Coronary microvascular dysfunction assessment: A comparative analysis of procedural aspects.

BACKGROUND: Full adoption of coronary microvascular dysfunction (CMD) assessment faces challenges due to its invasive nature and concerns about prolonged procedure time and increased contrast and/or radiation exposure. We compared procedural aspects of CMD invasive assessment to diagnostic left heart catheterization (DLHC) in patients with chest pain who were not found to have obstructive coronary artery disease. METHODS: A total of 227 patients in the Coronary Microvascular Disease Registry were compared to 1592 patients who underwent DLHC from August 2021 to November 2023. The two cohorts were compared using propensity-score matching; primary outcomes were fluoroscopy time and total contrast use. RESULTS: The participants' mean age was 64.1 ± 12.6 years. CMD-assessed patients were more likely to be female (66.5% vs. 45.2%, p < 0.001) and have hypertension (80.2% vs. 44.5%, p < 0.001), history of stroke (11.9% vs. 6.3%, p = 0.002), and history of myocardial infarction (20.3% vs. 7.7%, p < 0.001). CMD assessment was safe, without any reported adverse outcomes. A propensity-matched analysis showed that patients who underwent CMD assessment had slightly higher median contrast exposure (50 vs. 40 mL, p < 0.001), and slightly longer fluoroscopy time (6.9 vs. 4.7 min, p < 0.001). However, there was no difference in radiation dose (209.3 vs. 219 mGy, p = 0.58) and overall procedure time (31 vs. 29 min, p = 0.37). CONCLUSION: Compared to DLHC, CMD assessment is safe and requires only slightly additional contrast use (10 mL) and slightly longer fluoroscopy time (2 min) without clinical implications. These findings emphasize the favorable safety and feasibility of invasive CMD assessment.

A GLIS3-CD133-WNT-signaling axis regulates the self-renewal of adult murine pancreatic progenitor-like cells in colonies and organoids.

The existence and regenerative potential of resident stem and progenitor cells in the adult pancreas are controversial topics. A question that has been only minimally addressed is the capacity of a progenitor cell to self-renew, a key attribute that defines stem cells. Previously, our laboratory has identified putative stem and progenitor cells from the adult murine pancreas. Using an ex vivo colony/organoid culture system, we demonstrated that these stem/progenitor-like cells have self-renewal and multilineage differentiation potential. We have named these cells pancreatic colony-forming units (PCFUs) because they can give rise to three-dimensional colonies. However, the molecular mechanisms by which PCFUs self-renew have remained largely unknown. Here, we tested the hypothesis that PCFU self-renewal requires GLIS family zinc finger 3 (GLIS3), a zinc-finger transcription factor important in pancreas development. Pancreata from 2- to 4-month-old mice were dissociated, sorted for CD133highCD71low ductal cells, known to be enriched for PCFUs, and virally transduced with shRNAs to knock down GLIS3 and other proteins. We then plated these cells into our colony assays and analyzed the resulting colonies for protein and gene expression. Our results revealed a previously unknown GLIS3-to-CD133-to-WNT signaling axis in which GLIS3 and CD133 act as factors necessary for maintaining WNT receptors and signaling molecules in colonies, allowing responses to WNT ligands. Additionally, we found that CD133, but not GLIS3 or WNT, is required for phosphoinositide 3-kinase (PI3K)/AKT Ser/Thr kinase (AKT)-mediated PCFU survival. Collectively, our results uncover a molecular pathway that maintains self-renewal of adult murine PCFUs.

Activation of ductal progenitor-like cells from adult human pancreas requires extracellular matrix protein signaling.

Ductal progenitor-like cells are a sub-population of ductal cells in the adult human pancreas that have the potential to contribute to regenerative medicine. However, the microenvironmental cues that regulate their activation are poorly understood. Here, we establish a 3-dimensional suspension culture system containing six defined soluble factors in which primary human ductal progenitor-like and ductal non-progenitor cells survive but do not proliferate. Expansion and polarization occur when suspension cells are provided with a low concentration (5% v/v) of Matrigel, a sarcoma cell product enriched in many extracellular matrix (ECM) proteins. Screening of ECM proteins identified that collagen IV can partially recapitulate the effects of Matrigel. Inhibition of integrin α1ß1, a major collagen IV receptor, negates collagen IV- and Matrigel-stimulated effects. These results demonstrate that collagen IV is a key ECM protein that stimulates the expansion and polarization of human ductal progenitor-like and ductal non-progenitor cells via integrin α1ß1 receptor signaling.

From chest pain to coronary functional testing: Clinical and economic impact of coronary microvascular dysfunction.

BACKGROUND: Coronary functional testing to formally diagnose coronary microvascular dysfunction (CMD) reduces cardiovascular events and alleviates angina. This study aims to investigate the extensive and complex journey that patients with CMD undergo, from the onset of chest pain to eventual diagnosis. METHODS: Data from the Coronary Microvascular Disease Registry (CMDR) were analyzed, including information on the date of first documentation of chest pain, number of non-invasive and invasive tests the patient underwent, emergency department visits, and hospitalizations. In addition, we estimated the total cost per patient. A total of 61 patients with CMD diagnosis were included in this analysis. RESULTS: Most patients in our cohort were older than 50 years of age. The median time from initial chest pain symptoms to diagnosis was 0.62 (interquartile range [IQR]: 0.06-2.96) years. During this period, patients visited the emergency department a median of 1.0 (IQR: 0.0-2.0) times. Diagnostic tests included 3.0 (IQR: 2.0-6.0) electrocardiograms, 3.0 (IQR: 0.0-6.0) high-sensitivity troponin tests, and 1.0 (IQR: 1.0-2.0) echocardiograms. Prior to diagnosis of CMD, 13 (21.3 %) patients had left heart catheterization without coronary functional testing. Non-invasive testing for ischemia was conducted in 43 (70.5 %) patients. Alternative non-cardiac diagnoses were given to 11 (18.0 %) patients during the diagnostic process, with referrals made to gastroenterology for 16 (26.2 %) and pulmonology for 10 (16.4 %) patients. The cost was almost $2000/patient. CONCLUSION: Timely identification of CMD offers promising opportunities for prompt symptom alleviation, accompanied by reduced visits to the emergency department, cardiovascular testing, invasive medical procedures, and consequently reduced healthcare expenses.

Trefoil Factor 2 Expressed by the Murine Pancreatic Acinar Cells Is Required for the Development of Islets and for ß-Cell Function During Aging.

Exocrine-to-endocrine cross talk in the pancreas is crucial to maintain ß-cell function. However, the molecular mechanisms underlying this cross talk are largely undefined. Trefoil factor 2 (Tff2) is a secreted factor known to promote the proliferation of ß-cells in vitro, but its physiological role in vivo in the pancreas is unknown. Also, it remains unclear which pancreatic cell type expresses Tff2 protein. We therefore created a mouse model with a conditional knockout of Tff2 in the murine pancreas. We find that the Tff2 protein is preferentially expressed in acinar but not ductal or endocrine cells. Tff2 deficiency in the pancreas reduces ß-cell mass on embryonic day 16.5. However, homozygous mutant mice are born without a reduction of ß-cells and with acinar Tff3 compensation by day 7. When mice are aged to 1 year, both male and female homozygous and male heterozygous mutants develop impaired glucose tolerance without affected insulin sensitivity. Perifusion analysis reveals that the second phase of glucose-stimulated insulin secretion from islets is reduced in aged homozygous mutant compared with controls. Collectively, these results demonstrate a previously unknown role of Tff2 as an exocrine acinar cell-derived protein required for maintaining functional endocrine ß-cells in mice.

Extracellular vesicles: A dive into their role in the tumor microenvironment and cancer progression.

Extracellular vesicles (EVs) encompass a diverse set of membrane-derived particles released from cells and are found in numerous biological matrices and the extracellular space. Specific classes of EVs include apoptotic bodies, exosomes, and microvesicles, which vary in their size, origin, membrane protein expression, and interior cargo. EVs provide a mechanism for shuttling cargo between cells, which can influence cell physiology by transporting proteins, DNA, and RNA. EVs are an abundant component of the tumor microenvironment (TME) and are proposed to drive tumor growth and progression by communicating between fibroblasts, macrophages, and tumor cells in the TME. The cargo, source, and type of EV influences the pro- or anti-tumoral role of these molecules. Therefore, robust EV isolation and characterization techniques are required to ensure accurate elucidation of their association with disease. Here, we summarize different EV subclasses, methods for EV isolation and characterization, and a selection of current clinical trials studying EVs. We also review key studies exploring the role and impact of EVs in the TME, including how EVs mediate intercellular communication, drive cancer progression, and remodel the TME.

Methylcellulose colony assay and single-cell micro-manipulation reveal progenitor-like cells in adult human pancreatic ducts.

Progenitor cells capable of self-renewal and differentiation in the adult human pancreas are an under-explored resource for regenerative medicine. Using micro-manipulation and three-dimensional colony assays we identify cells within the adult human exocrine pancreas that resemble progenitor cells. Exocrine tissues were dissociated into single cells and plated into a colony assay containing methylcellulose and 5% Matrigel. A subpopulation of ductal cells formed colonies containing differentiated ductal, acinar, and endocrine lineage cells, and expanded up to 300-fold with a ROCK inhibitor. When transplanted into diabetic mice, colonies pre-treated with a NOTCH inhibitor gave rise to insulin-expressing cells. Both colonies and primary human ducts contained cells that simultaneously express progenitor transcription factors SOX9, NKX6.1, and PDX1. In addition, in silico analysis identified progenitor-like cells within ductal clusters in a single-cell RNA sequencing dataset. Therefore, progenitor-like cells capable of self-renewal and tri-lineage differentiation either pre-exist in the adult human exocrine pancreas, or readily adapt in culture.