Characterization of the redox activity and disulfide bond formation in apurinic/apyrimidinic endonuclease

Apurinic/apyrimidinic endonuclease (APE1) is an unusual nuclear redox factor in which the redox-active cysteines identified to date, C65 and C93, are surface inaccessible residues whose activities may be influenced by partial unfolding of APE1. To assess the role of the five remaining cysteines in APE1's redox activity, double-cysteine mutants were analyzed, excluding C65A, which is redox-inactive as a single mutant. C93A/C99A APE1 was found to be redox-inactive, whereas other double-cysteine mutants retained the same redox activity as that observed for C93A APE1. To determine whether these three cysteines, C65, C93, and C99, were sufficient for redox activity, all other cysteines were substituted with alanine, and this protein was shown to be fully redox-active. Mutants with impaired redox activity failed to stimulate cell proliferation, establishing an important role for APE1's redox activity in cell growth. Disulfide bond formation upon oxidation of APE1 was analyzed by proteolysis of the protein followed by mass spectrometry analysis. Within 5 min of exposure to hydrogen peroxide, a single disulfide bond formed between C65 and C138 followed by the formation of three additional disulfide bonds within 15 min; 10 total disulfide bonds formed within 1 h. A single mixed-disulfide bond involving C99 of APE1 was observed for the reaction of oxidized APE1 with thioredoxin (TRX). Disulfide-bonded APE1 or APE1-TRX species were further characterized by size exclusion chromatography and found to form large complexes. Taken together, our data suggest that APE1 is a unique redox factor with properties distinct from those of other redox factors.     

A Novel Brief Therapy for Patients Who Attempt Suicide: A 24-months Follow-Up Randomized Controlled Study of the Attempted Suicide Short Intervention Program (ASSIP)

Background

Attempted suicide is the main risk factor for suicide and repeated suicide attempts. However, the evidence for follow-up treatments reducing suicidal behavior in these patients is limited. The objective of the present study was to evaluate the efficacy of the Attempted Suicide Short Intervention Program (ASSIP) in reducing suicidal behavior. ASSIP is a novel brief therapy based on a patient-centered model of suicidal behavior, with an emphasis on early therapeutic alliance.

Methods and Findings

Patients who had recently attempted suicide were randomly allocated to treatment as usual (n = 60) or treatment as usual plus ASSIP (n = 60). ASSIP participants received three therapy sessions followed by regular contact through personalized letters over 24 months. Participants considered to be at high risk of suicide were included, 63% were diagnosed with an affective disorder, and 50% had a history of prior suicide attempts. Clinical exclusion criteria were habitual self-harm, serious cognitive impairment, and psychotic disorder. Study participants completed a set of psychosocial and clinical questionnaires every 6 months over a 24-month follow-up period.The study represents a real-world clinical setting at an outpatient clinic of a university hospital of psychiatry. The primary outcome measure was repeat suicide attempts during the 24-month follow-up period. Secondary outcome measures were suicidal ideation, depression, and health-care utilization. Furthermore, effects of prior suicide attempts, depression at baseline, diagnosis, and therapeutic alliance on outcome were investigated.During the 24-month follow-up period, five repeat suicide attempts were recorded in the ASSIP group and 41 attempts in the control group. The rates of participants reattempting suicide at least once were 8.3% (n = 5) and 26.7% (n = 16). ASSIP was associated with an approximately 80% reduced risk of participants making at least one repeat suicide attempt (Wald χ² ₁ = 13.1, 95% CI 12.4–13.7, p < 0.001). ASSIP participants spent 72% fewer days in the hospital during follow-up (ASSIP: 29 d; control group: 105 d; W = 94.5, p = 0.038). Higher scores of patient-rated therapeutic alliance in the ASSIP group were associated with a lower rate of repeat suicide attempts. Prior suicide attempts, depression, and a diagnosis of personality disorder at baseline did not significantly affect outcome. Participants with a diagnosis of borderline personality disorder (n = 20) had more previous suicide attempts and a higher number of reattempts.Key study limitations were missing data and dropout rates. Although both were generally low, they increased during follow-up. At 24 months, the group difference in dropout rate was significant: ASSIP, 7% (n = 4); control, 22% (n = 13). A further limitation is that we do not have detailed information of the co-active follow-up treatment apart from participant self-reports every 6 months on the setting and the duration of the co-active treatment.

Conclusions

ASSIP, a manual-based brief therapy for patients who have recently attempted suicide, administered in addition to the usual clinical treatment, was efficacious in reducing suicidal behavior in a real-world clinical setting. ASSIP fulfills the need for an easy-to-administer low-cost intervention. Large pragmatic trials will be needed to conclusively establish the efficacy of ASSIP and replicate our findings in other clinical settings.

Trial registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT02505373","term_id":"NCT02505373"}}NCT02505373     

Effects of farnesol and the off-flavor derivative geosmin on Streptomyces tendae.

Effects of the sesquiterpene farnesol (3,7,11-trimethyl-2,6,10-dodecatrien-1-ol) and the sesquiterpene derivative geosmin (1,10-trans-dimethyl-trans-9-decalol) were investigated in a geosmin-producing actinomycete, Streptomyces tendae. Exposure to 300 microM farnesol reduced biomass (fresh matter) accumulation by 97% compared with biomass accumulation by controls, whereas an equal amount of geosmin did not affect biomass accumulation. Increasing exposure to farnesol corresponded with reduced optical density of the culture, reduced levels of geosmin, and reduced metabolic heat production compared with controls, while exogenous geosmin did not affect these parameters. Geosmin dissipated from unioculated medium more rapidly than farnesol, indicating that in addition to the lower toxicity of geosmin, the actual exposure to geosmin over time may be less than exposure to an equal amount of farnesol. Cultures grown on Actinomyces-B medium contained 99.5% less geosmin and were more sensitive to farnesol than those grown on Hickey-Tresner medium, indicating that geosmin synthesis was associated with reduced sensitivity to farnesol. Consumption of farnesyl moieties during geosmin synthesis may reduce the potential for farnesol-induced inhibition of growth and metabolism.     

HIGH-RESOLUTION AIRBORNE REMOTE SENSING OF BLOOM-FORMING PHYTOPLANKTON1

Remote sensing of highly turbid finfish aquaculture impoundments using the Calibrated Airborne Multispectral Scanner (CAMS) mounted on a Lear jet flown at 900 m was conducted in central Mississippi on 16 May 1990. Concurrent in situ data consisted of phytoplankton pigment concentrations and standing crop, water color, turbidity, and surface-water temperature. Surface and near-surface assemblages of cyanophytes and chlorophytes varied dramatically among impoundments; total chlorophyll concentrations and standing crop values ranged from 8 to 483 mg·m^?3 and 8.0 × 10² to 2.2 × 10⁶ cells-mL^?1, respectively. Regression models fit to CAMS data provided reliable estimates for and produced accurate digital cartographs of total chlorophyll and carotenoid concentrations, phytoplankton standing crop, and turbidity. Although a model to effectively estimate in situ c-phycocyanin concentrations was not identified, the lack of a suitable model may have resulted from variability of pigment extraction during quantification rather than failure of remotely sensed imagery to detect c-phycocyanin. Models derived from imagery of impoundments directly beneath the aircraft sufficiently described in situ parameters in imagery of adjacent series of impoundments not directly below the aircraft. High-resolution airborne remote sensing provides a means for monitoring local phytoplankton dynamics in temporal and spatial scales analogous to biotic and abiotic processes affecting such dynamics and necessary for applications to ecological research and fisheries or aquacultural management.     

Mechanically induced c-fos expression is mediated by cAMP in MC3T3-E1 osteoblasts.

In serum-deprived MC3T3-E1 osteoblasts, mechanical stimulation caused by mild (287 x g) centrifugation induced a 10-fold increase in mRNA levels of the proto-oncogene, c-fos. Induction of c-fos was abolished by the cAMP-dependent protein kinase inhibitor H-89, suggesting that the transient c-fos mRNA increase is mediated by cAMP. Down-regulation of protein kinase C (PKC) activity by chronic TPA treatment failed to significantly reduce c-fos induction, suggesting that TPA-sensitive isoforms of PKC are not responsible for c-fos up-regulation. In addition, 287 x g centrifugation increased intracellular prostaglandin E2 (PGE2) levels 2.8-fold (P<0. 005). Since we have previously shown that prostaglandin E2 (PGE2) can induce c-fos expression via a cAMP-mediated mechanism, we asked whether the increase in c-fos mRNA was due to centrifugation-induced PGE2 release. Pretreatment with the cyclooxygenase inhibitors indomethacin and flurbiprofen did not hinder the early induction of c-fos by mechanical stimulation. We conclude that c-fos expression induced by mild mechanical loading is dependent primarily on cAMP, not PKC, and initial induction of c-fos is not necessarily dependent on the action of newly synthesized PGE2.     

Physical-Biological Coupling in Southern Lake Michigan: Influence of Episodic Sediment Resuspension on Phytoplankton

The influence of episodic, sediment resuspension on phytoplankton abundance/volume and composition, the photosynthetic maximum rate (P^B _max) and efficiency (^B), and chlorophyll-specific growth (_Chl) was evaluated during the spring isothermal period in southern Lake Michigan (Laurentian Great Lakes, USA). Resuspension altered the nutrient and light climate of nearshore waters; light attenuation (K_d) and phosphorus concentrations corresponded (p 0.0001 and p 0.001, respectively) with concentrations of suspended particulate matter (SPM). Phytoplankton cell volume and diatom cell abundance and volume were not associated with SPM concentrations (p > 0.05). Diatom composition displayed spatial dissimilarities corresponding with resuspension (p 0.001); small centric diatoms exhibiting meroplanktonic life histories and pennate diatoms considered benthic in origin were most abundant within SPM-impacted, nearshore waters whereas taxa typically comprising assemblages in optically-clear, offshore waters and the basin-wide, spring bloom were not. Values of P^B _max and ^B corresponded (p 0.0001) with both K_d coefficients and SPM concentrations, potentially reflecting increased light harvesting/utilization within impacted assemblages. However, integral production was inversely associated with K_d coefficients and SPM concentrations (p < 0.0001) and photosynthesis was light-limited (or nearly so) for most assemblages. Although _Chl values corresponded with K_d coefficients (p 0.05), values were quite low (x ± S.E., 0.10 ± 0.004 d^-1) throughout the study. Most likely, distinct rate processes between SPM- and non-impacted assemblages reflected short-term compositional (and corresponding physiological) variations due to infusion of meroplankton and/or tributary-derived phytoplankton. Overall, resuspension appears to have little, if any, long-term impact upon the structure and function of the lakes phytoplankton.