RRP1B is a metastasis modifier that regulates the expression of alternative mRNA isoforms through interactions with SRSF1.

RRP1B (ribosomal RNA processing 1 homolog B) was first identified as a metastasis susceptibility gene in breast cancer through its ability to modulate gene expression in a manner that can be used to accurately predict prognosis in breast cancer. However, the mechanism(s) by which RRP1B modulates gene expression is currently unclear. Many RRP1B binding candidates are involved in alternative splicing, a mechanism of gene expression regulation that is increasingly recognized to be involved in cancer progression and metastasis. One such target is SRSF1 (serine/arginine-rich splicing factor 1) (SF2/ASF, splicing factor 2/alternative splicing factor), an essential splicing regulator that also functions as an oncoprotein. Earlier studies demonstrated that splicing and transcription occur concurrently and are coupled processes. Given that RRP1B regulates transcriptional activity, we hypothesized that RRP1B also regulates the expression of alternative mRNA isoforms through its interaction with SRSF1. Interaction between RRP1B and SRSF1 was verified by coimmunoprecipitation and coimmunofluorescence. Treatment of cells with transcriptional inhibitors significantly increased this interaction, demonstrating that the association of these two proteins is transcriptionally regulated. To assess the role of RRP1B in the regulation of alternative isoform expression, RNA-sequencing data were generated from control and Rrp1b-knockdown cells. Knockdown of Rrp1b induced a significant change in isoform expression in over 600 genes compared with control cell lines. This was verified by quantitative reverse-transcription PCR using isoform-specific primers. Pathway enrichment analyses identified cell cycle and checkpoint regulation to be those most affected by Rrp1b knockdown. These data suggest that RRP1B suppresses metastatic progression by altering the transcriptome through its interaction with splicing regulators such as SRSF1.

Quantitative analysis of Pc 4 localization in mouse lymphoma (LY-R) cells via double-label confocal fluorescence microscopy.

Photodynamic therapy (PDT) is a novel cancer therapy that uses light-activated drugs (photosensitizers) to destroy tumor tissue. Reactive oxygen species produced during PDT are thought to cause the destruction of tumor tissue. However, the precise mechanism of PDT is not completely understood. To provide insight into the in vitro mechanisms of PDT, we studied the subcellular localization of the photosensitizer HOSiPcOSi(CH3)2-(CH2)3N(CH3)2 (Pc 4) in mouse lymphoma (LY-R) cells using double-label confocal fluorescence microscopy. This technique allowed us to observe the relative distributions of Pc 4 and an organelle-specific dye within the same cell via two, spectrally distinct, fluorescence images. To quantify the localization of Pc 4 within different organelles, linear correlation coefficients from the fluorescence data of Pc 4 and the organelle-specific dyes were calculated. Using this measurement, the subcellular spatial distributions of Pc 4 could be successfully monitored over an 18 h period. At early times (0-1 h) after introduction of Pc 4 to LY-R cells, the dye was found in the mitochondria, lysosomes and Golgi apparatus, as well as other cytoplasmic membranes, but not in the plasma membrane or the nucleus. Over the next 2 h, there was some loss of Pc 4 from the lysosomes as shown by the correlation coefficients. After an additional incubation period of 2 h Pc 4 slowly increased its accumulation in the lysosomes. The highest correlation coefficient (0.65) was for Pc 4 and BODIPY-FL C5 ceramide, which targets the Golgi apparatus, and also binds to other cytoplasmic membranes. The correlation coefficient was also high (0.60) for Pc 4 and a mitochondria-targeting dye (Mitotracker Green FM). Both of these correlation coefficients were higher than that for Pc 4 with the lysosome-targeting dye (Lysotracker Green DND-26). The results suggest that Pc 4 binds preferentially and strongly to mitochondria and Golgi complexes.

Effects of motion, ambient light, and hypoperfusion on pulse oximeter function.

STUDY OBJECTIVE: To compare the performance of five pulse oximeters during hypoperfusion, probe motion, and exposure to ambient light interference. DESIGN: Prospective study. SETTING: Laboratory facility at a university medical center. PATIENTS: 8 unanesthetized, ASA physical status I volunteers. INTERVENTIONS: We evaluated five common pulse oximeters with respect to three scenarios: (1) an operating room light was shone on oximeter probes, (2) a motion generator was used to generate 2 Hz and 4 Hz hand motion, and (3) a pneumatic compression device overlying the brachial artery was used to simulate hypoperfusion. Electrocardiographic (ECG) and arterial blood gas values were considered gold standards for heart rate (HR) and oxygen saturation (SpO2) respectively. SpO2 nondisplay and values greater than 4% from simultaneous arterial SaO2-oximeter values were defined as errors. Nondisplay of HR, or HR greater than 5% from ECG values, were also considered errors. MEASUREMENTS AND MAIN RESULTS: The Ohmeda and Nellcor N200 with finger probe had the highest total failure rates with respect to both SpO2 and HR due to ambient light interference (p < 0.05). The Nellcor N200 with finger probe and N200 with C lock were the most accurate with regard to SpO2 during 2 Hz and 4 Hz motion (p < 0.05). However, all oximeters failed dramatically during 4 Hz motion when measuring HR. In the hypoperfusion model, the Nellcor N200 with finger probe and the Nellcor C Lock oximeters performed significantly better than all others in terms of both HR and SpO2 (P < 0.05), while the Criticare oximeter failed 100% of the time. CONCLUSION: There are significant differences in the accuracy of commercially available pulse oximeters during nonideal circumstances, with failure rates varying from approximately 5% to 50% depending on the oximeter and source of interference. Furthermore, no single oximeter performed the best under all conditions.

Pulse oximeter performance during desaturation and resaturation: a comparison of seven models.

STUDY OBJECTIVE: To compare pulse oximeter performance during induced hypoxemia. DESIGN: Prospective investigation in human volunteers. SETTING: Laboratory facility at a university medical center. PATIENTS: 8 unanesthetized, healthy ASA physical status I volunteers. INTERVENTIONS: We evaluated the accuracy and response times of seven popular pulse oximeters during induced hypoxemia. Arterial blood fractional oxygen saturation (SaO2) measurements were performed simultaneously and considered a gold standard. MEASUREMENTS AND MAIN RESULTS: All oximeters were accurate (+/-2%) while subjects were breathing room air. During maximal hypoxemia (induced by breathing a FIO2 = 10% in nitrogen), large differences were noted between oxygen saturation as measured by pulse oximetry (SpO2) and SaO2 values, with pulse oximeters consistently underreporting SpO2 when actual SaO2 values were 75% or less. The Ohmeda 3740 (Ohmeda, Boulder, CO) using an ear probe was the first to detect desaturation (change in SpO2 > 3%) in 4 of 8 subjects (p < 0.05), and the Nellcor N200 reflectance oximeter (Nellcor, Inc., Pleasanton, CA) was first in 3 of 8 subjects (p < 0.05). During resaturation (after administering 100% oxygen), the Novametrix Oxypleth (Novametrix, Wallingford, CT) was significantly faster than other oximeters (p < 0.05) to return to baseline (SpO2 = 98%). CONCLUSION: Most models of oximeters tested performed well when hemoglobin oxygen saturation was high, but all were inaccurate when SaO2 was approximately 75%. During induced hypoxemia, there were significant differences in the response times of oximeters tested, with no model demonstrably superior to others in all measures of performance.

Headache prevention following accidental dural puncture in obstetric patients.

STUDY OBJECTIVE: To evaluate the efficacy of a prophylactic saline patch and a prophylactic blood patch in prevention of headache following accidental dural puncture in obstetric patients. DESIGN: Prospective, randomized, single-blind study on dural puncture headache occurrence and cessation. SETTING: Inpatient obstetric unit at a metropolitan medical center. PATIENTS: Seventy-four inpatients who underwent vaginal delivery or cesarean section following accidental dural puncture during administration of epidural anesthesia for labor and delivery. INTERVENTIONS: Group 1 (n = 24), the control group, received fluids and analgesics. Group 2 (n = 30) received prophylactic epidural saline (40 to 60 ml) through the epidural catheter following completion of the obstetric procedure. Group 3 (n = 20) received autologous blood (15 ml) via epidural catheter following completion of the obstetric procedure. MEASUREMENTS AND MAIN RESULTS: In Group 1, 21 of 24 patients (87.5%) developed headaches, with conservative management. In Group 2, 20 of 30 patients (66.7%) developed headaches, and in Group 3, 1 of 20 patients (5%) developed a headache. CONCLUSIONS: The results of this study suggest that the administration of a prophylactic epidural blood patch is highly effective in the prevention of headaches following dural puncture, with headache frequency reduced from 87.5% to 5%.

Interpleural block: a new technique for regional anaesthesia during percutaneous nephrostomy and nephrolithotomy.

Interpleural block was used in four patients undergoing percutaneous nephrostomy, one of whom also underwent percutaneous nephrolithotomy. Interpleural block was achieved with the standard technique using 30 ml of 0.5 per cent bupivacaine. All patients tolerated the procedure well and remained haemodynamically stable during the operative procedure. Mean pain relief from initiation of interpleural block was ten hours (SD = 4.32). Interpleural block was an effective method of obtaining anaesthesia for percutaneous nephrostomy and nephrolithotomy in these four patients.