Expression and regulation of type 2A protein phosphatases and alpha4 signalling in cardiac health and hypertrophy

Abstract Cardiac physiology and hypertrophy are regulated by the phosphorylation status of many proteins, which is partly controlled by a poorly defined type 2A protein phosphatase-alpha4 intracellular signalling axis. Quantitative PCR analysis revealed that mRNA levels of the type 2A catalytic subunits were differentially expressed in H9c2 cardiomyocytes (PP2ACb[PP2ACa[PP4C[PP6C), NRVM (PP2ACb[PP2ACa = PP4C = PP6C), and adult rat ventricular myocytes (PP2ACa[ PP2ACb[PP6C[PP4C). Western analysis confirmed that all type 2A catalytic subunits were expressed in H9c2 cardiomyocytes; however, PP4C protein was absent in adult myocytes and only detectable following 26S proteasome inhibition. Short-term knockdown of alpha4 protein expression attenuated expression of all type 2A catalytic subunits. Pressure overload-induced left ventricular (LV) hypertrophy was associated with an increase in both PP2AC and alpha4 protein expression. Although PP6C expression was unchanged, expression of PP6C regulatory subunits (1) Sit4-associated protein 1 (SAP1) and (2) ankyrin repeat domain (ANKRD) 28 and 44 proteins was elevated, whereas SAP2 expression was reduced in hypertrophied LV tissue. Co-immunoprecipitation studies demonstrated that the interaction between alpha4 and PP2AC or PP6C subunits was either unchanged or reduced in hypertrophied LV tissue, respectively. Phosphorylation status of phospholemman (Ser63 and Ser68) was significantly increased by knockdown of PP2ACa, PP2ACb, or PP4C protein expression. DNA damage assessed by histone H2A.X phosphorylation (cH2A.X) in hypertrophied tissue remained unchanged. However, exposure of cardiomyocytes to H2O2 increased levels of cH2A.X which was unaffected by knockdown of PP6C expression, but was abolished by the short-term knockdown of alpha4 expression. This study illustrates the significance and altered activity of the type 2A protein phosphatase-alpha4 complex in healthy and hypertrophied myocardium

Expression of type 2A protein phosphatases in cardiac health and disease

Cardiac physiology and hypertrophy is regulated by the phosphorylation status of many proteins, which is regulated (in part) by activity of the type 2A serine/threonine protein phosphatase family. Phosphatase activity of this family is conferred by the homologous PP2ACα/β, PP4C and PP6C catalytic subunits. Using quantitative PCR, gene expression of type 2A phosphatases showed that PP6C mRNA (5.07) expressed as relative units (RU) was higher than PP2ACα (3.52), PP2ACβ (3.24) and PP4C (4.54) in H9c2 cardiomyocytes. However, in adult rat ventricular myocytes (ARVM), PP4C mRNA (6.42) was higher than PP2ACα (2.52), PP2ACβ (3.93) and PP6C (4.96). Using Western immunoblotting, PP2ACα/β protein expression was similar in both H9c2 cardiomyocytes and ARVM. PP6C protein expression in ARVM was significantly higher (P<0.05) when compared to H9c2 cardiomyocytes, while PP4C protein expression in ARVM was undetectable. Additionally, we showed that 28 days of transverse aortic constriction in the mouse induced a ∼61% increase in the left ventricular (LV) mass index and a significant increase (p<0.01) in the expression of PP2ACα/β protein when compared to sham-operated LV myocardium. PP4C protein expression was not detectable in sham or hypertrophied LV myocardium, whereas PP6C protein expression was similar in both groups. This study illustrates the differences in the expression of the type 2A protein phosphatases in both H9c2 cardiomyocytes, ARVM and also highlights the importance of PP2A in cardiac pathological hypertrophy. Future work will aim to identify substrates of PP2ACα, PP2ACβ, PP4C and PP6C related to calcium regulation and hypertrophy in cardiomyocytes using small interfering RNA

Empirical seismic vulnerability assessment of Icelandic buildings affected by the 2000 sequence of earthquakes

Publisher's version (útgefin grein)In June 2000, two Mw6.5 earthquakes occurred within a 4-day interval in the largest agricultural region of Iceland causing substantial damage and no loss of life. The distance between the earthquake epicentres and the fault rupture was approximately 15 km. Nearly 5000 low-rise residential buildings were affected, some of which were located between the faults and exposed to strong ground motion from both events. The post-earthquakes damage and repair costs for every residential building in the epicentral region were assessed for insurance purposes. The database is detailed and complete for the whole region and represents one of the best quality post-earthquake vulnerability datasets used for seismic loss estimation. Nonetheless, the construction of vulnerability curves from this database is hampered by the fact that the loss values represent the cumulative damage from two sequential earthquakes in some areas, and single earthquakes in others. A novel methodology based on beta regression is proposed here in order to define the geographical limits on areas where buildings sustained cumulative damage and predict the seismic losses for future sequence of events in each area. The results show that the average building loss in areas affected by a single event is below 10% of the building replacement value, whilst this increases to an average of 25% in areas affected by the two earthquakes. The proposed methodology can be used to empirically assess the vulnerability in other areas which experienced sequence of events such as Emilia-Romagna (Italy) in 2012.The authors wish to offer their thanks to the Icelandic Catastrophe Insurance for placing the earthquake loss database and other relevant information at their disposal, and University of Iceland for a research Grant. Ioanna Ioannou and Tiziana Rossetto’s contribution to this study was Funded by the HORIZON2020 Project ‘IMPROVER’ (Grant Number: 653390). Ioannis Kosmidis was supported by The Alan Turing Institute under the EPSRC grant EP/N510129/1 (Turing award number TU/B/000082).Peer Reviewe

Developing a cosmetic series: Results from the ESSCA network, 2009-2018.

BACKGROUND There is considerable variability across European patch test centres as to which allergens are included in local and national cosmetics series. OBJECTIVES To propose a standardized, evidence-based cosmetic series for Europe based on up-to-date analysis of relevant contact allergens. METHODS We collated data from the European Surveillance System on Contact Allergies (ESSCA) from 2009 to 2018 to determine which cosmetic allergens produce a high yield of contact allergy. Contact allergens with a prevalence of >0.3% that were considered relevant were included. Rare contact allergens were excluded if deemed no longer relevant or added to a supplemental cosmetic series for further analysis. RESULTS Sensitization prevalences of 39 cosmetic contact allergens were tabulated. Thirty of these allergens yielded >0.3% positive reactions and are therefore included in our proposed European cosmetic series. Six were considered no longer relevant and therefore excluded. Three were included in a supplementary European cosmetic series. An additional nine allergens were included in either the core or supplemental European cosmetic series following literature review. CONCLUSION We have derived a potential European cosmetic series based upon the above methods. This will require ongoing investigation based upon the changing exposure profiles of cosmetic allergens as well as new and evolving substances