Sequence-Dependent Fluorescence of Cyanine Dyes on Microarrays

Cy3 and Cy5 are among the most commonly used oligonucleotide labeling molecules. Studies of nucleic acid structure and dynamics use these dyes, and they are ubiquitous in microarray experiments. They are sensitive to their environment and have higher quantum yield when bound to DNA. The fluorescent intensity of terminal cyanine dyes is also known to be significantly dependent on the base sequence of the oligonucleotide. We have developed a very precise and high-throughput method to evaluate the sequence dependence of oligonucleotide labeling dyes using microarrays and have applied the method to Cy3 and Cy5. We used light-directed in-situ synthesis of terminally-labeled microarrays to determine the fluorescence intensity of each dye on all 1024 possible 5′-labeled 5-mers. Their intensity is sensitive to all five bases. Their fluorescence is higher with 5′ guanines, and adenines in subsequent positions. Cytosine suppresses fluorescence. Intensity falls by half over the range of all 5-mers for Cy3, and two-thirds for Cy5. Labeling with 5′-biotin-streptavidin-Cy3/-Cy5 gives a completely different sequence dependence and greatly reduces fluorescence compared with direct terminal labeling

Expression of amphetamine sensitization is associated with recruitment of a reactive neuronal population in the nucleus accumbens core

Rationale: Repeated exposure to psychostimulant drugs causes a long-lasting increase in the psychomotor and reinforcing effects of these drugs and an array of neuroadaptations. One such alteration is a hypersensitivity of striatal activity such that a low dose of amphetamine in sensitized animals produces dorsal striatal activation patterns similar to acute treatment with a high dose of amphetamine. Objectives: To extend previous findings of striatal hypersensitivity with behavioral observations and with cellular activity in the nucleus accumbens and prefrontal cortex in sensitized animals. Materials and methods: Rats treated acutely with 0, 1, 2.5, or 5 mg/kg i.p. amphetamine and sensitized rats challenged with 1 mg/kg i.p. amphetamine were scored for stereotypy, rearing, and grooming, and locomotor activity recorded. c-fos positive nuclei were quantified in the nucleus accumbens and prefrontal cortex after expression of sensitization with 1 mg/kg i.p. amphetamine. Results: Intense stereotypy was seen in animals treated acutely with 5 mg/kg amphetamine, but not in the sensitized group treated with 1 mg/kg amphetamine. The c-fos response to amphetamine in the accumbens core was augmented in amphetamine-pretreated animals with a shift in the distribution of optical density, while no effect of sensitization was seen in the nucleus accumbens shell or prefrontal cortex. Conclusions A lack of stereotypy in the sensitized group indicates a dissociation of behavioral responses to amphetamine and striatal immediate-early gene activation patterns. The increase in c-fos positive nuclei and shift in the distribution of optical density observed in the nucleus accumbens core suggests recruitment of a new population of neurons during expression of sensitization

A Strawberry KNOX Gene Regulates Leaf, Flower and Meristem Architecture

The KNOTTED-LIKE HOMEODOMAIN (KNOX) genes play a central role in maintenance of the shoot apical meristem. They also contribute to the morphology of simple and compound leaves. In this report we characterize the FaKNOX1 gene from strawberry (Fragaria spp.) and demonstrate its function in trasgenic plants. The FaKNOX1 cDNA was isolated from a cultivated strawberry (F.×ananassa) flower EST library. The sequence is most similar to Class I KNOX genes, and was mapped to linkage group VI of the diploid strawberry genome. Unlike most KNOX genes studied, steady-state transcript levels were highest in flowers and fruits. Transcripts were also detected in emerging leaf primordia and the apical dome. Transgenic strawberry plants suppressing or overexpressing FaKNOX1 exhibited conspicuous changes in plant form. The FaKNOX1 RNAi plants presented a dwarfed phenotype with deeply serrated leaflets and exaggerated petiolules. They also exhibited a high level of cellular disorganization of the shoot apical meristem and leaves. Overexpression of FaKNOX1 caused dwarfed stature with wrinkled leaves. These gain- and loss-of-function assays in strawberry functionally demonstrate the contributions of a KNOX domain protein in a rosaceous species

Chronic kidney disease after liver, cardiac, lung, heart–lung, and hematopoietic stem cell transplant

Patient survival after cardiac, liver, and hematopoietic stem cell transplant (HSCT) is improving; however, this survival is limited by substantial pretransplant and treatment-related toxicities. A major cause of morbidity and mortality after transplant is chronic kidney disease (CKD). Although the majority of CKD after transplant is attributed to the use of calcineurin inhibitors, various other conditions such as thrombotic microangiopathy, nephrotic syndrome, and focal segmental glomerulosclerosis have been described. Though the immunosuppression used for each of the transplant types, cardiac, liver and HSCT is similar, the risk factors for developing CKD and the CKD severity described in patients after transplant vary. As the indications for transplant and the long-term survival improves for these children, so will the burden of CKD. Nephrologists should be involved early in the pretransplant workup of these patients. Transplant physicians and nephrologists will need to work together to identify those patients at risk of developing CKD early to prevent its development and progression to end-stage renal disease

Emerging concepts in biomarker discovery; The US-Japan workshop on immunological molecular markers in oncology

Supported by the Office of International Affairs, National Cancer Institute (NCI), the "US-Japan Workshop on Immunological Biomarkers in Oncology" was held in March 2009. The workshop was related to a task force launched by the International Society for the Biological Therapy of Cancer (iSBTc) and the United States Food and Drug Administration (FDA) to identify strategies for biomarker discovery and validation in the field of biotherapy. The effort will culminate on October 28th 2009 in the "iSBTc-FDA-NCI Workshop on Prognostic and Predictive Immunologic Biomarkers in Cancer", which will be held in Washington DC in association with the Annual Meeting. The purposes of the US-Japan workshop were a) to discuss novel approaches to enhance the discovery of predictive and/or prognostic markers in cancer immunotherapy; b) to define the state of the science in biomarker discovery and validation. The participation of Japanese and US scientists provided the opportunity to identify shared or discordant themes across the distinct immune genetic background and the diverse prevalence of disease between the two Nations

Intraperitoneal drain placement and outcomes after elective colorectal surgery: international matched, prospective, cohort study

Despite current guidelines, intraperitoneal drain placement after elective colorectal surgery remains widespread. Drains were not associated with earlier detection of intraperitoneal collections, but were associated with prolonged hospital stay and increased risk of surgical-site infections.Background Many surgeons routinely place intraperitoneal drains after elective colorectal surgery. However, enhanced recovery after surgery guidelines recommend against their routine use owing to a lack of clear clinical benefit. This study aimed to describe international variation in intraperitoneal drain placement and the safety of this practice. Methods COMPASS (COMPlicAted intra-abdominal collectionS after colorectal Surgery) was a prospective, international, cohort study which enrolled consecutive adults undergoing elective colorectal surgery (February to March 2020). The primary outcome was the rate of intraperitoneal drain placement. Secondary outcomes included: rate and time to diagnosis of postoperative intraperitoneal collections; rate of surgical site infections (SSIs); time to discharge; and 30-day major postoperative complications (Clavien-Dindo grade at least III). After propensity score matching, multivariable logistic regression and Cox proportional hazards regression were used to estimate the independent association of the secondary outcomes with drain placement. Results Overall, 1805 patients from 22 countries were included (798 women, 44.2 per cent; median age 67.0 years). The drain insertion rate was 51.9 per cent (937 patients). After matching, drains were not associated with reduced rates (odds ratio (OR) 1.33, 95 per cent c.i. 0.79 to 2.23; P = 0.287) or earlier detection (hazard ratio (HR) 0.87, 0.33 to 2.31; P = 0.780) of collections. Although not associated with worse major postoperative complications (OR 1.09, 0.68 to 1.75; P = 0.709), drains were associated with delayed hospital discharge (HR 0.58, 0.52 to 0.66; P < 0.001) and an increased risk of SSIs (OR 2.47, 1.50 to 4.05; P < 0.001). Conclusion Intraperitoneal drain placement after elective colorectal surgery is not associated with earlier detection of postoperative collections, but prolongs hospital stay and increases SSI risk