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Biomedical subjects

L Lim

Publications and source records attributed to L Lim.

At least 55 records · Page 3Linked to original sources
PubMed2000

Coupling of PAK-interacting exchange factor PIX to GIT1 promotes focal complex disassembly.

The p21-activated kinase PAK is targeted to focal complexes (FCs) through interactions with the SH3 domains of the PAK-interacting exchange factor PIX and Nck. PIX is a Rac GTP exchange factor that also binds the G-protein-coupled receptor kinase-interacting protein known as GIT1. Overexpression of GIT1 in fibroblasts or epithelial cells causes a loss of paxillin from FCs and stimulates cell motility. This is due to the direct interaction of a C-terminal 125-residue domain of GIT1 with paxillin, under the regulation of PIX. In its activated state, GIT1 can promote FC disassembly independent of actin-myosin contractile events. Additionally, GIT directly couples to a key component of FCs, focal adhesion kinase (FAK), via a conserved Spa2 homology domain. We propose that GIT1 and FAK cooperate to promote motility both by directly regulating focal complex dynamics and by the activation of Rac.

Adaptor Proteins, Signal Transducing↗
PubMed2000

A case series of acanthamoeba keratitis in Singapore.

Acanthamoeba keratitis has become an important cause of severe ocular inflammation and visual loss in the past two decades. Its prevalence has been linked to the increasing use of contact lens. Early diagnosis, effective treatment regimes and education on proper contact lens wear are important in the management and prevention of visual loss from this debilitating disease. We described a series of two cases of culture positive acanthamoeba keratitis and their subsequent management.

Acanthamoeba Keratitis↗
PubMed2000

Interferon-gamma counteracts reduced endotoxin responsiveness of whole blood following trauma and cardiopulmonary bypass.

Accidental as well as surgical trauma has been reported to cause reduced endotoxin responsiveness of blood in terms of cytokine production. In this study, the effect of interferon-gamma (IFN-gamma) on tumour necrosis factor-alpha (TNF-alpha)-producing capacity of whole blood after severe trauma and cardiac surgery was investigated. Blood samples of severely injured patients were collected at the first day after trauma and of cardiac surgery patients before, 4 h and 2 days after cardiopulmonary bypass (CPB). The blood samples were incubated with INF-gamma (0-100 U/ml) for 20 h and subsequently lipopolysaccharide (LPS)-induced TNF-alpha production was determined. Compared to healthy donors, LPS-induced TNF-alpha production was significantly reduced in blood cultures of trauma patients on day 1 after trauma and 4 h after CPB. Pre-incubation with IFN-gamma in vitro increased endotoxin-induced TNF-alpha production in volunteers' and all patients' blood specimens in a dose-dependent manner. IFN-gamma prompted an elevation of cytokine synthesis in CPB patients' blood which equalled that of volunteers, whereas it caused a lower rise in TNF-alpha production in blood of multiply injured patients, reaching levels of untreated donors only after incubation with 100 U/ml IFN-gamma. These experiments show that hyporesponsiveness of whole blood induced by trauma or cardiac surgery with CPB is not irreversible, but can be counteracted by the immunostimulant IFN-gamma. IFN-gamma, therefore, could be applied clinically in trauma patients or after cardiac surgery to prevent or to resolve infection complications.

Adjuvants, Immunologic↗
PubMed1999

HNF-3/forkhead homologue-4 influences lung morphogenesis and respiratory epithelial cell differentiation in vivo.

HNF-3/forkhead homologue 4 (HFH-4), a transcription factor of the winged helix/forkhead family, is expressed in various tissues including lung, brain, oviduct, testis, and embryonic kidney. In order to test whether the temporospatial expression of HFH-4 influences lung morphogenesis, HFH-4 was expressed in lungs of transgenic mice under control of the surfactant protein C (SP-C) promoter. The morphology of the lungs from SP-C/HFH-4 embryos (day 18 postconception) was distinctly abnormal, and the severity of the alterations correlated with the level of transgene expression as detected by in situ hybridization. At high levels of expression, HFH-4 altered epithelial cell differentiation and inhibited branching morphogenesis. Atypical cuboidal or columnar cells lined the lung periphery of SP-C/HFH-4 transgenic mice. The atypical epithelial cells seen in the SP-C/HFH-4 mice expressed thyroid transcription factor-1 and hepatocyte nuclear factor 3beta (HNF-3beta). However, surfactant proteins SP-B, SP-C, and Clara cell secretory protein, normally produced by nonciliated epithelial cells in lung parenchyma were lacking. beta-Tubulin IV, a marker of ciliated cells, stained the atypical columnar cells produced by expression of high levels of the SP-C/HFH-4 transgene. Ectopic expression of HFH-4 in developing mouse lung altered epithelial cell differentiation and morphology, restricting the expression of markers typical of nonciliated cells of the distal lung parenchyma.

Animals↗
PubMed1999

Variation in in-patient stroke management in ten centres in different countries: the INCLEN multicentre stroke collaboration.

BACKGROUND AND PURPOSE: Large within-country variations have been described in stroke management and there have been a few studies of between-country variation (in the USA and the UK). We designed a study to examine stroke management across a wide range of countries representing different stages of economic development. Large variations would suggest the need to explore methods of increasing the uptake of evidence-based stroke practice. METHODS: Members of the International Clinical Epidemiology Network (INCLEN) from 14 centres in ten countries agreed to review the records of the last 50 patients admitted to hospital with a clinical diagnosis of stroke. Information on demographic variables, the clinical diagnosis of stroke type, investigations performed and treatments given and the discharge destination of the patient were recorded and sent to the coordinating centre in Australia for analysis. RESULTS: There were statistically significant between-centre differences in the proportions of patients cared for by a neurologist, staying in hospital for at least ten days and having CT or MRI scans. Significant between-centre differences were also seen for treatment, for example, the use of aspirin in non-haemorrhagic stroke varied from 11 to 79%. The variation (for all interventions studied) was no longer statistically significant when examined within strata according to availability of facilities. CONCLUSIONS: The large variation between centres in the management of stroke is largely 'explained' by the availability of resources, even for interventions that do not depend on resource availability. It will be important to develop management guidelines that reflect evidence-based practice of relevance across a range of economic settings.

Age Factors↗
PubMed1999

The myotonic dystrophy kinase-related Cdc42-binding kinase is involved in the regulation of neurite outgrowth in PC12 cells.

The myotonic dystrophy kinase-related Cdc42-binding kinase (MRCKalpha) has been implicated in the morphological activities of Cdc42 in nonneural cells. Both MRCKalpha and the kinase-related Rho-binding kinase (ROKalpha) are involved in nonmuscle myosin light-chain phosphorylation and associated actin cytoskeleton reorganization. We now show that in PC12 cells, overexpression of the kinase domain of MRCKalpha and ROKalpha resulted in retraction of neurites formed on nerve growth factor (NGF) treatment, as observed with RhoA. However, introduction of kinase-dead MRCKalpha did not result in NGF-independent neurite outgrowth as observed with dominant negative kinase-dead ROKalpha or the Rho inhibitor C3. Neurite outgrowth induced by NGF or kinase-dead ROKalpha was inhibited by dominant negative Cdc42(N17), Rac1(N17), and the Src homology 3 domain of c-Crk, indicating the participation of common downstream components. Neurite outgrowth induced by either agent was blocked by kinase-dead MRCKalpha lacking the p21-binding domain or by a minimal C-terminal regulatory region consisting of the cysteine-rich domain/pleckstrin homology domain plus a region with homology to citron. The latter region alone was an effective blocker of NGF-induced outgrowth. These results suggest that although ROKalpha is involved in neurite retraction promoted by RhoA, the related MRCKalpha is conversely involved in neurite outgrowth promoted by Cdc42 and Rac.

Amino Acid Sequence↗
PubMed1999

Pak functions downstream of Dock to regulate photoreceptor axon guidance in Drosophila.

The SH2/SH3 adaptor protein Dock has been proposed to transduce signals from guidance receptors to the actin cytoskeleton in Drosophila photoreceptor (R cell) growth cones. Here, we demonstrate that Drosophila p21-activated kinase (Pak) is required in a Dock pathway regulating R cell axon guidance and targeting. Dock and Pak colocalize to R cell axons and growth cones, physically interact, and their loss-of-function phenotypes are indistinguishable. Normal patterns of R cell connectivity require Pak's kinase activity and binding sites for both Dock and Cdc42/Rac. A membrane-tethered form of Pak (Pak(myr) acts as a dominant gain-of-function protein. Retinal expression of Pak(myr) rescues the R cell connectivity phenotype in dock mutants. These data establish Pak as a critical regulator of axon guidance and a downstream effector of Dock in vivo.

Adaptor Proteins, Signal Transducing↗
PubMed1999

Structure of the small G protein Cdc42 bound to the GTPase-binding domain of ACK.

The proteins Cdc42 and Rac are members of the Rho family of small GTPases (G proteins), which control signal-transduction pathways that lead to rearrangements of the cell cytoskeleton, cell differentiation and cell proliferation. They do so by binding to downstream effector proteins. Some of these, known as CRIB (for Cdc42/Rac interactive-binding) proteins, bind to both Cdc42 and Rac, such as the PAK1-3 serine/threonine kinases, whereas others are specific for Cdc42, such as the ACK tyrosine kinases and the Wiscott-Aldrich-syndrome proteins (WASPs). The effector loop of Cdc42 and Rac (comprising residues 30-40, also called switch I), is one of two regions which change conformation on exchange of GDP for GTP. This region is almost identical in Cdc42 and Racs, indicating that it does not determine the specificity of these G proteins. Here we report the solution structure of the complex of Cdc42 with the GTPase-binding domain ofACK. Both proteins undergo significant conformational changes on binding, to form a new type of G-protein/effector complex. The interaction extends the beta-sheet in Cdc42 by binding an extended strand from ACK, as seen in Ras/effector interactions, but it also involves other regions of the G protein that are important for determining the specificity of effector binding.

Amino Acid Sequence↗
PubMed1999

Conjunctival rotation autograft for pterygium. An alternative to conjunctival autografting.

OBJECTIVE: To determine the safety and efficacy of conjunctival rotation autografting (CRA) as an alternative to conventional conjunctival autograft after pterygium excision. DESIGN: Prospective noncomparative case series. PARTICIPANTS: Consecutive patients seen at the Pterygium Clinic of the Singapore National Eye Centre who were thought to be unsuitable for conventional conjunctival autografting underwent a modified surgical procedure, which the authors describe as CRA. There were 51 rotation autografts performed on 45 eyes of 43 patients. INTERVENTION: In this procedure, the underlying fibrovascular pterygium tissue was removed and the original epithelium (with minimal subepithelial tissue included) replaced over the bare sclera with a 180 degrees rotation. Surgeries were performed by one surgeon (DT) from April 1995 to May 1996. MAIN OUTCOME MEASURE: Pterygium recurrence and complications of CRA were measured. RESULTS: The mean follow-up time was 12 months (range, 2-22 months). There were 46 primary and 5 recurrent pterygia. The indications for CRA were combined cataract and pterygium surgery (39.2%), double pterygia (31.4%), the need to preserve the superior conjunctiva (21.6%), and superior conjunctival scarring (7.8%). There were two recurrences (4% recurrence rate), one occurring at 4 months and the other occurring at 7 months after surgery. No significant complications were encountered. However, 50% of the grafts remained mildly injected for more than 3 months, and some remained injected for up to 13 months after surgery (average of 4 months). Pigmentary changes were also noted in six grafts (12%). CONCLUSION: Conjunctival rotation autografting is a useful technique of conjunctival grafting in cases in which it is not possible or desirable to use the superior conjunctiva as a donor source.

Adult↗
PubMed1999

Randomized clinical trial of a new dexamethasone delivery system (Surodex) for treatment of post-cataract surgery inflammation.

OBJECTIVE: To evaluate the safety of Surodex Drug Delivery System (Oculex Pharmaceuticals, Inc., Sunnyvale, CA) containing dexamethasone 60 micrograms, for use in cataract surgery, and to compare its anti-inflammatory efficacy with conventional dexamethasone 0.1% eyedrops. DESIGN: Randomized, masked, and partially controlled trial. PARTICIPANTS: Sixty eyes of 60 Asian patients undergoing extracapsular cataract extraction with intraocular lens implantation were examined. Of these, 28 eyes of 28 patients served as control eyes. Patients were stratified for age and presence of diabetes mellitus. INTERVENTION: Surodex was inserted in the anterior chamber of 32 eyes at the conclusion of surgery. These eyes received placebo eyedrops four times a day after surgery for 4 weeks. Control eyes received neither Surodex nor a placebo implant but were prescribed conventional 0.1% dexamethasone eyedrops four times a day for 4 weeks. MAIN OUTCOME MEASURES: Anterior chamber cells and flare were clinically graded at the slit lamp. Anterior chamber flare was objectively assessed with the Kowa FM500 Laser Flare Meter (Kowa Co. Ltd, Tokyo, Japan) for up to 3 months after surgery. Intraocular pressure and corneal endothelial specular microscopy with morphometric cell analysis were performed for up to 1 year after surgery. RESULTS: Clinical slit-lamp assessment of anterior chamber flare and cells showed no difference between Surodex-treated eyes and dexamethasone eyedrop-treated eyes. Flare meter readings showed lower flare levels in the Surodex group at all postoperative visits compared with the dexamethasone eyedrop group. Flare reduction in the Surodex group reached statistical significance at days 4, 8, 15, and 30 after surgery. At 3 months, flare was reduced to preoperative levels in the Surodex group but was still raised in the dexamethasone eyedrop group. Five eyes in the dexamethasone eyedrop group required augmentation of steroids and were deemed therapeutic failures as opposed to one eye in the Surodex group. One patient in the dexamethasone eyedrop group developed postoperative open-angle glaucoma with profound visual field loss and optic disc cupping, resulting in hand movements vision. No significant difference in endothelial cell loss was noted between Surodex-inserted eyes and dexamethasone eyedrop-treated eyes for up to 1 year after surgery. CONCLUSIONS: Intraocular placement of a single Surodex is a safe and effective treatment method to reduce intraocular inflammation after cataract surgery. There was no statistical difference in efficacy between Surodex and 0.1% dexamethasone eyedrops in reducing intraocular inflammation, as measured by clinical methods, while Surodex was clearly superior to eyedrops in reducing aqueous flare as objectively assessed with the laser flare meter.

Aged↗
PubMed1999

Melanoma chondroitin sulphate proteoglycan regulates cell spreading through Cdc42, Ack-1 and p130cas.

Melanoma chondroitin sulphate proteoglycan (MCSP) is a cell-surface antigen that has been implicated in the growth and invasion of melanoma tumours. Although this antigen is expressed early in melanoma progression, its biological function is unknown. MCSP can stimulate the integrin-alpha4 beta1-mediated adhesion and spreading of melanoma cells. Here we show that stimulated MCSP recruits tyrosine-phosphorylated p130 cas, an adaptor protein important in tumour cell motility and invasion. MCSP stimulation also results in a pronounced activation and recruitment of the Rho-family GTPase Cdc42. MCSP-induced spreading of melanoma cells is dependent upon active Cdc42, a Cdc42-associated tyrosine kinase (Ack-1) and tyrosine phosphorylation of p130cas. Furthermore, vectors inhibiting Ack-1 or Cdc42 expression and/or function abrogate MCSP-induced tyrosine phosphorylation and recruitment of p130cas. Our findings indicate that MCSP may modify tumour growth or invasion by a unique signal-transduction pathway that links Cdc42 activation to downstream tyrosine phosphorylation and subsequent cytoskeletal reorganization.

Cell Movement↗
PubMed1999

Molecular characterization of the Caenorhabditis elegans Rho GDP-dissociation inhibitor.

GDP-dissociation inhibitors (GDIs) form one of the classes of regulatory proteins that modulate the cycling of the Ras superfamily of GTPases between active GTP-bound and inactive GDP-bound states. We report here the characterization of the Caenorhabditis elegans RhoGDI (CeRhoGDI) as part of our investigations into Rho-GTPase signalling pathways that are involved in nematode development. CeRhoGDI is a 23-kDa protein that is localized predominantly in the cytosol. CeRhoGDI interacts only with the lipid-modified forms of C. elegans Rho-GTPases, CeRhoA, CeRac1 and Cdc42Ce, in vitro and is able to solubilize the membrane-bound forms of these GTPases. CeRhoGDI recognizes the GTPases in both GTP- and GDP-bound forms; hence it inhibits both the guanine-nucleotide dissociation and GTP-hydrolysis activities. The inhibitory activity towards the GTP-bound GTPases is weak compared with that towards GDP-bound GTPases. CeRhoGDI is expressed throughout development and is highly expressed in marginal and vulval epithelial cells, in sperm cells and spicules. Taken together, our results suggest that CeRhoGDI may be involved in specific morphogenetic events mediated by the C. elegans Rho-GTPases.

Amino Acid Sequence↗
PubMed1999

Influence of granulocyte-macrophage colony-stimulating factor (GM-CSF) on whole blood endotoxin responsiveness following trauma, cardiopulmonary bypass, and severe sepsis.

Major surgery, multiple injury, and severe sepsis lead to an impaired immune response. The suppressed status of the immune system is reflected by a reduced TNFalpha production of whole blood after stimulation with endotoxin in vitro and by a decreased HLA-DR expression on monocytes. In the present study, the effect of the immunostimulating hematopoetic growth factor GM-CSF on whole blood cultures of multiple injury, cardiac surgery, and severe sepsis patients was investigated. Endotoxin-induced TNFalpha production and HLA-DR expression was reduced in blood cultures of these patients compared to healthy donors. Preincubation with GM-CSF in vitro increased cytokine production in volunteers' and all patients' blood specimens in a dose-dependent manner. The elevation of cytokine response in cardiopulmonary bypass patients' blood, caused by in vitro preincubation with GM-CSF, equaled that of normal patients, whereas GM-CSF caused a lower rise of TNFalpha-producing capacity in blood of multiple-injury and sepsis patients. Further, GM-CSF treatment in vitro increased the down-regulated HLA-DR expression on monocytes prepared after cardiac surgery to a degree comparable to preoperative levels. Finally, GM-CSF incubation in vitro elevated TNFalpha synthesis in normal monocytes and in cells treated with a combination of the anti-inflammatory mediators IL-10, TGFbeta, and PGE2. These experiments show that hyporesponsiveness of whole blood induced by trauma, sepsis, or cardiac surgery is not irreversible but can be, at least in vitro, overridden by the immunostimulating compound GM-CSF.

Adolescent↗
PubMed1999

Symptoms during normal pregnancy: a prospective controlled study.

Symptoms of normal pregnancy have received scant attention in the literature and what is reported is largely unsubstantiated. Yet this is an important aspect of antenatal counselling and care which deserves further investigation if symptoms are to be interpreted correctly. Accordingly, we conducted a prospective controlled study of symptoms during normal pregnancy in both primigravidas and multigravidas. A total of 38 symptoms occurred with a significantly different frequency (mainly increased) in the pregnant subjects in the third trimester compared with the controls. Of these a mean of 24.2 symptoms was experienced by each pregnant woman, double that (mean, 11.2) experienced by healthy nonpregnant controls. The 5 symptoms reported most frequently by the pregnant subjects were frequency of micturition, fatigue, pelvic pressure, insomnia and lower backache. However, a wide range of symptoms involving most body systems were reported. This study has established that symptoms of pregnancy are more numerous than mentioned in current obstetric texts and that they can be attributed to the effects of pregnancy. The third trimester is associated with the greatest number of symptoms and there is a marked decline in their number after delivery.

Adolescent↗
PubMed1999

Participation of small GTPases in dorsal closure of the Drosophila embryo: distinct roles for Rho subfamily proteins in epithelial morphogenesis.

The Rho subfamily of Ras-related small GTPases participates in a variety of cellular events including organization of the actin cytoskeleton and signalling by c-Jun N-terminal kinase and p38 kinase cascades. These functions of the Rho subfamily are likely to be required in many developmental events. We have been studying the participation of the RHO subfamily in dorsal closure of the Drosophila embryo, a process involving morphogenesis of the epidermis. We have previously shown that Drac1, a Rho subfamily protein, is required for the presence of an actomyosin contractile apparatus believed to be driving the cell shape changes essential to dorsal closure. Expression of a dominant negative Drac1 transgene causes a loss of this contractile apparatus from the leading edge of the advancing epidermis and dorsal closure fails. We now show that two other Rho subfamily proteins, Dcdc42 and RhoA, as well as Ras1 are also required for dorsal closure. Dcdc42 appears to have conflicting roles during dorsal closure: establishment and/or maintenance of the leading edge cytoskeleton versus its down regulation. Down regulation of the leading edge cytoskeleton may be controlled by the serine/threonine kinase DPAK, a potential Drac1/Dcdc42 effector. RhoA is required for the integrity of the leading edge cytoskeleton specifically in cells flanking the segment borders. We have begun to characterize the interactions of the various small GTPases in regulating dorsal closure and find no evidence for the hierarchy of Rho subfamily activity described in some mammalian cell types. Rather, our results suggest that while all &Rgr; subfamily p21s tested are required for dorsal closure, they act largely in parallel.

Actins↗