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The table lists the phosphorylation values for the array of peptides containing tyrosine or serine/threonine phosphorylation sites. This array was used to compare lysates from HME2 parental cells, TGF-beta-induced EMT and lapatinib-induced EMT.

The ability of breast cancer cells to transiently transition between epithelial and mesenchymal states contributes to their metastatic potential. Therefore, driving tumor cells into a stable mesenchymal state, as opposed to complete tumor cell eradication, presents an opportunity to pharmacologically limit disease progression by promoting an asymptomatic state of dormancy. Here, we compare a reversible model of epithelial–mesenchymal transition (EMT) induced by TGFβ to a stable mesenchymal phenotype induced by chronic exposure to the ErbB kinase inhibitor lapatinib. Only cells capable of returning to an epithelial phenotype resulted in skeletal metastasis. Gene expression analyses of the two mesenchymal states indicated similar transition expression profiles. A potently downregulated gene in both datasets was spleen tyrosine kinase (SYK). In contrast to this similar diminution in mRNA, kinome analyses using a peptide array and DNA-conjugated peptide substrates showed a robust increase in SYK activity upon TGFβ-induced EMT only. SYK was present in cytoplasmic RNA processing depots known as P-bodies formed during the onset of EMT, and SYK activity was required for autophagy-mediated clearance of P-bodies during mesenchymal–epithelial transition (MET). Genetic knockout of autophagy-related 7 (ATG7) or pharmacologic inhibition of SYK activity with fostamatinib, a clinically approved inhibitor of SYK, prevented P-body clearance and MET, inhibiting metastatic tumor outgrowth. Overall, this study suggests assessment of SYK activity as a biomarker for metastatic disease and the use of fostamatinib as a means to stabilize the latency of disseminated tumor cells.Significance:These findings present inhibition of spleen tyrosine kinase as a therapeutic option to limit breast cancer metastasis by promoting systemic tumor dormancy.See related commentary by Farrington and Narla, p. 1756

This file contain supplemental figures S1 through S4. These figures present additional immunoblot and replicate immune fluorescent frames to further support the conclusions made from the data presented in the main manuscript.

The diagnosis of Q fever can be challenging and a high index of suspicion is necessary. Within this case series, we highlight the utility of the microbial cell-free DNA next-generation sequencing or Karius Test in the timely diagnosis and management of acute Q fever.

Background Methicillin-susceptible Staphylococcus aureus (MSSA) bacteremia requires weeks of IV antimicrobial therapy for treatment with antistaphylococcal penicillins (ASP) and first-generation cephalosporins being mainstays of therapy. While effective, use of these agents in the outpatient setting requires frequent dosing along with close toxicity monitoring through the duration of therapy. Ceftriaxone is an appealing alternative given daily dosing and tolerability, but efficacy in this setting remains uncertain. We sought to review outcomes of patients who received ceftriaxone versus an ASP and first-generation cephalosporins for definitive therapy of MSSA bacteremia. Methods Following IRB approval, we reviewed medical records of a retrospective adult cohort diagnosed with MSSA bacteremia treated with ceftriaxone, cefazolin, or an ASP between January 2018 and December 2019 at Mayo Clinic campuses. Treatment was defined as the antibiotic used in the outpatient setting. The primary outcome was 90-day treatment failure, defined as a composite of mortality and microbiologic relapse. Secondary outcomes were 30-day treatment failure and individual components of treatment failure. Multivariable Cox regression analysis was performed to analyze for an association between ceftriaxone use and the primary outcome after adjusting for factors that may also impact outcomes. Kaplan-Meier curves were compared utilizing a log-rank test for survival between groups. Results A total of 223 patients with the accompanying characteristics met study criteria (Table 1). Thirty-seven received ceftriaxone (16.5%) and 186 (83.4%) were treated with cefazolin or an ASP. Ninety and 30-day treatment failure were higher in those treated with ceftriaxone (27%, 8.1%) compared to cefazolin or an ASP (8.6%, 3.8%). Multivariable analysis for risk of 90-day treatment failure revealed a hazard ratio of 2.91 for ceftriaxone (Table 2) and a significant difference in event-free survival for those treated with ceftriaxone compared to cefazolin or an ASP (Figure 1). Conclusion Patients receiving definitive therapy with ceftriaxone for MSSA bacteremia experienced a higher rate of 90 and 30-day treatment failure and a lower rate of event-free survival as compared to those treated with cefazolin or an ASP. Disclosures All Authors: No reported disclosures.

Background Tobacco cessation among patients with head/neck cancer continues to be challenging despite evidence that cessation improves treatment outcomes. The purpose of this study was to understand barriers/facilitators to tobacco cessation among patients with head/neck cancer and health care providers and to obtain perspectives toward the development of a patient-centered tobacco cessation intervention. Materials and Methods In-depth qualitative interviews with 10 health care providers and 21 patients with head/neck cancer (12 inpatients and 9 outpatients) who were current or former smokers. Results Health was a common motivator to quit among patients. Although most patients indicated that their health care provider asked and advised them to quit, they were unaware of cessation resources. Suggestions for a tobacco cessation program included involvement of former smokers, health care provider involvement/counseling, supporting written materials, and incorporating follow-up and family support. Health care providers identified patients’ anger/frustration associated with the disease, social/demographic issues, and poor quality of life as the three most frequent challenges in treating patients. Although all providers reported asking about tobacco use, 70% emphasized a lack of formal training in tobacco cessation and lack of time. Their suggestions for a cessation program included having a “quarterback” responsible for this component with support from the entire health care team and continuity between outpatient and inpatient services to promote cessation, prevent relapse, and highlight the importance of follow-up and social support. Conclusion There is great interest and need, both from patients and providers, for tobacco cessation services in the oncology setting tailored for patients with head/neck cancer in the context of cancer care. Implications for Practice Although the combination of pharmacotherapy and cognitive-behavioral intervention is the standard evidence-based treatment for tobacco dependence, it must be adapted to meet the needs and wants of patients and providers to be effective. This study provides an in-depth examination of such needs among patients with head and neck cancer and providers in the context of cancer care. Providers and patients emphasized the need of having a trained health care provider dedicated to providing tobacco cessation through seamless integration between outpatient and inpatient services as well as follow-up with an emphasis on family involvement throughout the process.

Antispike monoclonal antibody treatment of 180 B-cell-depleted patients with mild-to-moderate coronavirus disease 2019 (COVID-19) resulted in good outcomes overall, with only 12.2% progressing to severe disease, 9.4% requiring hospitalization, 0.6% requiring mechanical ventilation, no deaths within 30 days, and 1.8% developing persistent COVID-19. Antispike monoclonal antibodies appear effective in this immunocompromised population.

Purpose: Human papillomavirus (HPV) vaccination uptake continues to be low in the United States. While a recommenda­tion from a health care provider (HCP) has been shown to be associated with vaccine acceptability among parents, little is known about factors associated with hesitancy despite HCP recommendation. We exam­ined factors associated with HPV vaccine hesitancy, despite a physician recommenda­tion, among Latina immigrant mothers of daughters aged 9-12 years. Methods: As part of a group randomized trial to promote HPV vaccination between 2013 and 2016, we conducted a baseline interviewer-administered survey of mothers to assess sociodemographics, knowledge and perceived risk of cervical cancer/HPV infection, self-efficacy, and intention to vaccinate their unvaccinated daughters. Hesitancy was defined as “don’t know/not sure” (DK/NS) in response to the question: “If your daughter’s doctor recommended that she gets the HPV vaccine, would you let her get it?” Results: Of the 317 participants, 35.3% indicated hesitancy to vaccinate their daughters if their physician recommended it. Although a number of variables were as­sociated with HPV vaccine hesitancy in the univariate model, five remained significant in the final multivariable model: daughter’s health insurance status; HPV awareness; perceived risk of HPV infection for their daughters; perceived self-risk of cervical cancer; and a self-efficacy score of ability to complete the HPV vaccination series. Conclusions: A recommendation by a health care provider may be not enough to motivate Latina immigrant mothers to vaccinate their daughters. Further efforts should focus on increasing awareness re­garding HPV and cervical cancer, heighten­ing perceived risk of HPV infection among daughters and boosting self-efficacy to get their children vaccinated against HPV. Ethn Dis. 2020; 30(4):661-670; doi:10.18865/ed.30.4.661

INTRODUCTION: An algorithmic approach, termed the prolonged clot time profile (PROCT), consisting of initial screening with prothrombin time (PT) and activated partial thromboplastin time (aPTT), reflexive mixing studies if indicated, and follow-up assays depending on initial testing results, offers an efficient approach to delineate the etiology of a prolonged PT/aPTT. Herein, we present the outcomes of the PROCT in the outpatient setting. METHODS: In this retrospective study, we reviewed medical records of consecutive outpatients who had prolonged PT and/or aPTT noted in the routine coagulation laboratory and who had PROCT ordered in our institutional Special Coagulation Laboratory between 2010 and 2017. RESULTS: One hundred and six patients, median age 55 years (IQR 30-67), met our study criteria. Twenty-nine patients had normal PT/aPTT, while 77 had persistent abnormalities and underwent reflexive testing. A prolonged PT, aPTT, or PT and aPTT was noted in 27 (35%), 27 (35%), and 23 (30%) respectively. Forty-nine (64%) had an acquired condition, 17 (22%) had a congenital condition, 7 (9%) had unclear etiology, and 4 (5%) were the result of laboratory artifact. The most common known cause of an isolated prolonged PT in our study was vitamin K deficiency in 8 (10%), the most common cause of an isolated prolonged aPTT was lupus anticoagulant in 4 (5%), and the most common cause of prolonged PT and aPTT was liver disease in 11 (14%). CONCLUSION: Prolonged PT/aPTT have a wide range of causes, including artifactual prolongation or abnormalities in secondary hemostasis due to both inherited and acquired conditions.

Objective: We conducted a scoping review of the literature to assess the prevalence of hospital policies regarding emergency contraception (EC) and the frequency that U.S. emergency departments (EDs) provide EC to sexual assault survivors. Study Design: We searched PubMed, Embase, the Cochrane Library, and CINAHL from inception to January 2019 for studies that assessed access to EC for sexual assault survivors in EDs. We included English-language studies that surveyed ED staff at U.S. hospitals and reported the weighted mean of the percentage of hospitals with an EC policy for sexual assault survivors and the percentage that provided EC counseling, offered or dispensed EC onsite. Secondary outcomes were rates of testing and prophylaxis for sexually transmitted infections (STIs) and HIV. Results: From 390 articles retrieved, 14 studies met the inclusion criteria; all studies were published between 2000 and 2013. Eligible studies surveyed staff at 3,314 hospitals. Overall, 60% (weighted mean) of hospitals had a policy on EC, 75% provided EC counseling, 44% offered EC, and 62% had EC available to dispense onsite. Four studies reported secondary outcomes: 81% of hospitals provided STI testing, 84% provided STI prophylaxis, 64% provided HIV testing, and 53% provided HIV prophylaxis. Conclusion: Existing literature demonstrates significant variability in EC policies and practices for sexual assault survivors in U.S. hospital EDs. Future research should assess whether EC access for survivors has improved in ED settings as well as evaluate persistent or new barriers to access.

To explore faculty perspectives on which characteristics of high-performing clerkship students are most important when determining an honors or top grade designation for clinical performance.In 2016-2017, the authors surveyed faculty (teaching ward attendings) for internal medicine clerkships and 1 pediatrics clerkship in inpatient settings at 5 U.S. academic medical centers. Survey items were framed around competencies, 24 student characteristics, and attitudes toward evaluation. Factor analysis examined constructs defining high-performing students.Of 516 faculty invited, 319 (62%) responded. The top 5 characteristics as rated by respondents were taking ownership, clinical reasoning, curiosity, dependability, and high ethical standards (in descending order). Twenty-one characteristics fit into 3 factors (Cronbach alpha, 0.81-0.87). Clinical reasoning did not fit into a factor. Factor 1 was the most important (mean rating, 8.7/10 [95% confidence interval (CI), 8.6-8.8]). It included professionalism components (ownership, curiosity, dependability, high ethical standards), presentation and interviewing skills, seeking feedback, and documentation. Factor 2 (mean, 7.9 [95% CI, 7.7-8.0]) included aspects of teamwork and communication, such as positive attitude and comments from others. Factor 3 (mean, 7.6 [95% CI, 7.4-7.7]) addressed systems-based thinking, including patient safety and care transitions.Professionalism components, clinical reasoning, and curiosity were among the most important characteristics distinguishing high-performing clerkship students. These may represent behaviors that are highly valued, observable, and relevant to training stage. Improved definition of the characteristics associated with clinical honors would assist students, faculty, and residency program directors when interpreting clinical performance within core clerkships.

The ability of breast cancer cells to transiently transition between epithelial and mesenchymal states contributes to their metastatic potential. Therefore, driving tumor cells into a stable mesenchymal state, as opposed to complete tumor cell eradication, presents an opportunity to pharmacologically limit disease progression by promoting an asymptomatic state of dormancy. Here, we compare a reversible model of epithelial–mesenchymal transition (EMT) induced by TGFβ to a stable mesenchymal phenotype induced by chronic exposure to the ErbB kinase inhibitor lapatinib. Only cells capable of returning to an epithelial phenotype resulted in skeletal metastasis. Gene expression analyses of the two mesenchymal states indicated similar transition expression profiles. A potently downregulated gene in both datasets was spleen tyrosine kinase (SYK). In contrast to this similar diminution in mRNA, kinome analyses using a peptide array and DNA-conjugated peptide substrates showed a robust increase in SYK activity upon TGFβ-induced EMT only. SYK was present in cytoplasmic RNA processing depots known as P-bodies formed during the onset of EMT, and SYK activity was required for autophagy-mediated clearance of P-bodies during mesenchymal–epithelial transition (MET). Genetic knockout of autophagy-related 7 (ATG7) or pharmacologic inhibition of SYK activity with fostamatinib, a clinically approved inhibitor of SYK, prevented P-body clearance and MET, inhibiting metastatic tumor outgrowth. Overall, this study suggests assessment of SYK activity as a biomarker for metastatic disease and the use of fostamatinib as a means to stabilize the latency of disseminated tumor cells. Significance: These findings present inhibition of spleen tyrosine kinase as a therapeutic option to limit breast cancer metastasis by promoting systemic tumor dormancy. See related commentary by Farrington and Narla, p. 1756

OBJECTIVE Racial and ethnic disparities persist in cervical cancer cases, 90% of which are caused by the human papillomavirus (HPV). Suboptimal vaccine uptake is problematic, particularly among Latinx women, who have the highest cervical cancer incidence compared with other racial/ethnic groups. We examined the association of self-efficacy and HPV vaccination intention among Latinx immigrant mothers of unvaccinated 9- to 12-year-old girls. METHODS An interview-administered survey assessed baseline sociodemographic information, knowledge and perceived risk of cervical cancer and HPV, self-efficacy, and intention to vaccinate among 313 Latinx immigrant mothers in Alabama from 2013 to 2017 before the implementation of an intervention to promote HPV vaccination. RESULTS Participants were, on average, 35 years old, with 9 years of education, and had lived in the United States for 12 years. Mothers who perceived their daughters were at risk of HPV infection were more likely to be vaccine intent than their hesitant counterparts (p

OBJECTIVES: Examine fundamental behaviors and characteristics that attending physicians in inpatient settings utilize to identify high-performing clerkship students. METHODS: We employed written comment data from a cross-sectional survey of Internal Medicine and Pediatrics attending physicians at a single academic medical center in the southern USA. Free-text responses regarding factors that faculty consider when assigning honors grades were analyzed by four trained researchers (interrater agreement 0.87) using conventional content analysis to identify themes. RESULTS: Seventy-nine of 141 (56%) attending physicians who were surveyed provided 90 comments. Four major theme areas for recognizing higher performing clerkship students were identified: Taking Ownership of Patient Care (35%), Medical Knowledge and Clinical Reasoning (20%), Team Orientation (15%), and Awareness of Opportunities for Growth and Progress (13%). CONCLUSION: Internal Medicine and Pediatric attending physicians identified characteristics that contributed to four themes in the determination of a high-performing medical student. These findings are particularly salient, as they highlight that commitment to patients, application of clinical knowledge and skills, teamwork, and awareness of growth and progress are valued by attending physicians for identifying top performing students in inpatient settings.

Objective Although considerable emphasis is placed on the attainment of honors in core medical school clerkships, little is known about what student characteristics are used by attending physicians to earn this designation. The purpose of this study was to evaluate what values and characteristics that attending physicians consider important in the evaluation of Pediatrics and Internal Medicine clerkship students for clinical honors designation. Methods This cross-sectional survey study was framed around Accreditation Council for Graduate Medical Education (ACGME) competencies. It was administered at three tertiary care hospitals associated with one large medical school in an urban setting. Teaching ward attendings in Pediatrics and Internal Medicine who evaluated third-year medical students between 2013 and 2016 were surveyed. Results Overall, Pediatric and Internal Medicine faculty demonstrated close agreement in which competencies were most important in designating clinical honors. Both groups believed that professionalism was the most important factor and that systems-based practice and patient care were among the least important factors. The only competency with a significant difference between the two groups was systems-based practice, with Internal Medicine placing more emphasis on the coordination of patient care and understanding social determinants of health. Conclusions Professionalism, communication skills, and medical knowledge are the most important characteristics when determining clinical honors on Pediatrics and Internal Medicine clerkships.

Herein, we report the discovery of 2-amino-4-bis(aryloxybenzyl)aminobutanoic acids as novel inhibitors of ASCT2(SLC1A5)-mediated glutamine accumulation in mammalian cells. Focused library development led to two novel ASCT2 inhibitors that exhibit significantly improved potency compared with prior art in C6 (rat) and HEK293 (human) cells. The potency of leads reported here represents a 40-fold improvement over our most potent, previously reported inhibitor and represents, to our knowledge, the most potent pharmacological inhibitors of ASCT2-mediated glutamine accumulation in live cells. These and other compounds in this novel series exhibit tractable chemical properties for further development as potential therapeutic leads.

Diagnosing patients simply with heart failure or nephrotic syndrome is insufficient, and clinicians should always search for the underlying causes of these syndromes. Amyloidosis represents a rare group of diseases in which abnormal protein, namely amyloid fibrils, build up in various organs. Presentation depends on which organ systems are involved, and symptoms could include breathlessness associated with fluid overload suggestive of cardiac and/or renal involvement and diarrhoea and weight loss, suggestive of gastrointestinal involvement. The authors present a case of congestive cardiac failure and nephrotic range proteinuria in a patient with persistent fluid overload secondary toamyloid light-chain (AL) amyloidosis.

OBJECTIVES/SPECIFIC AIMS: The purpose of this study was to examine factors associated with Latina immigrant mothers’ hesitancy in having their 9-12 year old daughters vaccinated against HPV despite a physician recommendation. METHODS/STUDY POPULATION: The data analyzed for this study was from a previous effort that examined the efficacy of an intervention to promote HPV vaccination among daughters (9-12 years of age) of Latina immigrants (N=317) through a randomized trial. Baseline data among Latina immigrant mothers with unvaccinated daughters was collected prior to this intervention. Participants were surveyed on sociodemographic characteristics, knowledge and perceptions of cervical cancer and HPV, and intention to vaccinate their daughters with the HPV vaccine if recommended by the daughter’s physician. RESULTS/ANTICIPATED RESULTS: Out of 317 participants, 205 reported willingness to vaccinate following a physician recommendation (64.7%) while 112 reported hesitancy (defined as an answer of “maybe”) to vaccinate their daughters (35.3%). None of the participants indicated “no”. No sociodemographic factors were significantly associated with vaccine hesitancy except for the daughter’s health insurance status (p = 0.03). A significant difference existed in the mother’s perceived risk of cervical cancer in herself based on hesitancy and willingness to vaccination (p < 0.001). Mother’s awareness of HPV (p < 0.0001), knowledge of HPV (p < 0.01), her perceived risk of HPV infection in herself (p < 0.01) and in her daughter (p < 0.0001), and her worry about her daughter being infected with HPV (p = 0.02) were also significant. Finally, there were differences among mothers in confidence of being able to have their daughters complete all three doses of the vaccine (p < 0.0001), and having the time (p < 0.0001) and the money (p < 0.0001) to complete the vaccination series. Factors that were significant in the univariate analysis were then incorporated into a binary logistic multivariable regression. HPV knowledge score was excluded from this analysis due to its limited sample size (n = 169). The five variables that were strongly associated with the outcome of vaccine hesitancy included: HPV awareness, mother’s perceived risk of HPV infection in their daughter, confidence in the ability to receive all three shots in vaccine series, confidence in being able to afford the vaccine series, and daughter’s health insurance. All were positively correlated with intention to vaccinate except for daughter’s health insurance status. DISCUSSION/SIGNIFICANCE OF IMPACT: Despite research showing Latinos are pro-vaccination and the association between HCP provider recommendation and vaccine acceptability among parents, over 35% of Latina immigrant mothers in our study were still hesitant about having their daughters vaccinated against HPV following a physician recommendation. Factors that strongly contributed to this hesitancy included HPV awareness, mother’s perceived risk of their daughter being infected with HPV, self-efficacy (ability to receive all required shots and finding time to complete the HPV vaccination series), and daughter’s health insurance status. These results indicate that HCP recommendation may be not enough of a driving factor to motivate Latina immigrant mothers to vaccinate their daughters against HPV. Further research efforts should focus on heightening perceived risk of HPV infection, improving knowledge of HPV, and boosting their self-efficacy to get their children vaccinated against HPV.

Background: Prothrombin time (PT) and activated partial thromboplastin time (aPTT) are screening tests of hemostasis commonly ordered for anticoagulant therapy monitoring, evaluation of bleeding, screening for acquired or inherited deficiencies of coagulation factors, and preoperative screening. Evaluation of a prolonged PT/aPTT is best accomplished in an algorithmic fashion consisting of mixing studies and factors assays or inhibitor testing as indicated. However, there is little data available on frequency of conditions that may result in abnormalities of PT/aPTT. The Special Coagulation Laboratory (SCL) at our institution offers an algorithmic approach termed Prolonged PT/aPTT Profile (PT/aPTT profile). Aim: To review our institutional experience with PT/aPTT profile testing (indications and outcomes), and to provide clinical correlation of abnormal PT/aPTT tests on patients seen at our institution in an effort to explain common causes of altered hemostatic testing in outpatient clinical practice. Methods: In this retrospective study, following IRB approval, we reviewed the medical records of patients who underwent outpatient PT/aPTT profile testing in the SCL at Mayo Clinic in Rochester, Minnesota between 2010 and 2017. Data abstracted included patient demographics and clinical presentation, indications for testing, use of anticoagulants, medical diagnoses which could potentially influence coagulation, and results of hemostatic testing (Table 1). Statistical analyses were completed using SAS 9.4 (SAS Institute Inc., Cary, North Carolina). Continuous variables are presented as median (range) while discrete variables are summarized as frequency (percentage). Comparisons of clinical characteristics between groups are made using the rank-sum test for continuous variables. Statistical significance is defined as a 2-tailed P value of less than 0.05. Results: A total of 107 patients met our study criteria, 58 (54 %) male, median age 55 years (range 1-89). Indications for testing included prior prolongation of PT (48%), PTT (29%), PT and aPTT (8%) on outside coagulation testing performed in a non-SCL setting, clinical bleeding episodes (10%) and preoperative screening (5%). Patients with established medical diagnoses known to influence coagulation testing including solid tumors, hematologic malignancies, liver disease, renal disease, autoimmune and rheumatologic conditions and pregnancy were documented along with other baseline patient characteristics (Table 1). SCL PT/aPTT profile testing revealed normal results in 29 of 107 (27%) patients on repeat testing while 78 patients warranted further evaluation of which 27 (35%) had an isolated prolonged PT, 28 (36%) had a prolonged APTT and 23 (29%) had prolongations of both PT and aPTT. After further workup and clinical correlation, causes of abnormal outpatient PT/aPTT profile testing were found to be related to an underlying congenital condition in 16 (21%) patients - most commonly factor VII deficiency (9%) and factor VIII deficiency (4%) - acquired conditions in 53 (68%) patients - most commonly anticoagulant therapy effects (31%) and liver disease (21%) - and of unclear etiology in 7 (9%) cases (Table 2). Specifically, the most common cause of an isolated prolonged PT in our study was found to be vitamin K deficiency (37%) whereas elevations in aPTT and both PT/aPTT were most frequently attributed to the effects of anticoagulant therapy (29% and 52% respectively). Other causes of prolongation on PT/aPTT profile testing following clinical correlation were also identified (Table 2). Discussion: While PT and aPTT are commonly ordered tests, there is little evidence regarding epidemiology of testing indications, distribution of test results, and the clinical correlation of test results. Our study demonstrates that abnormalities in hemostatic testing can be attributed to a wide range of causes including artefactual prolongation influenced by pretest variables or marked abnormalities in secondary hemostasis due to inherited and acquired conditions. Strengths of this study include the SCL algorithmic approach to testing to further elucidate coagulation cascade abnormalities to properly characterize conditions that pose severe hemostatic risk while limitations of this study reflect the experience of a single institution and possible referral bias. Disclosures No relevant conflicts of interest to declare.

In this study, 0.22 μm hydrophobic GVHP membrane was challenged with host bacteria, Escherichia coli (ED 8656) and bacteriophage (λ wild type). For investigation of filtration mechanism, we employed blocking laws, which are based on the profiles of t/v versus time (t) and t/v versus volume (v). For prominent demonstrating the linearity, the profiles were reconstructed. When feed contained bacteriophage or a mixture of bacteriophages and host bacteria, the profiles indicated that at the beginning of the filtration, standard blocking (pore plugging) was the dominant mechanism. Over time the bacteriophages filled the pores and formed a cake layer on the membrane surface. The presence of bacteria facilitated the cake layer build up. Nevertheless the filtration mechanism started with pore blockage and ended with cake formation. The comparison of the present study with another research project carried out by the authors indicates that the removal mechanisms do not rely upon the interactions between microorganisms. However the details of cake layer specifications depend on the nature of the microorganisms and operating conditions.

Translocation to the nucleus of diacylglycerol kinase (DGK)– ζ is dependent on a sequence homologous to the effector domain of Myristoylated Alanine Rich C-Kinase Substrate (MARCKS). These data would suggest that MARCKS could also localize to the nucleus. A single report demonstrated immunofluorescence staining of MARCKS in the nucleus; however, further experimental evidence confirming the specific domain responsible for this localization has not been reported. Here, we report that MARCKS is present in the nucleus in GBM cell lines. We then over-expressed wild-type MARCKS (WT) and MARCKS with the effector domain deleted (ΔED), both tagged with V5-epitope in a GBM cell line with low endogenous MARCKS expression (U87). We found that MARCKS-WT localized to the nucleus, while the MARCKS construct without the effector domain remained in the cytoplasm. We also found that over-expression of MARCKS-WT resulted in a significant increase in total cellular phosphatidyl-inositol (4,5) bisphosphate (PIP2) levels, consistent with prior evidence that MARCKS can regulate PIP2 levels. We also found increased staining for PIP2 in the nucleus with MARCKS-WT over-expression compared to MARCKS ΔED by immunofluorescence. Interestingly, we observed MARCKS and PIP2 co-localization in the nucleus. Lastly, we found changes in gene expression when MARCKS was not present in the nucleus (MARCKS ΔED). These data indicate that the MARCKS effector domain can function as a nuclear localization signal and that this sequence is critical for the ability of MARCKS to regulate PIP2 levels, nuclear localization, and gene expression. These data suggests a novel role for MARCKS in regulating nuclear functions such as gene expression.