Characterization of Active MMP9 in Chronic Inflammatory Diseases Using a Novel Anti-MMP9 Antibody.

Matrix metalloproteinase 9 (MMP9), a protease implicated in multiple diseases, is secreted as an inactive zymogen and requires proteolytic removal of the pro-domain for activation. The relative levels and functionality of the pro- and active-MMP9 isoforms in tissues are not characterized. We generated a specific antibody that distinguishes an active form of MMP9, F107-MMP9, from the inactive pro-MMP9 isoform. Using multiple in vitro assays and specimen types, we show that F107-MMP9 expression is localized and disease-specific compared with its more abundant parental pro-form. It is detected around sites of active tissue remodeling, including fistulae of inflammatory bowel and dermal fissures in hidradenitis suppurativa, and is expressed by myeloid cells, including macrophages and neutrophils. Together, our findings provide insights into the distribution and potential role of MMP9 in inflammatory diseases.

Radiographic prevalence of juvenile osteochondral conditions of the proximal interphalangeal joint of Australian Thoroughbred racehorse yearlings and associations with sales results and race performance.

Objectives: Ascertain the radiographic prevalence and variation in characteristics of juvenile osteochondral conditions (JOC) in the proximal interphalangeal joint (PIPJ) of Australian Thoroughbred racehorse yearlings. Establish whether there are any significant associations with public auction sale results and racing performance. Methods: Retrospective evaluation of 1,098 yearling repository radiograph sets. Comparison of sales results and whole career racing performance of the case group with two control groups: maternal siblings (N = 397) and yearlings without PIP JOC (N = 391). Results: 6.3% of yearlings had at least one PIPJ JOC lesion with 4.8% having subchondral lucencies of the proximal phalanx (P1SC), 0.6% with subchondral lucencies of the middle phalanx (P2SC) and 0.8% with osteochondral fragmentation (OCF). P1SC were more prevalent in forelimbs and P2SC and OCF were more commonly located in the hindlimbs. 51% of PIPJ JOC were not identified on a lateromedial projection (LM). A significantly lower proportion of horses with OCF were successfully sold at public auction (p ≤ 0.05) but there was no significant difference in sales price between the case group and controls. A lower proportion of horses with PIPJ JOC made it to the racetrack to race, although this was not statistically significant. There was no significant difference in racing performance between the case group and controls, although there was a trend toward case horses earning lower career prize money and lower prize money per race (p ≤ 0.1). Lesions located in a dorsal or palmar/plantar location on the LM projection earned a lower average prize money per race (p ≤ 0.05) than those in a central location, and showed a trend toward earning lower total prize money (p ≤ 0.1) and number of places (p ≤ 0.1). There was no significant difference in performance for horses with lesions at the medial, axial or lateral aspects of the articular surface. Clinical importance: Overall, the findings of this study indicate that the presence of PIPJ JOC in radiographs of Thoroughbred yearlings should be attributed a low to moderate risk to future racing performance, however certain lesion characteristics may be associated with decreased performance.

Uterine artery embolization during pregnancy.

Uterine artery pseudoaneurysms are very rare but serious malformations that can occur during pregnancy or postpartum. It is crucial to identify and treat them due to the morbid consequences associated with rupture. We present a case of a 27-year-old primigravid at 22 weeks 4 days with placenta previa and recent right salpingo-oophorectomy who presented with hematuria and right lower quadrant pain. A left uterine artery pseudoaneurysm was found on computed tomography, which grew from 1.3 to 1.8 cm over 2 days. During therapeutic endovascular embolization, the pseudoaneurysm was identified and the uterine artery was successfully embolized. The fetus was carried to 34 weeks 4 days. There is no medical treatment for pseudoaneurysms and the risk of rupture vs complication of embolization must be weighed on an individual basis. As shown in this case, interventions are generally recommended to prevent harm to both mother and fetus.

Using GenBank and SRA.

GenBank® and the Sequence Read Archive (SRA) are comprehensive databases of publicly available DNA sequences. GenBank contains data for 480,000 named organisms, more than 176,000 within the embryophyta, obtained through submissions from individual laboratories and batch submissions from large-scale sequencing projects. SRA contains reads from next-generation sequencing studies from over 110,000 species. Daily data exchange with the European Nucleotide Archive (ENA) in Europe and the DNA Data Bank of Japan (DDBJ) ensures worldwide coverage for both databases. GenBank and SRA data are accessible through the NCBI Entrez retrieval system that integrates these data with other data at NCBI, such as genomes, taxonomy, and the biomedical literature. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. Usage scenarios for both GenBank and SRA ranging from local and cloud analyses to online analyses supported by the NCBI web-based tools are discussed. Both GenBank and SRA, along with their related retrieval and analysis services, are available from the NCBI homepage at www.ncbi.nlm.nih.gov .

High-flow Nasal Oxygen Versus Standard Oxygen During Flexible Bronchoscopy in Lung Transplant Patients: A Randomized Controlled Trial.

BACKGROUND: Diagnostic and interventional flexible bronchoscopy (FB) is increasingly utilized in complex and high-risk patients. Patients are often sedated for comfort and procedure facilitation and hypoxia is commonly observed in this setting. We hypothesized that high-flow nasal oxygen (HFNO) would reduce the incidence of patients experiencing oxygen desaturation. METHODS: In this randomized controlled trial, postlung transplant patients booked for FB with transbronchial lung biopsy were assigned to either HFNO or low-flow nasal oxygen (LFNO). The patient and bronchoscopist were blinded to group allocation. The primary endpoint was the proportion of patients experiencing mild desaturation [peripheral oxygen saturation (SpO2)<94%]. Secondary endpoints included desaturation (SpO2<90%), the number of airway interventions required and procedure interruptions, the duration of oxygen desaturation and patient, bronchoscopist and anesthesiologist satisfaction scores. RESULTS: The trial analyzed data from 76 patients (LFNO, n=39; HFNO, n=37). HFNO reduced the proportion of patients experiencing SpO2<94% (43.2% vs. 89.7%, P<0.001) and SpO2<90% (16.2% vs. 69.2%, P<0.001). The FB was interrupted 11 times in 9 patients in the LFNO group, whereas there were no interruptions in the HFNO group. There were no differences in patient and bronchoscopist satisfaction scores between groups, anesthesiologists had higher satisfaction scores when using HFNO (P<0.001). CONCLUSION: Hypoxia occurred less commonly in postlung transplant patients receiving HFNO during FB. Further studies are warranted in other high-risk populations undergoing longer duration FB.

MMP-9 inhibition promotes anti-tumor immunity through disruption of biochemical and physical barriers to T-cell trafficking to tumors.

Matrix metalloproteinase-9 (MMP-9), whose expression is frequently dysregulated in cancer, promotes tumor growth, invasion, and metastasis by multiple mechanisms, including extracellular matrix remodeling and growth-factor and cytokine activation. We developed a monoclonal antibody against murine MMP-9, which we found decreased growth of established primary tumors in an orthotopic model of HER2-driven breast cancer (HC11-NeuT) in immunocompetent mice. RNA sequencing (RNAseq) profiling of NeuT tumors and additional mouse model tumors revealed that anti-MMP-9 treatment resulted in upregulation of immune signature pathways associated with cytotoxic T-cell response. As there is a need to boost the low response rates observed with anti-PDL1 antibody treatment in the clinical setting, we assessed the potential of anti-MMP-9 to improve T-cell response to immune checkpoint inhibitor anti-PDL1 in NeuT tumors. Anti-MMP-9 and anti-PDL1 cotreatment reduced T-cell receptor (TCR) clonality and increased TCR diversity, as detected by TCR sequencing of NeuT tumors. Flow cytometry analyses of tumors showed that the combination treatment increased the frequency of CD3+ T cells, including memory/effector CD4 and CD8 T cells, but not regulatory T cells, among tumor-infiltrating leukocytes. Moreover, in vitro enzymatic assays corroborated that MMP-9 cleaves key T-cell chemoattractant CXC receptor 3 ligands (CXC ligand [CXCL] 9, CXCL10, and CXCL11) and renders them inactive in T-cell migration assays. Consistent with our in vitro experiments, analysis of NeuT tumor protein lysates showed that anti-MMP-9 treatment increases expression of CXCL10 and other T cell-stimulating factors, such as interleukin (IL)-12p70 and IL-18. We show that inhibition of MMP-9, a key component of the tumor-promoting and immune-suppressive myeloid inflammatory milieu, increases T-helper cell 1 type cytokines, trafficking of effector/memory T cells into tumors, and intratumoral T-cell diversity.

Are Abused Women's Protective Actions Associated With Reduced Threats, Stalking, and Violence Perpetrated by Their Male Intimate Partners?

Victims of intimate partner violence may take various actions to protect themselves from their partner. This research examined the association between abused women's ( N = 755) protective strategies at baseline and her partner's threats, stalking, and moderate and severe violence 8 months later. Emergency domestic violence shelter and orders of protection significantly reduced subsequent abuse. Receiving medical treatment was associated with a significant increase in violence, and security devices (e.g., mace, changing locks) with an increase in stalking. Safety planning and other strategies had no statistical association with abuse at follow-up. Future research should continue to examine the efficacy of safety strategies.

Intimate partner sexual violence: a comparison of foreign- versus US-born physically abused Latinas.

Men's violence against women-particularly intimate partner sexual violence (IPSV)-is associated with the transmission of HIV. Men who physically abuse their female intimate partners often also sexually abuse them. Latinas are one of the fastest growing populations in the USA and at high-risk for contracting HIV, though little is known about IPSV against physically abused Latinas, including whether there is an association between nativity of the victim and the likelihood of sexual violence by intimate partners. This study examined the (1) prevalence of recent (past 6 months) IPSV against 555 physically abused, help-seeking Latinas and (2) relationship of nativity to recent IPSV. This study used data collected in 2002­2003 from participants in one major city on the East Coast and one West Coast county, who were involved in the Risk Assessment Validation (RAVE) Study. The RAVE Study assessed the accuracy of four different methods for predicting risk of future intimate partner violence. IPSV was defined as an abusive male partner physically forcing sex (rape) or making the woman have sex without a condom. Recent IPSV was reported by 38 % of the sample. Among those reporting recent IPSV, multiple assaults were common: 30%of women were raped and 51%were made to have unprotected sex six or more times during the past 6 months. IPSV was significantly associated with nativity. Physically abused Latinas who were foreign born had two times greater odds of reporting recent IPSV than physically abused Latinas born in the USA, after controlling for other demographic covariates. Exploratory post hoc analyses examining all pairwise comparisons of IPSV against Latinas born in the USA, Mexico, Central America, South America, and the Caribbean also revealed some significant differences that warrant further study with larger samples. HIV prevention efforts aimed at reducing IPSV in this population are needed.

Immigrant and nonimmigrant women: factors that predict leaving an abusive relationship.

This research used logistic regression to test components of Choice and Lamke's (1997) two-part decision-making model and Hamby's (2008) holistic risk assessment as predictors of the decision to separate from an abusive partner, comparing significant predictors for immigrant (n = 497) and nonimmigrant (n = 808) women. Findings demonstrated that immigrant women reported higher levels of perceived risks/barriers to leaving, provided some support for the use of a holistic risk assessment in understanding women's decisions to leave, and demonstrated that immigrant and nonimmigrant women have both similarities and differences in the factors that predict leaving. Clinical and policy implications are addressed.

Prevalence and correlates of suicidal behavior among adult female victims of intimate partner violence.

The prevalence and correlates of suicidal threats and attempts among 662 racially and ethnically diverse adult female victims of intimate partner violence (IPV) were studied. One in five women had threatened or attempted suicide during her lifetime. They observed that multiple logistic regression results indicated that women at greater risk of severe or potentially lethal assaults as measured by the Danger Assessment and those who reported having a chronic or disabling illness were more likely to have threatened or attempted suicide. A linear association was found between age and suicide threats/attempts, with younger women having increased odds. Finally, African American IPV victims were less likely to have threatened or attempted suicide as compared to Latina victims. Study implications are discussed.

Efficient generation and cryopreservation of cardiomyocytes derived from human embryonic stem cells.

AIM: Human embryonic stem cells (hESCs) represent a novel cell source to treat diseases such as heart failure and for use in drug screening. In this study, we aim to promote efficient generation of cardiomyocytes from hESCs by combining the current optimal techniques of controlled growth of undifferentiated cells and specific induction for cardiac differentiation. We also aim to examine whether these methods are scalable and whether the differentiated cells can be cryopreserved. METHODS & RESULTS: hESCs were maintained without conditioned medium or feeders and were sequentially treated with activin A and bone morphogenetic protein-4 in a serum-free medium. This led to differentiation into cell populations containing high percentages of cardiomyocytes. The differentiated cells expressed appropriate cardiomyocyte markers and maintained contractility in culture, and the majority of the cells displayed working chamber (atrial and ventricular) type electrophysiological properties. In addition, the cell growth and differentiation process was adaptable to large culture formats. Moreover, the cardiomyocytes survived following cryopreservation, and viable cardiac grafts were detected after transplantation of cryopreserved cells into rat hearts following myocardial infarctions. CONCLUSION: These results demonstrate that cardiomyocytes of high quality can be efficiently generated and cryopreserved using hESCs maintained in serum-free medium, a step forward towards the application of these cells to human clinical use or drug discovery.

Women with protective orders report failure to remove firearms from their abusive partners: results from an exploratory study.

AIMS: The purpose of this study was to describe the perceptions of women who sought court protection orders for domestic violence (PODV) about actions to implement laws intended to disarm their abusers. METHODS: We identified female victims of intimate partner violence (IPV) in New York and Los Angeles primarily through family courts and records of police calls for domestic violence. Of these, 782 were surveyed and asked about their experiences seeking PODV from courts, judges ordering the removal of firearms from defendants, and if firearms were actually surrendered or confiscated. RESULTS: Of the 542 victims who had obtained a PODV and knew whether their abuser owned a firearm, 82 (15%) reported that their abuser owned a firearm. Although state law either allowed or mandated judges issuing PODVs to require abusers to surrender their firearms, 21 victims (26%) reported that judges used this authority. Ten victims (12% of victims with armed abusers) reported that their abuser had either surrendered all of his firearms or had the firearms seized. When victims reported that the judge ordered their abuser to surrender his firearms, victims were more likely to report that all firearms were either surrendered by the abuser or confiscated by law enforcement. CONCLUSIONS: Based on the perceptions of the IPV victims in this study, laws designed to disarm domestic violence offenders were either poorly implemented or failed to inform victims when their abuser's firearms were surrendered or confiscated.

Intimate partner violence: development of a brief risk assessment for the emergency department.

OBJECTIVES: Women assaulted by intimate partners are frequently patients in emergency departments (EDs). Many victims and health care providers fail to take into account the potential risks of repeat partner violence. The objective of this study was to use data from a larger study of domestic violence risk assessment methods to develop a brief assessment for acute care settings to identify victims at highest risk for suffering severe injury or potentially lethal assault by an intimate partner or former partner. METHODS: Victims of intimate partner violence (IPV) were interviewed twice between 2002 and 2004. The baseline interview included the 20 items of Campbell's Danger Assessment (DA; predictor). The follow-up interview, conducted 9 months later on average, assessed abuse inflicted since the baseline interview (outcome). Multiple logistic regression was used to identify questions on the DA most predictive of severe abuse and potentially lethal assaults. Female IPV victims were recruited from New York City family courts, Los Angeles County Sheriff's Department 9-1-1 calls, New York City and Los Angeles shelters, and New York City hospitals; 666 women responded to the DA at baseline, and 60% participated in follow-up interviews. RESULTS: Severe injuries or potentially lethal assaults were experienced by 14.9% of retained study participants between the baseline and follow-up interviews. The best brief prediction instrument has five questions. A positive answer to any three questions has a sensitivity of 83% (95% confidence interval = 70.6% to 91.4%). CONCLUSIONS: This instrument can help predict which victims may be at increased risk for severe injury or potentially lethal assault and can aid clinicians in differentiating which patients require comprehensive safety interventions.

Cardiomyocytes derived from human embryonic stem cells in pro-survival factors enhance function of infarcted rat hearts.

Cardiomyocytes derived from human embryonic stem (hES) cells potentially offer large numbers of cells to facilitate repair of the infarcted heart. However, this approach has been limited by inefficient differentiation of hES cells into cardiomyocytes, insufficient purity of cardiomyocyte preparations and poor survival of hES cell-derived myocytes after transplantation. Seeking to overcome these challenges, we generated highly purified human cardiomyocytes using a readily scalable system for directed differentiation that relies on activin A and BMP4. We then identified a cocktail of pro-survival factors that limits cardiomyocyte death after transplantation. These techniques enabled consistent formation of myocardial grafts in the infarcted rat heart. The engrafted human myocardium attenuated ventricular dilation and preserved regional and global contractile function after myocardial infarction compared with controls receiving noncardiac hES cell derivatives or vehicle. The ability of hES cell-derived cardiomyocytes to partially remuscularize myocardial infarcts and attenuate heart failure encourages their study under conditions that closely match human disease.

Human embryonic stem cell-derived cardiomyocytes can be maintained in defined medium without serum.

Current procedures for the maintenance of cardiomyocytes from human embryonic stem (hES) cells rely on either co-culture with mouse cells or medium containing fetal bovine serum (FBS). Due to exposure to animal products, these methods carry the risk of potential pathogen contamination and increased immunogenicity. Additionally, FBS introduces inherent variability in the cultures due to the inevitable differences in serum lots. Here we investigated whether a defined serum-free medium containing creatine, carnitine, taurine, and insulin (CCTI) could maintain hES cell-derived cardiomyocytes. We show that hES cell-derived cardiomyocytes maintained in the CCTI medium in the absence of any feeders exhibit similar phenotypes to those maintained in serum, as indicated by the following observations: (1) comparable levels of cardiac gene transcription were found in cells grown in serum-containing medium versus those in the CCTI medium; (2) cardiomyocyte-associated proteins were expressed in cells cultured in the CCTI medium; (3) beating cells in the CCTI medium responded to pharmacological agents in a dose-dependent manner; and (4) the vast majority of the beating embryoid bodies displayed ventricular-like action potentials (APs), and the ventricular cells in serum-containing medium and the CCTI medium had indistinguishable AP properties. Therefore, culturing hES cell-derived cardiomyocytes in serum-free medium as described here should facilitate the use of the cells for in vitro and in vivo applications.

Outcomes of custody and visitation petitions when fathers are restrained by protection orders: the case of the New York family courts.

A random sample of custody and visitation petitions filed in New York City Family Courts in 1995 was used to examine outcomes of mothers' Order of Protection (OP) Petitions in relation to parents' custody and visitation petitions. Fathers restrained by OPs were more likely to secure visitation orders (64%) than not. In contrast, 80.8% of fathers' custody petitions were dismissed when they were restrained by OPs. Fathers' custody petitions were most likely to be ordered when mothers' OP petitions were withdrawn. Mothers were most likely to secure custody when their OP petitions were ordered or withdrawn. Courts rarely denied petitions. Those that did not result in court orders were either withdrawn by the petitioner or dismissed by the court (most likely because of failure of the petitioner to appear in court). This pattern has negative implications for battered women who may be vulnerable to pressure or threats from abusive ex-partners.

Basic fibroblast growth factor supports undifferentiated human embryonic stem cell growth without conditioned medium.

Previous studies have shown that prolonged propagation of undifferentiated human embryonic stem cells (hESCs) requires conditioned medium from mouse embryonic feeders (MEF-CM) as well as matrix components. Because hESCs express growth factor receptors, including those for basic fibroblast growth factor (bFGF), stem cell factor (SCF), and fetal liver tyrosine kinase-3 ligand (Flt3L), we evaluated these and other growth factors for their ability to maintain undifferentiated hESCs in the absence of conditioned medium. We found cultures maintained in bFGF alone or in combination with other factors showed characteristics similar to MEF-CM control cultures, including morphology, surface marker and transcription factor expression, telomerase activity, differentiation, and karyotypic stability. In contrast, cells in media containing Flt-3L, thrombopoietin, and SCF, individually or in combination, showed almost complete differentiation after 6 weeks in culture. These data demonstrate that hESCs can be maintained in nonconditioned medium using growth factors.

Conditionally replicative adenovirus driven by the human telomerase promoter provides broad-spectrum antitumor activity without liver toxicity.

The human telomerase reverse transcriptase (hTERT) promoter is known to selectively drive transgene expression in many human cancer cells expressing hTERT, the catalytic component of the telomerase ribonucleoprotein complex. We have created a conditionally replicative adenovirus where the viral E1A gene, which is required for viral replication, is under the control of the hTERT promoter (AdhTERTp-E1A). In vitro studies with AdhTERTp-E1A virus on a variety of normal and tumor cell lines have shown that viral genome replication and productive infection is primarily restricted to telomerase-positive tumor cells. Lytic replication was not observed in normal primary fibroblast and epithelial cell lines tested. In vivo administration of the virus into nude mice bearing human liver or prostate tumor xenografts produced significant tumor reduction and, in some cases, resulted in complete tumor regression. AdhTERTp-E1A virus did not actively express E1A in normal mouse liver, in contrast to a control oncolytic vector in which the CMV promoter (AdCMVp-E1A) was driving the E1A gene. In addition, AdhTERTp-E1A virus produced no apparent toxicity to the liver in systemically injected mice. The hTERT promoter-driven oncolytic virus also produced significantly less toxicity to freshly cultured human hepatocytes. These studies demonstrate that an oncolytic virus driven by the telomerase promoter can be used to effectively kill a wide variety of cancer cell types and has the potential to treat primary and metastatic cancer of diverse origins.