IS-PRM-based peptide targeting informed by long-read sequencing for alternative proteome detection.

Alternative splicing is a major contributor of transcriptomic complexity, but the extent to which transcript isoforms are translated into stable, functional protein isoforms is unclear. Furthermore, detection of relatively scarce isoform-specific peptides is challenging, with many protein isoforms remaining uncharted due to technical limitations. Recently, a family of advanced targeted MS strategies, termed internal standard parallel reaction monitoring (IS-PRM), have demonstrated multiplexed, sensitive detection of pre-defined peptides of interest. Such approaches have not yet been used to confirm existence of novel peptides. Here, we present a targeted proteogenomic approach that leverages sample-matched long-read RNA sequencing (LR RNAseq) data to predict potential protein isoforms with prior transcript evidence. Predicted tryptic isoform-specific peptides, which are specific to individual gene product isoforms, serve as "triggers" and "targets" in the IS-PRM method, Tomahto. Using the model human stem cell line WTC11, LR RNAseq data were generated and used to inform the generation of synthetic standards for 192 isoform-specific peptides (114 isoforms from 55 genes). These synthetic "trigger" peptides were labeled with super heavy tandem mass tags (TMT) and spiked into TMT-labeled WTC11 tryptic digest, predicted to contain corresponding endogenous "target" peptides. Compared to DDA mode, Tomahto increased detectability of isoforms by 3.6-fold, resulting in the identification of five previously unannotated isoforms. Our method detected protein isoform expression for 43 out of 55 genes corresponding to 54 resolved isoforms. This LR RNA seq-informed Tomahto targeted approach, called LRP-IS-PRM, is a new modality for generating protein-level evidence of alternative isoforms - a critical first step in designing functional studies and eventually clinical assays.

A transesophageal echocardiography technique to locate the kidney and monitor renal perfusion.

Monitoring the renal arterial Doppler flow velocity indices, the resistive index and pulsatility index, with ultrasound may help predict renal dysfunction. However, such monitoring has been done intermittently by transcutaneous ultrasound in the postoperative intensive care setting. In the operating room, transesophageal echocardiography (TEE) is an alternative to transcutaneous ultrasound for obtaining indices of renal perfusion. However, it is difficult to locate the right kidney using TEE. We propose a new technique to locate the left kidney that, in our experience, is simple and easy to perform. We believe, starting from a transgastric left ventricular short-axis view, turning left to locate the abdominal aorta, and following it to the origin of the left renal artery may help locate the left kidney faster than previously described techniques. We also propose a new technique to monitor these Doppler indices using TEE during the intraoperative period.

A rare case of a left atrial hemangioma mimicking a myxoma.

A 68-year-old gentleman presented with a recent history of exertional dyspnea associated with anginal chest pain. Transthoracic echocardiography revealed a well-circumscribed mass in the left atrium attached to the inter-atrial septum. A provisional diagnosis of left atrial (LA) myxoma was made. Coronary angiography revealed significant single-vessel disease. The patient underwent coronary artery bypass grafting and resection of the LA tumor. The histopathological diagnosis of the tumor came out to be a cardiac hemangioma.

Primary malignant fibrous histiocytoma involving the left pulmonary vein presenting as a left atrial tumor.

A 35-year-old woman presented with 4 months history of progressively increasing intermittent dyspnea and hemoptysis. Transthoracic echocardiography revealed a loculated mass in the left atrium (LA). A provisional diagnosis of LA myxoma was made. Intraoperatively the tumor was found extending into and closely adherent to the left pulmonary vein and could not be completely cleared off from the pulmonary venous wall. The histopathological examination of the tumor revealed it to be a myxoid malignant fibrous histiocytoma.