Does the National Bariatric Prioritization Tool predict health outcomes?

BACKGROUND: The National Bariatric Prioritization Tool (NBPT), developed in Aotearoa New Zealand (AoNZ), has not been validated using real patient data. The aim was to determine the predictive validity of the NBPT on health outcomes. METHODS: An observational study was undertaken of consecutive patients undergoing elective bariatric surgery at Middlemore Hospital using the NBPT from December 2014 to December 2016. The primary outcome was the correlation between prioritization score and percentage total weight loss (%TWL) at 18 months follow-up, with secondary outcomes being correlation with change in HbA1c, lipids, resolution of OSA, resolution of hypertension, and reduction in arthritis medications. Equity of access was measured by the relationship to age group, gender and ethnicity. RESULTS: There were 294 patients included. There was no correlation between %TWL and prioritization score (correlation -0.09, P = 0.14). The benefit score correlated with %TWL (correlation 0.25, P < 0.0001). There were correlations between prioritization score and HbA1c reduction (correlation 0.28, P < 0.0001), resolution of OSA (correlation 0.20, P < 0.001) and resolution of hypertension (correlation 0.20, P < 0.001). There was a significant difference in prioritization score based on ethnicity, with Maori and Pasifika scoring higher than New Zealand European (P = 0.0023). CONCLUSIONS: While the NBPT does not correlate with %TWL, it may have predictive validity through correlations with improvement of comorbidities such as diabetes, OSA and hypertension. Given higher rates of obesity and comorbidities in Maori and Pasifika, the higher scores may suggest the tool may be used to achieve equity of access. Further modifications should be considered to optimize outcomes.

Effect of Scleral Contact Lens Size and Duration of Wear on Intraocular Pressure.

OBJECTIVE: To evaluate the effects of scleral lens size and the duration of wear on intraocular pressure (IOP) during lens wear. METHODS: Healthy adults were recruited for this prospective and randomized study. Intraocular pressure measurements were performed using a pneumotonometer. A block randomization was used to assign the order of scleral lens diameter of either 15.6 mm or 18.0 mm for 5-hr bilateral wear over a course of two visits. Scleral IOP (sIOP) was measured during the predetermined intervals, 1.25 hr apart, during the 5-hr scleral lens wear. Corneal IOP (cIOP) was measured before and after the scleral lens wear. The primary outcome measure was the mean change in sIOP from prelens insertion baseline. RESULTS: Corneal IOP unchanged after scleral lens removal compared with the baseline measurements ( P =0.878). Smaller and larger lenses introduced significantly higher sIOP at 2.5 hr after lens insertion with the mean (95% CI) increase of 1.16 (0.54, 1.78) mm Hg and 1.37 (0.76, 1.99) mm Hg, respectively. There was no difference in IOP change between the smaller and larger diameter lenses ( P =0.590). CONCLUSIONS: Well-fitted scleral lenses do not result in clinically significant changes in intraocular pressure during 5-hr lens wear in young and healthy individuals.

Spanish-speaking Mexican-American parents' experiences while navigating the dental care system for their children.

OBJECTIVES: The purpose of this study is to describe Mexican-American parents' experiences navigating the dental care system for their children. METHODS: Thirty in-depth qualitative interviews were conducted with Spanish-speaking caregivers of young children in an urban county of Northern California, asking about their experiences navigating dental care for their children. Interviews were digitally recorded, translated, transcribed, coded, and analyzed using standard qualitative procedures. RESULTS: Caregivers reported challenges that highlight how various aspects of navigating the health care system are elemental to oral health literacy. These included making appointments, finding a provider they trust, using their dental insurance, and communicating with the dental care provider. CONCLUSIONS: When addressing oral health literacy, it is important to consider the navigational components to improve children's oral health literacy.

Home Spirometry Telemonitoring for Early Detection of Bronchiolitis Obliterans Syndrome in Patients with Chronic Graft-versus-Host Disease.

Early detection of bronchiolitis obliterans syndrome (BOS) after allogeneic hematopoietic cell transplantation (HCT) depends on recognition of subclinical spirometric changes, which is possible only with frequent interval spirometry. We evaluated the feasibility of home monitoring of weekly spirometry via a wireless handheld device and a web monitoring portal in a cohort of high-risk patients for the detection of lung function changes preceding BOS diagnosis. In this observational study, 46 patients with chronic graft-versus-host disease or a decline in forced expiratory volume in 1 second (FEV1) of unclear etiology after allogeneic HCT were enrolled to perform weekly home spirometry with a wireless portable spirometer for a period of 1 year. Measurements were transmitted wirelessly to a Cloud-based monitoring portal. Feasibility evaluation included adherence with study procedures and an assessment of the home spirometry measurements compared with laboratory pulmonary function tests. Thirty-six patients (78%) completed 1 year of weekly monitoring. Overall adherence with weekly home spirometry measurements was 72% (interquartile range, 47% to 90%), which did not meet the predetermined threshold of 75% for high adherence. Correlation of home FEV1 with laboratory FEV1 was high, with a bias of 0.123 L (lower limit, -0.294 L; upper limit, 0.541 L), which is within acceptable limits for reliability. Of the 12 patients who were diagnosed with BOS or suspected BOS during the study period, 9 had an antecedent FEV1 decline detected by home spirometry. Our data indicate that wireless handheld spirometry performed at home in a high-risk HCT cohort is feasible for close monitoring of pulmonary function and appears to facilitate early detection of BOS.

High Incidence of Herpes Zoster After Cord Blood Hematopoietic Cell Transplant Despite Longer Duration of Antiviral Prophylaxis.

BACKGROUND: Cord blood transplant (CBT) recipients have a high incidence of herpes zoster (HZ) in the context of short-term peritransplant antiviral prophylaxis. In 2009, international guidelines recommended HZ prophylaxis for at least 1 year after hematopoietic cell transplant. The impact of longer-term antiviral prophylaxis on HZ incidence after CBT is unknown. METHODS: We retrospectively analyzed varicella zoster virus (VZV)-seropositive CBT recipients who were transplanted between 2006 and 2016. We abstracted HZ events and other variables for up to 5 years post-CBT. We calculated the cumulative incidence of HZ and used Cox proportional hazards regression to identify variables associated with HZ. RESULTS: The study cohort consisted of 227 patients. Among 1-year survivors, 91% were still receiving prophylaxis, for a median duration of 20.6 months. HZ occurred in 44 patients (19%) at a median of 23.6 months. The cumulative incidence of HZ by 1 year after CBT was 1.8% (95% confidence interval [CI], .1%-4%), but increased to 26% (95% CI, 19%-33%) by 5 years. In a multivariable analysis, acute graft-vs-host disease was associated with increased risk, whereas antiviral prophylaxis was associated with reduced risk for HZ (adjusted hazard ratio, 0.19 [95% CI, .09-.4]). There was no association between CD4+ T-cell counts at 1 year post-CBT and subsequent risk for HZ. CONCLUSIONS: We found a high incidence of HZ after CBT despite antiviral prophylaxis for > 1 year. Based on these findings, we suggest longer duration of prophylaxis for HZ after CBT. Compliance with antiviral prophylaxis, VZV-specific immune monitoring, and vaccination to mitigate HZ after CBT also require further study.

Prospective Validation of the APPEND Clinical Prediction Rule for Appendicitis: A Cohort Study.

BACKGROUND: Right iliac fossa (RIF) pain is a common referral to general surgery as acute appendicitis is one of the most common underlying diagnoses. The clinical diagnosis of appendicitis continues to challenge clinicians. Clinical prediction rules (CPRs) are one method used to improve diagnostic accuracy and reduce negative appendicectomy rates. The APPEND score is a novel CPR that was developed at Middlemore Hospital. AIM: To prospectively evaluate the performance of the APPEND CPR within a pathway dedicated to the management of RIF pain. METHODS: A comparative cohort study of the clinical pathway incorporating the APPEND CPR pain was performed from January to July 2016. This was compared to the retrospective cohort used to develop the APPEND CPR. The primary end point was negative appendicectomy rate. RESULTS: The negative appendicectomy rate in the prospective cohort was 9.2% (95% CI: 5.3%, 13.2%) compared to 19.8% (CI 16.2, 23.4%) in the retrospective cohort that did not use the APPEND CPR. After adjusting for multiple variables, the odds ratio of a negative appendicectomy was 2.33 times higher (95% CI; 1.26, 4.3, P value 0.007) in the retrospective cohort compared to the prospective cohort. An APPEND score of ≥5 was 87 % specific for ruling in appendicitis (PPV 94%) and a score of ≥1 was 100% sensitive in ruling out appendicitis (NPV 100%). CONCLUSIONS: In a comparative cohort study of an RIF pain pathway incorporating the APPEND CPR, the rate of negative appendicectomy showed a significant reduction by more than 50%.

Importance of Content and Format of Oral Health Instruction to Low-income Mexican Immigrant Parents: A Qualitative Study.

PURPOSE: This study's purpose was to explore how content and format of children's oral health instruction in the dental clinic is perceived by parents and might affect parents' knowledge and behaviors. METHODS: Thirty low-income Mexican immigrant parents of children age five years and under were recruited from dental clinics in 2015 to 2016. In-person qualitative interviews in Spanish about their children's and their own experiences of dental care and home oral hygiene practices were conducted, digitally recorded, translated, and transcribed. Data analysis involved iteratively reading text data and developing and refining codes to find common themes. RESULTS: Twenty-five of 30 parents recalled receiving oral hygiene instruction, and 18 recalled receiving nutrition instruction and were included in analyses. The format and effectiveness of instruction varied. More engaging educational approaches were recalled and described in more detail than less engaging educational approaches. As a result of oral hygiene and nutritional instruction, most parents reported changing their oral hygiene home behaviors for their children; half aimed to reduce purchasing sugary foods and drinks. CONCLUSIONS: Most parents recalled receiving oral hygiene and nutrition instruction as part of their child's dental visit and reported incorporating the instruction and recommendations they received into their children's home routine.

Integrating a Nurse-Midwife-Led Oral Health Intervention Into CenteringPregnancy Prenatal Care: Results of a Pilot Study.

INTRODUCTION: National and professional organizations recommend oral health promotion in prenatal care to improve women's oral health. However, few prenatal programs include education about oral health promotion. The objective of this study was to determine if women receiving a brief, low-cost, and sustainable educational intervention entitled CenteringPregnancy Oral Health Promotion had clinically improved oral health compared to women receiving standard CenteringPregnancy care. METHODS: Women attending CenteringPregnancy, a group prenatal care model, at 4 health centers in the San Francisco Bay Area, participated in this nonrandomized controlled pilot study in 2010 to 2011. The intervention arm received the CenteringPregnancy Oral Health Promotion intervention consisting of two 15-minute skills-based educational modules addressing maternal and infant oral health, each module presented in a separate CenteringPregnancy prenatal care session. The present analysis focused on the maternal module that included facilitated discussions and skills-building activities including proper tooth brushing. The control arm received standard CenteringPregnancy prenatal care. Dental examinations and questionnaires were administered prior to and approximately 9 weeks postintervention. Primary outcomes included the Plaque Index, percent bleeding on probing, and percent of gingival pocket depths 4 mm or greater. Secondary outcomes were self-reported oral health knowledge, attitudes (importance and self-efficacy), and behaviors (tooth brushing and flossing). Regression models tested whether pre to post changes in outcomes differed between the intervention versus the control arms. RESULTS: One hundred and one women participated in the study; 49 were in the intervention arm, and 52 were in the control arm. The control and intervention arms did not vary significantly at baseline. Significant pre to post differences were noted between the arms with significant improvements in the intervention arm for the Plaque Index, bleeding on probing, and pocket depths 4 mm or greater. DISCUSSION: Providing brief oral health education and skills-building activities within prenatal care may be effective in improving women's oral health during pregnancy. These findings provide support for developing a full-scale randomized clinical trial of the CenteringPregnancy Oral Health Promotion intervention.

Use of a Targeted Urine Proteome Assay (TUPA) to identify protein biomarkers of delayed recovery after kidney transplant.

PURPOSE: Development of delayed graft function (DGF) following kidney transplant is associated with poor outcomes. An ability to rapidly identify patients with DGF versus those with immediate graft function (IGF) may facilitate the treatment of DGF and the research needed to improve prognosis. The purpose of this study was to use a Targeted Urine Proteome Assay to identify protein biomarkers of delayed recovery from kidney transplant. EXPERIMENTAL DESIGN: Potential biomarkers were identified using the Targeted Urine Proteome (MRM) Assay to interrogate the relative DGF/IGF levels of expression of 167 proteins in urine taken 12-18 h after kidney implantation from 21 DGF, 15 SGF (slow graft function), and 16 IGF patients. An iterative Random Forest analysis approach evaluated the relative importance of each biomarker, which was then used to identify an optimum biomarker panel that provided the maximum sensitivity and specificity with the least number of biomarkers. CONCLUSIONS AND CLINICAL RELEVANCE: Four proteins were identified that together distinguished DGF with a sensitivity of 77.4%, specificity of 82.6%, and AUC of 0.891. This panel represents an important step toward identifying DGF at an early stage so that more effective treatments can be developed to improve long-term graft outcomes.

Achieving Health Equity Through Community Engagement in Translating Evidence to Policy: The San Francisco Health Improvement Partnership, 2010-2016.

BACKGROUND: The San Francisco Health Improvement Partnership (SFHIP) promotes health equity by using a novel collective impact model that blends community engagement with evidence-to-policy translational science. The model involves diverse stakeholders, including ethnic-based community health equity coalitions, the local public health department, hospitals and health systems, a health sciences university, a school district, the faith community, and others sectors. COMMUNITY CONTEXT: We report on 3 SFHIP prevention initiatives: reducing consumption of sugar sweetened beverages (SSBs), regulating retail alcohol sales, and eliminating disparities in children's oral health. METHODS: SFHIP is governed by a steering committee. Partnership working groups for each initiative collaborate to 1) develop and implement action plans emphasizing feasible, scalable, translational-science-informed interventions and 2) consider sustainability early in the planning process by including policy and structural interventions. OUTCOME: Through SFHIP's efforts, San Francisco enacted ordinances regulating sale and advertising of SSBs and a ballot measure establishing a soda tax. Most San Francisco hospitals implemented or committed to implementing healthy-beverage policies that prohibited serving or selling SSBs. SFHIP helped prevent Starbucks and Taco Bell from receiving alcohol licenses in San Francisco and helped prevent state authorization of sale of powdered alcohol. SFHIP increased the number of primary care clinics providing fluoride varnish at routine well-child visits from 3 to 14 and acquired a state waiver to allow dental clinics to be paid for dental services delivered in schools. INTERPRETATION: The SFHIP model of collective impact emphasizing community engagement and policy change accomplished many of its intermediate goals to create an environment promoting health and health equity.

A novel approach for effective integration of new faculty leadership.

PURPOSE: We report on an accelerated and effective way of assimilating a new leader into a team at a large academic dental school department. METHODS: At University of California, San Francisco (UCSF), a new Chair was recruited through a national search to lead its largest department in the School of Dentistry. Two months after arrival, the new Chair embarked on a process of leadership assimilation among her executive team, facilitated by a professional consultant. Within four weeks, team members participated in one-on-one interviews with the professional facilitator consultant and then completed the leadership assimilation questionnaire and returned it electronically to the facilitator. The facilitator then summarized all answers into themes and met with the team members without the Chair to debrief. Thereafter, the facilitator met with the Chair to discuss the major themes. Next, the Chair met with the team members in a facilitated session to discuss the results and negotiate a path forward. RESULTS: Approximately half of the feedback described the "how" of leadership: comments on communication, building relationships, building trust, and understanding UCSF history. The remaining half described the "what": comments on vision, strategy, and operations. Team members indicated that the first debriefing session was helpful to alleviate initial anxiety and to start building team spirit. The session with the Chair was perceived as open and fruitful in which team members were able to express their concerns and hopes for the Department, while the Chair showed commitment to the team and the communication process. CONCLUSION: Leader assimilation allows teams to share their expectations and anxieties with the new leader early in the relationship in an open way, before new habits and beliefs are formed. Conversely, for the leader, it effectively and efficiently allows a window into the team members' thinking at a critical time period when otherwise first impressions occur. With a safe space created for open communication, the process allowed siloed individual division leaders to move toward a cohesive group while at the same time solidifying a commitment to the success of the new leader.

Phosphoproteomic Analysis Reveals a Novel Mechanism of CaMKIIα Regulation Inversely Induced by Cocaine Memory Extinction versus Reconsolidation.

UNLABELLED: Successful addiction treatment depends on maintaining long-term abstinence, making relapse prevention an essential therapeutic goal. However, exposure to environmental cues associated with drug use often thwarts abstinence efforts by triggering drug using memories that drive craving and relapse. We sought to develop a dual approach for weakening cocaine memories through phosphoproteomic identification of targets regulated in opposite directions by memory extinction compared with reconsolidation in male Sprague-Dawley rats that had been trained to self-administer cocaine paired with an audiovisual cue. We discovered a novel, inversely regulated, memory-dependent phosphorylation event on calcium-calmodulin-dependent kinase II α (CaMKIIα) at serine (S)331. Correspondingly, extinction-associated S331 phosphorylation inhibited CaMKIIα activity. Intra-basolateral amygdala inhibition of CaMKII promoted memory extinction and disrupted reconsolidation, leading to a reduction in subsequent cue-induced reinstatement. CaMKII inhibition had no effect if the memory was neither retrieved nor extinguished. Therefore, inhibition of CaMKII represents a novel mechanism for memory-based addiction treatment that leverages both extinction enhancement and reconsolidation disruption to reduce relapse-like behavior. SIGNIFICANCE STATEMENT: Preventing relapse to drug use is an important goal for the successful treatment of addictive disorders. Relapse-prevention therapies attempt to interfere with drug-associated memories, but are often hindered by unintentional memory strengthening. In this study, we identify phosphorylation events that are bidirectionally regulated by the reconsolidation versus extinction of a cocaine-associated memory, including a novel site on CaMKIIα. Additionally, using a rodent model of addiction, we show that CaMKII inhibition in the amygdala can reduce relapse-like behavior. Together, our data supports the existence of mechanisms that can be used to enhance current strategies for addiction treatment.

Development of a Targeted Urine Proteome Assay for kidney diseases.

PURPOSE: Since human urine is the most readily available biofluid whose proteome changes in response to disease, it is a logical sample for identifying protein biomarkers for kidney diseases. EXPERIMENTAL DESIGN: Potential biomarkers were identified by using a multiproteomics workflow to compare urine proteomes of kidney transplant patients with immediate and delayed graft function. Differentially expressed proteins were identified, and corresponding stable isotope labeled internal peptide standards were synthesized for scheduled MRM. RESULTS: The Targeted Urine Proteome Assay (TUPA) was then developed by identifying those peptides for which there were at least two transitions for which interference in a urine matrix across 156 MRM runs was <30%. This resulted in an assay that monitors 224 peptides from 167 quantifiable proteins. CONCLUSIONS AND CLINICAL RELEVANCE: TUPA opens the way for using a robust mass spectrometric technology, MRM, for quantifying and validating biomarkers from among 167 urinary proteins. This approach, while developed using differentially expressed urinary proteins from patients with delayed versus immediate graft function after kidney transplant, can be expanded to include differentially expressed urinary proteins in multiple kidney diseases. Thus, TUPA could provide a single assay to help diagnose, prognose, and manage many kidney diseases.

Molecular transitions from papillomavirus infection to cervical precancer and cancer: Role of stromal estrogen receptor signaling.

To study the multistep process of cervical cancer development, we analyzed 128 frozen cervical samples spanning normalcy, increasingly severe cervical intraepithelial neoplasia (CIN1- CIN3), and cervical cancer (CxCa) from multiple perspectives, revealing a cascade of progressive changes. Compared with normal tissue, expression of many DNA replication/repair and cell proliferation genes was increased in CIN1/CIN2 lesions and further sustained in CIN3, consistent with high-risk human papillomavirus (HPV)-induced tumor suppressor inactivation. The CIN3-to-CxCa transition showed metabolic shifts, including decreased expression of mitochondrial electron transport complex components and ribosomal protein genes. Significantly, despite clinical, epidemiological, and animal model results linking estrogen and estrogen receptor alpha (ERα) to CxCa, ERα expression declined >15-fold from normalcy to cancer, showing the strongest inverse correlation of any gene with the increasing expression of p16, a marker for HPV-linked cancers. This drop in ERα in CIN and tumor cells was confirmed at the protein level. However, ERα expression in stromal cells continued throughout CxCa development. Our further studies localized stromal ERα to FSP1+, CD34+, SMA- precursor fibrocytes adjacent to normal and precancerous CIN epithelium, and FSP1-, CD34-, SMA+ activated fibroblasts in CxCas. Moreover, rank correlations with ERα mRNA identified IL-8, CXCL12, CXCL14, their receptors, and other angiogenesis and immune cell infiltration and inflammatory factors as candidates for ERα-induced stroma-tumor signaling pathways. The results indicate that estrogen signaling in cervical cancer has dramatic differences from ERα+ breast cancers, and imply that estrogen signaling increasingly proceeds indirectly through ERα in tumor-associated stromal fibroblasts.

Early-Life Stress Perturbs Key Cellular Programs in the Developing Mouse Hippocampus.

Conflicting reports are available with regard to the effects of childhood abuse and neglect on hippocampal function in children. While earlier imaging studies and some animal work have suggested that the effects of early-life stress (ELS) manifest only in adulthood, more recent studies have documented impaired hippocampal function in maltreated children and adolescents. Additional work using animal modes is needed to clarify the effects of ELS on hippocampal development. In this regard, genomic, proteomic, and molecular tools uniquely available in the mouse make it a particularly attractive model system to study this issue. However, very little work has been done so far to characterize the effects of ELS on hippocampal development in the mouse. To address this issue, we examined the effects of brief daily separation (BDS), a mouse model of ELS that impairs hippocampal-dependent memory in adulthood, on hippocampal development in 28-day-old juvenile mice. This age was chosen because it corresponds to the developmental period in which human imaging studies have revealed abnormal hippocampal development in maltreated children. Exposure to BDS caused a significant decrease in the total protein content of synaptosomes harvested from the hippocampus of 28-day-old male and female mice, suggesting that BDS impairs normal synaptic development in the juvenile hippocampus. Using a novel liquid chromatography multiple reaction monitoring mass spectrometry (LC-MRM) assay, we found decreased expression of many synaptic proteins, as well as proteins involved in axonal growth, myelination, and mitochondrial activity. Golgi staining in 28-day-old BDS mice showed an increase in the number of immature and abnormally shaped spines and a decrease in the number of mature spines in CA1 neurons, consistent with defects in synaptic maturation and synaptic pruning at this age. In 14-day-old pups, BDS deceased the expression of proteins involved in axonal growth and myelination, but did not affect the total protein content of synaptosomes harvested from the hippocampus, or protein levels of other synaptic markers. These results add two important findings to previous work in the field. First, our findings demonstrate that in 28-day-old juvenile mice, BDS impairs synaptic maturation and reduces the expression of proteins that are necessary for axonal growth, myelination, and mitochondrial function. Second, the results suggest a sequential model in which BDS impairs normal axonal growth and myelination before it disrupts synaptic maturation in the juvenile hippocampus.

Transcriptional profiling of ectoderm specification to keratinocyte fate in human embryonic stem cells.

In recent years, several studies have shed light into the processes that regulate epidermal specification and homeostasis. We previously showed that a broad-spectrum γ-secretase inhibitor DAPT promoted early keratinocyte specification in human embryonic stem cells triggered to undergo ectoderm specification. Here, we show that DAPT accelerates human embryonic stem cell differentiation and induces expression of the ectoderm protein AP2. Furthermore, we utilize RNA sequencing to identify several candidate regulators of ectoderm specification including those involved in epithelial and epidermal development in human embryonic stem cells. Genes associated with transcriptional regulation and growth factor activity are significantly enriched upon DAPT treatment during specification of human embryonic stem cells to the ectoderm lineage. The human ectoderm cell signature identified in this study contains several genes expressed in ectodermal and epithelial tissues. Importantly, these genes are also associated with skin disorders and ectodermal defects, providing a platform for understanding the biology of human epidermal keratinocyte development under diseased and homeostatic conditions.

Haptoglobin enhances cardiac transplant rejection.

RATIONALE: Early graft inflammation enhances both acute and chronic rejection of heart transplants, but it is unclear how this inflammation is initiated. OBJECTIVE: To identify specific inflammatory modulators and determine their underlying molecular mechanisms after cardiac transplantation. METHODS AND RESULTS: We used a murine heterotopic cardiac transplant model to identify inflammatory modulators of early graft inflammation. Unbiased mass spectrometric analysis of cardiac tissue before and ≤72 hours after transplantation revealed that 22 proteins including haptoglobin, a known antioxidant, are significantly upregulated in our grafts. Through the use of haptoglobin-deficient mice, we show that 80% of haptoglobin-deficient recipients treated with perioperative administration of the costimulatory blocking agent CTLA4 immunoglobulin exhibited >100-day survival of full major histocompatibility complex mismatched allografts, whereas all similarly treated wild-type recipients rejected their transplants by 21 days after transplantation. We found that haptoglobin modifies the intra-allograft inflammatory milieu by enhancing levels of the inflammatory cytokine interleukin-6 and the chemokine MIP-2 (macrophage inflammatory protein 2) but impair levels of the immunosuppressive cytokine interleukin-10. Haptoglobin also enhances dendritic cell graft recruitment and augments antidonor T-cell responses. Moreover, we confirmed that the protein is present in human cardiac allograft specimens undergoing acute graft rejection. CONCLUSIONS: Our findings provide new insights into the mechanisms of inflammation after cardiac transplantation and suggest that, in contrast to its prior reported antioxidant function in vascular inflammation, haptoglobin is an enhancer of inflammation after cardiac transplantation. Haptoglobin may also be a key component in other sterile inflammatory conditions.

Development of a highly automated and multiplexed targeted proteome pipeline and assay for 112 rat brain synaptic proteins.

We present a comprehensive workflow for large scale (>1000 transitions/run) label-free LC-MRM proteome assays. Innovations include automated MRM transition selection, intelligent retention time scheduling that improves S/N by twofold, and automatic peak modeling. Improvements to data analysis include a novel Q/C metric, normalized group area ratio, MLR normalization, weighted regression analysis, and data dissemination through the Yale protein expression database. As a proof of principle we developed a robust 90 min LC-MRM assay for mouse/rat postsynaptic density fractions which resulted in the routine quantification of 337 peptides from 112 proteins based on 15 observations per protein. Parallel analyses with stable isotope dilution peptide standards (SIS), demonstrate very high correlation in retention time (1.0) and protein fold change (0.94) between the label-free and SIS analyses. Overall, our method achieved a technical CV of 11.4% with >97.5% of the 1697 transitions being quantified without user intervention, resulting in a highly efficient, robust, and single injection LC-MRM assay.

Data Pre-Processing for Label-Free Multiple Reaction Monitoring (MRM) Experiments.

Multiple Reaction Monitoring (MRM) conducted on a triple quadrupole mass spectrometer allows researchers to quantify the expression levels of a set of target proteins. Each protein is often characterized by several unique peptides that can be detected by monitoring predetermined fragment ions, called transitions, for each peptide. Concatenating large numbers of MRM transitions into a single assay enables simultaneous quantification of hundreds of peptides and proteins. In recognition of the important role that MRM can play in hypothesis-driven research and its increasing impact on clinical proteomics, targeted proteomics such as MRM was recently selected as the Nature Method of the Year. However, there are many challenges in MRM applications, especially data pre­processing where many steps still rely on manual inspection of each observation in practice. In this paper, we discuss an analysis pipeline to automate MRM data pre­processing. This pipeline includes data quality assessment across replicated samples, outlier detection, identification of inaccurate transitions, and data normalization. We demonstrate the utility of our pipeline through its applications to several real MRM data sets.