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Preclinical data have confirmed that human pluripotent stem cell-derived cardiomyocytes (PSC-CMs) can remuscularize the injured or diseased heart, with several clinical trials now in planning or recruitment stages. However, because ventricular arrhythmias represent a complication following engraftment of intramyocardially injected PSC-CMs, it is necessary to provide treatment strategies to control or prevent engraftment arrhythmias (EAs). Here, we show in a porcine model of myocardial infarction and PSC-CM transplantation that EAs are mechanistically linked to cellular heterogeneity in the input PSC-CM and resultant graft. Specifically, we identify atrial and pacemaker-like cardiomyocytes as culprit arrhythmogenic subpopulations. Two unique surface marker signatures, signal regulatory protein α (SIRPA) + CD90 - CD200 + and SIRPA + CD90 - CD200 - , identify arrhythmogenic and non-arrhythmogenic cardiomyocytes, respectively. Our data suggest that modifications to current PSC-CM-production and/or PSC-CM-selection protocols could potentially prevent EAs. We further show that pharmacologic and interventional anti-arrhythmic strategies can control and potentially abolish these arrhythmias., (© 2024. The Author(s).)

Aim: The aim of this work was to evaluate physicians' perceptions of ostomates' quality of life (QoL) and comfort of care among an international sample of physicians caring for ostomates., Method: This was a cross-sectional survey study. We conducted a survey of primary care physicians (PCP), gastroenterologists (GI), and general surgeons (GS) from three continents using the SERMO online physician platform. We piloted the survey for content, clarity and domain development using a pilot sample of physicians from each speciality before use. We summarized responses to questions related to physician comfort of ostomate care with descriptive statistics. We conducted multiple logistic regression with the primary outcome of physician perception of ostomate QoL., Results: A total of 617 physicians (PCP 264, GI 176, GS 177) completed the survey representing North America, Europe and Australia similarly. The average age was 46 years and 21% were women. Ninety per cent of physicians care for an ostomate at least once per month. Eighty eight per cent had access to enterostomal nurses. Eighty two per cent of physicians believed that ostomates have decreased QoL. Forty seven per cent believed that ostomates have decreased overall health. Almost half of respondents answered incorrectly to a 'bogus question' citing fake clinical evidence supporting a negative impact of ostomies on social relationships. Increased physician comfort in ostomy care (OR 1.30, p = 0.04) and US-based physicians (OR 1.75, p = 0.01) were associated with increased odds of answering that ostomates have no decreased QoL., Conclusion: Among a diverse international sample, most physicians believe that ostomates have decreased QoL but not overall health. Physician implicit bias, physician comfort and geographical variability account for these findings. Targeted efforts to increase physician comfort in ostomate care and establish universal best practices is needed., (© 2022 Association of Coloproctology of Great Britain and Ireland.)

Chinese hamster ovary (CHO) cells are the primary platform for the production of biopharmaceuticals. To increase yields, many CHO cell lines have been genetically engineered to resist cell death. However, the kinetics that governs cell fate in bioreactors are confounded by many variables associated with batch processes. Here, we used CRISPR-Cas9 to create combinatorial knockouts of the three known BCL-2 family effector proteins: Bak1, Bax, and Bok. To assess the response to apoptotic stimuli, cell lines were cultured in the presence of four cytotoxic compounds with different mechanisms of action. A population-based model was developed to describe the behavior of the resulting viable cell dynamics as a function of genotype and treatment. Our results validated the synergistic antiapoptotic nature of Bak1 and Bax, while the deletion of Bok had no significant impact. Importantly, the uniform application of apoptotic stresses permitted direct observation and quantification of a delay in the onset of cell death through Bayesian inference of meaningful model parameters. In addition to the classical death rate, a delay function was found to be essential in the accurate modeling of the cell death response. These findings represent an important bridge between cell line engineering strategies and biological modeling in a bioprocess context., (© 2022 The Authors. Biotechnology and Bioengineering published by Wiley Periodicals LLC.)

The reliable and cost-efficient manufacturing of monoclonal antibodies (mAbs) is essential to fulfil their ever-growing demand. Cell death in bioreactors reduces productivity and product quality, and is largely attributed to apoptosis. In perfusion bioreactors, this leads to the necessity of a bleed stream, which negatively affects the overall process economy. To combat this limitation, death-resistant Chinese hamster ovary cell lines were developed by simultaneously knocking out the apoptosis effector proteins Bak1, Bax, and Bok with CRISPR technology. These cell lines were cultured in fed-batch and perfusion bioreactors and compared to an unmodified control cell line. In fed-batch, the death-resistant cell lines showed higher cell densities and longer culture durations, lasting nearly a month under standard culture conditions. In perfusion, the death-resistant cell lines showed slower drops in viability and displayed an arrest in cell division after which cell size increased instead. Pertinently, the death-resistant cell lines demonstrated the ability to be cultured for several weeks without bleed, and achieved similar volumetric productivities at lower cell densities than that of the control cell line. Perfusion culture reduced fragmentation of the mAb produced, and the death-resistant cell lines showed increased glycosylation in the light chain in both bioreactor modes. These data demonstrate that rationally engineered death-resistant cell lines are ideal for mAb production in perfusion culture, negating the need to bleed the bioreactor whilst maintaining product quantity and quality.