Faculty Opinions recommendation of Ex vivo human pancreatic slice preparations offer a valuable model for studying pancreatic exocrine biology.

Abstract The lack of efficient gene transfer methods into primary human pancreatic exocrine cells hampers studies on the plasticity of these cells and their possible role in beta cell regeneration. Therefore, improved gene transfer protocols are needed. Lentiviral vectors are widely used to drive ectopic gene expression in mammalian cells, including primary human islet cells. Here we aimed to optimize gene transfer into primary human exocrine cells using modified lentiviral vectors or transduction conditions. We evaluated different promoters, viral envelopes, medium composition and transduction adjuvants. Transduction efficiency of a reporter vector was evaluated by fluorescence microscopy and flow cytometry. We show that protamine sulfate-assisted transduction of a VSV-G-pseudotyped vector expressing eGFP under the control of a CMV promoter in a serum-free environment resulted in the best transduction efficiency of exocrine cells, reaching up to 90% of GFP-positive cells 5 days after transduction. Our findings will enable further studies on pancreas (patho)physiology that require gene transfer such as gene overexpression, gene knockdown or lineage tracing studies.

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Slc26a6 is an apical membrane anion exchanger that drives HCO3−-dependent fluid secretion in murine pancreatic acinar cells

AJP Cell Physiology ◽

10.1152/ajpcell.00257.2019 ◽

2019 ◽

Vol 317 (6) ◽

pp. C1153-C1160

Author(s):

Takashi Munemasa ◽

Taro Mukaibo ◽

James E. Melvin

Keyword(s):

Anion Exchanger ◽

Apical Membrane ◽

Ex Vivo ◽

Acinar Cells ◽

Fluid Secretion ◽

Pancreatic Acinar Cells ◽

Anion Transporter ◽

Dependent Component ◽

Primary Driver ◽

Pancreatic Exocrine

The nonselective anion exchanger Slc26a6, also known as putative anion transporter 1 and chloride/formate exchanger, is thought to play a major role in [Formula: see text] transport in exocrine glands. In this study, Slc26a6 null mice were used to explore the function of Slc26a6 in the exocrine pancreas. Slc26a6 primarily localized to the apical membrane of pancreatic exocrine acinar cells. The volume of stimulated juice secretion by the ex vivo pancreas was significantly reduced ~35% in Slc26a6−/− mice, but no changes occurred in the gross structure or gland weights of Slc26a6 null mice. The secretion of pancreatic juice by Slc26a6+/+ mice was dependent on [Formula: see text] while, in contrast, fluid secretion by Slc26a6 −/− mice was independent of [Formula: see text], suggesting that Slc26a6 mediates the [Formula: see text]-dependent component of fluid secretion. Consistent with these observations, disruption of Slc26a6 also significantly reduced [Formula: see text] secretion by the pancreas ~35%. Taken together, these results demonstrate that the apical Slc26a6 anion exchanger in acinar cells is involved in [Formula: see text]-dependent fluid secretion but that another major [Formula: see text]-independent pathway is the primary driver of the fluid secretion process in the mouse pancreas.

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Ex vivohuman pancreatic slice preparations offer a valuable model for studying pancreatic exocrine biology

Journal of Biological Chemistry ◽

10.1074/jbc.m117.777433 ◽

2017 ◽

Vol 292 (14) ◽

pp. 5957-5969 ◽

Cited By ~ 26

Author(s):

Tao Liang ◽

Subhankar Dolai ◽

Li Xie ◽

Erin Winter ◽

Abrahim I. Orabi ◽

...

Keyword(s):

Pancreatic Exocrine ◽

Pancreatic Slice

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Correlation of γ-radioactivity and Scanning Electron Microscopy in measurement of retention of 111Indium-labeled canine endothelial cells on synthetic vascular grafts

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100156419 ◽

1989 ◽

Vol 47 ◽

pp. 886-887

Author(s):

E.J. Prendiville ◽

S. Laliberté Verdon ◽

K. E. Gould ◽

K. Ramberg ◽

R. J. Connolly ◽

...

Keyword(s):

Blood Flow ◽

Ex Vivo ◽

Vascular Grafts ◽

Retention Data ◽

Vascular Prostheses ◽

Group 4 ◽

Human Fibronectin ◽

Insufficient Time ◽

Group 3 ◽

Group 2

Endothelial cell (EC) seeding is postulated as a mechanism of improving patency in small caliber vascular grafts. However the majority of seeded EC are lost within 24 hours of restoration of blood flow in previous canine studies . We postulate that the cells have insufficient time to fully develop their attachment to the graft surface prior to exposure to hemodynamic stress. We allowed EC to incubate on fibronectin-coated ePTFE grafts for four different time periods after seeding and measured EC retention after perfusion in a canine ex vivo shunt circuit.Autologous canine EC, were enzymatically harvested, grown to confluence, and labeled with 30 μCi 111 Indium-oxine/80 cm 2 flask. Four groups of 5 cm x 4 mm ID ePTFE vascular prostheses were coated with 1.5 μg/cm.2 human fibronectin, and seeded with 1.5 x 105 EC/ cm.2. After seeding grafts in Group 1 were incubated in complete growth medium for 90 minutes, Group 2 were incubated for 24 hours, Group 3 for 72 hours and Group 4 for 6 days. Grafts were then placed in the canine ex vivo circuit, constructed between femoral artery and vein, and subjected to blood flow of 75 ml per minute for 6 hours. Continuous counting of γ-activity was made possible by placing the seeded graft inside the γ-counter detection crystal for the duration of perfusion. EC retention data after 30 minutes, 2 hours and 6 hours of flow are shown in the table.

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Elevated level of circulatory sTLT1 induces inflammation through SYK/MEK/ERK signalling in coronary artery disease

Clinical Science ◽

10.1042/cs20190999 ◽

2019 ◽

Vol 133 (22) ◽

pp. 2283-2299

Author(s):

Apabrita Ayan Das ◽

Devasmita Chakravarty ◽

Debmalya Bhunia ◽

Surajit Ghosh ◽

Prakash C. Mandal ◽

...

Keyword(s):

Risk Factors ◽

Coronary Artery Disease ◽

Coronary Artery ◽

Chronic Inflammation ◽

Ex Vivo ◽

Human Subjects ◽

Critical Role ◽

Atherosclerotic Cardiovascular Disease ◽

Tnf Α ◽

Artery Disease

Abstract The role of inflammation in all phases of atherosclerotic process is well established and soluble TREM-like transcript 1 (sTLT1) is reported to be associated with chronic inflammation. Yet, no information is available about the involvement of sTLT1 in atherosclerotic cardiovascular disease. Present study was undertaken to determine the pathophysiological significance of sTLT1 in atherosclerosis by employing an observational study on human subjects (n=117) followed by experiments in human macrophages and atherosclerotic apolipoprotein E (apoE)−/− mice. Plasma level of sTLT1 was found to be significantly (P<0.05) higher in clinical (2342 ± 184 pg/ml) and subclinical cases (1773 ± 118 pg/ml) than healthy controls (461 ± 57 pg/ml). Moreover, statistical analyses further indicated that sTLT1 was not only associated with common risk factors for Coronary Artery Disease (CAD) in both clinical and subclinical groups but also strongly correlated with disease severity. Ex vivo studies on macrophages showed that sTLT1 interacts with Fcɣ receptor I (FcɣRI) to activate spleen tyrosine kinase (SYK)-mediated downstream MAP kinase signalling cascade to activate nuclear factor-κ B (NF-kB). Activation of NF-kB induces secretion of tumour necrosis factor-α (TNF-α) from macrophage cells that plays pivotal role in governing the persistence of chronic inflammation. Atherosclerotic apoE−/− mice also showed high levels of sTLT1 and TNF-α in nearly occluded aortic stage indicating the contribution of sTLT1 in inflammation. Our results clearly demonstrate that sTLT1 is clinically related to the risk factors of CAD. We also showed that binding of sTLT1 with macrophage membrane receptor, FcɣR1 initiates inflammatory signals in macrophages suggesting its critical role in thrombus development and atherosclerosis.

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The Shikani HME: A New Tracheostomy Heat and Moisture Exchanger

Journal of Speech Language and Hearing Research ◽

10.1044/2020_jslhr-19-00107 ◽

2020 ◽

Vol 63 (9) ◽

pp. 2921-2929

Author(s):

Alan H. Shikani ◽

Elamin M. Elamin ◽

Andrew C. Miller

Keyword(s):

Ex Vivo ◽

Moisture Loss ◽

Speaking Valve ◽

Time Zero ◽

In Series ◽

Turbulent Airflow ◽

The Difference ◽

Loss Efficiency ◽

Heat And Moisture ◽

Lung Simulation

Purpose Tracheostomy patients face many adversities including loss of phonation and essential airway functions including air filtering, warming, and humidification. Heat and moisture exchangers (HMEs) facilitate humidification and filtering of inspired air. The Shikani HME (S-HME) is a novel turbulent airflow HME that may be used in-line with the Shikani Speaking Valve (SSV), allowing for uniquely preserved phonation during humidification. The aims of this study were to (a) compare the airflow resistance ( R airflow ) and humidification efficiency of the S-HME and the Mallinckrodt Tracheolife II tracheostomy HME (M-HME) when dry (time zero) and wet (after 24 hr) and (b) determine if in-line application of the S-HME with a tracheostomy speaking valve significantly increases R airflow over a tracheostomy speaking valve alone (whether SSV or Passy Muir Valve [PMV]). Method A prospective observational ex vivo study was conducted using a pneumotachometer lung simulation unit to measure airflow ( Q ) amplitude and R airflow , as indicated by a pressure drop ( P Drop ) across the device (S-HME, M-HME, SSV + S-HME, and PMV). Additionally, P Drop was studied for the S-HME and M-HME when dry at time zero (T 0 ) and after 24 hr of moisture testing (T 24 ) at Q of 0.5, 1, and 1.5 L/s. Results R airflow was significantly less for the S-HME than M-HME (T 0 and T 24 ). R airflow of the SSV + S-HME in series did not significant increase R airflow over the SSV or PMV alone. Moisture loss efficiency trended toward greater efficiency for the S-HME; however, the difference was not statistically significant. Conclusions The turbulent flow S-HME provides heat and moisture exchange with similar or greater efficacy than the widely used laminar airflow M-HME, but with significantly lower resistance. The S-HME also allows the innovative advantage of in-line use with the SSV, hence allowing concurrent humidification and phonation during application, without having to manipulate either device.

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