Rapid Detection of high-level oncogene amplifications in ultrasonic surgical aspirations of brain tumors

ABSTRACT: BACKGROUND: Genomic tumor information, such as identification of amplified oncogenes, can be used to plan treatment. The two sources of a brain tumor that are commonly available include formalin-fixed, paraffin-embedded (FFPE) sections from the small diagnostic biopsy and the ultrasonic surgical aspiration that contains the bulk of the tumor. In research centers, frozen tissue of a brain tumor may also be available. This study compared ultrasonic surgical aspiration and FFPE specimens from the same brain tumors for retrieval of DNA and molecular assessment of amplified oncogenes. METHODS: Surgical aspirations were centrifuged to separate erythrocytes from the tumor cells that predominantly formed large, overlying buffy coats. These were sampled to harvest nuclear pellets for DNA purification. Four glioblastomas, 2 lung carcinoma metastases, and an ependymoma were tested. An inexpensive PCR technique, multiplex ligation-dependent probe amplification (MLPA), quantified 79 oncogenes using 3 kits. Copy number (CN) results were normalized to DNA from non-neoplastic brain (NB) in calculated ratios, [tumor DNA]/[NB DNA]. Bland-Altman and Spearman rank correlative comparisons were determined. Regression analysis identified outliers. RESULTS: Purification of DNA from ultrasonic surgical aspirations was rapid (<3 days) versus FFPE (weeks) and yielded greater amounts in 6 of 7 tumors. Gene amplifications up to 15-fold corresponded closely between ultrasonic aspiration and FFPE assays in Bland-Altman analysis. Correlation coefficients ranged from 0.71 to 0.99 using 3 kit assays per tumor. Although normalized CN ratios greater than 2.0 were more numerous in FFPE specimens, some were found only in the ultrasonic surgical aspirations, consistent with tumor heterogeneity. Additionally, CN ratios revealed 9 high-level ([greater than or equal to] 6.0) gene amplifications in FFPE of which 8 were also detected in the ultrasonic aspirations at increased levels. The ultrasonic aspiration levels of these amplified genes were also greater than 6.0 CN ratio, except in one case (3.53 CN ratio). Ten of 17 mid-level ([greater than or equal to]3.0 & <6.0 CN ratio) amplifications detected in FFPE were also detected as being increased ([greater than or equal to] 2.0 CN ratio) in the aspirations. CONCLUSIONS: Buffy coats of centrifuged ultrasonic aspirations contained abundant tumor cells whose DNA permitted rapid, multiplex detection of high-level oncogene amplifications that were confirmed in FFPE. Virtual slides http://www.diagnosticpathology.diagnomx.eu/vs/1883718801686466.     

Occurrence,distribution and transport of pesticides into the Salton Sea Basin,California, 2001–2002

The Salton Sea is a hypersaline lake located in southeastern California. Concerns over the ecological impacts of sediment quality and potential human exposure to dust emissions from exposed lakebed sediments resulting from anticipated shrinking of shoreline led to a study of pesticide distribution and transport within the Salton Sea Basin, California, in 2001–2002. Three sampling stations—upriver, river mouth, and offshore—were established along each of the three major rivers that discharge into the Salton Sea. Large-volume water samples were collected for analysis of pesticides in water and suspended sediments at the nine sampling stations. Samples of the bottom sediment were also collected at each site for pesticide analysis. Sampling occurred in October 2001, March–April 2002, and October 2002, coinciding with the regional fall and spring peaks in pesticide use in the heavily agricultural watershed. Fourteen current-use pesticides were detected in water and the majority of dissolved concentrations ranged from the limits of detection to 151 ng/l. Diazinon, EPTC and malathion were detected at much higher concentrations (940–3,830 ng/l) at the New and Alamo River upriver and near-shore stations. Concentrations of carbaryl, dacthal, diazinon, and EPTC were higher in the two fall sampling periods, whereas concentrations of atrazine, carbofuran, and trifluralin were higher during the spring, which matched seasonal use patterns of these pesticides. Current-use pesticides were also detected on suspended and bed sediments in concentrations ranging from detection limits to 106 ng/g. Chlorpyrifos, dacthal, EPTC, trifluralin, and DDE were the most frequently detected pesticides on sediments from all three rivers. The number of detections and concentrations of suspended sediment-associated pesticides were often similar for the river upriver and near-shore sites, consistent with downstream transport of pesticides via suspended sediment. While detectable suspended sediment pesticide concentrations were more sporadic than detected aqueous concentrations, seasonal trends were similar to those for dissolved concentrations. Generally, the pesticides detected on suspended sediments were the same as those on the bed sediments, and concentrations were similar, especially at the Alamo River upriver site. With a few exceptions, pesticides were not detected in suspended or bed sediments from the off-shore sites. The partitioning of pesticides between water and sediment was not predictable from solely the physical–chemical properties of individual pesticide compounds, but appear to be a complicated function of the quantity of pesticide applied in the watershed, residence time of sediments in the water, and compound solubility and hydrophobicity. Sediment concentrations of most pesticides were found to be 100–1,000 times lower than the low-effects levels determined in human health risk assessment studies. However, maximum concentrations of chlorpyrifos on suspended sediments were approximately half the low-effects level, suggesting the need for further sediment characterization of lake sediments proximate to riverine inputs. Guest editor: S. H. Hurlbert The Salton Sea Centennial Symposium. Proceedings of a Symposium Celebrating a Century of Symbiosis Among Agriculture, Wildlife and People, 1905–2005, held in San Diego, California, USA, March 2005     

O2 uptake kinetics, pyruvate dehydrogenase activity, and muscle deoxygenation in young and older adults during the transition to moderate-intensity exercise

The adaptation of pulmonary O(2) uptake (Vo(2)(p)) kinetics is slowed in older compared with young adults during the transition to moderate-intensity exercise. In this study, we examined the relationship between Vo(2)(p) kinetics and mitochondrial pyruvate dehydrogenase (PDH) activity in young (n = 7) and older (n = 6) adults. Subjects performed cycle exercise to a work rate corresponding to approximately 90% of estimated lactate threshold. Phase 2 Vo(2)(p) kinetics were slower (P < 0.05) in older (tau = 40 +/- 17 s) compared with young (tau = 21 +/- 6 s) adults. Relative phosphocreatine (PCr) breakdown was greater (P < 0.05) at 30 s in older compared with young adults. Absolute PCr breakdown at 6 min was greater (P < 0.05) in older compared with young adults. In young adults, PDH activity increased (P < 0.05) from baseline to 30 s, with no further change observed at 6 min. In older adults, PDH activity during baseline exercise was similar to that seen in young adults. During the exercise transition, PDH activity did not increase (P > 0.05) at 30 s of exercise but was elevated (P < 0.05) after 6 min. The change in deoxyhemoglobin (HHb) was greater for a given Vo(2)(p) in older adults, and there was a similar time course of HHb accompanying the slower Vo(2)(p) kinetics in the older adults, suggesting a slower adaptation of bulk O(2) delivery in older adults. In conclusion, the slower adaptation of Vo(2)(p) in older adults is likely a result of both an increased metabolic inertia and lower O(2) availability.     

Population structure of Purple Sandpipers (Calidris maritima) as revealed by mitochondrial DNA and microsatellites

The Purple Sandpiper (Calidris maritima) is a medium‐sized shorebird that breeds in the Arctic and winters along northern Atlantic coastlines. Migration routes and affiliations between breeding grounds and wintering grounds are incompletely understood. Some populations appear to be declining, and future management policies for this species will benefit from understanding their migration patterns. This study used two mitochondrial DNA markers and 10 microsatellite loci to analyze current population structure and historical demographic trends. Samples were obtained from breeding locations in Nunavut (Canada), Iceland, and Svalbard (Norway) and from wintering locations along the coast of Maine (USA), Nova Scotia, New Brunswick, and Newfoundland (Canada), and Scotland (UK). Mitochondrial haplotypes displayed low genetic diversity, and a shallow phylogeny indicating recent divergence. With the exception of the two Canadian breeding populations from Nunavut, there was significant genetic differentiation among samples from all breeding locations; however, none of the breeding populations was a monophyletic group. We also found differentiation between both Iceland and Svalbard breeding populations and North American wintering populations. This pattern of divergence is consistent with a previously proposed migratory pathway between Canadian breeding locations and wintering grounds in the United Kingdom, but argues against migration between breeding grounds in Iceland and Svalbard and wintering grounds in North America. Breeding birds from Svalbard also showed a genetic signature intermediate between Canadian breeders and Icelandic breeders. Our results extend current knowledge of Purple Sandpiper population genetic structure and present new information regarding migration routes to wintering grounds in North America.     

Effective strategies for scaling up evidence-based practices in primary care: a systematic review

Background

While an extensive array of existing evidence-based practices (EBPs) have the potential to improve patient outcomes, little is known about how to implement EBPs on a larger scale. Therefore, we sought to identify effective strategies for scaling up EBPs in primary care.

Methods

We conducted a systematic review with the following inclusion criteria: (i) study design: randomized and non-randomized controlled trials, before-and-after (with/without control), and interrupted time series; (ii) participants: primary care-related units (e.g., clinical sites, patients); (iii) intervention: any strategy used to scale up an EBP; (iv) comparator: no restrictions; and (v) outcomes: no restrictions. We searched MEDLINE, Embase, PsycINFO, Web of Science, CINAHL, and the Cochrane Library from database inception to August 2016 and consulted clinical trial registries and gray literature. Two reviewers independently selected eligible studies, then extracted and analyzed data following the Cochrane methodology. We extracted components of scaling-up strategies and classified them into five categories: infrastructure, policy/regulation, financial, human resources-related, and patient involvement. We extracted scaling-up process outcomes, such as coverage, and provider/patient outcomes. We validated data extraction with study authors.

Results

We included 14 studies. They were published since 2003 and primarily conducted in low-/middle-income countries (n?=?11). Most were funded by governmental organizations (n?=?8). The clinical area most represented was infectious diseases (HIV, tuberculosis, and malaria, n?=?8), followed by newborn/child care (n?=?4), depression (n?=?1), and preventing seniors’ falls (n?=?1). Study designs were mostly before-and-after (without control, n?=?8). The most frequently targeted unit of scaling up was the clinical site (n?=?11). The component of a scaling-up strategy most frequently mentioned was human resource-related (n?=?12). All studies reported patient/provider outcomes. Three studies reported scaling-up coverage, but no study quantitatively reported achieving a coverage of 80% in combination with a favorable impact.

Conclusions

We found few studies assessing strategies for scaling up EBPs in primary care settings. It is uncertain whether any strategies were effective as most studies focused more on patient/provider outcomes and less on scaling-up process outcomes. Minimal consensus on the metrics of scaling up are needed for assessing the scaling up of EBPs in primary care.

Trial registration

This review is registered as PROSPERO CRD42016041461.

     

Effects of umbilical cord blood stem cells on healing factors for diabetic foot injuries

The use of stem or progenitor cells from bone marrow, or peripheral or umbilical cord blood is becoming more common for treatment of diabetic foot problems. These cells promote neovascularization by angiogenic factors and they promote epithelium formation by stimulating cell replication and migration under certain pathological conditions. We investigated the role of CD34 + stem cells from human umbilical cord blood in wound healing using a rat model. Rats were randomly divided into a control group and two groups with diabetes induced by a single dose of 55 mg/kg intraperitoneal streptozocin. Scarred areas 5 mm in diameter were created on the feet of all rats. The diabetic rats constituted the diabetes control group and a diabetes + stem cell group with local injection into the wound site of 0.5 × 106 CD34 + stem cells from human umbilical cord blood. The newly formed skin in the foot wounds following CD34 + stem cell treatment showed significantly improvement by immunohistochemistry and TUNEL staining, and were closer to the wound healing of the control group than the untreated diabetic animals. The increase in FGF expression that accompanied the local injection of CD34 + stem cells indicates that FGF stimulation helped prevent apoptosis. Our findings suggest a promising new treatment approach to diabetic wound healing.     

Effects of strobilurin containing fungicides on the deoxynivalenol content in winter wheat

The effects of the strobilurin fungicides Juwel, Juwel Top and Amistar on the deoxynivalenol contamination of winter wheat was studied in field experiments. In general, the application of strobilurins during stem elongation and inflorescence emergence of wheat resulted in increased deoxynivalenol contents in kernels as compared with the untreated control. This stimulating effect can be reversed by a following azole fungicide applied within a time period of one week during the stages of flowering.