The Dynamics of Cytoplasmic mRNA Metabolism.

For all but a few mRNAs, the dynamics of metabolism are unknown. Here, we developed an experimental and analytical framework for examining these dynamics for mRNAs from thousands of genes. mRNAs of mouse fibroblasts exit the nucleus with diverse intragenic and intergenic poly(A)-tail lengths. Once in the cytoplasm, they have a broad (1000-fold) range of deadenylation rate constants, which correspond to cytoplasmic lifetimes. Indeed, with few exceptions, degradation appears to occur primarily through deadenylation-linked mechanisms, with little contribution from either endonucleolytic cleavage or deadenylation-independent decapping. Most mRNA molecules degrade only after their tail lengths fall below 25 nt. Decay rate constants of short-tailed mRNAs vary broadly (1000-fold) and are larger for short-tailed mRNAs that have previously undergone more rapid deadenylation. This coupling helps clear rapidly deadenylated mRNAs, enabling the large range in deadenylation rate constants to impart a similarly large range in stabilities.

Conserved microRNA targeting reveals preexisting gene dosage sensitivities that shaped amniote sex chromosome evolution.

Mammalian X and Y Chromosomes evolved from an ordinary autosomal pair. Genetic decay of the Y led to X Chromosome inactivation (XCI) in females, but some Y-linked genes were retained during the course of sex chromosome evolution, and many X-linked genes did not become subject to XCI. We reconstructed gene-by-gene dosage sensitivities on the ancestral autosomes through phylogenetic analysis of microRNA (miRNA) target sites and compared these preexisting characteristics to the current status of Y-linked and X-linked genes in mammals. Preexisting heterogeneities in dosage sensitivity, manifesting as differences in the extent of miRNA-mediated repression, predicted either the retention of a Y homolog or the acquisition of XCI following Y gene decay. Analogous heterogeneities among avian Z-linked genes predicted either the retention of a W homolog or gene-specific dosage compensation following W gene decay. Genome-wide analyses of human copy number variation indicate that these heterogeneities consisted of sensitivity to both increases and decreases in dosage. We propose a model of XY/ZW evolution incorporating such preexisting dosage sensitivities in determining the evolutionary fates of individual genes. Our findings thus provide a more complete view of the role of dosage sensitivity in shaping the mammalian and avian sex chromosomes and reveal an important role for post-transcriptional regulatory sequences (miRNA target sites) in sex chromosome evolution.

Conventional immunomarkers stain a fraction of astrocytes in vitro: A comparison of rat cortical and spinal cord astrocytes in naïve and stimulated cultures.

Astrocytes are responsible for a wide variety of essential functions throughout the central nervous system. The protein markers glial fibrillary acidic protein (GFAP), glutamate aspartate transporter (GLAST), glutamate transporter-1 (GLT-1), glutamine synthetase (GS), 10-formyltetrahydrofolate dehydrogenase (ALDH1L1), and the transcription factor SOX9 are routinely used to label astrocytes in primary rodent cultures. However, GLAST, GLT-1, GS, and SOX9 are also produced by microglia and oligodendrocytes and GFAP, GLAST, GLT-1, and GS production levels are affected by astrocyte phenotypic changes associated with reactive astrogliosis. No group has performed a comprehensive immunocytochemical evaluation to quantify the percentage of cells labeled by these markers in vitro, nor compared changes in staining between cortex- and spinal cord-derived cells in naïve and stimulated cultures. Here, we quantified the percentage of cells positively stained for these six markers in astrocyte, microglia, and oligodendrocyte cultures isolated from neonatal rat cortices and spinal cords. Additionally, we incubated the astrocytes with transforming growth factor (TGF)-ß1 or TGF-ß3 to determine if the labeling of these markers is altered by these stimuli. We found that only SOX9 in cortical cultures and ALDH1L1 in spinal cord cultures labeled more than 75% of the cells in naïve and stimulated astrocyte cultures and stained less than 5% of the cells in microglia and oligodendrocyte cultures. Furthermore, significantly more cortical than spinal cord astrocytes stained for GFAP, GLAST, and ALDH1L1 in naïve cultures, whereas significantly more spinal cord than cortical astrocytes stained for GLAST and GS in TGF-ß1-treated cultures. These findings are important as variability in marker staining may lead to misinterpretation of the astrocyte response in cocultures, migration assays, or engineered disease models.

Real world treatment patterns in chronic myeloid leukemia patients newly initiated on tyrosine kinase inhibitors in an U.S. integrated healthcare system.

Purpose To evaluate treatment patterns in patients diagnosed with incident chronic myelogenous leukemia (CML) newly initiating therapy with imatinib, dasatinib, or nilotinib. Patients were followed to determine switching and discontinuation rates. Factors associated with switching or discontinuation from index TKI therapy, reasons for discontinuation based on electronic chart notes, and frequency of laboratory monitoring were assessed during the follow-up period. Methods A retrospective cohort study was conducted in chronic myelogenous leukemia patients aged ≥ 18 years who were identified from the Kaiser Permanente Southern California (KPSC) Cancer Registry database during the study time period of 1 January 2007 to 12 December 2013. The index date was defined as the date of the first TKI prescription (imatinib, dasatinib, or nilotinib) identified during the study time period with no prior history of TKI use within 12 months. Patients had to have continuous membership with drug benefit eligibility and no prior history of stem cell transplant (SCT) or other cancers during the 12 months prior to the index date. Baseline characteristics were identified during 12 months prior to the index date and outcomes were identified during the follow-up period after the index date. All patients were followed from index TKI therapy until end of study time period (12 December 2014), death, stem cell transplant, or disenrollment from the health plan unless one of the following occurred first: a patient switched their index therapy, or a patient discontinued their index therapy. Forward stepwise selection multivariable logistic regression models were used to evaluate factors associated with patients who continued therapy compared to those who switched or discontinued therapy with the index TKI. Chart notes were reviewed 30 days prior and 30 days post index TKI discontinuation to evaluate reasons for discontinuation. Molecular and cytogenetic testing frequency was also assessed during the follow-up period among the different patient groups. Results Two hundred sixteen patients were identified with incident chronic myelogenous leukemia and use of TKI therapy: 189 (87.5%) received imatinib, 19 (8.8%) received dasatinib, and 8 (3.7%) received nilotinib. The mean age on index date was 53 years and 63% were male; 103 patients (48%) continued on their index therapy, while 62 patients (28%) switched, and 51 patients (24%) discontinued.

A novel use of 2-octyl-cyanoacrylate: controlling post-hemodialysis site hemorrhage.

BACKGROUND: A common complication of hemodialysis is bleeding from the dialysis site. DISCUSSION: To demonstrate the use of 2-octyl-cyanoacrylate in controlling venous bleeding associated with hemodialysis access. CONCLUSION: 2-octyl-cyanoacrylate is effective in stopping venous bleeding from hemodialysis sites.

Neutralising antibody responses against SARS-CoV-2 Omicron BA.4/5 and wild-type virus in patients with inflammatory bowel disease following three doses of COVID-19 vaccine (VIP): a prospective, multicentre, cohort study.

Background: Patients with inflammatory bowel disease (IBD) receiving anti-TNF and JAK-inhibitor therapy have attenuated responses to COVID-19 vaccination. We aimed to determine how IBD treatments affect neutralising antibody responses against the Omicron BA.4/5 variant. Methods: In this multicentre cohort study, we prospectively recruited 340 adults (69 healthy controls and 271 IBD) at nine UK hospitals between May 28, 2021 and March 29, 2022. The IBD study population was established (>12 weeks therapy) on either thiopurine (n = 63), infliximab (n = 45), thiopurine and infliximab combination therapy (n = 48), ustekinumab (n = 45), vedolizumab (n = 46) or tofacitinib (n = 24). Patients were excluded if they were being treated with any other immunosuppressive therapies. Participants had two doses of either ChAdOx1 nCoV-19 or BNT162b2 vaccines, followed by a third dose of either BNT162b2 or mRNA1273. Pseudo-neutralisation assays against SARS-CoV-2 wild-type and BA.4/5 were performed. The half maximal inhibitory concentration (NT50) of participant sera was calculated. The primary outcome was anti-SARS-CoV-2 neutralising response against wild-type virus and Omicron BA.4/5 variant after the second and third doses of anti-SARS-CoV-2 vaccine, stratified by immunosuppressive therapy, adjusting for prior infection, vaccine type, age, and interval between vaccination and blood collection. This study is registered with ISRCTN (No. 13495664). Findings: Both heterologous (first two doses adenovirus vaccine, third dose mRNA vaccine) and homologous (three doses mRNA vaccine) vaccination strategies significantly increased neutralising titres against both wild-type SARS-CoV-2 virus and the Omicron BA.4/5 variant in healthy participants and patients with IBD. Antibody titres against BA.4/5 were significantly lower than antibodies against wild-type virus in both healthy participants and patients with IBD (p < 0.0001). Multivariable models demonstrated that neutralising antibodies against BA.4/5 after three doses of vaccine were significantly lower in patients with IBD on infliximab (Geometric Mean Ratio (GMR) 0.19 [0.10, 0.36], p < 0.0001), infliximab and thiopurine combination (GMR 0.25 [0.13, 0.49], p < 0.0001) or tofacitinib (GMR 0.43 [0.20, 0.91], p = 0.028), but not in patients on thiopurine monotherapy, ustekinumab, or vedolizumab. Breakthrough infection was associated with lower neutralising antibodies against wild-type (p = 0.037) and BA.4/5 (p = 0.045). Interpretation: A third dose of a COVID-19 mRNA vaccine based on the wild-type spike glycoprotein significantly boosts neutralising antibody titres in patients with IBD. However, responses are lower against the Omicron variant BA.4/5, particularly in patients taking anti-TNF and JAK-inhibitor therapy. Breakthrough infections are associated with lower neutralising antibodies and immunosuppressed patients with IBD may receive additional benefit from bivalent vaccine boosters which target Omicron variants. Funding: Pfizer.

Multiplex method for simultaneous serological detection of porcine reproductive and respiratory syndrome virus and porcine circovirus type 2.

Porcine circovirus type 2 (PCV2) and porcine reproductive and respiratory syndrome virus (PRRSV) are major contributors to the porcine respiratory disease complex (PRDC). Routine serological diagnosis and surveillance play an important role in the prevention of PRDC, as it is a leading cause of economic losses to the swine industry. We herein describe an advanced microsphere-based immunoassay that permits the simultaneous detection of antibodies to PCV2 and PRRSV, thereby reducing the time and effort involved in testing. Recombinant PRRSV nucleoprotein antigen and the PCV2 capsid antigen were coupled to fluorophore-dyed beads with distinct spectral addresses. Weekly serum samples from 72 pigs that were experimentally exposed to either PCV2, PRRSV, or both PCV2 and PRRSV were used to validate the microbead assay (MBA) in comparison with the "gold standard" enzyme-linked immunosorbent assays. The kinetics of the PCV2- and PRRSV-specific antibody responses measured by the microbead assay were comparable to those of the standard assays; Spearman's rank correlations were 0.72 (P < 0.001) for PRRSV and 0.80 (P < 0.001) for PCV2. Diagnostic sensitivity and specificity were determined using field sera whose positive or negative status was determined by the standard tests. The diagnostic sensitivity and specificity were both 98% for PCV2 and were 91% and 93%, respectively, for PRRSV (kappa coefficients, 0.85 and 0.67 for PCV2 and PRRSV, respectively). Multiplexing did not interfere with assay performance or diagnostic sensitivity. Therefore, the described study demonstrates proof of concept for the development of more versatile and economical microbead array-based multiplex serological test panels for veterinary use.

Overcoming Immunological Challenges Limiting Capsid-Mediated Gene Therapy With Machine Learning.

A key hurdle to making adeno-associated virus (AAV) capsid mediated gene therapy broadly beneficial to all patients is overcoming pre-existing and therapy-induced immune responses to these vectors. Recent advances in high-throughput DNA synthesis, multiplexing and sequencing technologies have accelerated engineering of improved capsid properties such as production yield, packaging efficiency, biodistribution and transduction efficiency. Here we outline how machine learning, advances in viral immunology, and high-throughput measurements can enable engineering of a new generation of de-immunized capsids beyond the antigenic landscape of natural AAVs, towards expanding the therapeutic reach of gene therapy.

Culture-Confirmed Staphylococcus aureus Infection after Elective Hysterectomy: Burden of Disease and Risk Factors.

Background: Our study sought to describe the incidence of culture-confirmed postsurgical Staphylococcus aureus infection after elective hysterectomy and evaluate patient characteristics, risk factors, and economic consequences associated with Staphylococcus aureus infection. Methods: This was a retrospective cohort study of patients in the United States (≥18 years old; Kaiser Permanente health plan members) who underwent elective hysterectomy from 2007 to 2013. Hysterectomies were categorized by surgical setting (inpatient vs. outpatient) and procedure (abdominal, laparoscopic, or vaginal). We estimated the cumulative incidence of culture-confirmed Staphylococcus aureus infection (90 days post-surgery) and compared healthcare resource utilization and costs (within 120 days post-surgery) among patients with/without Staphylococcus aureus infection or with other infection. Results: Among 30,960 patients identified, 20,675 underwent inpatient hysterectomy (abdominal: 47.8%; laparoscopic: 24.8%; vaginal: 27.3%), and 10,285 underwent outpatient hysterectomy (laparoscopic: 86.1%; vaginal: 13.9%). The incidence of culture-confirmed Staphylococcus aureus infection was 0.8% and 0.4% for inpatient (abdominal: 1.2%; laparoscopic: 0.5%; vaginal: 0.2%) and outpatient (laparoscopic: 0.5%; vaginal: 0.1%) surgery, respectively. Patients with Staphylococcus aureus infection had more emergency department visits, hospitalizations, and re-operations compared with patients without infection or with non-Staphylococcus aureus infection. Mean total costs for patients with Staphylococcus aureus infection were higher (inpatient: $18,261; outpatient: $4,422) compared with patients without infection (inpatient: $6,171; p < 0.0001; outpatient: $905; p = 0.0023) or non-Staphylococcus aureus infection (inpatient: $11,207; p = 0.0117; outpatient: $3,005; p = 0.2117). Conclusions: Culture-confirmed postsurgical Staphylococcus aureus infection incidence was predominately associated with procedure type rather than surgical setting. Patients with post-surgical Staphylococcus aureus infection had higher health care utilization and costs than those without infection or with other infection types. Additional effective infection control strategies are needed to reduce the morbidity and costs associated with Staphylococcus aureus infection.

The biochemical basis of microRNA targeting efficacy.

MicroRNAs (miRNAs) act within Argonaute proteins to guide repression of messenger RNA targets. Although various approaches have provided insight into target recognition, the sparsity of miRNA-target affinity measurements has limited understanding and prediction of targeting efficacy. Here, we adapted RNA bind-n-seq to enable measurement of relative binding affinities between Argonaute-miRNA complexes and all sequences ≤12 nucleotides in length. This approach revealed noncanonical target sites specific to each miRNA, miRNA-specific differences in canonical target-site affinities, and a 100-fold impact of dinucleotides flanking each site. These data enabled construction of a biochemical model of miRNA-mediated repression, which was extended to all miRNA sequences using a convolutional neural network. This model substantially improved prediction of cellular repression, thereby providing a biochemical basis for quantitatively integrating miRNAs into gene-regulatory networks.

Safety and efficacy of intravenous immunoglobulin (Flebogamma® 10% DIF) in patients with immune thrombocytopenic purpura.

AIM: To evaluate the safety and efficacy of 10% intravenous immunoglobulin (IVIG; Flebogamma® 10% DIF) in individuals with chronic immune thrombocytopenic purpura (ITP). PATIENTS & METHODS: Patients aged 3-70 years, diagnosed with chronic ITP, received 1 g/kg IVIG over two consecutive days. RESULTS: 64 evaluable patients (51 adults, 13 children) with chronic ITP received IVIG. The primary efficacy end point (increased platelet counts from ≤20 × 109/l to ≥50 × 109/l by day 8) was achieved by 81.3% of patients; mean time to response was 1.7 days (all responders). Adverse events, mostly mild or moderate, were reported in 59 patients (92.2%). CONCLUSION: Flebogamma® 10% DIF administered over two consecutive days was safe and effective in adults and children with chronic ITP.

Primary Nonadherence to Overactive Bladder Medications in an Integrated Managed Care Health Care System.

BACKGROUND: Treatment for overactive bladder (OAB) remains suboptimal, in part because of patient nonadherence to medications. Primary nonadherence is when patients fail to pick up their initial prescriptions. OBJECTIVE: To measure primary nonadherence to OAB medications within 30 days of a first OAB prescription order using electronic medical records from a U.S. managed care health care system METHODS: A retrospective cohort study was conducted using electronic medical records from the Kaiser Permanente Southern California (KPSC) database to identify patients with new OAB prescriptions between January 1, 2007, and December 31, 2013. The index date was defined as the first order of an OAB prescription. Patients had to be aged ≥ 18 years on the index date and were required to have 12 months of continuous membership with drug benefit eligibility before, during, and after the index date. Patients were defined as primary nonadherent if they did not pick up their new OAB prescriptions within 30 days of the order date. Descriptive statistics and a multivariable logistic regression analysis with backward selection were conducted to identify factors associated with patients who were primary nonadherent versus adherent. RESULTS: There were 9,050 patients with a new OAB prescription order; 1,662 (18%) of these were primary nonadherent. Patients with primary nonadherence were younger in age (56.9 [SD ± 16.0] years vs. 63.9 [SD ± 14.8] years; P < 0.001) and more likely to have commercial insurance (65.9% vs. 46.2%; P < 0.001). They also had lower mean Charlson Comorbidity Index (CCI) scores (1.99 vs. 2.70; P < 0.001), fewer OAB-related comorbidities, fewer concomitant medications (P < 0.005), and fewer overall prescriptions dispensed in the previous 12 months (P < 0.001) compared with adherent patients. Significant factors such as commercial insurance (P = 0.013), race other than white (P = 0.020), CCI = 0 versus CCI ≥ 2 (P = 0.001), urinary tract infections (P < 0.001), and falls (P = 0.047) were associated with a higher likelihood of primary nonadherence versus adherence. CONCLUSIONS: Nearly 1 in 5 patients did not pick up their new OAB medications within 30 days of the order date. Knowledge of factors associated with primary nonadherence may inform strategies for improving management of OAB. DISCLOSURES: This study was supported by a research grant provided by Astellas Pharma Global Development. Rashid and Lin do not have any financial interests or potential conflict of interest with regard to the work. Vassilakis, Kristy, and Ng were employees of Astellas Pharma Global Development when this study was conducted. Study concept and design were contributed by Rashid and Ng, along with the other authors. Rashid and Lin collected the data, and data interpretation was performed by Rashid, Ng, and Lin, along with Vassilakis and Kristy. The manuscript was written by Rashid and Ng, along with Vassilakis and Lin, and revised by Rashid, Ng, and Lin.

All-Cause and Acute Pancreatitis Health Care Costs in Patients With Severe Hypertriglyceridemia.

OBJECTIVE: The aim of this study was to assess health care utilization and costs related to acute pancreatitis (AP) in patients with severe hypertriglyceridemia (sHTG) levels. METHODS: Patients with sHTG levels 1000 mg/dL or higher were identified from January 1, 2007, to June 30, 2013. The first identified incident triglyceride level was labeled as index date. All-cause, AP-related health care visits, and mean total all-cause costs in patients with and without AP were compared during 12 months postindex. A generalized linear model regression was used to compare costs while controlling for patient characteristics and comorbidities. RESULTS: Five thousand five hundred fifty sHTG patients were identified, and 5.4% of these patients developed AP during postindex. Patients with AP had significantly (P < 0.05) more all-cause outpatient visits, hospitalizations, longer length of stays during the hospital visits, and emergency department visits versus patients without AP. Mean (SD) unadjusted all-cause health care costs in the 12 months postindex were $25,343 ($33,139) for patients with AP compared with $15,195 ($24,040) for patients with no AP. The regression showed annual all-cause costs were 49.9% higher (P < 0.01) for patients with AP versus without AP. CONCLUSIONS: Patients who developed AP were associated with higher costs; managing patients with sHTG at risk of developing AP may help reduce unnecessary costs.

Severe hypertriglyceridemia and factors associated with acute pancreatitis in an integrated health care system.

OBJECTIVE: To evaluate patient characteristics, treatment patterns, comorbidities, and risk factors associated with the development of acute pancreatitis (AP) in patients with severe hypertriglyceridemia (HTG) in an integrated health care delivery system. METHODS: We identified a retrospective cohort of severe HTG patients with a fasting triglyceride level ≥ 1000 mg/dL during January 1, 2007 to June 30, 2013 (index date) in an integrated health care delivery system. Patients were aged ≥18 years on index date and had 12 months of continuous membership and drug eligibility before the index date and during postindex including index date. Baseline patient characteristics, comorbidities, and risk factors were evaluated during 12-month preindex. Outcomes such as development of AP, treatment patterns, adherence to index therapy, and change in triglyceride (TG) laboratory levels were evaluated during postindex. Descriptive statistics were used to identify differences between patients developing AP vs no development of AP. A stepwise multivariate logistic regression and backward elimination method were used to assess statistically significant predictive factors associated with development of AP vs no AP. RESULTS: We identified 5550 patients with severe HTG, and 5.4% of these patients developed AP during postindex. Patients were mostly male (≥70%) in both groups; however, younger in the AP group (45 years ± 10.6) vs no AP group (50 years ± 11.4) with P value < .0001. The AP group had higher baseline Charslon Comorbidity Index score, alcohol abuse history (42.2%), any pancreatitis history (51.5%), diabetes (47%), and hypertension (55%), vs the no AP group (P values < .05). Patients in the AP group had higher baseline mean TG levels (2148, SD ± 1578) vs the no AP group (1559, SD ± 861), P value < .0001. Over 50% of the patients were prescribed their index therapy by a primary care provider. Predictive factors associated with the development of AP included younger age, alcohol use, and prior history of any pancreatitis, hypertension, renal disease stage 4, and other prescriber specialty. From parameters estimates, for each 100 mg/dL unit of increase in the TG level above 1000 mg/dL, there was a 3 percent increase in risk of developing AP. CONCLUSIONS: Patients with severe HTG are at a higher risk of developing AP. A number of comorbidities, risk factors, and baseline TG levels are associated with an increased incidence of AP. Patients with severe HTG are underdiagnosed, undertreated and are nonadherent to their index lipid therapy. There is a need to better define optimal approaches to treating severe HTG so as to reduce the incidence of AP. Economic studies are also needed to evaluate the burden of AP on various health care systems.

Rates, factors, reasons, and economic impact associated with switching in rheumatoid arthritis patients newly initiated on biologic disease modifying anti-rheumatic drugs in an integrated healthcare system.

Objectives To identify how many RA patients newly-initiated on bDMARD therapy switch to another bDMARD during the first year of treatment; to evaluate the factors and reasons associated with bDMARD switching; and to compare the RA-related healthcare resource utilization (HCRU) and costs between switchers vs non-switchers during the post-index period. Methods A retrospective cohort study was conducted in RA patients using the Kaiser Permanente Southern California (KPSC) database with the study time period of January 1, 2007 to December 31, 2012. The index date was defined as the date of the first bDMARD prescription. Patients had to have continuous membership eligibility with drug benefit and no prior history of bDMARD during the 24 months prior to the index date. bDMARD switching was defined as a different bDMARD claim during post-index. A multivariable logistic regression model was used to evaluate factors associated with switchers vs non-switchers. Chart notes were reviewed to evaluate reasons for switching from index bDMARD. RA-related HCRU use and costs were evaluated using a generalized linear model (GLM) with gamma distribution and log link function. Results Two hundred and fifty-one patients (12%) switched from their index bDMARD to a different bDMARD during the post-index period. bDMARD switchers were more likely to be female, of Asian/Pacific race, younger than ≤65 years of age, overweight, CCI score ≤2, initiating etanercept or adalimumab, and have a commercial insurance plan compared to non-switchers. Reasons for switching were related mostly to lack or loss of efficacy (∼51%); bDMARD switchers had overall mean adjusted RA related total costs that were 25% higher (p = 0.04) compared to non-switchers. Conclusion It is important for RA patients to receive appropriate therapy and consider bDMARD with different mechanisms of action to decrease subsequent switching, and decrease overall RA related costs as shown in this study.

Economic burden related to chemotherapy-related adverse events in patients with metastatic breast cancer in an integrated health care system.

BACKGROUND: Breast cancer is treated with many different modalities, including chemotherapy that can be given as a single agent or in combination. Patients often experience adverse events from chemotherapy during the cycles of treatment which can lead to economic burden. OBJECTIVE: The objective of this study was to evaluate costs related to chemotherapy-related adverse events in patients with metastatic breast cancer (mBC) in an integrated health care delivery system. METHODS: Patients with mBC newly initiated on chemotherapy were identified and the first infusion was defined as the index date. Patients were ≥18 years old at time of index date, had at least 6 months of health plan membership and drug eligibility prior to their index date. The chemotherapy adverse events were identified after the index date and during first line of chemotherapy. Episodes of care (EOC) were created using healthcare visits. Chart review was conducted to establish whether the adverse events were related to chemotherapy. Costs were calculated for each visit, including medications related to the adverse events, and aggregated to calculate the total EOC cost. RESULTS: A total of 1,682 patients with mBC were identified after applying study criteria; 54% of these patients had one or more adverse events related to chemotherapy. After applying the EOC method, there were a total of 5,475 episodes (4,185 single episodes [76.4%] and 1,290 multiple episodes [23.6%]) related to chemotherapy-related adverse events. Within single episodes, hematological (1,387 EOC, 33.1%), musculoskeletal/pain related (1,070 EOC, 25.6%), and gastrointestinal (775 EOC, 18.5%) were the most frequent adverse events. Patients with adverse events related to single EOC with anemia and neutropenia had the highest total outpatient costs with 901 EOC ($81,991) and 187 EOC ($17,017); these patients also had highest total inpatient costs with 46 EOC ($542,798) and 16 EOC ($136,768). However, within multiple episodes, hematological (420 EOC, 32.6%), followed by infections/pyrexia (335 EOC, 25.9%) and gastrointestinal (278 EOC, 22.6%) were the most frequent adverse events. CONCLUSION: The economic burden related to chemotherapy adverse events in patients with mBC is substantial.

ORF1 but not ORF2 dependent differences are important for in vitro replication of PCV2 in porcine alveolar macrophages singularly or coinfected with PRRSV.

The objective of this study was to investigate cytokine expression and in vitro replication of porcine circovirus type 2 (PCV2) and porcine reproductive and respiratory syndrome virus (PRRSV) in pulmonary alveolar macrophages (PAMs) emphasizing PCV2 open-reading frame (ORF) origin (PCV2a or PCV2b) and PRRSV strain. Chimeric PCV2 viruses composed of different combinations of ORF1 and ORF2 of PCV2a or PCV2b (chimera PCV2a-2b and chimera PCV2b-2a) were constructed and five different PRRSV isolates were utilized: Type 1 (SD 01-08) or type 2 (NC16845b, VR-2332, MN-184, JA-142). PAMs were infected singularly or with combinations of PCV2b, PCV2a, chimera PCV2a-2b, and chimera PCV2b-2a, and one of the five PRRSV isolates. Real-time PCR was used to test PAMs (PCV2 mRNA) and supernatants (PRRSV RNA, PCV2 DNA, PCV2 mRNA) harvested at 24, 48, 72 and 96h post inoculation (hpi). Levels of IFN-γ, TNF-α and IL-10 were determined by quantitative ELISAs. PCV2 replication in PAMs was limited to groups inoculated with PCV2 strains containing ORF1 of PCV2a (PCV2a, chimera PCV2a-2b). Furthermore, in supernatants, PCV2 mRNA was only detected in groups coinfected with PRRSV regardless of strain at 48hpi supporting an enhancing effect of PRRSV on PCV2 infection. Changes in cytokine levels were minimal and associated with PRRSV strain for TNF-α. In summary, in vitro differences in PCV2 replication in PAMs inoculated with different PCV2-PRRSV combinations were independent of PCV2 ORF2 origin with minimal effects of concurrent PRRSV infection perhaps indicating that PCV2-specific changes in ORF1 may be more important than those in ORF2.

Predictive value of T-wave abnormalities at the time of emergency department presentation in patients with potential acute coronary syndromes.

OBJECTIVES: T-wave abnormalities on electrocardiograms (ECGs) are common, but their ability to predict 30-day cardiovascular outcomes at the time of emergency department (ED) presentation is unknown. The authors determined the association between T-wave abnormalities on the presenting ECG and cardiovascular outcomes within 30 days of presentation in patients with potential acute coronary syndromes (ACSs). METHODS: This was a secondary analysis of a prospective cohort study of ED patients that presented with a potential ACS. Patients were excluded if they had a prior myocardial infarction, ST-segment elevation or depressions, right or left bundle branch block, or Q-waves on the initial ECG. Data included demographics, medical and cardiac history, and ECG findings including the presence or absence of T-wave flattening, inversions of 1-5 mm, and inversions >5 mm. Investigators followed the hospital course for admitted patients, and 30-day follow-up was performed on all patients. The main outcome was a composite of death, acute myocardial infarction, revascularization, coronary stenosis greater than 50%, or a stress test with reversible ischemia. RESULTS: Of 8,298 patient visits, 5,582 met criteria for inclusion: 4,166 (74.6%) had no T-wave abnormalities, 721 (12.9%) had T-wave flattening in two or more leads, 659 (11.8%) had T-wave inversions of 1-5 mm, and 36 (0.64%) had T-wave inversions >5 mm. The composite endpoint was more common in patients with T-wave flattening (8.2% vs. 5.7%; p = 0.0001; relative risk [RR] = 1.4; 95% confidence interval [CI] = 1.1 to 1.9), T-wave inversions 1-5 mm (13.2% vs. 5.7%; p = 0.0001; RR = 2.4; 95% CI = 1.8 to 3.1), and T-wave inversions >5 mm (19.4% vs. 5.7%; p = 0.0001; RR = 3.4; 95% CI = 1.7 to 6.1), or any T-wave abnormality (10.8% vs. 5.7%; p = 0.0001; RR = 1.9; 95% CI = 1.6 to 2.3), even after adjustment for initial troponin. This association also existed in the subset of patients without known coronary artery disease. CONCLUSIONS: In patients with potential ACS presenting to the ED, T-wave abnormalities are associated with higher rates of 30-day cardiovascular events.

Changes in mortality for extremely low birth weight infants in the 1990s: implications for treatment decisions and resource use.

OBJECTIVE: Much has changed in neonatal intensive care unit (NICU) care over the past decade. High-frequency oscillation, inhaled nitric oxide, and antenatal corticosteroids are now widely available. We wondered how these medical advances had affected both the epidemiology and ethics of life and death for extremely low birth weight (ELBW) infants in the NICU. METHODS: We identified 1142 ELBW infants (birth weight [BW] < 1000 g) consecutively admitted to our NICU between 1991 and 2001. We abstracted BW, gestational age, survival or death, and length of stay in the NICU. Statistical analyses were performed by using linear regression and 2-way analysis of variance. RESULTS: Both increasing BW and later year were significantly associated with improved survival. However, for larger ELBW infants, survival was approximately 90% for the entire decade, and large-scale improvement was hardly possible. For smaller infants, greater improvements were both possible and observed, at least early in the decade. From 1991 to 1997, overall ELBW survival increased steadily (approximately 4% per year). However, from 1997 to 2001, there was no significant improvement in survival for ELBW infants. There was no change in the distribution of deaths accounted for by BW subgroups within the ELBW population from 1991 to 2001. Median length of stay for infants who eventually expired before discharge rose from 2 days in 1991 to 10 days in 2001. As a consequence, during the past decade, the percentage of infants whose outcome was "undeclared" by day of life 4 rose from 10% to 20% for ELBW infants overall and to 33% for infants with BWs of 450 to 700 g. The percentage of ELBW NICU bed-days occupied by nonsurvivors remained very low (approximately 7%) from 1991 to 2001. CONCLUSIONS: 1) Fewer infants in all ELBW subgroups are dying, compared with a decade ago, and the improvement has been most prominent for BWs of 450 to 700 g, at which mortality was and remains to be greatest. 2) This progress seems to have slowed, or even stopped, by the end of the decade. 3) Although most NICU nonsurvivors still expire early, doomed infants are lingering longer. 4) Nonsurvivors continue to occupy a constant (and extremely small) fraction of NICU bed-days.