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Nicholas F LaRusso

Publications and source records attributed to Nicholas F LaRusso.

17 recordsLinked to original sources
PubMed2003

Agonist-induced coordinated trafficking of functionally related transport proteins for water and ions in cholangiocytes.

We previously proposed that ductal bile formation is regulated by secretin-responsive relocation of aquaporin 1 (AQP1), a water-selective channel protein, from an intracellular vesicular compartment to the apical membrane of cholangiocytes. In this study, we immunoisolated AQP1-containing vesicles from cholangiocytes prepared from rat liver; quantitative immunoblotting revealed enrichment in these vesicles of not only AQP1 but also cystic fibrosis transmembrane regulator (CFTR) and AE2, a Cl- channel and a Cl-/HCO3- exchanger, respectively. Dual labeled immunogold electron microscopy of cultured polarized mouse cholangiocytes showed significant colocalization of AQP1, CFTR, and AE2 in an intracellular vesicular compartment; exposure of cholangiocytes to dibutyryl-cAMP (100 microm) resulted in co-redistribution of all three proteins to the apical cholangiocyte plasma membrane. After administration of secretin to rats in vivo, bile flow increased, and AQP1, CFTR, and AE2 co-redistributed to the apical cholangiocyte membrane; both events were blocked by pharmacologic disassembly of microtubules. Based on these in vitro and in vivo observations utilizing independent and complementary approaches, we propose that cholangiocytes contain an organelle that sequesters functionally related proteins that can account for ion-driven water transport, that this organelle moves to the apical cholangiocyte membrane in response to secretory agonists, and that these events account for ductal bile secretion at a molecular level.

Animals↗
PubMed2003

Hepatic artery and portal vein remodeling in rat liver: vascular response to selective cholangiocyte proliferation.

Three-dimensional reconstruction of the biliary tree, hepatic artery, and portal vein in normal rats and rats fed alpha-naphthylisothiocyanate (ANIT), a compound that causes selective proliferation of epithelial cells (ie, cholangiocytes) that line the bile ducts, was performed. All hepatic structures in ANIT-fed rats branched 1.5 times more often than in normal rats, reflecting an increased number of segments, whereas the length of the biliary tree, hepatic artery, and portal vein remain unchanged. The length of the proximal vessel segments was uniform in both groups of rats whereas the length of distal segments decreased twofold in ANIT-fed rats, suggesting that small vessels preferentially undergo proliferation. In contrast, the length of all bile duct segments decreased twofold, suggesting that ANIT induced proliferation of all compartments of the biliary tree. The total volume of the biliary tree, hepatic artery, and portal vein was increased 18, 4, and 3 times, respectively, after ANIT feeding. The diameters of the bile ducts (range, 20 to 259 microm) and arterial (range, 21 to 276 microm) segments in ANIT-fed rats did not differ from normal rats (range, 21 to 245 microm and 20 to 265 microm, respectively). In contrast, the diameters of proximal venous segments in ANIT-fed rats were significantly less (316 +/- 68 micro m versus 488 +/- 89 micro m, P < 0.001). The data suggest that after experimentally induced cholangiocyte proliferation, the hepatic artery and portal vein also undergo marked proliferation, presumably to support the increased nutritional and functional demands of the proliferated bile ducts. The molecular mechanisms of these vascular changes remain to be determined.

1-Naphthylisothiocyanate↗
PubMed2003

Somatostatin stimulates ductal bile absorption and inhibits ductal bile secretion in mice via SSTR2 on cholangiocytes.

With an in vitro model using enclosed intrahepatic bile duct units (IBDUs) isolated from wild-type and somatostatin receptor (SSTR) subtype 2 knockout mice, we tested the effects of somatostatin, secretin, and a selective SSTR2 agonist (L-779976) on fluid movement across the bile duct epithelial cell layer. By RT-PCR, four of five known subtypes of SSTRs (SSTR1, SSTR2A/2B, SSTR3, and SSTR4, but not SSTR5) were detected in cholangiocytes in wild-type mice. In contrast, SSTR2A/2B were completely depleted in the SSTR2 knockout mice whereas SSTR1, SSTR3 and SSTR4 were expressed in these cholangiocytes. Somatostatin induced a decrease of luminal area of IBDUs isolated from wild-type mice, reflecting net fluid absorption; L-779976 also induced a comparable decrease of luminal area. No significant decrease of luminal area by either somatostatin or L-779976 was observed in IBDUs from SSTR2 knockout mice. Secretin, a choleretic hormone, induced a significant increase of luminal area of IBDUs of wild-type mice, reflecting net fluid secretion; somatostatin and L-779976 inhibited (P < 0.01) secretin-induced fluid secretion. The inhibitory effect of both somatostatin and L-779976 on secretin-induced IBDU secretion was absent in IBDUs of SSTR2 knockout mice. Somatostatin induced an increase of intracellular cGMP and inhibited secretin-stimulated cAMP synthesis in cholangiocytes; depletion of SSTR2 blocked these effects of somatostatin. These data suggest that somatostatin regulates ductal bile formation in mice not only by inhibition of ductal fluid secretion but also by stimulation of ductal fluid absorption via interacting with SSTR2 on cholangiocytes, a process involving the intracellular cAMP/cGMP second messengers.

Absorption↗
PubMed2002

Specific inhibition of AQP1 water channels in isolated rat intrahepatic bile duct units by small interfering RNAs.

Cholangiocytes express water channels (i.e. aquaporins (AQPs)), proteins that are increasingly recognized as important in water transport by biliary epithelia. However, direct functional studies demonstrating AQP-mediated water transport in cholangiocytes are limited, in part because of the lack of specific AQP inhibitors. To address this issue, we designed, synthesized, and utilized small interfering RNAs (siRNAs) selective for AQP1 and investigated their effectiveness in altering AQP1-mediated water transport in intrahepatic bile duct units (IBDUs) isolated from rat liver. Twenty-four hours after transfection of IBDUs with siRNAs targeting two different regions of the AQP1 transcript, both AQP1 mRNA and protein expression were inhibited by 76.6-92.0 and 57.9-79.4%, respectively. siRNAs containing the same percent of base pairs as the AQP1-siRNAs but in random sequence (i.e. scrambled siRNAs) had no effect. Suppression of AQP1 expression in cholangiocytes resulted in a decrease in water transport by IBDUs in response to both an inward osmotic gradient (200 mosm) or a secretory agonist (forskolin), the osmotic water permeability coefficient (P(f)) decreasing up to 58.8% and net water secretion (J(v)) decreasing up to 87%. A strong correlation between AQP1 protein expression and water transport in IBDUs transfected with AQP1-siRNAs was consistent with the decrease in water transport by IBDUs resulting from AQP1 gene silencing by AQP1-siRNAs. This study is the first to demonstrate the feasibility of utilizing siRNAs to specifically reduce the expression of AQPs in epithelial cells and provides direct evidence of the contribution of AQP1 to water transport by biliary epithelia.

Animals↗
PubMed2002

A novel incentive system for faculty in an academic medical center.

The need to contain health care costs has led some physicians to become salaried employees of health care organizations. However, the use of nonfinancial incentives for physicians in such an environment has not been broadly explored. The authors describe a novel incentive system that is designed to promote continuing high-quality care and to increase patient access to health care while enhancing clinical and academic productivity and physician satisfaction. Key components of this system include annual targets, flexibility in meeting these targets, and ability to convert clinical productivity generated in excess of what was necessary to meet the target to support scholarly activities. This system led to increased faculty productivity, improved patient access, enhanced scholarly activity, and overall enhanced career satisfaction.

Academic Medical Centers↗
PubMed2002

Cryptosporidiosis.

Explore the source record for details and available documents.

AIDS-Related Opportunistic Infections↗
PubMed2002

Expression and localization of aquaporin water channels in rat hepatocytes. Evidence for a role in canalicular bile secretion.

Although bile formation requires that large volumes of water be rapidly transported across liver epithelia, including hepatocytes, the molecular mechanisms by which water is secreted into bile are obscure. The aquaporins are a family of 10 channel-forming, integral membrane proteins of approximately 28 kDa numbered 0-9 that allow water to rapidly traverse epithelial barriers in several organs including kidney, eye, and brain. We found transcripts of three of 10 aquaporins in hepatocytes (aquaporin 8 aquaporin 9 > aquaporin 0) by reverse transcription-polymerase chain reaction and quantitative ribonuclease protection assays; immunohistochemistry confirmed the presence of these three proteins in liver. Immunoblots of subcellular fractions of hepatocytes showed enrichment of aquaporins 0 and 8 in microsomes and canalicular plasma membranes; aquaporin 9 was enriched only in basolateral plasma membranes. Immunofluorescence of hepatocyte couplets confirmed the intracellular/canalicular localization of aquaporins 0 and 8 and the basolateral localization of aquaporin 9. Upon exposure of couplets to a choleretic stimulus (i.e. dibutyryl cAMP), aquaporin 8 redistributed to the canalicular plasma membrane; the subcellular distributions of aquaporins 0 and 9 were unaffected. In addition, exposure of couplets to dibutyryl cAMP caused an increase in canalicular water transport in the presence and absence of an osmotic gradient, an effect that was blocked by aquaporin inhibitors. These results provide evidence that aquaporins are present in hepatocytes and that aquaporins are involved in agonist-stimulated canalicular bile secretion.

Animals↗
PubMed2002

Cryptosporidiosis and the pathogenesis of AIDS-cholangiopathy.

Cryptosporidium, an intracellular parasite that infects the gastrointestinal epithelium and other mucosal surfaces, causes self-limited diarrhea in immunocompetent subjects and potentially life-threatening syndromes in immunocompromised individuals, primarily those with acquired immunodeficiency syndrome (AIDS). Cryptosporidium is also the single most common identifiable pathogen in the biliary tract in patients with AIDS-cholangiopathy, an important biliary disorder caused by opportunistic infection of the biliary epithelium and resulting in significant morbidity and mortality in AIDS patients. The organism stimulates periductal inflammation in the biliary tree, induces biliary epithelial cell apoptosis, and thus contributes to the pathogenesis of AIDS-cholangiopathy. Currently, there is no fully effective medical therapy for both of the diseases. A better understanding of the parasitology of Cryptosporidium and the pathophysiology of biliary cryptosporidiosis should aid not only our understanding of the pathogenesis of AIDS-cholangiopathy but also the development of effective therapies and control of this ubiquitous, highly infectious threat.

AIDS-Related Opportunistic Infections↗
PubMed2002

Intrahepatic bile ducts transport water in response to absorbed glucose.

The physiological relevance of the absorption of glucose from bile by cholangiocytes remains unclear. The aim of this study was to test the hypothesis that absorbed glucose drives aquaporin (AQP)-mediated water transport by biliary epithelia and is thus involved in ductal bile formation. Glucose absorption and water transport by biliary epithelia were studied in vitro by microperfusing intrahepatic bile duct units (IBDUs) isolated from rat liver. In a separate set of in vivo experiments, bile flow and absorption of biliary glucose were measured after intraportal infusion of D-glucose or phlorizin. IBDUs absorbed D-glucose in a dose- and phlorizin-dependent manner with an absorption maximum of 92.8 +/- 6.2 pmol. min(-1). mm(-1). Absorption of D-glucose by microperfused IBDUs resulted in an increase of water absorption (J(v) = 3-10 nl. min(-1). mm(-1), P(f) = 40 x 10(-3) cm/sec). Glucose-driven water absorption by IBDUs was inhibited by HgCl(2), suggesting that water passively follows absorbed D-glucose mainly transcellularly via mercury-sensitive AQPs. In vivo studies showed that as the amount of absorbed biliary glucose increased after intraportal infusion of D-glucose, bile flow decreased. In contrast, as the absorption of biliary glucose decreased after phlorizin, bile flow increased. Results support the hypothesis that the physiological significance of the absorption of biliary glucose by cholangiocytes is likely related to regulation of ductal bile formation.

Animals↗
PubMed2002

Channel-mediated water movement across enclosed or perfused mouse intrahepatic bile duct units.

We previously reported the development of reproducible techniques for isolating and perfusing intact intrahepatic bile duct units (IBDUs) from rats. Given the advantages of transgenic and knockout mice for exploring ductal bile formation, we report here the adaptation of those techniques to mice and their initial application to the study of water transport across mouse intrahepatic biliary epithelia. IBDUs were isolated from livers of normal mice by microdissection combined with enzymatic digestion. After culture, isolated IBDUs sealed to form intact, polarized compartments, and a microperfusion system employing those isolated IBDUs developed. A quantitative image analysis technique was used to observe a rapid increase of luminal area when sealed IBDUs were exposed to a series of inward osmotic gradients reflecting net water secretion; the choleretic agonists secretin and forskolin also induced water secretion into IBDUs. The increase of IBDU luminal area induced by inward osmotic gradients and choleretic agonists was reversibly inhibited by HgCl2, a water channel inhibitor. With the use of a quantitative epifluorescence technique in perfused mouse IBDUs, a high osmotic water permeability (P(f) = 2.5-5.6 x 10(-2) cm/s) was found in response to osmotic gradients, further supporting the presence of water channels. These findings suggest that, as in the rat, water transport across intrahepatic biliary epithelia in mice is water channel mediated.

Animals↗
PubMed2002

Experimental models to study cholangiocyte biology.

Cholangiocytes-the epithelial cells which line the bile ducts-are increasingly recognized as important transporting epithelia actively involved in the absorption and secretion of water, ions, and solutes. This recognition is due in part to the recent development of new experimental models. New biologic concepts have emerged including the identification and topography of receptors and flux proteins on the apical and/or basolateral membrane which are involved in the molecular mechanisms of ductal bile secretion. Individually isolated and/or perfused bile duct units from livers of rats and mice serve as new,physiologically relevant in vitro models to study cholangiocyte transport. Biliary tree dimensions and novel insights into anatomic remodeling of proliferating bile ducts have emerged from three-dimensional reconstruction using CT scanning and sophisticated software. Moreover, new pathologic concepts have arisen regarding the interaction of cholangiocytes with pathogens such as Cryptosporidium parvum. These concepts and associated methodologies may provide the framework to develop new therapies for the cholangiopathies, a group of important hepatobiliary diseases in which cholangiocytes are the target cell.

Animals↗
PubMedDate not supplied

Optimization of a research web environment for academic internal medicine faculty.

Usability evaluations are a powerful tool that can assist developers in their efforts to optimize the quality of their web environment. This underutilized, experimental method can serve to move applications toward true user-centered design. This article describes the usability methodology and illustrates its importance and application by describing a usability study undertaken at the Mayo Clinic for the purpose of improving an academic research web environment. Academic institutions struggling in an era of declining reimbursements are finding it difficult to maintain academic enterprises on the back of clinical revenues. This may result in declining amounts of time that clinical investigators have to spend in non-patient-related activities. For this reason, we have undertaken to design a web environment, which can minimize the time that a clinician-investigator needs to spend to accomplish academic instrumental activities of daily living. Usability evaluation is a powerful application of human factors engineering, which can improve the utility of web-based Informatics applications.

Computer Simulation↗