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Biomedical subjects

Glenn Prestwich

Publications and source records attributed to Glenn Prestwich.

4 recordsLinked to original sources
PubMed2006

Caspase-3-dependent activation of calcium-independent phospholipase A2 enhances cell migration in non-apoptotic ovarian cancer cells.

Calcium-independent phospholipase A(2) (iPLA(2)) plays a pivotal role in phospholipid remodeling and many other biological processes, including inflammation and cancer development. iPLA(2) can be activated by caspase-3 via a proteolytic process in apoptotic cells. In this study we identify novel signaling and functional loops of iPLA(2) activation leading to migration of non-apoptotic human ovarian cancer cells. The extracellular matrix protein, laminin-10/11, but not collagen I, induces integrin- and caspase-3-dependent cleavage and activation of overexpressed and endogenous iPLA(2). The truncated iPLA(2) (amino acids 514-806) generates lysophosphatidic acid and arachidonic acid. Arachidonic acid is important for enhancing cell migration toward laminin-10/11. Lysophosphatidic acid activates Akt that in turn acts in a feedback loop to block the cleavage of poly-(ADP-ribose) polymerase and DNA fragmentation factor as well as prevent apoptosis. By using pharmacological inhibitors, blocking antibodies, and genetic approaches (such as point mutations, dominant negative forms of genes, and siRNAs against specific targets), we show that beta(1), but not beta(4), integrin is involved in iPLA(2) activation and cell migration to laminin-10/11. The role of caspase-3 in iPLA(2) activation and cell migration are supported by several lines of evidence. 1) Point mutation of Asp(513) (a cleavage site of caspase-3 in iPLA(2)) to Ala blocks laminin-10/11-induced cleavage and activation of overexpressed iPLA(2), whereas mutation of Asp(733) to Ala has no such effect, 2) treatment of inhibitors or a small interfering RNA against caspase-3 results in decreased cell migration toward laminin-10/11, and 3) selective caspase-3 inhibitor blocks cleavage of endogenous iPLA(2) induced by laminin-10/11. Importantly, small interfering RNA-mediated down-regulation of endogenous iPLA(2) expression in ovarian carcinoma HEY cells results in decreased migration toward laminin, suggesting that our findings are pathophysiologically important.

Apoptosis↗
PubMed2006

Cross-linked hydrogels for middle ear packing.

OBJECTIVE: To develop an ideal supportive packing material for ossiculoplasty, tympanoplasty, or other otologic procedures. MATERIALS AND METHODS: Several materials, namely, Carbylan-SX (P-C; Sentrx Surgical, Inc., Salt Lake City, UT), Gelfoam (P-GF; Pharmacia & Upjohn, Kalamazoo, MI), and Merogel (P-MG; Medtronics, Inc., Minneapolis, MN), were prepared and then placed into a Hartley guinea pig's (Elm Hill, Chelmsford, MA) middle ear cavities through a large myringotomy incision. The contralateral ear underwent a large myringotomy without packing material being placed. Preoperative and posteroperative auditory brainstem response studies were performed using Intelligent Hearing system software. The animals were examined weekly. Two weeks after packing placement, the animals were killed, and the temporal bones were harvested. Whole temporal bone sectioning was performed to analyze the presence of implant, surrounding inflammation, presence of osteoneogenesis and fibrosis, or adhesions. RESULTS: All the materials, except the P-MG, were easy to place into the middle ear cavity. The P-MG contains woven strands that are difficult to trim into the small sizes needed for placement. The P-MG group had a smaller average amount of implant present compared with the other groups at 2 weeks. The degree of osteoneogenesis was similar among the P-GF, P-C, and P-MG groups. The P-MG and P-C groups contained the lowest amount of fibrosis between the implant and surrounding middle ear structures. CONCLUSION: This study demonstrates promising results with P-C as a potential supportive packing material for otologic procedures. P-C compares favorably with P-MG and P-GF in a guinea pig model with respect to ease of placement and amount of fibrosis.

Animals↗
PubMed2003

Use of expression constructs to dissect the functional domains of the CHS/beige protein: identification of multiple phenotypes.

The Chediak-Higashi Syndrome (CHS) and the orthologous murine disorder beige are characterized at the cellular level by the presence of giant lysosomes. The CHS1/Beige protein is a 3787 amino acid protein of unknown function. To determine functional domains of the CHS1/Beige protein, we generated truncated constructs of the gene/protein. These truncated proteins were transiently expressed in Cos-7 or HeLa cells and their effect on membrane trafficking was examined. Beige is apparently a cytosolic protein, as are most transiently expressed truncated Beige constructs. Expression of the Beige construct FM (amino acids 1-2037) in wild-type cells led to enlarged lysosomes. Similarly, expression of a 5.5-kb region (amino acids 2035-3787) of the carboxyl terminal of Beige (22B) also resulted in enlarged lysosomes. Expression of FM solely affected lysosome size, whereas expression of 22B led to alterations in lysosome size, changes in the Golgi and eventually cell death. The two constructs could be used to further dissect phenotypes resulting from loss of the Beige protein. CHS or beigej fibroblasts show an absence of nuclear staining using a monoclonal antibody directed against phosphatidylinositol 4,5 bisphosphate [PtdIns(4,5) P2]. Transformation of beige j fibroblasts with a YAC containing the full-length Beige gene resulted in the normalization of lysosome size and nuclear PtdIns(4,5)P2 staining. Expression of the carboxyl dominant negative construct 22B led to loss of nuclear PtdIns(4,5)P2 staining. Expression of the FM dominant negative clone did not alter nuclear PtdIns(4,5) P2 localization. These results suggest that the Beige protein interacts with at least two different partners and that the Beige protein affects cellular events, such as nuclear PtdIns(4,5)P2 localization, in addition to lysosome size.

Animals↗