Evaluating the Effectiveness of Proton Beam Therapy Compared to Conventional Radiotherapy in Non-Metastatic Rectal Cancer: A Systematic Review of Clinical Outcomes.

Background and Objectives: Conventional radiotherapies used in the current management of rectal cancer commonly cause iatrogenic radiotoxicity. Proton beam therapy has emerged as an alternative to conventional radiotherapy with the aim of improving tumour control and reducing off-set radiation exposure to surrounding tissue. However, the real-world treatment and oncological outcomes associated with the use of proton beam therapy in rectal cancer remain poorly characterised. This systematic review seeks to evaluate the radiation dosages and safety of proton beam therapy compared to conventional radiotherapy in patients with non-metastatic rectal cancer. Materials and Methods: A computer-assisted search was performed on the Medline, Embase and Cochrane Central databases. Studies that evaluated the adverse effects and oncological outcomes of proton beam therapy and conventional radiotherapy in adult patients with non-metastatic rectal cancer were included. Results: Eight studies were included in this review. There was insufficient evidence to determine the adverse treatment outcomes of proton beam therapy versus conventional radiotherapy. No current studies assessed radiotoxicities nor oncological outcomes. Pooled dosimetric comparisons between proton beam therapy and various conventional radiotherapies were associated with reduced radiation exposure to the pelvis, bowel and bladder. Conclusions: This systematic review demonstrates a significant paucity of evidence in the current literature surrounding adverse effects and oncological outcomes related to proton beam therapy compared to conventional radiotherapy for non-metastatic rectal cancer. Pooled analyses of dosimetric studies highlight greater predicted radiation-sparing effects with proton beam therapy in this setting. This evidence, however, is based on evidence at a moderate risk of bias and clinical heterogeneity. Overall, more robust, prospective clinical trials are required.

The nutrient sensor CRTC and Sarcalumenin/thinman represent an alternate pathway in cardiac hypertrophy.

CREB-regulated transcription co-activator (CRTC) is activated by Calcineurin (CaN) to regulate gluconeogenic genes. CaN also has roles in cardiac hypertrophy. Here, we explore a cardiac-autonomous role for CRTC in cardiac hypertrophy. In Drosophila, CRTC mutants exhibit severe cardiac restriction, myofibrillar disorganization, fibrosis, and tachycardia. Cardiac-specific CRTC knockdown (KD) phenocopies mutants, and cardiac overexpression causes hypertrophy. CaN-induced hypertrophy in Drosophila is reduced in CRTC mutants, suggesting that CRTC mediates the effects. RNA sequencing (RNA-seq) of CRTC-KD and -overexpressing hearts reveals contraregulation of metabolic genes. Genes with conserved CREB sites include the fly ortholog of Sarcalumenin, a Ca2+-binding protein. Cardiac manipulation of this gene recapitulates the CRTC-KD and -overexpression phenotypes. CRTC KD in zebrafish also causes cardiac restriction, and CRTC KD in human induced cardiomyocytes causes a reduction in Srl expression and increased action potential duration. Our data from three model systems suggest that CaN-CRTC-Sarcalumenin signaling represents an alternate, conserved pathway underlying cardiac function and hypertrophy.

Mitochondrial MICOS complex genes, implicated in hypoplastic left heart syndrome, maintain cardiac contractility and actomyosin integrity.

Hypoplastic left heart syndrome (HLHS) is a severe congenital heart disease (CHD) with a likely oligogenic etiology, but our understanding of the genetic complexities and pathogenic mechanisms leading to HLHS is limited. We performed whole genome sequencing (WGS) on 183 HLHS patient-parent trios to identify candidate genes, which were functionally tested in the Drosophila heart model. Bioinformatic analysis of WGS data from an index family of a HLHS proband born to consanguineous parents prioritized 9 candidate genes with rare, predicted damaging homozygous variants. Of them, cardiac-specific knockdown (KD) of mitochondrial MICOS complex subunit dCHCHD3/6 resulted in drastically compromised heart contractility, diminished levels of sarcomeric actin and myosin, reduced cardiac ATP levels, and mitochondrial fission-fusion defects. These defects were similar to those inflicted by cardiac KD of ATP synthase subunits of the electron transport chain (ETC), consistent with the MICOS complex's role in maintaining cristae morphology and ETC assembly. Five additional HLHS probands harbored rare, predicted damaging variants in CHCHD3 or CHCHD6. Hypothesizing an oligogenic basis for HLHS, we tested 60 additional prioritized candidate genes from these patients for genetic interactions with CHCHD3/6 in sensitized fly hearts. Moderate KD of CHCHD3/6 in combination with Cdk12 (activator of RNA polymerase II), RNF149 (goliath, E3 ubiquitin ligase), or SPTBN1 (ß-Spectrin, scaffolding protein) caused synergistic heart defects, suggesting the likely involvement of diverse pathways in HLHS. Further elucidation of novel candidate genes and genetic interactions of potentially disease-contributing pathways is expected to lead to a better understanding of HLHS and other CHDs.

Multiplatform modeling of atrial fibrillation identifies phospholamban as a central regulator of cardiac rhythm.

Atrial fibrillation (AF) is a common and genetically inheritable form of cardiac arrhythmia; however, it is currently not known how these genetic predispositions contribute to the initiation and/or maintenance of AF-associated phenotypes. One major barrier to progress is the lack of experimental systems to investigate the effects of gene function on rhythm parameters in models with human atrial and whole-organ relevance. Here, we assembled a multi-model platform enabling high-throughput characterization of the effects of gene function on action potential duration and rhythm parameters using human induced pluripotent stem cell-derived atrial-like cardiomyocytes and a Drosophila heart model, and validation of the findings using computational models of human adult atrial myocytes and tissue. As proof of concept, we screened 20 AF-associated genes and identified phospholamban loss of function as a top conserved hit that shortens action potential duration and increases the incidence of arrhythmia phenotypes upon stress. Mechanistically, our study reveals that phospholamban regulates rhythm homeostasis by functionally interacting with L-type Ca2+ channels and NCX. In summary, our study illustrates how a multi-model system approach paves the way for the discovery and molecular delineation of gene regulatory networks controlling atrial rhythm with application to AF.

Idiopathic ventricular fibrillation associated with long-coupled Purkinje ectopy.

INTRODUCTION: Idiopathic ventricular fibrillation (IVF) is mainly associated with and triggered by short-coupled (R-on-T) ventricular ectopics. However, little is known about the risk of VF associated with long-coupled premature ventricular complexes (LCPVCs). OBJECTIVE: To examine the prevalence and characteristics of IVF patients presenting with LCPVCs. METHODS: Consecutive patients with IVF and PVCs from five arrhythmia referral centers were reviewed. We included patients presenting LCPVCs, defined as PVCs falling after the end of the T wave, with a normal QTc interval. We evaluated demographics, medical history, and clinical circumstances associated with PVCs and VF episodes. The origin of PVCs was determined by invasive mapping. RESULTS: Seventy-nine patients with IVF were reviewed. Among them, 12 (15.2%) met the inclusion criteria (8 women, age 36 ± 14 years). Eleven patients had documented LCPVCs initiating repetitive PVCs or sustained VF, whereas 1 had only documented isolated PVCs. In 10 of 12 patients, PVCs were recorded showing both long and short coupling intervals of 418 ± 46 and 304 ± 33 ms, respectively. Mapping showed that PVCs originated from the left Purkinje in 10 patients, from the right Purkinje in 1 patient, and both in 1 patient. Compared to other patients from the initial cohort, IVF with LCPVCs was associated with a left-sided origin of PVCs (92% in long-coupled IVF vs. 46% of left Purkinje PVCs in short-coupled IVF, p = .004). CONCLUSION: Long-coupled fascicular PVCs, traditionally recognized as benign, can be associated with IVF in a subset of patients. They can induce IVF by themselves or in association with short-coupled PVCs.

The effective use of blebbistatin to study the action potential of cardiac pacemaker cells of zebrafish (Danio rerio) during incremental warming.

Blebbistatin potently inhibits actin-myosin interaction, preventing contractile activity of excitable cells including cardiac myocytes, despite electrical excitation of an action potential (AP). We collected intracellular microelectrode recordings of pacemaker cells located in the sinoatrial region (SAR) of the zebrafish heart at room temperature and during acute warming to investigate whether or not blebbistatin inhibition of contraction significantly alters pacemaker cell electrophysiology. Changes were evaluated based on 16 variables that characterized the AP waveform. None of these AP variables nor the spontaneous heart rate were significantly modified with the application of 10 µM blebbistatin when recordings were made at room temperature. Compared with the control group, the blebbistatin-treated group showed minor changes in the rate of spontaneous diastolic depolarization (P = 0.027) and the 50% and 80% repolarization (P = 0.008 and 0.010, respectively) in the 26°C-29°C temperature bin, but not at higher temperatures. These findings suggest that blebbistatin is an effective excitation-contraction uncoupler that does not appreciably affect APs generated in pacemaking cells of the SAR and can, therefore, be used in zebrafish cardiac studies.

Membrane and calcium clock mechanisms contribute variably as a function of temperature to setting cardiac pacemaker rate in zebrafish Danio rerio.

Here, we show that heart rate in zebrafish Danio rerio is dependent upon two pacemaking mechanisms and it possesses a limited ability to reset the cardiac pacemaker with temperature acclimation. Electrocardiogram recordings, taken from individual, anaesthetised zebrafish that had been acclimated to 18, 23 or 28°C were used to follow the response of maximum heart rate (fHmax ) to acute warming from 18°C until signs of cardiac failure appeared (up to c. 40°C). Because fHmax was similar across the acclimation groups at almost all equivalent test temperatures, warm acclimation was limited to one significant effect, the 23°C acclimated zebrafish had a significantly higher (21%) peak fHmax and reached a higher (3°C) test temperature than the 18°C acclimated zebrafish. Using zatebradine to block the membrane hyperpolarisation-activated cyclic nucleotide-gated channels (HCN) and examine the contribution of the membrane clock mechanisms to cardiac pacemaking, f Hmax was significantly reduced (by at least 40%) at all acute test temperatures and significantly more so at most test temperatures for zebrafish acclimated to 28°C vs. 23°C. Thus, HCN channels and the membrane clock were not only important, but could be modified by thermal acclimation. Using a combination of ryanodine (to block sarcoplasmic calcium release) and thapsigargin (to block sarcoplasmic calcium reuptake) to examine the contribution of sarcoplasmic reticular handling of calcium and the calcium clock, f Hmax was again consistently reduced independent of the test temperature and acclimation temperature, but to a significantly lesser degree than zatebradine for zebrafish acclimated to both 28 and 18°C. Thus, the calcium clock mechanism plays an additional role in setting pacemaker activity that was independent of temperature. In conclusion, the zebrafish cardiac pacemaker has a limited temperature acclimation ability compared with known effects for other fishes and involves two pacemaking mechanisms, one of which was independent of temperature.

Calcium response of KCl-excited populations of ventricular myocytes from the European sea bass (Dicentrarchus labrax): a promising approach to integrate cell-to-cell heterogeneity in studying the cellular basis of fish cardiac performance.

Climate change challenges the capacity of fishes to thrive in their habitat. However, through phenotypic diversity, they demonstrate remarkable resilience to deteriorating conditions. In fish populations, inter-individual variation in a number of fitness-determining physiological traits, including cardiac performance, is classically observed. Information about the cellular bases of inter-individual variability in cardiac performance is scarce including the possible contribution of excitation-contraction (EC) coupling. This study aimed at providing insight into EC coupling-related Ca(2+) response and thermal plasticity in the European sea bass (Dicentrarchus labrax). A cell population approach was used to lay the methodological basis for identifying the cellular determinants of cardiac performance. Fish were acclimated at 12 and 22 °C and changes in intracellular calcium concentration ([Ca(2+)]i) following KCl stimulation were measured using Fura-2, at 12 or 22 °C-test. The increase in [Ca(2+)]i resulted primarily from extracellular Ca(2+) entry but sarcoplasmic reticulum stores were also shown to be involved. As previously reported in sea bass, a modest effect of adrenaline was observed. Moreover, although the response appeared relatively insensitive to an acute temperature change, a difference in Ca(2+) response was observed between 12- and 22 °C-acclimated fish. In particular, a greater increase in [Ca(2+)]i at a high level of adrenaline was observed in 22 °C-acclimated fish that may be related to an improved efficiency of adrenaline under these conditions. In conclusion, this method allows a rapid screening of cellular characteristics. It represents a promising tool to identify the cellular determinants of inter-individual variability in fishes' capacity for environmental adaptation.