Subcellular Distribution of Two Spin Trapping Agents in Rat Heart: Possible Explanation for Their Different Protective Effects Against Doxorubicin-Induced Cardiotoxicity

Previous investigations, performed on isolated rat atria, showed that the lipophylic spin-trapping agent N-tert-butyl-alpha-phenylnitrone (PBN) is able to prevent the acute cardiotoxic effects produced by doxorubicin (DXR), whereas the hydrophylic compound 5,5-dimethyl-pyrroline-N-oxide (DMPO) is inactive. The present study was designed to ascertain whether differences in the pharmacological effects of the two spin traps are related to their different subcellular distribution. Langendorff rat hearts were perfused for 60 minutes with [^I4C]-DXR and either PBN or DMPO. The subcellular mapping of the three compounds was performed by measuring DXR by liquid scintillation counting, PBN by GC/MS, and DMPO by HPLC in the following isolated fractions: nuclei, mitochondria, sarcoplasmic reticulum, sarcolemma, cytosol. DMPO was shown to accumulate in the cytosolic compartment; both PBM and DXR are taken up by nuclei and mitochondria, while only trace amounts of DXR were detected in the sarcoplasmic reticulum. These results suggest that mitochondrial (and not sarcoplasmic) enzymes are mainly involved in DXR-induced free radical production, which is thought to cause the acute cardiotoxic effects of DXR. An involvement of DXR-induced free radical generation in the nuclear compartment seems unlikely in the short-term “in vitro” effects observed with the experimental model adopted for these studies, although it may play a role in the delayed pathology.     

The survey on cellular and tissue-engineered therapies in Europe and neighboring Eurasian countries in 2014 and 2015

Background aims

With the support of five established scientific organizations, this report, the seventh of its kind, describes activity in Europe for the years 2014 and 2015 in the area of cellular and tissue-engineered therapies, excluding hematopoietic stem cell (HSC) treatments for the reconstitution of hematopoiesis.

Anaerobic contribution to the time to exhaustion at the minimal exercise intensity at which maximal oxygen uptake occurs in elite cyclists, kayakists and swimmers

Using 23 elite male athletes (8 cyclists, 7 kayakists, and 8 swimmers), the contribution of the anaerobic energy system to the time to exhaustion (t _lim) at the minimal exercise intensity (speed or power) at which maximal oxygen uptake (V˙O_{2 max}) occurs (I _{V˙O2 max}) was assessed by analysing the relationship between the t _lim and the accumulated oxygen deficit (AOD). After 10-min warming up at 60% of V˙O_{2 max}, the exercise intensity was increased so that each subject reached his I _V˙O2max in 30 s and then continued at that level until he was exhausted. Pre-tests included a continuous incremental test with 2 min steps for determining the I _V˙O2max and a series of 5-min submaximal intensities to collect the data that would allow the estimation of the energy expenditure at I _V˙O2max . The AOD for the t _lim exercise was calculated as the difference between the above estimation and the accumulated oxygen uptake. The mean percentage value of energy expenditure covered by anaerobic metabolism was 15.2 [(SD 6)%, range 8.9–24.1] with significant differences between swimmers and kayakists (16.8% vs 11.5%, P≤0.05) and cyclists and kayakists (16.4% vs 11.5%, P≤0.05). Absolute AOD values ranged from 26.4 ml · kg⁻¹ to 83.6 ml · kg⁻¹ with a mean value of 45.9 (SD 18) ml · kg⁻¹. Considering all the subjects, the t _lim was found to have a positive and significant correlation with AOD (r = 0.62, P≤0.05), and a negative and significant correlation with V˙O_{2 max} (r = −0.46, P≤0.05). The data would suggest that the contribution of anaerobic processes during exercise performed at I _V˙O2max should not be ignored when t _lim is used as a supplementary parameter to evaluate specific adaptation of athletes. Accepted: 17 December 1996     

Subcellular localization of the spindle proteins Aurora A, Mad2, and BUBR1 assessed by immunohistochemistry.

The spindle checkpoint, the primary mechanism to ensure that two daughter cells receive the same amount of DNA, is compromised in many malignant tumors and has been implicated as a contributor to aneuploidy and carcinogenesis. The extent of expression and subcellular localization of the spindle proteins Aurora A, Mad2, and BUBR1 varies considerably in different immunohistochemical (IHC) reports from archival tumor tissues. Given the conflicting reports in the literature about the localization of these proteins, we examined the subcellular localization of Aurora kinase A, Mad2, and BUBR1 in normal and cancerous human tissues by IHC. In normal tissues, Aurora A was mainly localized to the nucleus when monoclonal or purified polyclonal antibodies were used, and Mad2 was localized to the nucleus, whereas BUBR1 was localized to the cytoplasm. In malignant tissues, Aurora A showed additional staining in the cytoplasm in the majority of tumors analyzed. Furthermore, BUBR1 was also localized to both the nucleus and cytoplasm in a significant fraction of tumors. Subcellular localization of Mad2 was similar in normal and malignant tissues. Thus, the validity of some earlier IHC studies of Aurora A, Mad2, and BUBR1 should be reconsidered, indicating that high-quality antibodies and a high-alkaline antigen-retrieval technique are required to achieve optimal results. We conclude that the subcellular localizations of these spindle proteins are different, although they have overlapping biological functions, and that Aurora A and BUBR1 undergo a shift in the subcellular localization during malignant transformation.