Gamma-carboxylated isoforms of recombinant human protein S with different biologic properties

Human protein S (HPS), a regulator of hemostasis, is a vitamin K- dependent plasma protein with potential clinical utility. We have obtained high-level expression of the cDNA for HPS in two mammalian cell lines. Both cell lines secreted single chain recombinant HPS (rHPS) in serum-free medium as determined by Western blot analysis. The ability of the rHPS from both cell lines to act as a cofactor for human protein C (HPC) was determined; the rHPS secreted from the human 293 cell line had an activity six times that of the rHPS from the AV12-664 Syrian hamster cell line. Furthermore, the relative specific cofactor activity of rHPS from the 293 cell line was actually 2.5-fold higher than that of single-chain human plasma-derived HPS. Essentially all of the rHPS secreted from the 293 cell line exhibited a calcium-dependent elution profile on anion exchange chromatography, whereas only 25% to 35% of the hamster cell-derived rHPS exhibited this profile. However, the calcium-eluted rHPS from the AV12 cell line had a high specific cofactor activity, equivalent to that of the 293-derived rHPS. A NaCl- elutable rHPS fraction (calcium nondependent) was isolated from the recombinant AV12-664 cell line, further purified, and found to have reduced activity, only 40% that of the calcium-dependent rHPS. The only observable difference in the calcium-dependent and nondependent rHPS molecules was in the content of gamma-carboxyglutamic acid (Gla); the calcium-dependent material contained approximately 10 mol Gla/mol protein whereas the calcium-nondependent material contained only approximately 8 mol Gla/mol of protein. In addition, the calcium- nondependent rHPS had reduced ability to interact with phospholipid vesicles as evidenced by an eightfold increase in the apparent kd. Our data demonstrate the isolation of rHPS with high specific activity, and show that a reduction in as few as two Gla residues dramatically decreases its functional cofactor activity for HPC, due to a reduction in ability to interact with the phospholipid bilayer.     

Quantitative measurement of lead in paint by XRF analysis without manual substrate correction

X-ray fluorescence analysis has been used for measurement of lead in paint for more than a decade. The early systems provided a nondestructive alternative technology to laboratory-based technologies, but were somewhat time consuming and often led to inconclusive results. The procedure required manual substrate correction, multiple measurements, operator's discretion in validating a measurement due to interfering elements and laboratory analysis of inconclusive samples. A new instrument, the RMD LPA-1 system, has been developed based on X-ray fluorescence technology that addresses all of the drawbacks to the older systems. This new system uses a carefully designed and controlled geometry and modern microprocessor technology to automatically provide a rapid quantitative measurement of lead in paint with a 95% confidence level. The improved precision and accuracy achieved with this system are due to geometric enhancements and a mathematical approach which incorporates corrections for both random and systematic errors such as matrix effects and Compton scatter. This technology has been incorporated in a hand-held X-ray fluorescence lead paint analyzer system. A key design philosophy for this system was to maintain a very narrow, task-specific focus, the system was not designed to be an all purpose XRF analyzer, rather it is optimized to meet regulatory requirements of lead paint testing in the most efficient manner. The development of the LPA-1 system is an example of what can be accomplished by listening to the needs and desires of the users, rethinking the design of an existing technique and incorporating modern microprocessor technology.     

Development of a multi-layered psychosocial care system for children in areas of political violence

Few psychosocial and mental health care systems have been reported for children affected by political violence in low- and middle income settings and there is a paucity of research-supported recommendations. This paper describes a field tested multi-layered psychosocial care system for children (focus age between 8-14 years), aiming to translate common principles and guidelines into a comprehensive support package. This community-based approach includes different overlapping levels of interventions to address varying needs for support. These levels provide assessment and management of problems that range from the social-pedagogic domain to the psychosocial, the psychological and the psychiatric domains. Specific intervention methodologies and their rationale are described within the context of a four-country program (Burundi, Sri Lanka, Indonesia and Sudan). The paper aims to contribute to bridge the divide in the literature between guidelines, consensus & research and clinical practice in the field of psychosocial and mental health care in low- and middle-income countries.     

The effect of PGI2 and theophylline on the response of platelets subjected to shear stress

A specially designed rotational viscometer was used to investigate the effects of the antiplatelet agent PGI2 in combination with theophylline on the response of human platelets subjected to shear stress. Samples of citrated platelet-rich plasma (PRP) were exposed to shear stress in the viscometer for a period of 5 min at 23 degrees C. The levels of stress studied ranged from 50 to 300 dynes/sq cm. Pretreatment of the platelets with 0.01 microM PGI2 and 500 microM theophylline before exposure to shear stress caused a large reduction in shear-induced platelet aggregation. However, it was also observed that the PGI2-- theophylline pretreatment concomitantly caused a large increase in shear-induced platelet lysis and serotonin release at stress levels equal to or greater than 150 dynes/sq cm. This observed increase in platelet fragility may have important implications for clinical applications of PGI2. The results are discussed and compared to those obtained in prior work in which platelets were pretreated with acetylsalicylic acid or with PGE1.     

Impact of Breast Density Legislation on Breast Cancer Risk Assessment and Supplemental Screening: A Survey of 110 Radiology Facilities

Breast density notification laws, passed in 19 states as of October 2014, mandate that patients be informed of their breast density. The purpose of this study is to assess the impact of this legislation on radiology practices, including performance of breast cancer risk assessment and supplemental screening studies. A 20‐question anonymous web‐based survey was emailed to radiologists in the Society of Breast Imaging between August 2013 and March 2014. Statistical analysis was performed using Fisher's exact test. Around 121 radiologists from 110 facilities in 34 USA states and 1 Canadian site responded. About 50% (55/110) of facilities had breast density legislation, 36% of facilities (39/109) performed breast cancer risk assessment (one facility did not respond). Risk assessment was performed as a new task in response to density legislation in 40% (6/15) of facilities in states with notification laws. However, there was no significant difference in performing risk assessment between facilities in states with a law and those without (p < 0.831). In anticipation of breast density legislation, 33% (16/48), 6% (3/48), and 6% (3/48) of facilities in states with laws implemented handheld whole breast ultrasound (WBUS), automated WBUS, and tomosynthesis, respectively. The ratio of facilities offering handheld WBUS was significantly higher in states with a law than in states without (p < 0.001). In response to breast density legislation, more than 33% of facilities are offering supplemental screening with WBUS and tomosynthesis, and many are performing formal risk assessment for determining patient management.     

Hyperbaric bupivacaine 2.5 mg prolongs analgesia compared with plain bupivacaine when added to intrathecal fentanyl 25 microg in advanced labor

We investigated the effect of sequential administration of intrathecal (IT) hyperbaric bupivacaine (after the initial administration of IT hypobaric fentanyl) on the duration of spinal analgesia. Thirty-seven nulliparous parturients with a cervical dilation >/= 5 cm were randomized to receive either IT fentanyl 25 micro g and plain bupivacaine 2.5 mg (group P; n = 19) or IT fentanyl 25 micro g and hyperbaric (with 8% glucose) bupivacaine 2.5 mg (group H; n = 18). The two components of the IT injectate were administered sequentially (fentanyl 25 micro g diluted in 2 mL of normal saline, immediately followed by 0.5 mL of 0.5% bupivacaine). Patients were then positioned with their torso elevated at 30 degrees for 30 min. Pain scores using 0-100 visual analog scales were collected before combined spinal/epidural analgesia and at 5, 15, and 30 min after the block. Patients in Group H had a longer median duration of analgesia (122 min; range, 80-210 min) than Group P (95 min; range, 75-125 min) (P < 0.01). Group H also had a more limited dermatomal spread (median highest sensory level of T8 versus T4 in group P; P < 0.05). The side-effect profile was similar. Under these circumstances, hyperbaric bupivacaine conferred an increased duration of IT analgesia compared with plain bupivacaine.