No Direct Postconditioning Effect of Poloxamer 188 on Mitochondrial Function after Ischemia Reperfusion Injury in Rat Isolated Hearts.

Myocardial infarction is a leading cause for morbidity and mortality worldwide. The only viable treatment for the ischemic insult is timely reperfusion, which further exacerbates myocardial injury. Maintaining mitochondrial function is crucial in preserving cardiomyocyte function in ischemia reperfusion (IR) injury. Poloxamer (P) 188 has been shown to improve cardiac IR injury by improving cellular and mitochondrial function. The aim of this study was to show if P188 postconditioning has direct protective effects on mitochondrial function in the heart. Langendorff prepared rat hearts were subjected to IR injury ex-vivo and reperfused for 10 min with 1 mM P188 vs. vehicle. Cardiac mitochondria were isolated with 1 mM P188 vs. 1 mM polyethylene glycol (PEG) vs. vehicle by differential centrifugation. Mitochondrial function was assessed by adenosine triphosphate synthesis, oxygen consumption, and calcium retention capacity. Mitochondrial function decreased significantly after ischemia and showed mild improvement with reperfusion. P188 did not improve mitochondrial function in the ex-vivo heart, and neither further P188 nor PEG induced direct mitochondrial protection after IR injury in this model.

Video laryngoscopic oral intubation in rats: a simple and effective method.

Endotracheal intubation is a vital component of many rat in vivo experiments to secure the airway and allow controlled ventilation. Even in the hands of experienced researchers, however, the procedure remains technically challenging. The safest and most reliable way for human intubation is by video laryngoscopy. Previous attempts to apply this technique in rodents have been complicated and expensive. We, hereby, describe a novel, noninvasive method to safely intubate rats orally by video laryngoscopy, thus avoiding the need for a surgical tracheostomy. By repurposing a commercially available ear wax removal device, visualization of the rat larynx can be significantly enhanced. Because of its small diameter, integrated illumination, and a powerful camera with adequate focal length, the device has all of the necessary properties for exploring the upper airway of a rat. After identifying the vocal cords by video laryngoscopy, the insertion of an endotracheal tube (a 14G intravenous catheter) into the trachea under constant visual control is facilitated by using PE50 polyethylene tubing as a stylet (Seldinger technique). The procedure has been performed more than 60 times in our laboratory; all intubations were successful on the first attempt, and no adverse events were observed. We conclude that the described procedure is a simple and effective way to intubate a rat noninvasively, using inexpensive and commercially available equipment.

Cardioprotection by Poloxamer 188 is Mediated through Increased Endothelial Nitric Oxide Production.

Ischemia/reperfusion (I/R) injury significantly contributes to the morbidity and mortality associated with cardiac events. Poloxamer 188 (P188), a nonionic triblock copolymer, has been proposed to mitigate I/R injury by stabilizing cell membranes. However, the underlying mechanisms remain incompletely understood, particularly concerning endothelial cell function and nitric oxide (NO) production. We employed human induced pluripotent stem cell (iPSC)-derived cardiomyocytes (CMs) and endothelial cells (ECs) to elucidate the effects of P188 on cellular survival, function, and NO secretion under simulated I/R conditions. iPSC-CMs contractility and iPSC-ECs' NO production were assessed following exposure to P188. Further, an isolated heart model using Brown Norway rats subjected to I/R injury was utilized to evaluate the ex-vivo cardioprotective effects of P188, examining cardiac function and NO production, with and without the administration of a NO inhibitor. In iPSC-derived models, P188 significantly preserved CM contractile function and enhanced cell viability after hypoxia/reoxygenation. Remarkably, P188 treatment led to a pronounced increase in NO secretion in iPSC-ECs, a novel finding demonstrating endothelial protective effects beyond membrane stabilization. In the rat isolated heart model, administration of P188 during reperfusion notably improved cardiac function and reduced I/R injury markers. This cardioprotective effect was abrogated by NO inhibition, underscoring the pivotal role of NO. Additionally, a dose-dependent increase in NO production was observed in non-ischemic rat hearts treated with P188, further establishing the critical function of NO in P188 induced cardioprotection. In conclusion, our comprehensive study unveils a novel role of NO in mediating the protective effects of P188 against I/R injury. This mechanism is evident in both cellular models and intact rat hearts, highlighting the potential of P188 as a therapeutic agent against I/R injury. Our findings pave the way for further investigation into P188's therapeutic mechanisms and its potential application in clinical settings to mitigate I/R-related cardiac dysfunction.

Design and Implementation of a Rat Ex Vivo Lung Perfusion Model.

Ex vivo lung preparations are a useful model that can be translated to many different fields of research, complementing corresponding in vivo and in vitro models. Laboratories wishing to use isolated lungs need to be aware of important steps and inherent challenges to establish a setup that is affordable, reliable, and that can be easily adapted to fit the topic of interest. This paper describes a DIY (do it yourself) model for ex vivo rat lung ventilation and perfusion to study drug and gas effects on pulmonary vascular tone, independent of changes in cardiac output. Creating this model includes a) the design and construction of the apparatus, and b) the lung isolation procedure. This model results in a setup that is more cost-effective than commercial alternatives and yet modular enough to adapt to changes in specific research questions. Various obstacles had to be resolved to ensure a consistent model that is capable of being used for a variety of different research topics. Once established, this model has proven to be highly adaptable to different questions and can easily be altered for different fields of study.

Buffer glucose adjustment affects myocardial function after ischemia-reperfusion in long-term diabetic rat isolated hearts.

Due to its comorbidities type 2 diabetes mellitus (T2DM) and hypertension, the Zucker Spontaneous Hypertensive Fatty (ZSF1) rat is a clinically relevant animal model when assessing ischemia-reperfusion (IR) injury. Most IR studies in hearts isolated from diabetic animals have been conducted at normal glucose concentrations, providing a different environment compared to in-vivo. We hypothesized IR injury to be attenuated in isolated hearts of diabetic ZSF1 rats when adjusting the Krebs-buffer (KB) to their in-vivo, i.e., elevated blood glucose (BG) levels. Diabetic and non-diabetic ZSF1 rats were anesthetized, hearts isolated and Langendorff-prepared. While standard KB was used for the non-diabetic and diabetic unadjusted groups, KB with glucose levels increased to each rat's prior BG level was used for the adjusted diabetic group. All hearts underwent 30 min ischemia and 120 min reperfusion. Diastolic contracture during ischemia and early reperfusion was delayed and temporarily attenuated in the adjusted compared to the unadjusted diabetic and the non-diabetic groups. The decrease in coronary flow on reperfusion was attenuated in diabetic animals. Left ventricular developed pressure and contractility were not different among the three groups. Infarct size was significantly lower in non-diabetic animals; buffer adjustment made no difference in diabetic animals. In our study, T2DM did not worsen myocardial function in ZSF1 rat isolated hearts. Since our results reveal that hearts with an adjusted glucose level exhibit an at least temporary improvement of function following IR, further studies should consider adapting glucose levels to create more realistic conditions in isolated, perfused hearts.