Reliability of the surgical Pleth index for assessment of postoperative pain: a pilot study.

BACKGROUND: Postoperative pain control is essential and may have a beneficial effect on postoperative outcome and morbidity. Analgesia quality is controlled using tools such as a Numerical Rating Scale (NRS). These tools require cooperation and often fail in the presence of reduced awareness. The Surgical Pleth index (SPI) has been introduced as a monitoring tool for intraoperative pain under general anaesthesia. OBJECTIVE: We investigated the correlation between SPI and pain intensity, analgesic consumption and fitness for discharge in the postanaesthesia care unit. DESIGN: An observational study. SETTING: The central postanaesthesia care unit of our tertiary care hospital. PATIENTS: Written informed consent was obtained from 100 patients scheduled for elective surgery under general anaesthesia. Patients below the age of 18 years and those with an abnormal cardiac rhythm were excluded from the study. INTERVENTION: Patients were interviewed every 10 min for 2 h. MAIN OUTCOME MEASURES: Pain intensity measured by NRS, discomfort and Aldrete and Post-Anaesthetic Discharge Scoring System (PADSS) scores were noted. SPI and total dose of opioids administered were recorded. RESULTS: A total of 1300 pain measurements were recorded; 482 (37%) reflected no or mild pain (NRS 0 to 3), 532 (41%) moderate pain (NRS 4 to 6) and 286 (22%) severe pain (NRS 7 to 10). Both NRS (r = 0.62, P < 0.001) and SPI (r = 0.38, P < 0.001) correlated significantly with total opioid consumption. SPI showed a moderate correlation with NRS (r = 0.49, P < 0.001). Receiver operating characteristic analysis showed moderate sensitivity and specificity for discrimination between low and moderate pain (NRS ≤3) (sensitivity 67%, specificity 69% for SPI ≤45), and between moderate and severe pain (NRS >6) (sensitivity 72%, specificity 72% for SPI ≥57). SPI and NRS showed weak negative correlations with Aldrete and PADSS scores. CONCLUSION: Sensitivity and specificity of SPI to discriminate between low, moderate and severe pain levels was moderate. Both NRS and SPI correlated significantly with total opioid consumption.

EASY-HIT: HIV full-replication technology for broad discovery of multiple classes of HIV inhibitors.

HIV replication assays are important tools for HIV drug discovery efforts. Here, we present a full HIV replication system (EASY-HIT) for the identification and analysis of HIV inhibitors. This technology is based on adherently growing HIV-susceptible cells, with a stable fluorescent reporter gene activated by HIV Tat and Rev. A fluorescence-based assay was designed that measures HIV infection by two parameters relating to the early and the late phases of HIV replication, respectively. Validation of the assay with a panel of nine reference inhibitors yielded effective inhibitory concentrations consistent with published data and allowed discrimination between inhibitors of early and late phases of HIV replication. Finer resolution of the effects of reference drugs on different steps of HIV replication was achieved in secondary time-of-addition assays. The EASY-HIT assay yielded high Z' scores (>0.9) and signal stabilities, confirming its robustness. Screening of the LOPAC(1280) library identified 10 compounds (0.8%), of which eight were known to inhibit HIV, validating the suitability of this assay for screening applications. Studies evaluating anti-HIV activities of natural products with the EASY-HIT technology led to the identification of three novel inhibitory compounds that apparently act at different steps of HIV-1 replication. Furthermore, we demonstrate successful evaluation of plant extracts for HIV-inhibitory activities, suggesting application of this technology for the surveillance of biological extracts with anti-HIV activities. We conclude that the EASY-HIT technology is a versatile tool for the discovery and characterization of HIV inhibitors.

Long-term HIV-1 infection of neural progenitor populations.

BACKGROUND: HIV can reside in the brain for many years. While astrocytes are known to tolerate long-term HIV infection, the potential of other neural cell types to harbour HIV is unclear. OBJECTIVE: To investigate whether HIV can persist in neural progenitor cell populations. DESIGN: A multipotent human neural stem cell line (HNSC.100) was used to compare HIV infection in neural progenitor and astrocyte cell populations. METHODS: Expression of cellular genes/proteins was analysed by real-time reverse transcriptase PCR, Western blot, immunocytochemistry and flow cytometry. Morphological properties of cells were measured by quantitative fluorescent image analysis. Virus release by cells exposed to HIV-1IIIB was monitored by enzyme-linked immunosorbent assay for Gag. Proviral copy numbers were determined by real-time PCR and early HIV transcripts by reverse transcriptase PCR. Rev activity was determined with a fluorescent-based reporter assay. RESULTS: Progenitor populations differed from astrocyte populations by showing much lower glial fibrillary acidic protein (GFAP) production, higher cell-surface expression of the CXCR4 chemokine receptor, higher Rev activity and distinct cell morphologies. HIV-exposed progenitor cultures released moderate amounts of virus for over 2 months and continued to display cell-associated HIV markers (proviral DNA, early HIV transcripts) during the entire observation period (115 days). Differentiation of HIV-infected progenitor cells to astrocytes was associated with transient activation of virus production. Long-term HIV infection of progenitor populations led to upregulation of GFAP and changes in cell morphology. CONCLUSION: These studies suggest that neural progenitor populations can contribute to the reservoir for HIV in the brain and undergo changes as a consequence of HIV persistence.

Analysis of the influence of subcellular localization of the HIV Rev protein on Rev-dependent gene expression by multi-fluorescence live-cell imaging.

The human immunodeficiency virus Rev protein is a post-transcriptional activator of HIV gene expression. Rev is a nucleocytoplasmic shuttle protein that displays characteristic nuclear/nucleolar subcellular localization in various cell lines. Cytoplasmic localization of Rev occurs under various conditions disrupting Rev function. The goal of this study was to investigate the relationship between localization of Rev and its functional activity in living cells. A triple-fluorescent imaging assay, called AQ-FIND, was established for automatic quantitative evaluation of nucleocytoplasmic distribution of fluorescently tagged proteins. This assay was used to screen 500 rev genes generated by error-prone PCR for Rev mutants with different localization phenotypes. Activities of the Rev mutants were determined with a second quantitative, dual-fluorescent reporter assay. In HeLa cells, the majority of nuclear Rev mutants had activities similar to wild-type Rev. The activities of Rev mutants with abnormal cytoplasmic localization ranged from moderately impaired to nonfunctional. There was no linear correlation between subcellular distribution and levels of Rev activity. In astrocytes, nuclear Rev mutants showed similar impaired activities as the cytoplasmic wild-type Rev. Our data suggest that steady-state subcellular localization is not a primary regulator of Rev activity but may change as a secondary consequence of altered Rev function. The methodologies described here have potential for studying the significance of subcellular localization for functions of other regulatory factors.