Fluid phase endocytosis by isolated rabbit lacrimal gland acinar cells

Gierow, J. Peter; Lambert, Robert; Mircheff, Austin K.

doi:10.1016/s0014-4835(05)80066-1

Cited by 42 publications

(44 citation statements)

References 28 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…All animal procedures were in accordance with the Guide for the Care and Use of Laboratory Animals, published by the National Institutes of Health (NIH Publication no. 85-23, revised 1996) (9,17,18) and were approved by the University of Southern California Institutional Animal Care and Use Committee. Acinar cells isolated from LG from New Zealand White rabbits (1.8 -2.2 kg) (Irish Farms, Norco, CA) were sequentially incubated in Mg 2ϩ -and Ca 2ϩ -free HBSS supplemented with EDTA and Ham's medium supplemented with collagenase, hyaluronidase, and DNAse, and cultured for 2 days in Peter's serum-free culture medium (18).…”

Section: Methodsmentioning

confidence: 99%

Rab27b regulates exocytosis of secretory vesicles in acinar epithelial cells from the lacrimal gland

Chiang¹,

Ngo²,

Schechter

et al. 2011

American Journal of Physiology-Cell Physiology

View full text Add to dashboard Cite

mechanisms largely yet uncharacterized. We investigated the role of Rab27b in the terminal release of these secretory vesicles. Confocal fluorescence microscopy analysis of primary cultured rabbit lacrimal gland acinar cells revealed that Rab27b was enriched on the membrane of large subapical vesicles that were significantly colocalized with Rab3D and Myosin 5C. Stimulation of cultured acinar cells with the secretagogue carbachol resulted in apical fusion of these secretory vesicles with the plasma membrane. Evaluation of morphological changes by transmission electron microscopy of lacrimal glands from Rab27b Ϫ/Ϫ and Rab27 ash/ash /Rab27b Ϫ/Ϫ mice, but not ashen mice deficient in Rab27a, showed changes in abundance and organization of secretory vesicles, further confirming a role for this protein in secretory vesicle exocytosis. Glands lacking Rab27b also showed increased lysosomes, damaged mitochondria, and autophagosomelike organelles. In vitro, expression of constitutively active Rab27b increased the average size but retained the subapical distribution of Rab27b-enriched secretory vesicles, whereas dominant-negative Rab27b redistributed this protein from membrane to the cytoplasm. Functional studies measuring release of a cotransduced secretory protein, syncollin-GFP, showed that constitutively active Rab27b enhanced, whereas dominant-negative Rab27b suppressed, stimulated release. Disruption of actin filaments inhibited vesicle fusion to the apical membrane but did not disrupt homotypic fusion. These data show that Rab27b participates in aspects of lacrimal gland acinar cell secretory vesicle formation and release. actin; Rab3d; exocrine secretion; syncollin; mouse models A FUNCTIONING LACRIMAL GLAND (LG) is critical for a healthy ocular surface. This exocrine gland is the primary source of tear proteins and fluid, which, released up on the gland's exposure to sympathetic or parasympathetic agonists provided by innervating nerves, contribute to the middle aqueous layer of the precorneal film. Much of the LG's secretions originate from acinar cells, which constitute over 80% of the mass of the LG (13). These LG acinar cells produce a diverse array of secretory proteins that are internally sorted into large pools of serous and mucous secretory vesicles (SV) sized ϳ1 m in diameter and stored beneath the apical plasma membrane (APM) in preparation for regulated exocytosis. SV contents include nutrients and growth factors (lacritin and EGF), antibacterial and antiviral factors (secretory IgA and lactoferrin), and an array of proteases and lysosomal hydrolases (10, 39, 47). Despite its physiological importance, however, few studies have focused on the mechanisms of secretory membrane trafficking in LG acinar cells, in part due to the fragility and heterogeneity of these LG acinar SV relative to those in other acinar secretory cells, e.g., pancreatic acini (7).The current schematic of exocytosis in the LG acinar cell suggests multiple participants. Mature LG acinar cell SV localize beneath an actin-rich network und...

show abstract

Section: Methodsmentioning

confidence: 99%

Rab27b regulates exocytosis of secretory vesicles in acinar epithelial cells from the lacrimal gland

Chiang¹,

Ngo²,

Schechter

et al. 2011

American Journal of Physiology-Cell Physiology

View full text Add to dashboard Cite

show abstract

“…Individual cells within these structures display distinct apical and basal-lateral domains, a polarized cytoskeleton, secretory vesicles and also release protein stores in response to secretagogues. 21,23 To determine the transduction efficiency of the recombinant Ad vectors, acinar cells were analyzed by fluorescence-activated cell sorting (FACS). The addition of Ad 5-containing green fluorescent protein (Ad-GFP) to reconstituted acini over a range of multiplicity of infections (MOIs) demonstrated that exposure at an MOI of 5 for 16-18 h resulted in high transduction efficiency.…”

Section: Ad-mediated Gene Transfer Into Lacrimal Acinar Epithelial Cellsmentioning

confidence: 99%

“…Lacrimal gland acinar cells were isolated and cultured for 2-3 days as described previously. [21][22][23] …”

Section: Cell Isolation and Culturementioning

confidence: 99%

“…The cell pellet was then exposed to a cocktail of collagenase, hyaluronidase and DNAse 21,22 at 371C for 10 min, cells were again rinsed with DPBS and collected by centrifugation as described above. Cells were resuspended in DPBS, and GFP-positive cells were quantified by a Becton Dickinson FAC-Scant analyzer (Becton Dickinson, Franklin Lakes, NJ, USA) using a 15 mW air-cooled argon laser set at 488 nm and recorded with a 530 nm emission filter in the FL1 emission channel.…”

Section: Flow Cytometry Analysismentioning

confidence: 99%

“…As an initial step toward this goal, we investigated the effect of replication-defective adenovirus serotype 5 (Ad5) containing either green fluorescent protein (GFP) or b-galactosidase (LacZ) on the organization and function of the lacrimal acinar secretory pathway in reconstituted acinus-like structures formed in vitro from acinar cells from rabbit lacrimal gland. [21][22][23] At conditions associated with high-efficiency transduction (480%), Ad elicited specific changes in the organization and function of the stimulated secretory pathway, changes that could be largely recapitulated by ultraviolet light (UV)-inactivated Ad. These findings suggest that ocular gene therapy utilizing Ad or Adderived products may alter secretory membrane-trafficking functions in epithelial cells of the ocular surface.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Adenoviral capsid modulates secretory compartment organization and function in acinar epithelial cells from rabbit lacrimal gland

et al. 2004

View full text Add to dashboard Cite

Although adenovirus (Ad) exhibits tropism for epithelial cells, little is known about the cellular effects of adenoviral binding and internalization on epithelial functions. Here, we examine its effects on the secretory acinar epithelial cells of the lacrimal gland, responsible for stimulated release of tear proteins into ocular fluid. Exposure of reconstituted rabbit lacrimal acini to replication-defective Ad for 16-18 h under conditions that resulted in 480% transduction efficiency did not alter cytoskeletal filament or biosynthetic/endosomal membrane compartment organization. Transduction specifically altered the organization of the stimulated secretory pathway, eliciting major dispersal of rab3D immunofluorescence from apical stores normally associated with mature secretory vesicles. Biochemical studies revealed that this dispersal was not associated with altered rab3D expression nor its release from cellular membranes. Ultraviolet (UV)-inactivated Ad elicited similar dispersal of rab3D immunofluorescence. In acini exposed to replication-defective or UVinactivated Ad, carbachol-stimulated release of bulk protein and b-hexosaminidase were significantly (Pp0.05) inhibited to an extent proportional to the loss of rab3D-enriched mature secretory vesicles associated with these treatments. We propose that the altered secretory compartment organization and function caused by Ad reflects changes in the normal maturation of secretory vesicles, and that these changes are caused by exposure to the Ad capsid.

show abstract

Sjögren's autoimmunity: how perturbation of recognition in endomembrane traffic may provoke pathological recognition at the cell surface

et al. 1998

Self Cite

View full text Add to dashboard Cite

CD4 T cell antigen recognition requires presentation by major histocompatibility complex Class II molecules (MHC II). B cell surface immunoglobulins recognize antigens independently of MHC II, but activation typically requires CD4 cell cytokines as accessory signals. Plasma membrane-endomembrane traffic in lacrimal gland acinar cells, targets of autoimmune activity in Sjögren's syndrome, may satisfy both requirements. The Golgi protein galactosyltransferase and the lysosomal proteins cathepsin B and cathepsin D appear at the plasma membranes during sustained secretomotor stimulation. The RNA transcription termination factor La, a frequent target of Sjögren's autoantibodies, appears in the acinar cell cytoplasm and plasma membranes during viral infection and during in vitro exposure to cytokines. MHC II cycle through endomembrane compartments which contain La, galactosyltransferase, cathepsin B and cathepsin D and which are sites of proteolysis. This traffic may permit trilateral interactions in which B cells recognize autoantigens at the surface membranes, CD4 T cells recognize peptides presented by MHC II, B cells provide accessory signals to CD4 T cells, and CD4 T cells provide cytokines that activate B cells. Acinar cells stimulate lymphocyte proliferation in autologous mixed cell reactions, confirming that they are capable of provoking autoimmune responses.

show abstract

Fluid phase endocytosis by isolated rabbit lacrimal gland acinar cells

Cited by 42 publications

References 28 publications

Rab27b regulates exocytosis of secretory vesicles in acinar epithelial cells from the lacrimal gland

Rab27b regulates exocytosis of secretory vesicles in acinar epithelial cells from the lacrimal gland

Adenoviral capsid modulates secretory compartment organization and function in acinar epithelial cells from rabbit lacrimal gland

Sjögren's autoimmunity: how perturbation of recognition in endomembrane traffic may provoke pathological recognition at the cell surface

Contact Info

Product

Resources

About