had no effect upon apoptotic susceptibility to UVB or SS. However, cells treated with 4-methylumbelliferone to inhibit HA synthesis were sensitized to apoptosis induced by SS or UVB. When fibroblasts were transfected with Has2-specific siRNA that lowered Has2 mRNA and HA levels by 90%, both Has1/3 null and WT cells became significantly more sensitive to apoptosis. The exogenous addition of high molecular weight HA failed to reverse this effect. We conclude that Has1/3 null skin fibroblasts (which have higher levels of Has2 gene expression) are resistant to stress-induced apoptosis.
Structure of Pneumococcal Peptidoglycan Hydrolase LytB Reveals Insights into the Bacterial Cell Wall Remodeling and Pathogenesis*

Background: The pneumococcal endo-β-N-acetylglucosaminidase LytB is required for cell division and colonization. Results: Structural analysis revealed that the catalytic domain of LytB consists of three structurally distinct modules. Conclusion: All three modules of LytB are necessary for its optimal activity toward peptidoglycan hydrolysis and for pneumococcal adhesion to respiratory epithelial cells. Significance: Provided is the structural insight into LytB-mediated pneumococcal cell wall remodeling and pathogenesis. Streptococcus pneumoniae causes a series of devastating infections in humans. Previous studies have shown that the endo-β-N-acetylglucosaminidase LytB is critical for pneumococcal cell division and nasal colonization, but the biochemical mechanism of LytB action remains unknown. Here we report the 1.65 Å crystal structure of the catalytic domain (residues Lys-375–Asp-658) of LytB (termed LytBCAT), excluding the choline binding domain. LytBCAT consists of three structurally independent modules: SH3b, WW, and GH73. These modules form a “T-shaped” pocket that accommodates a putative tetrasaccharide-pentapeptide substrate of peptidoglycan. Structural comparison and simulation revealed that the GH73 module of LytB harbors the active site, including the catalytic residue Glu-564. In vitro assays of hydrolytic activity indicated that LytB prefers the peptidoglycan from the lytB-deficient pneumococci, suggesting the existence of a specific substrate of LytB in the immature peptidoglycan. Combined with in vitro cell-dispersing and in vivo cell separation assays, we demonstrated that all three modules are necessary for the optimal activity of LytB. Further functional analysis showed that the full catalytic activity of LytB is required for pneumococcal adhesion to and invasion into human lung epithelial cells. Structure-based alignment indicated that the unique modular organization of LytB is highly conserved in its orthologs from Streptococcus mitis group and Gemella species. These findings provided structural insights into the pneumococcal cell wall remodeling and novel hints for the rational design of therapeutic agents against pneumococcal growth and thereby the related diseases.
Greater effect of stroke thrombolysis in the presence of arterial obstruction

Recanalization of arterial obstruction is associated with improved clinical outcomes. There are no controlled data demonstrating whether arterial obstruction status predicts the treatment effect of intravenous (IV) tissue plasminogen activator (tPA). We aimed to determine if the presence of arterial obstruction improves the treatment effect of IV tPA over placebo in attenuating infarct growth.
Got milk?

The article by Mirzaei and colleagues 1 is another thoughtful and clever use of the Nurse’s Health Study. The evaluation of the mother’s data from this study is carefully conducted with an innovative approach to taking self-reported outcomes and validating their approach to estimating actual serum 25(OH)D levels. Further, by focusing on the gestational period, the problems of telescoping and sequencing effects in retrospective studies are mitigated to a degree. There are careful considerations in the handling of design issues and the authors acknowledge the obvious limitations of such studies, such as differential recall bias by cases vs controls. Acknowledging it, of course, does not eliminate it, but there is equally no evidence to demonstrate the findings result from recall bias. Thus, the authors should be congratulated on a very thoughtful and well done work. Nevertheless, we want to express some cautions, limitations, and overinterpretations of the data. It is easy to see the potential headlines that could result from this work or the public health perceptions that all pregnant mothers should be consuming greater amounts of milk to prevent multiple sclerosis (MS). These concerns can be directly traced to language in the paper such as ‘‘33% of the MS cases among mothers who drank less than 2 glasses of milk per day could have been hypothetically prevented. ’’ While this is carefully worded and analytically correct in its statement, it is too easy to take the current fervor for vitamin D and a cross-sectional retrospective analysis and demand pregnant women not drink or smoke, and now drink more milk. The concern over an overly simplistic message is opposite the article’s extremely complex nature and analyses of the evidence. It incorporates not only direct variables, but also covariates that may be highly correlated. It is difficult to assess the interrelated variables and isolate their impact on the results. For example, milk consumption has north to south gradients. The Northeast on average consumes 335ml per day; North Central 291ml/ day; West 276ml/day, and the South 214ml/day. While the authors adjusted for location at birth, they did so as North, Middle, South, which may have left unadjusted some of the milk consumption and latitudinal effects in a combination that with highly correlated environmental, genetic, and vitamin D effects give rise to an association with the milk consumption data. The evidence for the vitamin D effect is mostly through a gradient as individual levels of milk consumption are not significantly different except at the highest levels of consumption. Furthermore, the gradient effect may not be the appropriate paradigm for the impact on disease. With rickets, as with many other nutritional diseases, it is a threshold that is key and while there is some evidence for this, the basic association rests with the gradient. In addition, what is the biological basis by which the estimated level of vitamin D associated with the selfreported intake could work to prevent, in utero, subsequent MS? It is not totally clear what level of vitamin D intake is required to sustain an adequate serum level.
DJ‐1 modulates the expression of Cu/Zn‐superoxide dismutase‐1 through the Erk1/2–Elk1 pathway in neuroprotection

Loss of function mutations of Park7/DJ‐1 gene increase the susceptibility of dopaminergic cells to reactive oxygen species and cause early onset familial Parkinson disease (PD). However, the mechanisms underlying dopaminergic neuron loss related to DJ‐1 mutation remain undefined. Therefore, it is important to find the new mechanisms underlying the antioxidative functions of DJ‐1.
Cerebral amyloid angiopathy in the elderly

Cerebral amyloid angiopathy (CAA) results from deposition of β‐amyloid in the media and adventitia of small arteries and capillaries of the leptomeninges and cerebral cortex and is a major cause of lobar intracerebral hemorrhage and cognitive impairment in the elderly. CAA is associated with a high prevalence of magnetic resonance imaging markers of small vessel disease, including cerebral microbleeds and white matter hyperintensities. Although advanced CAA is present in approximately ¼ of brains with Alzheimer disease (AD), fewer than half of CAA cases meet pathologic criteria for AD. This review will discuss the pathophysiology of CAA and focus on new imaging modalities and laboratory biomarkers that may aid in the clinical diagnosis of individuals with the disease. Ann Neurol 2011;70:871–880
The RpfB-Dependent Quorum Sensing Signal Turnover System Is Required for Adaptation and Virulence in Rice Bacterial Blight Pathogen Xanthomonas oryzae pv. oryzae.

Xanthomonas oryzae pv. oryzae, the bacterial blight pathogen of rice, produces diffusible signal factor (DSF) family quorum sensing signals to regulate virulence. The biosynthesis and perception of DSF family signals require components of the rpf (regulation of pathogenicity factors) cluster. In this study, we report that RpfB plays an essential role in DSF family signal turnover in X. oryzae pv. oryzae PXO99A. The production of DSF family signals was boosted by deletion of the rpfB gene and was abolished by its overexpression. The RpfC/RpfG-mediated DSF signaling system negatively regulates rpfB expression via the global transcription regulator Clp, whose activity is reversible in the presence of cyclic diguanylate monophosphate. These findings indicate that the DSF family signal turnover system in PXO99A is generally consistent with that in Xanthomonas campestris pv. campestris. Moreover, this study has revealed several specific roles of RpfB in PXO99A. First, the rpfB deletion mutant produced high levels of DSF family signals but reduced extracellular polysaccharide production, extracellular amylase activity, and attenuated pathogenicity. Second, the rpfB/rpfC double-deletion mutant was partially deficient in xanthomonadin production. Taken together, the RpfB-dependent DSF family signal turnover system is a conserved and naturally presenting signal turnover system in Xanthomonas spp. , which plays unique roles in X. oryzae pv. oryzae adaptation and pathogenesis.
cAMP-responsive Element-binding Protein (CREB) and cAMP Co-regulate Activator Protein 1 (AP1)-dependent Regeneration-associated Gene Expression and Neurite Growth*

Background: The inability to up-regulate “regeneration-associated genes” contributes to the failure of axonal regeneration after injury. Results: Expression of activated CREB and cAMP up-regulation co-regulate AP1-dependent regeneration-associated gene expression and enhance axon growth. Conclusion: Targeting transcription factor activity and the associated second messengers facilitates regenerative growth. Significance: This transcription factor plus small molecule approach may be an efficient means to stimulate axon regeneration. To regenerate damaged axons, neurons must express a cassette of regeneration-associated genes (RAGs) that increases intrinsic growth capacity and confers resistance to extrinsic inhibitory cues. Here we show that dibutyrl-cAMP or forskolin combined with constitutive-active CREB are superior to either agent alone in driving neurite growth on permissive and inhibitory substrates. Of the RAGs examined, only arginase 1 (Arg1) expression correlated with the increased neurite growth induced by the cAMP/CREB combination, both of which were AP1-dependent. This suggests that cAMP-induced AP1 activity is necessary and interacts with CREB to drive expression of RAGs relevant for regeneration and demonstrates that combining a small molecule (cAMP) with an activated transcription factor (CREB) stimulates the gene expression necessary to enhance axonal regeneration.
Sample preparation and image acquisition using optical‐reflective microscopy in the measurement of fiber orientation in thermoplastic composites

A complete sample preparation procedure used to determine three‐dimensional fiber orientation from optical micrographs of glass fiber‐reinforced thermoplastic composites is presented. Considerations for elimination of irregularities in the elliptical footprints, contrast enhancement between fibers and surrounding polymer matrix, controlled‐etching that allows the identification of small shadows where fiber recedes into the matrix, and topographical reconstruction of the elliptical footprint are described in the procedure. This procedure has produced high‐quality optical micrographs employed to obtain accurate fiber orientation data for thermoplastic composites using the method of ellipses. The optimal definition of the nonelliptical footprints’ borders allows an accurate measurement of orientation in small sampling areas.
Macrophages prevent hemorrhagic infarct transformation in murine stroke models

Inflammation is increasingly viewed as a new therapeutic target in subacute stages of brain infarction. However, apart from causing secondary damage, inflammation could equally promote beneficial lesion remodeling and repair. Distinct subpopulations of monocytes/macrophages (MOs/MPs) may critically determine the outcome of lesion‐associated inflammation.
Carotid plaque inflammation on 18F‐fluorodeoxyglucose positron emission tomography predicts early stroke recurrence

Symptomatic carotid stenosis is associated with a 3‐fold risk of early stroke recurrence compared to other stroke subtypes. Current carotid imaging techniques rely on estimating plaque‐related lumen narrowing but do not evaluate intraplaque inflammation, a key mediator of plaque rupture and thromboembolism. Using combined 18F‐fluorodeoxyglucose positron‐emission tomography (FDG‐PET)/computed tomography, we investigated the relation between inflammation‐related FDG uptake and stroke recurrence.
RecO Protein Initiates DNA Recombination and Strand Annealing through Two Alternative DNA Binding Mechanisms*

Background: RecO anneals DNA and initiates homologous recombination. Results: Binding of RecO to DNA/SSB is regulated by zinc during annealing and by RecR during recombination. Conclusion: Alternative DNA repair reactions are supported by different DNA binding mechanisms of RecO. Significance: Results explain how the same RMP supports multiple reactions during DNA repair and chromosome maintenance. Recombination mediator proteins (RMPs) are important for genome stability in all organisms. Several RMPs support two alternative reactions: initiation of homologous recombination and DNA annealing. We examined mechanisms of RMPs in both reactions with Mycobacterium smegmatis RecO (MsRecO) and demonstrated that MsRecO interacts with ssDNA by two distinct mechanisms. Zinc stimulates MsRecO binding to ssDNA during annealing, whereas the recombination function is zinc-independent and is regulated by interaction with MsRecR. Thus, different structural motifs or conformations of MsRecO are responsible for interaction with ssDNA during annealing and recombination. Neither annealing nor recombinase loading depends on MsRecO interaction with the conserved C-terminal tail of single-stranded (ss) DNA-binding protein (SSB), which is known to bind Escherichia coli RecO. However, similarly to E. coli proteins, MsRecO and MsRecOR do not dismiss SSB from ssDNA, suggesting that RMPs form a complex with SSB-ssDNA even in the absence of binding to the major protein interaction motif. We propose that alternative conformations of such complexes define the mechanism by which RMPs initiate the repair of stalled replication and support two different functions during recombinational repair of DNA breaks.
Building capacity for cross-sectorial approaches to the care of families where a parent has a mental illness

A collaborative approach to care of parents with a mental illness is essential to address the needs of the whole family. However, a number of constraints within adult mental health and child-focused services prevent a unified approach to the care of families where a parent has a mental illness (FaPMI). This paper reports on a cross-sector workforce development initiative aimed to build skill and knowledge in practitioners to work with FaPMI. The initiative included training in the ‘Keeping Families and Children in Mind’ e-resource to mental health, child and family services. The resource was combined with face to face training and small group meetings over six weeks, to help develop emerging champions/practice leaders to promote FaPMI-focused practice. A significant decline in the perceived ability to engage in FaPMI-focused practice emerged immediately post-training. The lack of perceived organisational supports was the major barrier to the implementation of FaPMI-focused practice across sectors. However following continued support of practitioners, a significant increase in FaPMI-focused practice across sectors, and a return to pre-survey levels of perceived organizational support, was observed at six-month post-training. The importance of providing additional implementation supports and time to consolidate practice change following training is highlighted. Recommended implementation and training strategies that will improve FaPMI-focused cross-sector approaches to better meet the need of FaPMI are described.
Ligands for FKBP12 Increase Ca2+ Influx and Protein Synthesis to Improve Skeletal Muscle Function*

Background: Rapamycin is a known inhibitor of protein synthesis but also modulates the activity of RyR1. Results: FKBP12 deficiency and low doses of either rapamycin or SLF increase, rather than decrease, protein synthesis and improve muscle function. Conclusion: In skeletal muscle, FKBP12 regulates Ca2+ influx, Ca2+ store refilling, and protein synthesis. Significance: This study lays the groundwork for the development of interventions to slow muscle fatigue. Rapamycin at high doses (2–10 mg/kg body weight) inhibits mammalian target of rapamycin complex 1 (mTORC1) and protein synthesis in mice. In contrast, low doses of rapamycin (10 μg/kg) increase mTORC1 activity and protein synthesis in skeletal muscle. Similar changes are found with SLF (synthetic ligand for FKBP12, which does not inhibit mTORC1) and in mice with a skeletal muscle-specific FKBP12 deficiency. These interventions also increase Ca2+ influx to enhance refilling of sarcoplasmic reticulum Ca2+ stores, slow muscle fatigue, and increase running endurance without negatively impacting cardiac function. FKBP12 deficiency or longer treatments with low dose rapamycin or SLF increase the percentage of type I fibers, further adding to fatigue resistance. We demonstrate that FKBP12 and its ligands impact multiple aspects of muscle function.
Pathogenesis of intracranial atherosclerosis

Using magnetic resonance imaging of middle cerebral artery, Xu et al report low prevalence of hemorrhage in advanced atherosclerotic lesions.1 This interesting finding is consistent with our pathologic analysis of basilar artery atherosclerosis.2 In that study, we proposed a new model of pathogenesis of basilar atherosclerosis in which the lesion is relatively benign, compared to extracranial/systemic lesions, due to paucity of neovascularity. The limited neovascularity of the intracranial atherosclerotic lesion is likely due to infrequent or absent vasa vasorum of these vessels.3,4 Reduced prevalence of intraplaque hemorrhage is a logical consequence of a paucity of plaque neovascularity. Moreover, the recent findings of good outcome for medically treated patients with intracranial atherosclerosis5 may well reflect a relatively benign lesion.
Domestic violence: solution‐focused practice with men and women who are violent

This article reviews the progress of sixty-eight adults (fifty-two men and sixteen women) on a solution-focused programme aimed at reducing domestic violence. The practice principles are described, using examples of participants' experiences. The outcomes over a three-and-a-half-year period are presented and difficulties in assessing effectiveness of outcomes are discussed.