The intrauterine device (IUD) is gaining popularity as a reversible form of contraception. Ultrasonography serves as first-line imaging for the evaluation of IUD position in patients with pelvic pain, abnormal bleeding, or absent retrieval strings. This review highlights the imaging of both properly positioned and malpositioned IUDs. The problems associated with malpositioned IUDs include expulsion, displacement, embedment, and perforation. Management considerations depend on the severity of the malposition and the presence or absence of symptoms. Three-dimensional ultrasonography has proven to be more sensitive in the evaluation of more subtle findings of malposition, particularly side-arm embedment. Familiarity with the ultrasonographic features of properly positioned and malpositioned IUDs is essential.
OBJECTIVE: Adipose tissue (AT) inflammation is associated with systemic insulin resistance and hyperinsulinemia in obese rodents and humans. A longstanding concept is that hyperinsulinemia may promote systemic insulin resistance through downregulation of its receptor on target tissues. Here we tested the novel hypothesis that insulin also impairs systemic insulin sensitivity by specifically enhancing adipose inflammation.
METHODS: Circulating insulin levels were reduced by about 50% in diet-induced and genetically obese mice by treatments with diazoxide or streptozotocin, respectively. We then examined AT crown-like structures, macrophage markers and pro-inflammatory cytokine expression in AT. AT lipogenesis and systemic insulin sensitivity was also monitored. Conversely, insulin was infused into lean mice to determine its affects on the above parameters.
RESULTS: Lowering circulating insulin levels in obese mice by streptozotocin treatment decreased macrophage content in AT, enhancing insulin stimulated Akt phosphorylation and de novo lipogenesis (DNL). Moreover, responsiveness of blood glucose levels to injected insulin was improved by streptozotocin and diazoxide treatments of obese mice without changes in body weight. Remarkably, even in lean mice, infusion of insulin under constant euglycemic conditions stimulated expression of cytokines in AT. Consistent with these findings, insulin treatment of 3T3-L1 adipocytes caused a 10-fold increase in CCL2 mRNA levels within 6 h, which was blocked by the ERK inhibitor PD98059.
CONCLUSION: Taken together, these results indicate that obesity-associated hyperinsulinemia unexpectedly drives AT inflammation in obese mice, which in turn contributes to factors that suppress insulin-stimulated adipocyte DNL and systemic insulin sensitivity.
Murine immune responses to virus-like particle-associated pre- and postfusion forms of the respiratory syncytial virus F protein
Virus-like particles (VLPs) built on the Newcastle disease virus (NDV) core proteins, NP and M, and containing two chimeric proteins, F/F and H/G, composed of respiratory syncytial virus (RSV) fusion protein (F) and glycoprotein (G) ectodomains fused to the transmembrane and cytoplasmic domains of the NDV F and HN proteins, respectively, stimulate durable, protective RSV neutralizing antibodies in mice. Here, we report the properties of VLPs constructed to contain mutant RSV F protein ectodomains stabilized in prefusion (pre-F/F) or postfusion (post-F/F) configurations. The structures of the chimeric proteins assembled into VLPs were verified immunologically by their reactivities with a conformationally restricted anti-F protein monoclonal antibody. Following immunization of mice, without adjuvant, pre-F/F-containing VLPs induced significantly higher neutralizing antibody titers than the post-F/F-containing VLPs or the wild-type F/F-containing VLPs after a single immunization but not after prime and boost immunization. The specificities of anti-F IgG induced by the two mutant VLPs were assessed by enzyme-linked immunosorbent assay (ELISA) using soluble forms of the prefusion and postfusion forms of the F protein as targets. While both types of VLPs stimulated similar levels of IgG specific for the soluble postfusion F protein, titers of IgG specific for prefusion F induced by the pre-F/F-containing VLPs were higher than those induced by post-F/F-containing VLPs. Thus, VLPs containing a stabilized prefusion form of the RSV F protein represent a promising RSV vaccine candidate.
IMPORTANCE: The development of vaccines for respiratory syncytial virus has been hampered by a lack of understanding of the requirements for eliciting high titers of neutralizing antibodies. The results of this study suggest that particle-associated RSV F protein containing mutations that stabilize the structure in a prefusion conformation may stimulate higher titers of protective antibodies than particles containing F protein in a wild-type or postfusion conformation. These findings indicate that the prefusion F protein assembled into VLPs has the potential to produce a successful RSV vaccine candidate.
Effectiveness of Rituximab for the Treatment of Rheumatoid Arthritis in Patients with Prior Exposure to Anti-TNF: Results from the CORRONA Registry
OBJECTIVE: To characterize the real-world effectiveness of rituximab (RTX) in patients with rheumatoid arthritis.
METHODS: Clinical effectiveness at 12 months was assessed in patients who were prescribed RTX based on the Clinical Disease Activity Index (CDAI). Change in CDAI was calculated (CDAI at 12 mos minus at initiation). Achievement of remission or low disease activity (LDA; CDAI < /= 10) among those with moderate/high disease activity at the time of RTX initiation was compared based on prior anti-tumor necrosis factor agent (anti-TNF) use (1 vs > /= 2) using logistic regression models.
RESULTS: Patients (n = 265) were followed for 12 months with a mean change in CDAI of -8.1 (95% CI -9.8 - -6.4). Of the 218 patients with moderate/high disease activity at baseline, patients with 1 prior anti-TNF (baseline CDAI 25.0) demonstrated a mean change in CDAI of -10.1 (95% CI -13.2 - -7.0); patients with > /= 2 prior anti-TNF (baseline CDAI 30.0) demonstrated a mean change of -10.5 (95% CI -12.9 - -8.0). The unadjusted OR for achieving LDA/remission in patients with moderate/high disease activity at baseline exposed to > /= 2 versus 1 prior anti-TNF was 0.40 (95% CI 0.22-0.73), which was robust to 4 different adjusted models (OR range 0.38-0.44).
CONCLUSION: A good clinical response was observed in all patients; however, patients previously treated with 1 anti-TNF, who had lower baseline CDAI and a greater opportunity for clinical improvement compared with patients previously treated with > /= 2 anti-TNF, were more likely to achieve LDA/remission.
IACUC protocols can be reviewed by either the full committee or designated members. Both review methods use the principles of the 3 Rs (reduce, refine, replace) as the overarching paradigm, with federal regulations and policies providing more detailed guidance. The primary goal of this study was to determine the frequency of topics discussed by IACUC during full-committee reviews and whether the topics included those required for consideration by IACUC (for example, pain and distress, number of animals used, availability of alternatives, skill and experience of researchers). We recorded and transcribed 87 protocol discussions undergoing full-committee review at 10 academic institutions. Each transcript was coded to capture the key concepts of the discussion and analyzed for the frequency of the codes mentioned. Pain and distress was the code mentioned most often, followed by the specific procedures performed, the study design, and the completeness of the protocol form. Infrequently mentioned topics were alternatives to animal use or painful or distressful procedures, the importance of the research, and preliminary data. Not all of the topics required to be considered by the IACUC were openly discussed for all protocols, and many of the discussions were limited in their depth.
Structural analysis of a novel rabbit monoclonal antibody R53 targeting an epitope in HIV-1 gp120 C4 region critical for receptor and co-receptor binding
The fourth conserved region (C4) in the HIV-1 envelope glycoprotein (Env) gp120 is a structural element that is important for its function, as it binds to both the receptor CD4 and the co-receptor CCR5/CXCR4. It has long been known that this region is highly immunogenic and that it harbors B-cell as well as T-cell epitopes. It is the target of a number of antibodies in animal studies, which are called CD4-blockers. However, the mechanism by which the virus shields itself from such antibody responses is not known. Here, we determined the crystal structure of R53 in complex with its epitope peptide using a novel anti-C4 rabbit monoclonal antibody R53. Our data show that although the epitope of R53 covers a highly conserved sequence (433)AMYAPPI(439), it is not available in the gp120 trimer and in the CD4-bound conformation. Our results suggest a masking mechanism to explain how HIV-1 protects this critical region from the human immune system.
The diversity of antigen receptors and the specificity it underlies are the hallmarks of the cellular arm of the adaptive immune system. T and B lymphocytes are indeed truly unique in their ability to generate receptors capable of recognizing virtually any pathogen. It has been known for several decades that T lymphocytes recognize short peptides derived from degraded proteins presented by major histocompatibility complex (MHC) molecules at the cell surface. Interaction between peptide-MHC (pMHC) and the T cell receptor (TCR) is central to both thymic selection and peripheral antigen recognition. It is widely assumed that TCR diversity is required, or at least highly desirable, to provide sufficient immune coverage. However, a number of immune responses are associated with the selection of predictable, narrow, or skewed repertoires and public TCR chains. Here, we summarize the current knowledge on the formation of the TCR repertoire and its maintenance in health and disease. We also outline the various molecular mechanisms that govern the composition of the pre-selection, naive and antigen-specific TCR repertoires. Finally, we suggest that with the development of high-throughput sequencing, common TCR 'signatures' raised against specific antigens could provide important diagnostic biomarkers and surrogate predictors of disease onset, progression and outcome.
Conclusion: As physicians involved at multiple levels in the treatment of breast cancer patients, it is our duty to understand the basics of diagnostic and treatment options for our patients, but also to promote specific counseling about the potential benefits of reconstruction within the medical and general communities. Raising awareness about breast reconstruction enables more women to consider it as an option that for many will translate into an improved quality of life as a breast cancer survivor.
Introduction: Photoreceptors are highly specialized neurons that have evolved to optimally capture photons. Understanding their demise, in order to develop therapeutic strategies, has the potential to improve the quality of life of millions of patients.
Packaging of genomic DNA into nucleosomes is nearly universally conserved in eukaryotes, and many features of the nucleosome landscape are quite conserved. Nonetheless, quantitative aspects of nucleosome packaging differ between species because, for example, the average length of linker DNA between nucleosomes can differ significantly even between closely related species. We recently showed that the difference in nucleosome spacing between two Hemiascomycete species-Saccharomyces cerevisiae and Kluyveromyces lactis-is established by trans-acting factors rather than being encoded in cis in the DNA sequence. Here, we generated several S. cerevisiae strains in which endogenous copies of candidate nucleosome spacing factors are deleted and replaced with the orthologous factors from K. lactis. We find no change in nucleosome spacing in such strains in which H1 or Isw1 complexes are swapped. In contrast, the K. lactis gene encoding the ATP-dependent remodeler Chd1 was found to direct longer internucleosomal spacing in S. cerevisiae, establishing that this remodeler is partially responsible for the relatively long internucleosomal spacing observed in K. lactis. By analyzing several chimeric proteins, we find that sequence differences that contribute to the spacing activity of this remodeler are dispersed throughout the coding sequence, but that the strongest spacing effect is linked to the understudied N-terminal end of Chd1. Taken together, our data find a role for sequence evolution of a chromatin remodeler in establishing quantitative aspects of the chromatin landscape in a species-specific manner.
The small GTPase Rab8 interacts with VAMP-3 to regulate the delivery of recycling T-cell receptors to the immune synapse
IFT20, a component of the intraflagellar transport (IFT) system that controls ciliogenesis, regulates immune synapse assembly in the non-ciliated T-cell by promoting T-cell receptor (TCR) recycling. Here, we have addressed the role of Rab8 (for which there are two isoforms Rab8a and Rab8b), a small GTPase implicated in ciliogenesis, in TCR traffic to the immune synapse. We show that Rab8, which colocalizes with IFT20 in Rab11(+) endosomes, is required for TCR recycling. Interestingly, as opposed to in IFT20-deficient T-cells, TCR(+) endosomes polarized normally beneath the immune synapse membrane in the presence of dominant-negative Rab8, but were unable to undergo the final docking or fusion step. This could be accounted for by the inability of the vesicular (v)-SNARE VAMP-3 to cluster at the immune synapse in the absence of functional Rab8, which is responsible for its recruitment. Of note, and similar to in T-cells, VAMP-3 interacts with Rab8 at the base of the cilium in NIH-3T3 cells, where it regulates ciliary growth and targeting of the protein smoothened. The results identify Rab8 as a new player in vesicular traffic to the immune synapse and provide insight into the pathways co-opted by different cell types for immune synapse assembly and ciliogenesis.
Luciferin Amides Enable in Vivo Bioluminescence Detection of Endogenous Fatty Acid Amide Hydrolase Activity
Firefly luciferase is homologous to fatty acyl-CoA synthetases. We hypothesized that the firefly luciferase substrate d-luciferin and its analogs are fatty acid mimics that are ideally suited to probe the chemistry of enzymes that release fatty acid products. Here, we synthesized luciferin amides and found that these molecules are hydrolyzed to substrates for firefly luciferase by the enzyme fatty acid amide hydrolase (FAAH). In the presence of luciferase, these molecules enable highly sensitive and selective bioluminescent detection of FAAH activity in vitro, in live cells, and in vivo. The potency and tissue distribution of FAAH inhibitors can be imaged in live mice, and luciferin amides serve as exemplary reagents for greatly improved bioluminescence imaging in FAAH-expressing tissues such as the brain.
Human cytomegalovirus (HCMV) exhibits surprisingly high genomic diversity during natural infection although little is known about the limits or patterns of HCMV diversity among humans. To address this deficiency, we analyzed genomic diversity among congenitally infected infants. We show that there is an upper limit to HCMV genomic diversity in these patient samples, with approximately 25% of the genome being devoid of polymorphisms. These low diversity regions were distributed across 26 loci that were preferentially located in DNA-processing genes. Furthermore, by developing, to our knowledge, the first genome-wide mutation and recombination rate maps for HCMV, we show that genomic diversity is positively correlated with these two rates. In contrast, median levels of viral genomic diversity did not vary between putatively single or mixed strain infections. We also provide evidence that HCMV populations isolated from vascular compartments of hosts from different continents are genetically similar and that polymorphisms in glycoproteins and regulatory proteins are enriched in these viral populations. This analysis provides the most highly detailed map of HCMV genomic diversity in human hosts to date and informs our understanding of the distribution of HCMV genomic diversity within human hosts.
Quantitative Correlation of in Vivo Properties with in Vitro Assay Results: The in Vitro Binding of a Biotin-DNA Analogue Modifier with Streptavidin Predicts the in Vivo Avidin-Induced Clearability of the Analogue-Modified Antibody
Quantitative prediction of in vivo behavior using an in vitro assay would dramatically accelerate pharmaceutical development. However, studies quantitatively correlating in vivo properties with in vitro assay results are rare because of the difficulty in quantitatively understanding the in vivo behavior of an agent. We now demonstrate such a correlation as a case study based on our quantitative understanding of the in vivo chemistry. In an ongoing pretargeting project, we designed a trifunctional antibody (Ab) that concomitantly carried a biotin and a DNA analogue (hereafter termed MORF). The biotin and the MORF were fused into one structure prior to conjugation to the Ab for the concomitant attachment. Because it was known that avidin-bound Ab molecules leave the circulation rapidly, this design would theoretically allow complete clearance by avidin. The clearability of the trifunctional Ab was determined by calculating the blood MORF concentration ratio of avidin-treated Ab to non-avidin-treated Ab using mice injected with these compounds. In theory, any compromised clearability should be due to the presence of impurities. In vitro, we measured the biotinylated percentage of the Ab-reacting (MORF-biotin) superset-NH2 modifier, by addition of streptavidin to the radiolabeled (MORF-biotin) superset-NH2 samples and subsequent high-performance liquid chromatography (HPLC) analysis. On the basis of our previous quantitative understanding, we predicted that the clearability of the Ab would be equal to the biotinylation percentage measured via HPLC. We validated this prediction within a 3% difference. In addition to the high avidin-induced clearability of the trifunctional Ab (up to approximately 95%) achieved by the design, we were able to predict the required quality of the (MORF-biotin) superset-NH2 modifier for any given in vivo clearability. This approach may greatly reduce the steps and time currently required in pharmaceutical development in the process of synthesis, chemical analysis, in vitro cell study, and in vivo validation.
Sorting juvenile offenders by risk requires valid risk assessment instruments, critical tools for facilitating matching the right services to the right youth at the right time. Juvenile justice systems need to consider risk in conjunction with juveniles’ mental health, to ensure that appropriate services are provided.
FSHD2 is a rare form of facioscapulohumeral muscular dystrophy (FSHD) characterized by the absence of a contraction in the D4Z4 macrosatellite repeat region on chromosome 4q35 that is the hallmark of FSHD1. However, hypomethylation of this region is common to both subtypes. Recently, mutations in SMCHD1 combined with a permissive 4q35 allele were reported to cause FSHD2. We identified a novel p.Lys275del SMCHD1 mutation in a family affected with FSHD2 using whole-exome sequencing and linkage analysis. This mutation alters a highly conserved amino acid in the ATPase domain of SMCHD1. Subject III-11 is a male who developed asymmetrical muscle weakness characteristic of FSHD at 13 years. Physical examination revealed marked bilateral atrophy at biceps brachii, bilateral scapular winging, some asymmetrical weakness at tibialis anterior and peroneal muscles, and mild lower facial weakness. Biopsy of biceps brachii in subject II-5, the father of III-11, demonstrated lobulated fibers and dystrophic changes. Endomysial and perivascular inflammation was found, which has been reported in FSHD1 but not FSHD2. Given the previous report of SMCHD1 mutations in FSHD2 and the clinical presentations consistent with the FSHD phenotype, we conclude that the SMCHD1 mutation is the likely cause of the disease in this family.
SIGNIFICANCE: Aberrant epigenetic regulation is an integral aspect of many diseases and complex disorders. Facioscapulohumeral muscular dystrophy (FSHD), a progressive myopathy that afflicts individuals of all ages, is caused by disrupted genetic and epigenetic regulation of a macrosatellite repeat. FSHD provides a powerful model to investigate disease-relevant epigenetic modifiers and general mechanisms of epigenetic regulation that govern gene expression.
RECENT ADVANCES: In the context of a genetically permissive allele, the one aspect of FSHD that is consistent across all known cases is the aberrant epigenetic state of the disease locus. In addition, certain mutations in the chromatin regulator SMCHD1 (structural maintenance of chromosomes hinge-domain protein 1) are sufficient to cause FSHD2 and enhance disease severity in FSHD1. Thus, there are multiple pathways to generate the epigenetic dysregulation required for FSHD.
CRITICAL ISSUES: Why do some individuals with the genetic requirements for FSHD develop disease pathology, while others remain asymptomatic? Similarly, disease progression is highly variable among individuals. What are the relative contributions of genetic background and environmental factors in determining disease manifestation, progression, and severity in FSHD? What is the interplay between epigenetic factors regulating the disease locus and which, if any, are viable therapeutic targets?
FUTURE DIRECTIONS: Epigenetic regulation represents a potentially powerful therapeutic target for FSHD. Determining the epigenetic signatures that are predictive of disease severity and identifying the spectrum of disease modifiers in FSHD are vital to the development of effective therapies.
In this panel discussion on the future of data science, practicing librarians address the needs of librarians today and how to provide new generation librarians with the skills to become data librarians.
In this panel discussion on the future of data science, current library educators address the needs of librarians today and how to provide new generation librarians with the skills to become data librarians.
Objective: Publisher policies have long guided scientists on how to cite publications, but now many publishers have adopted policies for data citation and sharing. This project examines the data sharing and citation practices of MIT authors in atmospheric sciences, a field that has seen a recent rise of publisher data policies. Through a multipronged approach, we sought to understand how research groups in this discipline find and cite data used in their research, share their own produced research data, and what variables (specifically funder and publisher mandates) may alter or influence this behavior.
Methods: Through group interviews with research labs in the atmospheric sciences and a bibliometric study of their publications over the last 4 years (~200 papers), we looked to create a more holistic understanding of our researchers’ activity and data-sharing perceptions in this area.
Results: Preliminary results show that while researchers are not yet consistently providing persistent identifiers to datasets or making full data sets publicly available, they perceive their data sharing efforts to be in line with the expectations and needs of their community. Such inconsistencies in publisher language and researcher behavior highlight the need to further explore definitions of data sharing, underlying data, etc., across stakeholders.
Conclusions: This work expands our knowledge of how our researchers are interpreting and acting upon their data sharing obligations and serves as an initial partnership between our data management services and subject liaisons in order to increase our understanding of data sharing at the discipline level.