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Biomedical subjects

P L Lim

Publications and source records attributed to P L Lim.

At least 19 recordsLinked to original sources
PubMed2006

Outpatient parenteral antibiotic therapy in Singapore.

Outpatient parenteral antibiotic therapy (OPAT) remains in its infancy in Singapore, with the first patients enrolled 4 years ago. Singapore's three largest hospitals, with over 3000 inpatient beds, now have designated and approved OPAT services. This study reviews the demographic, clinical and cost data of all patients enrolled in 2005 to facilitate benchmarking between services in Singapore and abroad and also to identify common needs for further development. In 2005, 225 OPAT enrollments in 208 different patients resulted in 4050 days of OPAT care. Orthopaedic diagnoses constituted 40% of admissions. Vancomycin was the most frequently used antibiotic (34%). The re-admission rate was 8.9%, but complications of OPAT care were only occasionally implicated. An estimated $207,200 was saved by patients despite there being significant financial disincentives to subsidised patients. OPAT is a safe, cost-efficient system that is becoming increasingly accepted in Singapore by patients, clinicians and management. Our three services have evolved independently into very similar practices. There is potential for further innovation, including outreach and carer-delivered dosing. However, major financial disincentives require review.

Adolescent↗
PubMed2006

Extremely low exposure of a community to severe acute respiratory syndrome coronavirus: false seropositivity due to use of bacterially derived antigens.

Estimates of seropositivity to a new infectious agent in a community are useful to public health. For severe acute respiratory syndrome (SARS), the figures are conflicting. Herein, we screened 12,000 people in a community stricken by SARS 10 months previously and found 53 individuals (0.44%) who had immunoglobulin G antibodies to the SARS coronavirus (SARS-CoV) nucleocapsid (N) produced in bacteria. However, only seven of these (group 1) had sera which also reacted with the native N antigen expressed in SARS-CoV-infected Vero cells, N-transfected 293T cells, and tissues of infected SARS patients. Of these, six individuals had had SARS previously. The remaining person, as well as the 46 other individuals (group 2), were healthy and had no history of SARS. Group 1 antibodies recognized epitopes located slightly differently in N from those of group 2 antibodies, and a mouse hybridoma antibody resembling the former type was generated. Unusually, group 2 antibodies appeared to recognize cross-reactive bacterial epitopes that presumably were posttranslationally modified in eukaryotes and hence were probably not induced by SARS-CoV or related coronaviruses but rather by bacteria. The N antigen is thus highly unique. The extremely low rate (0.008%) of asymptomatic SARS infection found attests to the high virulence of the SARS-CoV virus.

Animals↗
PubMed2005

Hippocampal involvement in dengue fever.

Flaviviruses are among the most important emerging viruses known to man. Dengue is the most common flavivirus infection in Singapore, and is transmitted between humans by the Aedes mosquito. We report a 25-year-old man with dengue fever complicated by selective hippocampal involvement manifesting as amnesia. This has not been described in the literature previously. Dengue polymerase chain reaction and serology were positive. Magnetic resonance imaging of the brain showed bilateral hippocampal involvement.

Adult↗
PubMed2005

Wound infections in tsunami survivors: a commentary.

Survivors of the December 2004 Indian Ocean tsunami sustained a variety of wound infections, ranging from common pathogens to rarely seen organisms. This article discusses the likely microbiology potentially seen in wound infections with exposure to freshwater, seawater, soil, faecal or other contamination, and attempts to provide an organising framework for choosing empiric antibiotics for such infections. Therapy for less frequently encountered clinical entities is also discussed, including tetanus, cutaneous and septicaemic melioidosis, post-traumatic mucormycosis, Vibrio vulnificus and Aeromonas hydrophila.

Disasters↗
PubMed2001

Expression of human BRE in multiple isoforms.

BRE, a putative stress-modulating gene, found able to down-regulate TNF-alpha-induced NF-kappaB activation upon overexpression, is now shown in human cells expressed as multiple mRNA isoforms. A total of six isoforms are produced by alternative splicing predominantly at either end of the gene. Predicted from the cDNA sequences of these isoforms, three of them (alpha(a), alpha(b), and alpha(c)) code for BRE of different C-terminus, and the other three (beta(a), beta(b), and beta(c)) may possibly be the nonfunctional counterparts. All human cells examined coexpress all the predominant splice variants, albeit at different ratios. Comparing with normal cells, immortalized human cell lines uniformly express higher levels of BRE. Interestingly, peripheral blood monocytes responded to LPS by down-regulating the expression of all the BRE isoforms, which was however less obvious in the cell line counterpart, THP-1. Isoform alpha(a), which codes for the canonical BRE with a C-terminal peroxisomal targeting sequence, is the most abundant transcript. We propose that the function of BRE and its isoforms is to regulate peroxisomal activities.

Animals↗
PubMed2001

Molecular mimicry: anti-DNA antibodies may arise inadvertently as a response to antibodies generated to microorganisms.

The origin of anti-DNA antibodies remains speculative. We argue that some of these antibodies may arise inadvertently in nature during the course of a normal immune response due to their induction by antibodies which bear structures (mimotopes) that mimic DNA. These antibodies are not necessarily DNA specific but, like the T15 idiotype (id)-positive antibodies which bind to phosphorylcholine, are produced normally to some environmental or microbial antigen. Such a mimotope was found in a T15(+) antibody at the highly specific region encoded principally by the D gene, DFL16.1. This mimotope was also found in human antibodies that are encoded by DXP'1, the human counterpart of DFL16.1 and which is used commonly in anti-DNA antibodies. The mimotope is closely related to the epitope responsible for the T15 id and appears to be cryptic or normally hidden in the native protein. The existence of such a common, conserved sequence raises questions about how easily anti-DNA antibodies can be generated in nature and what purpose these proteins may serve. Molecular mimicry with regard to autoimmunity must thus be viewed as existing not necessarily between the infectious agent and self-antigens, but also between the antibodies induced by the organism and the self-antigens.

Animals↗
PubMed2000

A human and a mouse anti-idiotypic antibody specific for human T14(+) anti-DNA antibodies reconstructed by phage display.

Little is known about human anti-idiotypic antibodies. Phage display methodology was used to reconstruct these antibodies from lupus patients, which recognize a subset (T14(+)) of anti-DNA antibodies. Antigen-specific B cells were isolated from the blood using a peptide based on a complementarity determining region (V(H)CDR3) of the prototypic T14(+) antibody. cDNA fragments of the V(H) and V(L) genes prepared from the cells were expressed as phage displayed single chain Fv (scFv) fragments using the pCANTAB-5E phagemid vector. From a reactive clone obtained, the Ig genes used were identified to be V(H)3, D5-D3, J(H)4b, V(kappa)I and J(kappa)2. The heavy chain was highly mutated, especially in CDR3, which bears mutations mostly of the replacement type; this region is also unusual in being extremely long due to a D-D fusion. In contrast, a mouse hybridoma antibody, made to the same T14(+) peptide and transformed as a scFv fragment, uses a short V(H)CDR3 comprising five amino acids, three of which are tyrosines. Tyrosines may be important for antigen binding because two of these also exist in the human V(H)CDR3. The light chains of both antibodies may also contribute to the specificity of the protein, because their V(L) segments, including the CDRs, are highly homologous to each other.

Amino Acid Sequence↗
PubMed2000

Isolation of leptin-binding peptides from a random peptide phage library.

Leptin plays a role in regulating the body weight in mice. Injection of recombinant mouse leptin expressed in Escherichia coli reduced the food intake and body weight in normal, ob/ob and diet-induced obesity mice. Hyperglycemia, hyperinsulinemia and hypothermia can also be corrected in ob/ob mice after leptin injection. Leptin is a 16-kDa secretory protein comprising 167 amino acids produced in adipose tissue and is secreted to blood stream. In this study, a recombinant mouse leptin was generated and purified from a baculovirus expression system. This protein was used to identify putative ligands using a phage library of random peptides. Three leptin-binding phage clones were found, which were characterized by DNA sequencing and ELISA methods. The amino acid sequences of the reactive peptides are: LAYCSDPVRCLVWWY, MFWISAVSFVDHALV and LVLVLSAFLCCGVG. All three clones bound to recombinant human and mouse leptins. These peptides may be useful tools to study leptin-receptor interaction, food intake and body weight regulation.

Amino Acid Sequence↗
PubMed2000

Strand bias in Ig somatic hypermutation is determined by signal sequence within the variable region.

Ig genes undergo hypermutation with a nucleotide preference of A over T for mutation on the coding strand. As only with concomitant strand bias can such nucleotide bias be observed, Ig gene hypermutation is generally accepted as a strand-specific process, for which the mechanistic basis remains unknown. It has previously been shown that different non-Ig sequences replacing the LVJ region of an Ig transgene to various extents are targeted for hypermutation with similar mutation frequencies. However, the nucleotide bias characteristic of Ig hypermutation was not found in two of the three such sequences studied. To test whether it is the DNA sequences of the non-Ig substrates that determine the pattern of nucleotide bias in hypermutation or whether the LVJ sequence may contain element(s) that confer strand bias, we have added back all the replaced LVJ sequences to one of the transgenes, L(kappa)-Vgpt*, that expresses no strand bias in hypermutation and studied the outcome. The results show that the gpt sequence in the presence of the complete LVJ sequence hypermutates differently from the same sequence in L(kappa)-Vgpt* where 84% of the LVJ was replaced. The main difference is the resumption of strand bias characteristic of Ig hypermutation. Thus, whether or not a substrate sequence manifests strand bias in hypermutation is not inherently determined by the substrate DNA sequence. This indicates the presence of special element(s) within the LVJ that confer strand bias.

Animals↗
PubMed2000

The effect of lifestyle changes on results of 24-h ambulatory oesophageal pH monitoring.

OBJECTIVES: Twenty-four-hour pH monitoring is the 'gold standard' investigation of gastro-oesophageal reflux disease. It has been suggested that results may be influenced by lifestyle alteration during the study. The aim of this study was to determine the influence of lifestyle alteration and anxiety on outcome in pH monitoring. METHODS: One hundred consecutive patients attending for pH monitoring were recruited. Post-intubation anxiety was quantified (STAI form Y-1). On completion a questionnaire was recorded comprising six questions regarding alteration to eating/drinking (FOOD) (score 0-6) and six regarding posture and activity (ACTIVITY) (score 0-6). The higher the score the greater the alteration of lifestyle. A 10-point visual analogue scale (VAS) scored the degree of distress caused by the experience. pH studies were positive if exposure times at pH < 4 were greater than established normal values or if the Symptom Index was > 50%. Comparisons were made between patients with positive and negative studies regarding FOOD, ACTIVITY, VAS and STAI scores. RESULTS: Patients with positive pH studies had mean FOOD (2 vs 2; P> 0.05), ACTIVITY (2 vs 2; P> 0.05), VAS (3 vs 3; P> 0.05) and STAI (31 vs 32; P> 0.05) scores which were not significantly different from those of patients with negative pH studies. CONCLUSIONS: Lifestyle alteration was minimal in patients undergoing pH monitoring, nor was the experience distressing. There was no difference in lifestyle alteration and anxiety between those with positive and negative pH profiles.

Adolescent↗
PubMed1999

A frequent activated smoothened mutation in sporadic basal cell carcinomas.

Basal-cell carcinomas (BCCs) are the most common cancer in Caucasians. It has been reported that the patched gene is inactivated in 30-40% sporadic BCCs and 20% sporadic medulloblastomas via loss of heterozygosity and nonsense mutations. Recently, two activating smoothened mutations have been found in the sporadic basal cell carcinomas. One, at base pair 1604 (G-to-T transversion) of exon 9, changes codon 535 from tryptophan to leucine, and the other, at base pair 1685 (G-to-A transition) of exon 10, changes codon 562 from arginine to glutamine (Xie et al., 1998). In our study, 1604G-->T was found in 20 out of 97 (20.6%) sporadic BCCs. The high prevalence indicates that 1604G is the mutation hot spot in our tumor samples. This mutation was detected in all three histological subtypes of BCCs, suggesting that smoothened mutation is an early event during the development of the tumor. Our finding of a high smoothened mutation rate, together with high frequent patched gene mutations reported recently, indicates that activation of the hedgehog signal transduction pathway is the most common and early event in the development of sporadic BCCs. Additionally, to determine whether smoothened, like patched, is also involved in the carcinogenesis of medulloblastomas, we screened medulloblastoma samples for these two mutations by restriction analysis. We have found the 1604G-->T mutation in 1 out of 21 medulloblastomas. This result confirmed smoothened gene involvement in the carcinogenesis of medulloblastoma.

Basal Cell Carcinoma↗
PubMed1998

One-step 2-minute test to detect typhoid-specific antibodies based on particle separation in tubes.

Typhoid fever is caused by Salmonella typhi. Detection of anti-S. typhi antibodies in the patient is a useful diagnostic aid. Among the various methods developed over the years for this purpose, the Widal test, based on bacterial agglutination, has remained the most widely used, even though it is neither specific nor sensitive. Its popularity stems from the fact that it is simple to use and inexpensive. We describe a new test which also uses a simple one-step procedure but is more rapid and accurate than the Widal. The new test (TUBEX) detects anti-Salmonella O9 (both immunoglobulin M [IgM] and IgG) antibodies in patients by inhibiting the binding between an anti-O9 IgM monoclonal antibody (MAb) conjugated to colored latex particles and S. typhi lipopolysaccharide (LPS) conjugated to magnetic latex particles. The reactants are mixed in a specially designed microtube for 2 min, and the result is read based on the resultant color of the supernatant following forced sedimentation of the magnetic beads. In the absence of inhibitory antibodies, there is a color change (from blue to red) due to cosedimentation of the indicator particles with the magnetic particles, whereas if these antibodies are present, they prevent such a change to a degree dependent on their concentration. Preliminary examination of TUBEX using the anti-O9 MAb and irrelevant MAbs as inhibitors revealed the test to be specific and reproducible, with an analytical sensitivity of 16 micrograms per ml of antibody. The reagents remained stable for at least 9 months when kept at 4 degrees C. In the examination of 16 stored sera obtained from 14 patients with proven cases of typhoid fever and 78 serum samples from 75 subjects without typhoid fever, TUBEX was found to be 100% sensitive and 100% specific. The nontyphoid group comprised 26 healthy blood donors, 30 antinuclear antibody (ANA)-negative patients, 9 ANA-positive patients, of whom 1 was positive for anti-DNA antibody, 4 typhus patients, and 6 septicemic patients. In addition, the sera obtained from 11 patients clinically diagnosed as having typhoid fever were all positive in the test. The TUBEX results correlated to some extent, albeit insignificantly (r = 0.38, P = 0.07), with those of an enzyme-linked immunoassay (ELISA) which used a similar detection format (inhibition) and reagents (S. typhi LPS and anti-O9 antibody). TUBEX correlated very well with ELISAs which detected anti-S. typhi LPS IgM (r = 0.58, P = 0.003) or IgG (r = 0.54, P = 0.006) antibodies from the typhoid patients. There was no correlation with the Widal test. The TUBEX test, if performed on slides (instead of tubes) or with soluble antigen (instead of antigen-conjugated magnetic beads), suffered significantly in sensitivity. Direct agglutination tests using LPS-conjugated indicator particles performed either on slides or in microwells also failed to detect antibodies from the majority of typhoid patients. Thus, TUBEX appears to be well designed and well suited for use in the laboratory or by the bedside as a simple, rapid aid to the routine diagnosis of typhoid fever.

Antibodies, Bacterial↗
PubMed1997

Phage-displayed La/SS-B antigen as a diagnostic reagent.

BACKGROUND: The detection of antibodies to La/SS-B, a nuclear RNA-binding protein in mammalian cells, aids in the diagnosis of Sjogren's syndrome and systemic lupus erythematosus (SLE). This is performed conventionally by immunoprecipitation using a crude splenic extract and more recently, by the more sensitive and rapid enzyme-linked immunosorbent assay (ELISA) which uses a purified La/SS-B antigen. The latter antigen is obtained from cellular extracts of the antigen or from bacterial cell lysates containing the recombinant antigen usually by affinity chromatographic method. OBJECTIVE: To produce a La/SS-B antigen for use in ELISA that can be obtained easily and inexpensively without the need for extensive purification (including affinity chromatography). STUDY DESIGN: The antigen was produced as a fusion protein of the minor coat protein of M13 bacteriophage and used in this phage-associated form in an ELISA. La/SS-B cDNA derived from Hep-2 cells was cloned into the phagemid, pCANTAB-5E, and transfected to Escherichia coli. Phage clones selected for the presence of insert both by gene and antigenic analyses were used in the ELISA to detect anti-La/SS-B antibodies from patients with Sjogren's syndrome and SLE. RESULTS: A phage clone was obtained which contained a La/SS-B cDNA fragment truncated at the C-terminal end (after base-pair 631). The phage-displayed antigen derived from this clone was obtained by precipitation of the phage particles from the bacterial culture supernatant with polyethylene glycol. Used in the ELISA, this antigen detected 27 of 28 precipitin-positive sera and was negative for 50 control sera. The soluble (phage-free) form of the antigen was obtained from a nonsuppressor host as a cell lysate which could not be used in this form in an ELISA for antibody detection. It was useable, however, in Western blot analysis which confirmed the reactivity of the recombinant antigen. CONCLUSION: Phage-displayed antigens may be used in place of soluble forms of these antigens in detection assays which have the advantage that they are easy and inexpensive to produce.

Autoantigens↗
PubMed1997

Identification of a surrogate TSL-1 antigen of Trichinella spiralis from a phage library of random peptides.

We sought to find a peptide analogue of an important antigen (TSL-1) of Trichinella spiralis which is recognized by the 7C2C5 antibody. A phage library which displays a short (15-mer) randomly-generated peptide at the filament of the minor coat protein of the virion was used for selection by the 7C2C5 antibody. A peptide thus identified, ICDASGLGCWCWSLSP, was found to be a true surrogate since its binding to the antibody could be blocked by the native antigen and, conversely, an antiserum made to the peptide could recognize the native antigen. In addition, the peptide appeared to detect T. spiralis-infected pigs although it was less discriminatory than the native antigen.

Amino Acid Sequence↗