Activity (P30): Analysis of activity over 4 repeated measurements showed an overall increase in A) locomotion [F(1,71) = 7.12, p = 9×10⁻³] and B) distance [F(1,71) = 7.36, p = 8.37×10^−<b>3</b>], but no change on C) speed [F(1,74) = 1.92, p = 1.69] in propofol treated animals.

<p>Propofol treatment did not alter D) anxiety related behavior in adolescent animals [F(1,74) = 0.02, p = 0.89]. An overall change in activity was observed over individual measurements, resulting in a significant decrease in locomotion [F(3,71) = 13.6, p = 4.08×10⁻⁷] and distance [F(3,71) = 5.35, p = 2.23×10⁻³] and a significant increase in speed [F(3,74) = 15.7, p = 5.53×10⁻⁸] and the index of anxiety [F(3,74) = 7.25, p = 3×10⁻⁴]. (n_controls = 12 animals, n_propofol = 8 animals).</p

Novel object recognition on P30 and P120: At the age of 30 days, both propofol treated animals (t(7) = 7.45, ***q = 4.3×10⁻⁴) and controls (t(10) = 6.30, ***q = 3.6×10⁻⁴) spent significantly more time with the novel object indicating their ability to discriminate the novel from the old object.

<p>Propofol (t(7) = −1.44, q = 0.192) as well as control animals (t(10) = −1.92, q = 0.168) failed to do so after a 24 hrs inter-trial interval. At P120 both groups spent a random amount of time with either of the objects after 6 hrs and also after a 24 hrs interval, indicating that they were unable to remember the old object. (n_controls = 12 animals, n_propofol = 8 animals).</p

Activity (P120): Analysis of activity over 4 repeated measurements showed no treatment effects on A) locomotion [F(1,73) = 0.94, p = 0.33], B) distance [F(1,74) = 1.86, p = 0.18], C) speed [F(1,74) = 0.44, p = 0.51] or D) anxiety related behavior [F(1,62) = 0.57, p = 0.45] in propofol treated animals.

<p>Apart from a transient effect on locomotion [F(3,71) = 5.92, p = 1.13×10⁻³], no significant changes over repeated measurements were observed in adult aged animals. (n_controls = 12 animals, n_propofol = 8 animals).</p

Impact of propofol on neurotrophins.

<p>Densitometric quantifications of mRNA levels of BDNF and NT-3 in cortex and thalamus of P6 rats, analysed by qRT-PCR. Values represent mean normalised ratios of the densities of BDNF and NT-3 bands compared to the density of the control group (n = 6–7/point+SE). There was an effect of propofol treatment with a decrease of BDNF levels over time, which was significant after 6 hrs in the cortex [F(1,30) = 66.5, p<0.001]. There was also a decrease in NT-3 levels, which was significant in the cortex after 6 hrs [F(1,28) = 12.7, p = 0.004] and after 12 hrs in the thalamus [F(1,24) = 3.5, p = 0.06].</p

Impact of propofol on survival promoting proteins.

<p>Densitometric quantifications of pAKT and pERK1/2 in the cortex and thalamus of P6 rats, analysed by Western blotting. Values represent mean normalised ratios of the densities of pAKT and pERK1/2 bands compared to the density of the control group (n = 6/point+SE). There was an effect of propofol treatment in decrease of pAKT levels over time in the thalamus, which was significant after 12 hrs [F(1,28) = 5.6, p = 0.06]. Post-hoc analysis showed most pronounced decrease after 12 hrs (2-sample t-test). In the cortex there was a significant decrease of pERK1/2 levels over the time, which was significant after 6, 12 and 24 hrs [F(1,29) = 12.7, p = 0.013].</p

Impact of propofol on key proteins involved in apoptotic signalling.

<p>Densitometric quantifications of caspase-3 and AIF in cortex and thalamus of P6 rats as analysed by Western blotting. Values represent mean normalised ratios of the densities of caspase-3 and AIF bands compared to densities of the control group (n = 5–6/point+SE). There was an effect of propofol treatment on caspase-3 levels over time, which was significant after 24 hrs in the cortex [F(1,29) = 3.63, p = 0.06] and after 12 hrs in the thalamus [F(1,28) = 3.1, p = 0.09).</p

Metabolic effects of fasting on human and mouse blood in vivo

Starvation is a strong physiological stimulus of macroautophagy/autophagy. In this study, we addressed the question as to whether it would be possible to measure autophagy in blood cells after nutrient deprivation. Fasting of mice for 48 h (which causes ∼20% weight loss) or starvation of human volunteers for up to 4 d (which causes + puncta per cell in response to nutrient deprivation, only neutrophils from starved volunteers showed signs of activated autophagy (as determined by a combination of multi-color immunofluorescence, cytofluorometry and image analysis). Altogether, these results suggest that white blood cells are suitable for monitoring autophagic flux. In addition, we propose that the evaluation of protein acetylation in circulating leukocytes can be adopted as a biochemical marker of organismal energetic status.</p

Combined evaluation of LC3B puncta and HMGB1 expression predicts residual risk of relapse after adjuvant chemotherapy in breast cancer

<p>In spite of adjuvant chemotherapy, a significant fraction of patients with localized breast cancer (BC) relapse after optimal treatment. We determined the occurrence of cytoplasmic MAP1LC3B/LC3B (microtubule-associated protein 1 light chain 3B)-positive puncta, as well as the presence of nuclear HMGB1 (high mobility group box 1) in cancer cells within surgical BC specimens by immunohistochemistry, first in a test cohort (152 patients) and then in a validation cohort of localized BC patients who all received adjuvant anthracycline-based chemotherapy (1646 patients). Cytoplasmic LC3B⁺ puncta inversely correlated with the intensity of SQSTM1 staining, suggesting that a high percentage cells of LC3B⁺ puncta reflects increased autophagic flux. After setting optimal thresholds in the test cohort, cytoplasmic LC3B⁺ puncta and nuclear HMGB1 were scored as positive in 27.2% and 28.6% of the tumors, respectively, in the validation cohort, while 8.7% were considered as double positive. LC3B⁺ puncta or HMGB1 expression alone did not constitute independent prognostic factors for metastasis-free survival (MFS) in multivariate analyses. However, the combined positivity for LC3B⁺ puncta and nuclear HMGB1 constituted an independent prognostic factor significantly associated with prolonged MFS (hazard ratio: 0.49 95% confidence interval [0.26–0.89]; <i>P</i> = 0.02), and improved breast cancer specific survival (hazard ratio: 0.21 95% confidence interval [0.05–0.85]; <i>P</i> = 0.029). Subgroup analyses revealed that within patients with poor-prognosis BC, HMGB1⁺ LC3B⁺ double-positive tumors had a better prognosis than BC that lacked one or both of these markers. Altogether, these results suggest that the combined positivity for LC3B⁺ puncta and nuclear HMGB1 is a positive predictor for longer BC survival.</p