Treatment with a low pH processing aid to reduce Campylobacter counts on broiler parts.

New regulations and performance standards for Campylobacter have been implemented by the USDA - Food Safety and Inspection Service (FSIS). The objective of this study was to evaluate treatment with a low pH processing aid (CMS1 PoultrypHresh), a formulated low pH processing aid, to reduce numbers of Campylobacter which could help companies meet regulatory requirements. Two experiments (3 replicates each) were conducted. Experiment 1, in each of 3 replicates, skin-on split chicken breasts (n = 15) were obtained from a local grocery and divided into groups of 5. The skin of each part was inoculated with approximately 107 cells of a gentamicin resistant C. coli (CCGR) marker strain in an area of approximately 6.5 cm2. CCGR cells were allowed to attach for 5 min prior to treatment. Ten inoculated breasts were individually placed into separate 6 L plastic storage boxes containing either 3.5 L deionized water or PoultrypHresh solution at a pH of 1.4. Parts were subjected to agitation (bubbled air) for 25 s. After treatment, each part was removed, allowed to drain for 5 s, and placed into a plastic bag prior to mechanical rinsing with 150 mL of buffered peptone water for 60 s. Five inoculated breasts served as controls, were untreated with a dip or agitation and sampled as above. Experiment 2 procedures were repeated using skin-on thighs under the same conditions. Rinsates were collected from each chicken part, serially diluted, and plated onto Campy Cefex agar with 200 ppm gentamicin (CCGen). All plates were incubated microaerobically (5% O2, 10% CO2, 85% N2) for 48 h at 42°C, colonies were counted and the cfu/mL was log transformed. The use of PoultrypHresh on split breast produced a 99.6% reduction compared to untreated controls, while thighs showed a 99.4% reduction. This study demonstrated an approximate 3 log reduction (P < 0.05) using a 25 s air agitation treatment in PoultrypHresh at pH 1.4 with no observable damage, which will help processors meet FSIS regulations.

HIV viraemia during hepatitis B vaccination shortens the duration of protective antibody levels.

OBJECTIVES: Individuals with HIV infection often have early waning of protective antibody following hepatitis B virus (HBV) vaccination. HIV viraemia at the time of vaccination may limit the durability of serum anti-HBV surface antibody (HBsAb) levels. We investigated the relationship of HIV plasma viral load (VL) and duration of HBsAb among vaccinees enrolled in the US Military HIV Natural History Study. METHODS: We included in the study participants who had no history of prior HBV infection, who had received all HBV vaccine doses after HIV diagnosis, and who had demonstrated an initial vaccine response, defined as HBsAb ≥ 10 IU/L. Responders were retrospectively followed with serial HBV serology from the time of the last vaccine dose until the development of waning (HBsAb < 10 IU/L) or the last HBsAb measurement. Time to and risk for waning were evaluated with Kaplan-Meier survival methods and Cox proportional hazards models, respectively. RESULTS: A total of 186 initial vaccine responders were identified. During 570 person-years of observation, HBsAb waned in 52 of 186 participants (28%). The cumulative proportion maintaining HBsAb ≥ 10 IU/L was 83% at 2 years and 56% at 5 years. Participants with an undetectable VL [hazard ratio (HR) 0.37; 95% confidence interval (CI) 0.18-0.76] or with detectable VL of ≤ 10 000 copies/mL (HR 0.46; 95% CI 0.21-1.00) had reduced risk of waning. Other factors including age, number of vaccine doses, CD4 count, and receipt of highly active antiretroviral therapy (HAART) were not significantly associated with risk of waning HBsAb. CONCLUSIONS: Undetectable or low HIV VL at the time of HBV vaccination is associated with greater durability of vaccine response in patients with HIV infection.

Determinants of incident chronic kidney disease and progression in a cohort of HIV-infected persons with unrestricted access to health care.

OBJECTIVES: As socioeconomic factors may impact the risk of chronic kidney disease (CKD), we evaluated the incidence and risk factors of incident CKD among an HIV-infected cohort with universal access to health care and minimal injecting drug use (IDU). METHODS: Incident CKD was defined as an estimated glomerular filteration rate (eGFR) <60 ml/min/1.73 m(2) for ≥ 90 days. eGFR was calculated using the Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) equation. Rates were calculated per 1000 person-years (PY). Associations with outcomes were assessed using two separate Cox proportional hazard models, adjusting for baseline and time-updated covariates. RESULTS: Among 3360 participants [median age 29 years; 92% male; 44% African American (AA)] contributing 23,091 PY of follow-up, 116 developed incident CKD [5.0/1000 PY; 95% confidence interval (CI) 4.2-6.0/1000 PY]. The median first eGFR value was 97.0 mL/min/1.73 m(2) [interquartile range (IQR) 85.3-110.1 mL/min/1.73 m(2)]. Baseline factors associated with CKD included older age, lower CD4 count at HIV diagnosis [compared with CD4 count ≥ 500 cells/µL, hazard ratio (HR) 2.1 (95% CI 1.2-3.8) for CD4 count 350-499 cells/µL; HR 3.6 (95% CI 2.0-6.3) for CD4 count 201-349 cells/µL; HR 4.3 (95% CI 2.0-9.4) for CD4 count ≤ 200 cells/µL], and HIV diagnosis in the pre-highly active antiretroviral therapy (HAART) era. In the time-updated model, low nadir CD4 counts, diabetes, hepatitis B, hypertension and less HAART use were also associated with CKD. AA ethnicity was not associated with incident CKD in either model. CONCLUSIONS: The low incidence of CKD and the lack of association with ethnicity observed in this study may in part be attributable to unique features of our cohort such as younger age, early HIV diagnosis, minimal IDU, and unrestricted access to care. Lower baseline CD4 counts were significantly associated with incident CKD, suggesting early HIV diagnosis and timely introduction of HAART may reduce the burden of CKD.

Skin and soft tissue infections among HIV-infected persons in the late combination antiretroviral therapy era.

Skin and soft tissue infections (SSTIs) occur at higher rates among HIV-infected persons, but current trends and risk factors are largely undefined. We evaluated SSTIs among a prospective cohort of HIV-infected persons during the late combination antiretroviral therapy (cART) era (2006-2010). Of the 1918 HIV-infected persons evaluated, 379 (20%) developed an SSTI during a median of 3.7 years of follow-up; of these, 118 (31%) developed at least one recurrent SSTI. The incidence rate of SSTIs was 101 (95% confidence interval [CI] 93-109) cases per 1000 person-years, and rates did not significantly change during the study period. Compared with not receiving cART and having an HIV RNA level >1000 copies/mL, patients receiving cART with an HIV RNA level <1000 copies/mL had a reduced risk of an SSTI (hazard ratio 0.64, 95% CI 0.48-0.86, P < 0.01). In summary, initial and recurrent SSTIs are common among HIV-infected persons, and HIV control is associated with a lower risk of SSTIs.

Syphilis co-infection does not affect HIV disease progression.

HIV and syphilis are often seen as co-infections since they share a common mode of transmission. During episodes of syphilis, CD4 counts transiently decrease and HIV viral loads increase; however, the effect of syphilis co-infection on HIV disease progression (time to AIDS or death) is unclear. We analysed prospectively collected information on 2239 persons with estimated dates of HIV seroconversion (205 [9.2%] with confirmed syphilis and 66 [2.9%] with probable syphilis) in order to determine the effect of syphilis co-infection on HIV disease progression. In multivariate models censored at highly active antiretroviral therapy (HAART) initiation or last visit, adjusting for CD4 count, age, race, gender, and hepatitis B and C status, syphilis (confirmed + probable) was not associated with increased hazard of AIDS or death (hazard ratio 0.99, 95% CI 0.73-1.33). Treating HAART as a time-varying covariate or limiting the analysis to only confirmed syphilis cases did not significantly alter the results. Despite transient changes in CD4 counts and viral loads, syphilis does not appear to affect HIV disease progression.

Multiple imputation in a large-scale complex survey: a practical guide.

The Cancer Care Outcomes Research and Surveillance (CanCORS) Consortium is a multisite, multimode, multiwave study of the quality and patterns of care delivered to population-based cohorts of newly diagnosed patients with lung and colorectal cancer. As is typical in observational studies, missing data are a serious concern for CanCORS, following complicated patterns that impose severe challenges to the consortium investigators. Despite the popularity of multiple imputation of missing data, its acceptance and application still lag in large-scale studies with complicated data sets such as CanCORS. We use sequential regression multiple imputation, implemented in public-available software, to deal with non-response in the CanCORS surveys and construct a centralised completed database that can be easily used by investigators from multiple sites. Our work illustrates the feasibility of multiple imputation in a large-scale multiobjective survey, showing its capacity to handle complex missing data. We present the implementation process in detail as an example for practitioners and discuss some of the challenging issues which need further research.

A genome-wide analysis of biomineralization-related proteins in the sea urchin Strongylocentrotus purpuratus.

Biomineralization, the biologically controlled formation of mineral deposits, is of widespread importance in biology, medicine, and engineering. Mineralized structures are found in most metazoan phyla and often have supportive, protective, or feeding functions. Among deuterostomes, only echinoderms and vertebrates produce extensive biomineralized structures. Although skeletons appeared independently in these two groups, ancestors of the vertebrates and echinoderms may have utilized similar components of a shared genetic "toolkit" to carry out biomineralization. The present study had two goals. First, we sought to expand our understanding of the proteins involved in biomineralization in the sea urchin, a powerful model system for analyzing the basic cellular and molecular mechanisms that underlie this process. Second, we sought to shed light on the possible evolutionary relationships between biomineralization in echinoderms and vertebrates. We used several computational methods to survey the genome of the purple sea urchin Strongylocentrotus purpuratus for gene products involved in biomineralization. Our analysis has greatly expanded the collection of biomineralization-related proteins. We have found that these proteins are often members of small families encoded by genes that are clustered in the genome. Most of the proteins are sea urchin-specific; that is, they have no apparent homologues in other invertebrate deuterostomes or vertebrates. Similarly, many of the vertebrate proteins that mediate mineral deposition do not have counterparts in the S. purpuratus genome. Our findings therefore reveal substantial differences in the primary sequences of proteins that mediate biomineral formation in echinoderms and vertebrates, possibly reflecting loose constraints on the primary structures of the proteins involved. On the other hand, certain cellular and molecular processes associated with earlier events in skeletogenesis appear similar in echinoderms and vertebrates, leaving open the possibility of deeper evolutionary relationships.

The treating physician as active gatekeeper in the recruitment of research subjects.

BACKGROUND: Institutional Review Boards vary in regard to the conditions imposed on investigators concerning contacting potential subjects to participate in health-services research studies. OBJECTIVE: The impact of more active involvement of the treating physician was examined in the approval process for recruiting study subjects. DESIGN: In recruiting subjects for a Massachusetts-based, multihospital (n = 17), health-services research study of treatment patterns for early stage breast cancer that required patient interviews, four hospitals stipulated that the treating surgeon provide written permission to the investigators to allow any contact with a potential study subject for the purpose of recruitment (active physician involvement group); the remaining 13 hospitals stipulated that the treating surgeon need only respond to the investigators if contact with a potential subject was forbidden (passive physician involvement group). SUBJECTS: Of the 1401 potential subjects treated for early stage breast cancer, 697 were in the active physician involvement group and 704 were in the passive physician involvement group. MEASURES: The percentages of patients for whom contact was allowed for recruitment purposes and who enrolled in the study were determined for the active physician involvement group and the passive physician involvement group, respectively. Logistic regression models were used to assess the independent effect of physician involvement on study enrollment. RESULTS: Of the 697 patients in the active physician involvement group, contact was approved by the treating surgeon for 72% (n = 505), compared with 91% (n = 638) of the passive physician involvement group (P <0.001). After adjustment for a variety of patient, physician, and hospital-level variables, patients in the passive physician involvement group were found to be significantly more likely to be enrolled in the study (adjusted OR 2.61; 95% CI, 1.53-4.45). However, among those patients approved for investigator contact, there were no significant differences between patients who were enrolled and patients who were not enrolled in the study with regard to physician involvement in the recruitment process (adjusted OR 1.13; 95% CI, 0.70-1.81). CONCLUSION: Our findings demonstrate that more stringent IRB requirements on health services researchers to verify permission from the treating physician to access patients for recruitment purposes adversely impact on the enrollment of patients even in noninterventional research studies. Current procedures for involving the treating physician as a gatekeeper in the recruitment of research subjects may limit access to patient participation in research studies from the perspectives of both researchers and potential subjects.

A multiple imputation strategy for incomplete longitudinal data.

Longitudinal studies are commonly used to study processes of change. Because data are collected over time, missing data are pervasive in longitudinal studies, and complete ascertainment of all variables is rare. In this paper a new imputation strategy for completing longitudinal data sets is proposed. The proposed methodology makes use of shrinkage estimators for pooling information across geographic entities, and of model averaging for pooling predictions across different statistical models. Bayes factors are used to compute weights (probabilities) for a set of models considered to be reasonable for at least some of the units for which imputations must be produced, imputations are produced by draws from the predictive distributions of the missing data, and multiple imputations are used to better reflect selected sources of uncertainty in the imputation process. The imputation strategy is developed within the context of an application to completing incomplete longitudinal variables in the so-called Area Resource File. The proposed procedure is compared with several other imputation procedures in terms of inferences derived with the imputations, and the proposed methodology is demonstrated to provide valid estimates of model parameters when the completed data are analysed. Extensions to other missing data problems in longitudinal studies are straightforward so long as the missing data mechanism can be assumed to be ignorable.

Discussion of treatment options for early-stage breast cancer: effect of provider specialty on type of surgery and satisfaction.

OBJECTIVES: To describe the physicians with whom breast cancer patients discuss treatment options and assess whether discussing surgical options with a medical oncologist is associated with type of surgery and satisfaction. RESEARCH DESIGN: Medical record abstraction and survey. SUBJECTS: Women with early-stage breast cancer numbering 2,426 in two states-Massachusetts, where the rate of breast-conserving surgery is high, and Minnesota, where it is lower. MEASURES: Receipt of breast-conserving surgery and satisfaction. RESULTS: Women in Massachusetts discussed breast cancer treatments with more physicians than women in Minnesota (mean 3.5 vs. 2.8; P <0.001) and more often discussed surgical options with a medical oncologist (52% vs. 28%; P <0.001). Using propensity score analyses, in Massachusetts, discussing surgical options with a medical oncologist was not related to type of surgery (adjusted difference in rate of breast-conserving surgery: 3.9%, 95% CI -3.6% to 11.5%) but was associated with greater satisfaction (adjusted difference: 8.1, 95% CI 2.0% to 14.2%). In Minnesota, discussing surgical options with a medical oncologist was associated with breast-conserving surgery (adjusted difference: 12.6%, 95% CI 5.6% to 19.7%) with no difference in satisfaction (adjusted difference: -1.5%, 95% CI -6.8% to 3.8%). CONCLUSIONS: Outcomes associated with discussing surgical treatments with a medical oncologist vary with local care patterns. Where breast-conserving surgery is standard care, seeing a medical oncologist is not related to type of surgery, but is associated with greater satisfaction. Where it is not the standard, seeing a medical oncologist is associated with more breast-conserving surgery and equivalent satisfaction. These findings suggest that collaborative care may benefit women with respect to treatment selection or satisfaction.

Quality of ambulatory care after myocardial infarction among Medicare patients by type of insurance and region.

PURPOSE: To evaluate use of effective cardiac medications and rehabilitation after myocardial infarction in the ambulatory setting in health maintenance organizations (HMOs) and fee-for-service care, and by region. SUBJECTS AND METHODS: We surveyed elderly Medicare patients during 1996 and 1997 in California (n = 516), Florida (n = 304), and the Northeast (n = 220; Massachusetts, New York, and Pennsylvania) approximately 18 months after myocardial infarction. We assessed use of cardiac medications and rehabilitation for HMO (n = 520) and fee-for-service (n = 520) patients matched by age, sex, month of infarct, and region. RESULTS: Across all regions, similar proportions of HMO and fee-for-service patients were using aspirin (72%, n = 374 vs. 74%, n = 387), beta-blockers (38%, n = 195 vs. 32%, n = 168), angiotensin-converting enzyme inhibitors (31%, n = 159 vs. 29%, n = 148), cholesterol-lowering agents (28%, n = 146 vs. 30%, n = 157), and calcium channel blockers (31%, n = 162 vs. 31%, n = 159; all P >0.07), except in California where more HMO patients received beta-blockers (36%, n = 93 vs. 26%, n = 66, P = 0.01). In adjusted analyses, use of these drugs did not differ significantly between HMO and fee-for-service patients. Substantial regional differences were evident in the use of beta-blockers (Northeast 46%, n = 102; Florida 34%, n = 102; California 31%, n = 159) and cholesterol-lowering agents (California 35%, n = 182; Florida 24%, n = 73; Northeast 22%, n = 48; each P <0.001). Fee-for-service patients were more likely than HMO patients to receive cardiac rehabilitation in unadjusted (32%, n = 167, vs. 22%, n = 141, P = 0.001) and adjusted analyses. CONCLUSIONS: Both HMO and fee-for-service patients would likely benefit from greater use of beta-blockers and cholesterol-lowering agents. Professional fees for cardiac rehabilitation may promote increased use among fee-for-service patients. Future studies should assess the quality of ambulatory cardiac care in different types of HMOs and the reasons for geographic variations in cardiac drug use.

Impact of underuse, overuse, and discretionary use on geographic variation in the use of coronary angiography after acute myocardial infarction.

BACKGROUND: Geographic variation in the use of medical procedures has been well documented. However, it is not known whether this variation is due to differences in use when procedures are indicated, discretionary, or contraindicated. OBJECTIVES: To examine whether use of coronary angiography after acute myocardial infarction (AMI) according to appropriateness criteria varied across geographic regions and whether underuse, overuse, or discretionary use accounted for variation in overall use. DESIGN: Retrospective cohort study using data from the Cooperative Cardiovascular Project. SETTING: Ninety-five hospital referral regions. PATIENTS: There were 44,294 Medicare patients hospitalized with AMI during 1994 or 1995, classified according to appropriateness for angiography. MAIN OUTCOME MEASURE: Variation in use of angiography, as measured by the difference between high and low rates of use across regions. RESULTS: Across regions, variation in the use of angiography was similar for indications judged necessary; appropriate, but not necessary; or uncertain. Variation was lowest for indications judged unsuitable (difference between high rate and low rate across regions = 16.3%; 95% CI = 12.6%; 20.6%). The primary cause of variation in the overall rate of angiography was due to use for indications judged appropriate, but not necessary or uncertain. When variation associated with these indications was accounted for, the difference between the resulting high and low overall rates was 10.8% (9.4%, 12.4%). In contrast, variation in the overall rate remained high when underuse in necessary situations or overuse in unsuitable situations was accounted for. CONCLUSIONS: Across regions, practice was more similar for patients categorized unsuitable for angiography than for patients with other indications. Variation in overall use of angiography appeared to be driven by utilization for discretionary indications rather than by underuse or overuse. If equivalent rates across geographic areas are judged desirable, then greater effort must be directed toward defining care for patients with discretionary indications.

Validating recommendations for coronary angiography following acute myocardial infarction in the elderly: a matched analysis using propensity scores.

We determined whether adherence to recommendations for coronary angiography more than 12 h after symptom onset but prior to hospital discharge after acute myocardial infarction (AMI) resulted in better survival. Using propensity scores, we created a matched retrospective sample of 19,568 Medicare patients hospitalized with AMI during 1994-1995 in the United States. Twenty-nine percent, 36%, and 34% of patients were judged necessary, appropriate, or uncertain, respectively, for angiography while 60% of those judged necessary received the procedure during the hospitalization. The 3-year survival benefit was largest for patients rated necessary [mean survival difference (95% CI): 17.6% (15.1, 20.1)] and smallest for those rated uncertain [8.8% (6.8, 10.7)]. Angiography recommendations appear to select patients who are likely to benefit from the procedure and the consequent interventions. Because of the magnitude of the benefit and of the number of patients involved, steps should be taken to replicate these findings.

Appropriateness of coronary angiography after myocardial infarction among Medicare beneficiaries. Managed care versus fee for service.

BACKGROUND: Previous studies have documented that cardiac procedures are performed less frequently in patients enrolled in managed-care plans than in those with fee-for-service coverage. However, it is not known whether this difference is due to less frequent use of cardiac procedures when they are indicated or to less frequent use when they are not indicated. METHODS: We compared the use of coronary angiography after acute myocardial infarction among Medicare beneficiaries who had traditional fee-for-service coverage with the use among Medicare beneficiaries enrolled in managed-care plans. The analysis was adjusted for differences in demographic and clinical characteristics of the patients and for characteristics of the hospitals to which they were admitted. We studied more than 50,000 beneficiaries in seven states and evaluated their care according to guidelines proposed by the American College of Cardiology and the American Heart Association (ACC-AHA). RESULTS: Among the 44 percent of patients in both groups who had ACC-AHA class I indications (those for which angiography is useful and effective), more fee-for-service beneficiaries than managed-care enrollees underwent angiography (46 percent vs. 37 percent, P<0.001). The rate of angiography was very low among patients with class I indications who were admitted to hospitals without angiography facilities (31 percent in the fee-for-service group and 15 percent in the managed-care group, P<0.001). Among patients with class III indications (those for which angiography is not effective), the rate of use was low in both groups (approximately 13 percent). CONCLUSIONS: In situations in which angiography is thought to be useful, it is used less often among Medicare beneficiaries enrolled in managed-care plans than among those with fee-for-service coverage. Moreover, rates of use among patients with class I indications are fairly low in both groups, suggesting that there is room for improving the care of elderly patients with myocardial infarction, especially those admitted to hospitals without angiography facilities.

Single repeat deletion in ApoA-I blocks cholesterol esterification and results in rapid catabolism of delta6 and wild-type ApoA-I in transgenic mice.

The deletion mutation Delta6 apolipoprotein A-I lacks residues 143-164 or repeat 6 in the mature apoA-I protein. In vitro studies show this mutation dramatically reduces the rate of lecithin:cholesterol acyltransferase (LCAT) catalyzed cholesterol esterification. The present study was initiated to investigate the effect of this mutation on in vivo high density lipoprotein (HDL) cholesterol esterification and metabolism. Transgenic mice expressing human Delta6 apoA-I (TgDelta6 +/+) were created and then crossed with apoA-I knockout mice (-/-) to generate mice expressing only human Delta6 apoA-I (TgDelta6 -/-). Human Delta6 apoA-I was associated with homogeneous sized alpha-HDL, when wild-type mouse apoA-I was present (in TgDelta6 +/+ and +/- mice). However, in the absence of endogenous mouse apoA-I, Delta6 apoA-I was found exclusively in cholesterol ester-poor HDL, and lipid-free HDL fractions. This observation coincides with the 6-fold lower cholesterol ester mass in TgDelta6 -/- mouse plasma compared with control. Structural studies show that despite the structural perturbation of a domain extending from repeat 5 to repeat 8 (137-178), Delta6 apoA-I binds to spherical unilamellar vesicles with only 2-fold less binding affinity. In summary, these data show a domain corresponding to apoA-I repeat 6 is responsible for providing an essential conformation for LCAT catalyzed generation of cholesterol esters. Deletion of apoA-I repeat 6 not only blocks normal levels of cholesterol esterification but also exerts a dominant inhibition on the ability of wild-type apoA-I to activate LCAT in vivo.

Understanding variability in physician ratings of the appropriateness of coronary angiography after acute myocardial infarction.

We examined variability in ratings of the appropriateness of coronary angiography for 890 clinical scenarios (indications) after an acute myocardial infarction (AMI) from a nine-member multispecialty panel as a function of panel characteristics and the attributes of the clinical indications. We documented a substantial degree of reliability in the ratings. However, key differences among the experts in terms of both their overall propensity to score high and their beliefs regarding the impact of clinical factors on appropriateness were identified. Age, cardiac complications, post-AMI angina, and noninvasive test results were the clinical factors most strongly related to appropriateness ratings for coronary angiography. Further research on the effectiveness of coronary angiography in older patients and in patients with shock, pulmonary edema, and silent ischemia is needed to improve our knowledge about the appropriateness of this procedure in these patients.

Applying Bayesian ideas to the development of medical guidelines.

Measurements of the quality of health care, in particular the underuse and overuse of medical therapies and diagnostic tests, often involve employment of medical practice guidelines to assess the appropriateness of treatments. This paper presents a case study of a Bayesian analysis for the development of medical guidelines based on expert opinion, using ordinal categorical rater data. We develop guidelines for the use of coronary angiography following an acute myocardial infarction (AMI) for 890 clinical indications using statistical models fit to appropriateness ratings obtained from a nine-member expert panel. The main foci of our analyses were on the estimation of an appropriateness score for each of the clinical indications, an associated measure of precision, and functions of the underlying score. We considered two classes of models that assume the ratings are either in the form of grouped normal data or are ungrouped variables arising from a normal distribution, while permitting rater effects and indication heterogeneity in both. We estimated models using Markov chain Monte Carlo methods and constructed indices quantifying appropriateness based on posterior probabilities of selected model parameters. We compared our model-based approach to the standard approach currently employed in medical guideline development and found that the standard approach correctly identified 99 per cent of the appropriate indications while overestimating appropriateness 18 per cent of the time compared to our model-based approach.

The hydrophobic face orientation of apolipoprotein A-I amphipathic helix domain 143-164 regulates lecithin:cholesterol acyltransferase activation.

Apolipoprotein A-I (apoA-I) activates the plasma enzyme lecithin:cholesterol acyltransferase (LCAT), catalyzing the rapid conversion of lipoprotein cholesterol to cholesterol ester. Structural mutants of apoA-I have been used to study the details of apoA-I-LCAT-catalyzed cholesterol ester formation. Several studies have shown that the alpha-helical segments corresponding to amino acids 143-164 and 165-186 (repeats 6 and 7) are essential for LCAT activation. In the present studies, we examined how the orientation of the hydrophobic face, independent of an increase in overall hydrophobicity, affects LCAT activation. We designed, expressed, and characterized a mutant, reverse of 6 apoA-I (RO6 apoA-I), in which the primary amino acid sequence of repeat 6 (amino acids 143-164) was reversed from its normal orientation. This mutation rotates the hydrophobic face of repeat 6 approximately 80 degrees. Lipid-free RO6 apoA-I showed a marked stabilization when denatured by guanidine hydrochloride, but showed significant destabilization to guanidine hydrochloride denaturation in the lipid-bound state compared with wild-type apoA-I. Recombinant high density lipoprotein discs (rHDL) formed from RO6 apoA-I, sn-1-palmitoyl-sn-2-oleoyl phosphati-dylcholine, and cholesterol were approximately 12 A smaller than wild-type apoA-I rHDL. The reduced size suggests that one of the repeats did not effectively participate in phospholipid binding and organization. The sn-1-palmitoyl-sn-2-oleoyl phosphatidylcholine RO6 rHDL were a less effective substrate for LCAT. Mapping the entire lipid-free and lipid-bound RO6 apoA-I with a series of monoclonal antibodies revealed that both the lipid-free and lipid-bound RO6 apoA-I displayed altered or absent epitopes in domains within and adjacent to repeat 6. Together, these results suggest that the proper alignment and orientation of the hydrophobic face of repeat 6 is an important determinant for maintaining and stabilizing helix-bilayer and helix-helix interactions.

Rating the appropriateness of coronary angiography--do practicing physicians agree with an expert panel and with each other?

BACKGROUND: Evaluations of the appropriateness of medical care are important to monitor the quality of care and to contain costs and enhance safety by reducing inappropriate care. Experts' views are usually incorporated into evaluations of appropriateness. However, practicing physicians may not concur with these views, and physicians' clinical backgrounds may influence their beliefs. METHODS: We asked 1058 internists, family practitioners, and cardiologists in California, Florida, New York, Pennsylvania, and Texas to rate the appropriateness of coronary angiography after acute myocardial infarction for 20 common indications. Nine clinical experts also rated these indications using an established consensus method. RESULTS: For 17 of the 20 indications, median ratings of surveyed physicians and the expert panel agreed within 1 unit on a 9-unit scale. Patients' older age had a negative effect on ratings by the expert panel but not on ratings by surveyed physicians. In multivariable analyses of surveyed physicians, cardiologists rated angiography as significantly more appropriate than did primary care physicians for complicated indications, and for uncomplicated indications cardiologists who performed invasive procedures gave higher appropriateness ratings for angiography than did cardiologists who did not perform such procedures and primary care physicians. For uncomplicated indications, physicians from hospitals providing coronary angioplasty and bypass surgery rated angiography as more appropriate than physicians from other hospitals. Physicians from New York and those employed by health maintenance organizations rated angiography as less appropriate than other physicians. CONCLUSIONS: Surveyed physicians agreed with clinical experts about the appropriateness of coronary angiography after myocardial infarction for most indications, indicating that well-designed expert panels can closely reflect the views of practicing physicians. Variations in beliefs among practicing physicians suggest that evaluations of medical practice should incorporate the views of a range of relevant types of physicians.

Recombinant adeno-associated virus (AAV-CFTR) vectors do not integrate in a site-specific fashion in an immortalized epithelial cell line.

Adeno-associated virus-2 (AAV) can integrate in a site-specific manner to human chromosome 19 and is currently in phase I clinical trials for cystic fibrosis (CF) at Johns Hopkins Hospital. The goal of this study was to determine the fate of recombinant AAV containing the CFTR cDNA (AAV-CFTR) in an immortalized pseudotetraploid CF bronchial epithelial cell line (IB3-1) established from a patient with CF. Fluorescence in situ hybridization (FISH) and Southern blotting of DNA from IB3-1 cells infected with wild-type (wt) or recombinant AAV-CFTR were performed. CFRH2, an IB3-1 cell line with an estimated 15-20 integrated copies of CFTR cDNA, was used to test FISH sensitivity. All metaphase spreads had integrated copies: a single site in 36 of 56 (64.3%) and two sites within the same metaphase spread in 20 of 56 (35.7%). 3-CF-8, an IB3-1 cell line with integration of a partial CFTR cDNA (3.9 kb) was also analyzed by FISH. Integration was observed in 56 of 157 (35.7%) metaphase spreads examined. IB3-1 cells infected with wild-type AAV showed integration in 51 of 86 (59%) metaphase spreads examined. Of 51 integrations, 48 (94%) were to chromosome 19. Examination of 67 metaphase chromosome spreads of IB3-1 cells infected with AAV-CFTR vector (Azero) identified four integrations (6%) to different chromosomes. No integration was to chromosome 19 which differs significantly (P < 0.0001) from wild-type AAV. We then analyzed the A35 cell line, a clone of Azero selected for stable CFTR expression. Genomic DNA from A35 cells did not show a single site of integration; however episomal AAV-CFTR sequences were abundant in the low molecular weight DNA fraction. Examination of 68 metaphase chromosome preparations identified eight distinct integrations, none to chromosome 19. These studies show that FISH is sensitive for the detection of a partial CFTR cDNA integration. Wild-type AAV integrates in a predominantly site-specific fashion. Recombinant AAV-CFTR integrates at low frequency in a nonspecific manner and persists in episomal form in this epithelial cell line.