Characterization of Agfa Structurix series D4 and D3sc x-ray films in the 0.7-4.6 keV energy range.

X-ray films remain a key asset for high-resolution x-ray spectral imaging in high-energy-density experiments conducted at the National Ignition Facility (NIF). The soft x-ray Opacity Spectrometer (OpSpec) fielded at the NIF has an elliptically shaped crystal design that measures x rays in the 900-2100 eV range and currently uses an image plate as the detecting medium. However, Agfa D4 and D3sc x-ray films' higher spatial resolution provides increased spectral resolution to the data over the IP-TR image plates, driving the desire for regular use of x-ray film as a detecting medium. The calibration of Agfa D4 x-ray film for use in the OpSpec is communicated here. These calibration efforts are vital to the accuracy of the NIF opacity measurements and are conducted in a previously un-studied x-ray energy range under a new film development protocol required by NIF. The absolute response of Agfa D4 x-ray film from 705 to 4620 eV has been measured using the Nevada National Security Site Manson x-ray source. A broader range of energies was selected to compare results with previously published data. The measurements were taken using selected anodes, filters, and applied voltages to produce well-defined energy lines.

A double-blinded randomised controlled study to investigate the effect of intraperitoneal levobupivacaine on post laparoscopic pain.

BACKGROUND: Laparoscopic surgery is the cornerstone of modern gynaecological surgery, with shorter hospital stays and a quicker return to normal activities. However postoperative pain remains problematic. No strategy to reduce phrenic nerve irritation, including heating or humidifying the insufflating gas, alternatives to CO2, and intraperitoneal analgesics, has shown superiority. METHODS: 100 women undergoing laparoscopic surgery were randomly allocated, having either 40ml of 0.25% levobupivacaine or 40ml 0.9% sodium chloride solution administered into the peritoneal cavity following surgery. The patients and the main researcher were blinded. All women received standardised anaesthetic and laparoscopic technique, and postoperative pain control including nursing position and nature of analgesia. Postoperative pain was assessed 3 hours, 8 hours, day 1 and day 4/5 postoperatively. RESULTS: 100 patients were recruited undergoing surgery for benign causes aged 19-73(mean 40.3±13). There was no difference between the groups for age(p=0.64) or length of operation(p=0.56). There were no adverse events related to use of intraperitoneal instillation. There was a significant reduction in shoulder-tip pain scores in the levobupivacaine group at 3 hours(p=0.04). Furthermore, there was a significant reduction in wound-pain scores in the levobupivacaine group at 8hrs(p=0.04) and at day 4(p=0.04). No difference was found in pelvic pain between the two groups. No significant difference was found in the use of post-operative analgesia. CONCLUSIONS: Intraperitoneal instillation of 40ml of levobupivacaine has some benefit in reducing postoperative pain and need for analgesia in the initial hours following gynaecological surgery. However, further well-designed randomised control trials are required to decide the optimum route and concentration of administering local anaesthetic.

Perfluorinated alkyl acids in the serum and follicular fluid of UK women with and without polycystic ovarian syndrome undergoing fertility treatment and associations with hormonal and metabolic parameters.

Women with polycystic ovarian syndrome (PCOS) undergoing treatment for infertility could be a sensitive subpopulation for endocrine effects of exposure to perfluorinated alkyl acids (PFAAs), persistent organic pollutants with potential endocrine activity. Women with, PCOS (n = 30) and age- and BMI-matched controls (n = 29) were recruited from a UK fertility clinic in 2015. Paired serum and follicular fluid samples were collected and analysed for 13 PFAAs. Sex steroid and thyroid hormones, and metabolic markers were measured and assessed for associations with serum PFAAs. Four PFAAs were detected in all serum and follicular fluid samples and concentrations in the two matrices were highly correlated (R2 > 0.95): perfluorooctane sulfonate (PFOS), perfluorooctanoic acid (PFOA), perfluorohexane sulfonate (PFHxS), and perfluorononanoic acid (PFNA). Serum PFOS was positively associated with age (1 ng/mL per yr, p < 0.05) and was higher in PCOS cases than controls (geometric mean [GM] 3.9 vs. 3.1 ng/mL, p < 0.05) and in women with irregular vs. regular menstrual cycles (GM 3.9 vs. 3.0 ng/mL, p = 0.01). After adjustment for confounders, serum testosterone was significantly associated with PFOA, PFHxS, PFNA, and the molar sum of the four frequently detected serum PFAAs (approximately 50 percent increase per ln-unit) among controls but not PCOS cases. HbA1c in PCOS cases was inversely associated with serum PFOA, PFHxs, and sum of PFAAs (2-3 mmol/mol per ln-unit). In controls, fasting glucose was positively associated with serum PFOA and sum of PFAAs (0.25 nmol/L per ln-unit increase in PFAAs). Few other associations were observed. The analyses and findings here should be considered exploratory in light of the relatively small sample sizes and large number of statistical comparisons conducted. However, the data do not suggest increased sensitivity to potential endocrine effects of PFAAs in PCOS patients.

Clinical outcomes following ICSI cycles using surgically recovered sperm and the impact of maternal age: 2004-2015 SART CORS registry.

PURPOSE: The aims of this study were (1) to evaluate clinical outcomes after ICSI cycles using surgically recovered sperm and (2) to assess the influence of maternal age on those outcomes. METHODS: A retrospective cohort study of 24,763 IVF cycles of fresh autologous oocytes and ICSI using surgically recovered sperm reported to the SART CORS database from 2004 to 2015. RESULTS AND CONCLUSIONS: Older women had significantly longer stimulation (p < 0.001), a lower number of oocytes retrieved (p < 0.001), a lower number of 2PN zygotes (p < 0.001), a lower chance of having a blastocyst transferred (p < 0.001), and a higher number of fresh embryos transferred (p < 0.001). There was no significant association between the number of 2PNs per oocyte retrieved and maternal age (p = 0.214). Both clinical pregnancy rates and live birth rates (LBR) decreased with advanced maternal age (p < 0.001). LBR ranged from 50.4% in women < 30 to 7.2% in women > 42 years, and for cleavage-stage transfers, the LBR ranged from 47.3% in women< 30 to 6.3% in women > 42 years. There were no differences in gestational age at delivery, proportion of term deliveries, preterm deliveries, neonatal birth weight < 2500 g, neonatal birth weight > 4000 g and average birthweight of neonates for singleton pregnancies according to age. For twin pregnancies, women < 30 years had significantly higher number of live births, term deliveries, and lower preterm deliveries than older women. There was a similar number of female (6051) and male neonates (5858; p = 0.2). Overall, pregnancy outcomes with ICSI using surgically recovered sperm are reassuring and comparable to those of ICSI with ejaculated sperm.

Comparison of osteoporosis in US adults with type 1 and type 2 diabetes mellitus.

PURPOSE: We examined bone mineral density (BMD) and osteoporosis prevalence in those with type 1 compared to type 2 diabetes derived from a nationally representative sample from the civilian community in the United States. METHODS: Data from the National Health and Nutrition Examination Survey (NHANES) for 2005-2006, 2007-2008, 2009-2010, and 2013-2014 were merged to obtain a large sample of diabetics at least 20 years of age with participation in the interview and medical examination. Osteoporosis status was defined by BMD at the total femur, femoral neck, or total lumbar spine. Self-reported diabetics that were prescribed insulin within the first year of diagnosis, are currently taking insulin, and reported no prescriptions for any diabetic pills were classified as type 1. Remaining self-reported diabetics were deemed as having type 2. RESULTS: A total of 2050 diabetics were included in which 87 (4%) were classified as type 1. Type 1 diabetics were found to have a significantly lower BMD at the total femur and femoral neck, but not at the lumbar spine in the adjusted models. Diabetics with type 1 were 4.7 times more likely to have osteoporosis than those with type 2. There was no significant relationship between diabetes type and BMD or osteoporosis prior to adjustment for confounders. CONCLUSIONS: Although our results show an increased likelihood of osteoporosis among those with type 1 diabetes, future studies including a larger sample from a community population are needed. It may benefit diabetics, especially those with type 1, to initiate osteoporosis screening methods including evaluation of fracture risk, bone quality, and BMD measurements at multiple sites earlier than recommended.

On the use of the Cu2+-iminodiacetic acid complex for double histidine based distance measurements by pulsed ESR.

Cu2+ based distance measurements using the double-histidine (dHis) motif by pulsed ESR present an attractive strategy to obtain precise, narrow distance distributions that can be easily related to protein backbone structure (Cunningham et al., Angew. Chem., Int. Ed., 2015, 54, 633). The Cu2+-ion is introduced as a complex with the iminodiacetic acid (IDA) chelating agent, which enhances binding selectivity to the two histidine residues that are site-selectively placed on the protein through mutagenesis. However, initial results of this method produced weak dipolar modulations. To enhance applicability of the double histidine motif using IDA, we perform a systematic examination of the possible causes of these weak dipolar modulations. We examine the efficiency of the Cu2+-ion to form the Cu2+-IDA complex in solution. In addition, we analyze the selectivity of Cu2+-IDA binding to dHis sites at both α-helical and ß-strand environments. Our results indicate that the dHis motif on the ß-sheet sites have high affinity towards Cu2+-IDA while the dHis sites on α-helices show poor affinity for the metal-ion complex. We are able to use our new findings to optimize conditions to maximize dHis loading while minimizing both free Cu2+ and unbound Cu2+-IDA complex in solution, allowing us to double the sensitivity of the Double Electron-Electron Resonance (DEER) experiment. Finally, we illustrate how Cu2+-based CW-ESR and DEER can be combined to obtain information on populations of different Cu2+-complexes in solution.

The impact of osteoporotic fractures compared with other health conditions in older adults living in Virginia, United States.

UNLABELLED: This study compared length of stay, hospital costs, 30-day readmission, and mortality for patients admitted primarily for osteoporotic fractures to those admitted for five other common health conditions. The results indicated that osteoporotic fractures were associated with highest hospital charges and the second highest hospital stay after adjusting for confounders. INTRODUCTION: This study aimed to compare the effect of osteoporotic fractures and other common hospitalized conditions in both men and women age 55 years and older on a large in-patient sample. METHODS: De-identified patient level and readmission and transfer data from the Virginia Health Information (VHI) system for 2008 through 2014 were merged. Logistic regression models were used to assess mortality and 30-day readmission, while generalized linear models were fitted to assess LOS and hospital charges. RESULTS: After adjustment for confounders, osteoporotic fractures had the second longest LOS (6.0 days, 95 % CI = 5.9-6.0) and the highest average total hospital charges ($47,386.0, 95 % CI = $46,707.0-$48,074.0) compared to the other five common health problems. CONCLUSION: Recognizing risk and susceptibility to osteoporotic fractures is an important motivator for individual behaviors that mitigate this disease. Furthermore, acknowledging the economic impact and disabling burden of osteoporotic fractures on society are compelling reasons to promote bone health as well as to prevent, diagnose, and manage osteoporosis.

Association of CMV, HBV, or HCV co-infection with vaccine response in adults with well-controlled HIV infection.

Even after CD4 count recovery on antiretroviral therapy, HIV infection is associated with decreased response to most vaccines compared to the general population. Chronic infections with viruses such as cytomegalovirus (CMV), hepatitis B virus (HBV), and hepatitis C virus (HCV), which are more prevalent in HIV-infected populations, have been linked to immune dysfunction and decreased vaccine response in the general population. However, whether co-infection with these other viruses contributes to the decreased vaccine response seen in adults with well-controlled HIV infection is unknown. We conducted a secondary analysis of data and serum from adults with well-controlled HIV infection from an inactivated polio vaccine trial (224 subjects) and a pneumococcal conjugate vaccine study (128 subjects). We evaluated the association of CMV, HBV, or HCV co-infection with post-vaccination antibody levels using both univariate and multivariate analyses, controlling for factors such as age, race, CD4 count, comorbidities, smoking status, and baseline antibody levels. Ninety-three percent, 7%, and 14% of subjects were co-infected with CMV, HBV, and HCV respectively. On both univariate and multivariate analysis, neither CMV nor HCV co-infection were significantly associated with post-vaccination antibody levels to either vaccine. HBV co-infection was significantly associated with post-vaccination antibody concentrations for pneumococcal serotype 7F on univariate analysis and 6A on multivariate analysis, but the association was with higher antibody concentrations. In conclusion, co-infection with CMV, HBV, or HCV does not appear to contribute to the decreased vaccine response seen in adults with well-controlled HIV infection.

Comparison of tongue muscle characteristics of preterm and full term infants during nutritive and nonnutritive sucking.

BACKGROUND: Independent oral feeding requires coordination of suck, swallow and breathe and the lingual musculature plays a significant role in this coordinative action. However, clinical benchmarks of lingual function fundamental to successful feeding have not been explored. AIMS: The present study tests our model for quantifying infant lingual force and size and compares the muscle measures of interest in two cohorts: healthy full-term infants (FT) (N=5) and healthy preterm infants (PT) (N=6). METHOD: Using an instrumented pacifier and bottle nipple, we determined the resultant compressive forces applied to the nipple by the tongue during nutritive (NS) and nonnutritive sucking (NNS). Muscle size was estimated from measures of posterior tongue thickness using ultrasonography. RESULTS: After controlling for weight and post menstrual age, statistically significant differences were found between FT and PT infants beginning to feed for NNS frequency and NS tongue force. Clinically significant differences were detected for NNS tongue force and posterior tongue thickness. Additionally, PT infants demonstrated a significant difference in mean tongue force between NS and NNS and FT infants did not. FT infants demonstrated a significant difference in mean frequency between NS and NNS and PT infants did not. Linear regression indicated that mean posterior tongue thickness alone predicted 55% of the variance in NS force. CONCLUSIONS: Results demonstrate the feasibility of our approach and suggest that infant tongue muscle characteristics necessary for successful feeding differ between healthy full term infants and preterm infants who are beginning oral feeding.

Osteoporosis treatment disparities: a 6-year aggregate analysis from national survey data.

UNLABELLED: We studied factors to determine the receipt of osteoporosis treatment in individuals with osteoporosis. Treatment was associated with age, gender, race, body mass index (BMI), family history, arthritis and thyroid problems, daily glucocorticoid use, number of prescriptions and healthcare visits, and insurance type. INTRODUCTION: Osteoporosis is underrecognized and undertreated. Few studies have examined factors associated with osteoporosis treatment in a large, national sample of men and women. METHODS: We aggregated National Health and Nutrition Examination Survey (NHANES) data from 2005 to 2010 and created a subsample which included individuals 50 years or older who were identified to have osteoporosis either by self-report data or by bone density measurements. The primary outcome was the receipt of osteoporosis treatment either from self-report or from prescription records. Covariates included sociodemographics, clinical characteristics, and access to healthcare variables. Logistic regression analyses were performed to determine factors that associate with osteoporosis treatment. RESULTS: From a sample of 31,0134 participants, 1,133 subjects (3.65 %) met the study criteria. Treatment was associated with age (odds ratio (OR) = 1.14), gender (OR = 13.25), race (OR = 2.23, White vs. Black; OR = 1.76, other vs. Black), BMI (OR = 1.67, normal vs. obese; OR = 2.68, overweight vs. obese), family history of osteoporosis (OR = 1.94), arthritis (OR = 1.43), daily glucocorticoid use (OR = 1.43), number of prescriptions (OR = 1.01), and number of healthcare visits in the past year (OR = 1.44, 4-9 vs. 0-3 visits). All odds ratios were statistically significant. CONCLUSION: A large number of individuals diagnosed with osteoporosis above the age of 50 remain untreated. It is important for healthcare providers to better assess older adults with osteoporosis, including individuals who frequently receive medical care.

Peripheral bone mineral density in patients with distal radial fractures.

Fractures of the distal forearm are widely regarded as the result of "fragility". We have examined the extent to which patients with Colles' fractures have osteopenia. We measured the bone mineral density (BMD) in the contralateral radius of 235 women presenting with Colles' fractures over a period of two years. While women of all ages had low values for ultra-distal BMD, the values, in age-matched terms, were particularly low among premenopausal women aged less than 45 years. This result was not due to the presence of women with an early menopause. This large survey confirms and extends the findings from earlier small studies. We consider that it is particularly important to investigate young patients with fractures of the distal forearm to identify those with osteoporosis, to seek an underlying cause and to consider treatment.

Randomized, placebo-controlled, double-blind study of a cytomegalovirus-specific monoclonal antibody (MSL-109) for prevention of cytomegalovirus infection after allogeneic hematopoietic stem cell transplantation.

MSL-109 is a monoclonal antibody specific to the cytomegalovirus (CMV) glycoprotein H with high neutralizing capacity. In a prospective, randomized, double-blind study, allogeneic hematopoietic stem cell transplantation (HSCT) recipients with positive donor and/or recipient serology for CMV before transplantation received either 60 mg/kg MSL-109 (n = 59), 15 mg/kg MSL-109 (n = 60), or placebo (n = 60) intravenously every 2 weeks from day -1 until day 84 after transplantation. CMV pp65 antigenemia, CMV-DNA load in plasma, and viremia by culture were tested weekly. Primary end points were development of pp65 antigenemia at any level and/or viremia for which ganciclovir was given. There was no statistically significant difference in CMV pp65 antigenemia or viremia among patients in the 60-mg group (pp65 antigenemia, 47%; viremia, 15%), the 15-mg group (52%; 23%), and the placebo group (45%; 17%). There was also no difference in maximum levels of pp65 antigenemia, time to clearance of pp65 antigenemia after start of ganciclovir, CMV disease, invasive bacterial and fungal infections, time to neutrophil and platelet engraftment, acute graft-versus-host disease, days of hospitalization, and overall survival rate among the 3 groups. However, a subgroup analysis of CMV-seronegative recipients with a seropositive donor (D+/R-) showed a transiently improved survival rate by day 100 in MSL-109 recipients (mortality: 60-mg group, 1/13; 15-mg group, 1/12; placebo group, 6/10 [P = .02 for 60-mg versus placebo groups; P = .08 for 15-mg versus placebo groups]); by the end of follow-up, the difference was no longer statistically significant. The improved survival rate in D+/R- patients could not be attributed to a reduction in CMV disease; however, MSL-109 was associated with improved platelet engraftment and less grade III to IV acute graft-versus-host disease in this subgroup. In a subgroup analysis of CMV-seropositive recipients of MSL-109 (D+/R+ and D-/R+), overall mortality was increased compared to that of the placebo group (P = .12 for the 60-mg versus placebo groups, P = .05 for the 15-mg versus placebo groups, and P = .04 for the dose levels combined versus placebo). MSL-109 was well tolerated and no immune response to the drug was observed. Thus, MSL-109 was safe but did not reduce CMV infection in allogeneic HSCT recipients. The transient survival advantage seen early after transplantation in CMV D+/R- patients and the negative effect on survival in seropositive patients remain unexplained. Thus, there is no evidence that MSL-109 is beneficial in CMV-seropositive HSCT recipients.

The level of DAL80 expression down-regulates GATA factor-mediated transcription in Saccharomyces cerevisiae.

Nitrogen-catabolic gene expression in Saccharomyces cerevisiae is regulated by the action of four GATA family transcription factors: Gln3p and Gat1p/Nil1p are transcriptional activators, and Dal80 and Deh1p/Gzf3p are repressors. In addition to the GATA sequences situated upstream of all nitrogen catabolite repression-sensitive genes that encode enzyme and transport proteins, the promoters of the GAT1, DAL80, and DEH1 genes all contain multiple GATA sequences as well. These GATA sequences are the binding sites of the GATA family transcription factors and are hypothesized to mediate their autogenous and cross regulation. Here we show, using DAL80 fused to the carbon-regulated GAL1,10 or copper-regulated CUP1 promoter, that GAT1 expression is inversely regulated by the level of DAL80 expression, i.e., as DAL80 expression increases, GAT1 expression decreases. The amount of DAL80 expression also dictates the level at which DAL3, a gene activated almost exclusively by Gln3p, is transcribed. Gat1p was found to partially substitute for Gln3p in transcription. These data support the contention that regulation of GATA-factor gene expression is tightly and dynamically coupled. Finally, we suggest that the complicated regulatory circuit in which the GATA family transcription factors participate is probably most beneficial as cells make the transition from excess to limited nitrogen availability.

Calreticulin binding and other biological activities of survival peptide Y-P30 including effects of systemic treatment of rats.

Neuron survival-promoting peptide Y-P30, purified from oxidatively stressed neural cell lines, inhibits the appearance of microglia and rescues neurons 1 week after direct application to lesions of the rat cerebral cortex (7). Y-P30 affinity matrices treated with solubilized membranes from a variety of cell lines including human neuroblastoma SY5Y, mouse hippocampal cells HN 33.1, and human promonocytes HL-60, as well as with cerebral cortex tissue from both humans and rats, showed highly specific binding to calreticulin, a ubiquitous calcium binding protein that may be critical for integrin function. Treatment of cultures with 0.1 nM Y-P30 stabilized all these cell types whether differentiated or not, while 1 microM peptide also inhibited the morphological differentiation of the HL-60 cells into macrophages. Western analysis of the medium of SY5Y cell cultures suggested Y-P30-stimulated release of calreticulin, a result consistent with its other biological activities. Likewise, single dose systemic application of Y-P30 in unoperated rats and in rats with cerebral cortex lesions produced significant reductions in cerebral cortex membrane-associated calreticulin. Both direct and intravenous treatment with peptide also reduced cortical neuron atrophy 4 days after these lesions but only direct application consistently inhibited the appearance of ED-1(+) monocyte derivatives. We suggest that in vitro and in vivo mechanisms of Y-P30 effects are similar and involve the targeting of calreticulin. The results also suggest that some of these activities are apparent in the cerebral cortex after systemic application of this peptide.

Nitrogen catabolite repression of DAL80 expression depends on the relative levels of Gat1p and Ure2p production in Saccharomyces cerevisiae.

GATA family activators (Gln3p and Gat1p) and repressors (Dal80p and Deh1p) regulate nitrogen catabolite repression (NCR)-sensitive transcription in Saccharomyces cerevisiae presumably via their competitive binding to the GATA sequences upstream of NCR-sensitive genes. Ure2p, which is not a GATA family member, inhibits Gln3p/Gat1p from functioning in the presence of good nitrogen sources. We show that NCR-sensitive DAL80 transcription can be influenced by the relative levels of GAT1 and URE2 expression. NCR, normally observed with ammonia or glutamine, is severely diminished when Gat1p is overproduced, and this inhibition is overcome by simultaneously increasing URE2 expression. Further, overproduction of Ure2p nearly eliminates NCR-sensitive transcription under derepressive growth conditions, i.e. with proline as the sole nitrogen source. Enhanced green fluorescent protein-Gat1p is nuclear when Gat1p-dependent transcription is high and cytoplasmic when it is inhibited by overproduction of Ure2p.

Using online and offline change models to improve ICU access and revenues.

BACKGROUND: Hospital operational problems that span departments often present formidable challenges because they involve both processes and organizational relationships. Many improvement efforts fail because of relationship issues rather than a lack of understanding of system processes. Reflection on a recent change initiative led to the development of an integrated change model that includes both online and offline components. The online component draws on performance improvement models that provide concepts and tools for use in team meetings to improve processes. The offline component borrows from an earlier tradition of change management that offers guidelines for individuals or teams desiring to be change agents. METHODS: The integrated change model was applied in 1997 at The Johns Hopkins Hospital, Baltimore, to reduce ambulance bypass hours, a chronic problem resulting in $6.7 million in lost revenue annually. The goal was to reduce red alert hours per month by 50%. Three Plan-Do-Study-Act (PDSA) cycles were implemented to test change concepts. RESULTS: There was a significant reduction in red alert hours after the change initiative, with an estimated $6 million in additional hospital revenue. DISCUSSION: The integrated change model may serve as a prototype for improving complex problems in which improving organizational relationships may be as difficult as improving processes and is likely to require a significant amount of work offline. For example, this approach may be particularly helpful for improving processes that span departments or functional units such as reducing cycle times for admissions, first-dose medications, as well as in building and improving integrated delivery systems. The model awaits further testing and evolution.

Overlapping positive and negative GATA factor binding sites mediate inducible DAL7 gene expression in Saccharomyces cerevisiae.

Allantoin pathway gene expression in Saccharomyces cerevisiae responds to two different environmental stimuli. The expression of these genes is induced in the presence of allantoin or its degradative metabolites and repressed when a good nitrogen source (e. g. asparagine or glutamine) is provided. Three types of cis-acting sites and trans-acting factors are required for allantoin pathway gene transcription as follows: (i) UAS(NTR) element associated with the transcriptional activators Gln3p and Gat1p, (ii) URS(GATA) element associated with the repressor Dal80p, and (iii) UIS(ALL) element associated with the Dal82 and Dal81 proteins required for inducer-dependent transcription. Most of the work leading to the above conclusions has employed inducer-independent allantoin pathway genes (e.g. DAL5 and DAL3). The purpose of this work is to extend our understanding of these elements and their roles to inducible allantoin pathway genes using the DAL7 (encoding malate synthase) as a model. We show that eight distinct cis-acting sites participate in the process as follows: a newly identified GC-rich element, two UAS(NTR), two UIS(ALL), and three URS(GATA) elements. The two GATA-containing UAS(NTR) elements are coincident with two of the three GATA sequences that make up the URS(GATA) elements. The remaining URS(GATA) GATA sequence, however, is not a UAS(NTR) element but appears to function only in repression. The data provide insights into how these cis- and trans-acting factors function together to accomplish the regulated expression of the DAL7 gene that is observed in vivo.

Comparison of features of human breast cancer cell lines and their corresponding tumors.

Although human tumor-derived cell lines play an important role in the investigation of cancer biology and genetics, there is no comprehensive study comparing tumor cell line properties with those of the individual tumors from which they were derived. We compared the properties of a series of 18 human breast cancer cell lines that were cultured for a median period of 25 months (range, 9-60 months) and their corresponding archival tumor tissues. We compared morphological characteristics, ploidy, and immunohistochemical expression of estrogen receptors, progesterone receptors, and HER2/neu and p53 proteins. For 17 of these cases, we also tested for allelic losses at 18 chromosomal regions frequently deleted in breast tumors using 51 polymorphic microsatellite markers, and we determined the TP53 gene mutation status in exons 5 to 10. There was an excellent correlation between the breast tumor cell lines and their corresponding tumor tissues for morphological features (100%); presence of aneuploidy (87%); immunohistochemical expression of estrogen receptors (87%), progesterone receptors (73%), and HER2/neu (93%) and p53 proteins (100%); allelic loss at all of the chromosomal regions analyzed (82-100% concordance); and TP53 gene mutations (75%). The same parental allele was lost in 279 (99%) of 281 of the comparisons of allelic losses. The fractional allelic loss indices (a reflection of the total allelic loss) of the cell lines and their corresponding tumor tissues were identical or similar in 15 (88%) of 17 paired comparisons. Although our previous studies (A. Gazdar et al., Int. J. Cancer, in press) indicated that only a subset of primary breast carcinomas that have several features indicative of advanced tumors with poor prognosis can be successfully cultured, the cell lines retain the properties of their parental tumors for lengthy culture periods and, thus, provide suitable model systems for biomedical studies.

Identification of a survival-promoting peptide in medium conditioned by oxidatively stressed cell lines of nervous system origin.

A survival-promoting peptide has been purified from medium conditioned by Y79 human retinoblastoma cells and a mouse hippocampal cell line (HN 33.1) exposed to H2O2. A 30 residue synthetic peptide was made on the basis of N-terminal sequences obtained during purification, and it was found to exhibit gel mobility and staining properties similar to the purified molecules. The peptide maintains cells and their processes in vitro for the HN 33.1 cell line treated with H2O2, and in vivo for cortical neurons after lesions of the cerebral cortex. It has weak homology with a fragment of a putative bacterial antigen and, like that molecule, binds IgG. The peptide also contains a motif reminiscent of a critical sequence in the catalytic region of calcineurin-type phosphatases; surprisingly, like several members of this family, the peptide catalyzes the hydrolysis of para-nitrophenylphosphate in the presence of Mn2+. Application of the peptide to one side of bilateral cerebral cortex lesions centered on area 2 in rats results in an increase in IgG immunoreactivity in the vicinity of the lesions 7 d after surgery. Microglia immunopositive for IgG and ED-1 are, however, dramatically reduced around the lesions in the treated hemisphere. Furthermore, pyramidal neurons that would normally shrink, die, or disintegrate were maintained, as determined by MAP2 immunocytochemistry and Nissl staining. These survival effects were often found in both hemispheres. The results suggest that this peptide operates by diffusion to regulate the immune response and thereby rescue neurons that would usually degenerate after cortical lesions. The phosphatase activity of this molecule also suggests the potential for direct neuron survival-promoting effects.