Intrauterine fetal death occurs in approximately 1 in every 160 pregnancies; postmortem evaluation often fails to find an underlying cause.  The objective of this paper was to determine the spectrum and prevalence of mutations in the three most common Long QT syndrome (LQTS) susceptible genes in a cohort of cases of unexplained intrauterine death.

Retrospective post-mortem genetic testing was carried out on 91 unexplained intrauterine fetal deaths collected between 2006 and 2012.  Publically available exome databases were assessed for the general population frequency of identified genetic variants.  Mutations analysis was performed by liquid chromatography and DNA sequencing.  Functional analysis of novel mutations was performed using heterologous expression and patch-clamp recording.

The three LQTS missense mutations (KCNQ1,p.A283T; KCNQ1, p.R397W; and KCNH2[1b], p.R25W) were discovered in three intrauterine fetal deaths.  Both KV7.1-A283T (16-week male) and KV7.1- R397W (16-week female) mutations were associated with marked KV7.1 loss-of- function consistent with in utero LQTS type 1, while the HERG1b-R25W mutation (33.2-week male) exhibited a loss of function consistent with in utero LQTS type 2. Furthermore, 5 intrauterine fetal deaths hosted SCN5A rare nonsynonymous genetic variants that conferred in vitro electrophysiological characteristics consistent with potentially pro-arrhythmic phenotypes.

Conclusions:

This analysis found that 3 of 91 unexplained intrauterine deaths were associated with missense LQTS mutations, and genetic variants leading to dysfunctional LQTS-associated ion channels were found in 8 cases.  Although preliminary, these results may provide insights into possible mechanisms of stillbirth.

• Crotti L, Tester DJ, White WM et al.  Long QT Syndrome–Associated Mutations in Intrauterine Fetal Death.  JAMA. 2013;309(14):1473-1482

The development of atherosclerotic diseases involves the proliferation and migration of a variety of cell types.  It has been generally accepted that vascular smooth muscle cells in the arterial wall play a crucial role in this process through a process of ‘de-differentiation’ from a normal contractile phenotype to a proliferative and synthetic state that drives remodelling and disease development.  In this study, Tang et al re-examine this paradigm and provide compelling evidence to challenge the status quo.  Using sophisticated in-vitro lineage tracing techniques, they initially identified a novel type of stem cell which they dub a multipotent vascular stem cell (MVSC).  Under normal physiological conditions these MVSCs replenish normal smooth muscle cells to maintain vascular integrity and function, but in response to vascular injury, the MVSCs undergo proliferative change and produce phenotypically abnormal smooth muscle cells.  Therefore, rather than a process of ‘de-differentiation’  of mature cells, atherosclerosis  is elegantly demonstrated to be a disease of stem cells, switching attention to a new target in understanding the basic pathophysiology behind myocardial infarction and stroke.

Conclusion

Multipotent vascular stem cells affected by vascular injury undergo differentiation into abnormal proliferative smooth muscle cells characteristic of atherosclerosis.  This new evidence re-targets atherosclerosis as a stem-cell disease, not only providing fundamental new insights into vascular biology but potentially providing new therapeutic targets in cardiovascular disease

•  Tang Z, Wang A, Yuan F, Yan Z, Liu B, Chu JS, Helms JA, Li S.  Differentiation of multipotent vascular stem cells contributes to vascular diseases.  Nat Commun. 2012 Jun 6;3:875.

Serum PCSK9 plays an active role in controlling the expression of LDL receptors by targeting them for lysosomal destruction.  REGN727/SAR236553 (REGN727) is a novel human monoclonal antibody which inhibits PCSK9 binding to the LDL receptor; a previous phase 1 proof of concept trial suggested the potential for significant reductions in LDL in familial hypercholesterolaemia.

The goal of this phase 2 study was to assess the efficacy and safety of REGN727 in a larger, more diverse, and more severely affected group of patients with familial hypercholesterolaemia.  The trial was carried out at 16 lipid clinics in the USA and Canada, and recruited 77 patients with heterozygous familial hypercholesterolaemia and LDL concentrations of 2·6 mmol/L or higher on a stable diet and statin dose, with or without ezetimibe.  Patients were randomly assigned to receive REGN727 150 mg, 200 mg, or 300 mg every 4 weeks, or 150 mg every 2 weeks, or placebo every 2 weeks.  The primary endpoint was mean percent reduction in LDL-C from baseline at week 12.

Least squares mean LDL-C reduction for patients on placebo was 10.65% compared to 42.53% for 300mg every 4 weeks (p<0.0001), and all treatment groups showed significant reductions in LDL, regardless of the use of ezetimibe.  No significant CK or liver enzyme rises were noted, although one patient stopped treatment due to an injection site reaction.

Conclusions:

The monoclonal antibody REGN727 achieved significant further LDL-C reduction in patients with familial heterozygous familial hypercholesterolaemia and elevated LDL-C treated with high-dose statins, with or without ezetimibe.

• Stein EA, Gipie D, Bergeron J et al.  Effect of a monoclonal antibody to PCSK9, REGN727/SAR236553, to reduce low-density lipoprotein cholesterol in patients with heterozygous familial hypercholesterolaemia on stable statin dose with or without ezetimibe therapy: a phase 2 randomised controlled trial.  Lancet 2012; 380, 29-36.

Ischaemia reperfusion injury (IR) is characterised by  abnormal increases in intracellular calcium during myocardial reperfusion that lead to cardiomyocyte death.  In addition, IR is accompanied by an increase in the expression of microRNAs (miRNAs) such as miR-214, which has previously been identified as a sensitive marker of cardiac stress.

In this study, Aurora et al. found that genetic deletion of miR-214 in mice was associated with an increase in cardiomyocyte apoptosis, loss of cardiac contractility, and excessive fibrosis following IR injury.  Subsequent work demonstrated that the cardioprotective role of miR-214 was due to its repression the mRNA encoding a key regulator of calcium influx, the Ncx1 sodium/calcium exchanger, and also to repression of several downstreatm effectors of calcium signalling that mediate cell death.

Conclusions:

This paper reveals a key role for the microRNA miR-214 in the response to ischaemia-reperfusion injury through its influence on calcium homeostasis.  Therefore, increasing levels of miR-214 to attentuate calcium overload – and therefore cell death – could be a new therapeutic strategy for limiting ischaemia-reperfusion injury.

• Aurora AB, Mahmoud AI, Luo X et al.  MicroRNA-214 protects the mouse heart from ischemic injury by controlling Ca²⁺ overload and cell death.  J Clin Invest 2012;122:1222-1232.

Despite the impressive gains in cardiovascular morbidity and mortality that have occurred over the past decades, a veritable elephant still exists in the room in the shape of the tidal wave of obesity sweeping the globe, which may reverse these advances.  Obesity has proved fundamentally difficult to manage, with few therapies demonstrating sustained weight loss over time be they pharmacological or behavioural.  Paradoxically, that other great modifiable risk factor for cardiovascular disease, smoking, is highly effective at controlling weight, with smokers having notably lower BMIs than non-smokers, some smokers smoking as a method of weight control and weight gain being common when quitting.  These anorexic effects have been attributed to nicotine as it effectively decreases feeding in animal models, but while nicotine has some effect on peripheral energy metabolism, little is known about its effect on central pathways determining food intake and body mass.

To investigate this the authors of this study performed a series of elegant experiments using a combination of pharmacological, molecular genetic, electrophysiological, and feeding studies in mice, to first demonstrate that specific neurons in the arcuate nucleus are acted on by nicotine and that these in turn influence pro-opiomelanocortin cells.  These constituents of the hypothalamic melanocortin system are an essential mechanism involved in the regulation of energy balance and food intake and as is demonstrated here, are influenced by nicotine to increase their firing rate and thus reduce appetite and weight gain.  Notably, naturally occurring strains of obese mice demonstrate defects in these same systems and so provide further evidence to support this mechanism as being significant in promoting weight gain.

Conclusions

These important data convincingly demonstrate the neurobiological mechanisms behind nicotine’s anorexic effects.  Further work may demonstrate new drug targets both to control smoking behaviours and to combat obesity.

• Mineur YS, Abizaid A, Rao Y, Salas R, DiLeone RJ, Gündisch D, Diano S, De Biasi M, Horvath TL, Gao XB, Picciotto MR. Nicotine decreases food intake through activation of POMC neurons. Science. 2011 Jun 10;332(6035):1330-2.

The quest to regenerate functional myocardium from stem cells following myocardial infarction has garnered a great deal of media attention and has become something of a holy grail in cardiovascular medicine.  Despite the basic science of stem cells still being at an early stage, the first human trials have already been conducted – but the results thus far have been neutral at best with little or no evidence of clinical benefit.  A significant bottleneck is the identification of a viable source of stem/progenitor cells that could contribute new muscle with the therapeutic ideal being to stimulate a resident source, thus avoiding the caveats of limited graft survival, restricted homing to the site of injury and host immune rejection.

In contrast to the bone marrow derived cells that have been generally used thus far the authors of this study, done in various experimental mouse models, demonstrate that the adult heart contains a resident stem or progenitor cell population, which has the potential to contribute bona fide terminally differentiated cardiomyocytes after myocardial infarction.  By priming these cells with a peptide called thymosin β4 they were able to induce embryonic reprogramming resulting in the mobilization of this population and subsequent differentiation to give rise to de novo cardiomyocytes.  Following experimentally induced myocardial infarction, these cells were shown to migrate to the site of injury and then differentiate without any evidence of cellular fusion into structurally and functionally active cardiomyocytes that showed evidence of gap junction formation with adjacent cells, synchronous calcium transients and the formation of operational contractile apparatus.  Despite a low overall fraction of these cells being present at the site of injury and a relatively poor overall efficiency of differentiation, serial MRI scans revealed significant improvements in ejection fraction, cardiac volumes and scar size in comparison to sham treated animals.  Unfortunately, only pre-treatment with thymosin β4 demonstrated these effects and further work will be needed translate these findings into potential therapies.

Conclusions

The peptide thymosin β4 is able to induce a resident population of cardiac progenitor cells to trans-differentiate into functioning cardiomyocytes following myocardial infarction leading to improvements in cardiac function.  Future work may uncover viable cardiac regenerative therapies for use in clinical practice.

• Smart N, Bollini S, Dubé KN, Vieira JM, Zhou B, Davidson S, Yellon D, Riegler J, Price AN, Lythgoe MF, Pu WT, Riley PR.  De novo cardiomyocytes from within the activated adult heart after injury.  Nature. 2011 Jun 8. doi: 10.1038/nature10188. [Epub ahead of print]

Although the relationship between cholesterol, triglycerides, and cardiovascular disease is well established, little is known about the role of the third major class of lipids – phospholipids.  In this regard, the intestinal microflora play a crucial role in that they digest and absorb many crucial nutrients, including lipids.  Therefore in this study the authors attempted to discover whether a link exists between gut-flora-dependent phospholipid metabolism and atherosclerosis risk through generation of pro-atherosclerotic metabolites.

Three metabolites of the dietary lipid phosphatidylcholine — choline, trimethylamine N-oxide (TMAO) and betaine — were firstly identified from a clinical cohort prior to the onset of a clinical cardiovascular event. Subsequently, a mouse model was used where dietary supplementation with choline, TMAO or betaine was found to upregulate multiple macrophage scavenger receptors linked to atherosclerosis, and supplementation with choline or TMAO promoted atherosclerosis. Lastly, studies using germ-free mice showed that suppression of intestinal microflora in atherosclerosis-prone mice inhibited dietary-choline-enhanced atherosclerosis.

Conclusion:

Gut flora play an important role in the production of metabolites that induce atherosclerosis.  The results suggest that an appropriately targeted probiotic intervention may be of use in preventing cardiovascular disease.

• Wang Z, Klipfell E, Bennett BJ et al.  Gut flora metabolism of phosphatidylcholine promotes cardiovascular disease.  Nature 2011;472:57-65.

Metabolomics is the study of small-molecule metabolite profiles formed by cellular processes.

The RESOLUTE All Comers trial showed that the Resolute zotarolimus-eluting stent was non-inferior to the Xience V everolimus eluting stent in terms of target lesion failure (revascularisation, myocardial infarction, or cardiac death).  However, long-term follow-up data from drug-eluting stent trials are generally scarce.  Silber et al. report the 2-year clinical outcomes from the original RESOLUTE trial.

All patients in the study had at least one coronary lesion between 2.25 and 4.0mm in diameter causing more than 50% stenosis.  There were no restrictions as to the number of stents implanted, nor the the number of vessels or lesions treated.  Safety and efficacy outcomes examined at two years included death, myocardial infarction, revascularisation and stent-related composite outcomes.

1140 patients were assigned to Resolute and 1152 to Xience stents, of which 1121 and 1128, respectively, completed 2-year follow-up.  No difference was noted between patient-related outcomes (death, MI, revascularisation, 20.6% vs 20.5%; p=0.958) and stent-related outcomes (11.2% vs 10.7%; p=0.736) in the two patient groups.  In additon, only three patients in each group had stent thrombosis after one year (very late stent thrombosis).  Therefore despite a low rate of problems with the stents themselves, many patients went on to have further cardiac events.

Conclusions:

At 2-year follow-up, no significant safety or efficacy differences existed between Resolute and Xience stents.  However, a high rate of recurrent events emphasises the need for effective secondary prevention in patients with established coronary disease.

• Silber S, Windecker SS, Vranckx P et al.  Unrestricted randomised use of two new generation drug-eluting coronary stents: 2-year patient-related versus stent-related outcomes from the RESOLUTE All Comers trial. Lancet 2011. DOI: 10.1016/S0140-6736(11)60395-4

Patient-related vs. stent-related outcomes in the RESOLUTE trial

Using invasive imaging techniques such as intra-vascular ultrasound, the burden of atherosclerotic plaque disease in any given individual has previously been shown to correlate with the likelihood of future cardivascular events .  Magnetic Resonance Imaging (MRI) has the advantage of being non-invasive and, compared to other currently available molecular imaging modalities, it has good spatial resolution, making it ideal for imaging the thin arterial wall.  However, compared to positron emission tomography and optical imaging, it has a lower sensitivity.

To try and overcome this limitation, the authors of this study designed a novel elastin-specific magnetic resonance contrast agent (ESMA), to help to define atherosclerotic plaque burden in an Apoe-/- mouse model.  Elastin shows increased expression during plaque development and also has biological signalling functions.  Furthermore, using magnetic resonance signal values the amount of elastin can be quantified.

After administration of the ESMA, the authors noted a gradual increase in MRI signal from plaque, which was diminished in those mice treated with pravastatin.  This signal increased with time, and the authors correlated their MRI signal findings with immunoblotting and histology to confirm the amount of elastin present.
Conclusions:

Elastin represents a feasible target for molecular imaging of atherosclerosis using MRI.  The ability to quantify the amount of elastin present means that this technique has the potential both to assess atherosclerotic plaque burden and how it responds to treatment.

• Makowksi MR, Wiethoff AJ, Blume U et al.  Assessment of atherosclerotic plaque burden with an elastin-specific magnetic resonance contrast agent.  Nature Med 2011;17(3):383-389.