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Biomedical subjects

H L Tan

Publications and source records attributed to H L Tan.

At least 73 records · Page 4Linked to original sources
PubMed1994

Hevein, a lectin-like protein from Hevea brasiliensis (rubber tree) is involved in the coagulation of latex.

Hevein, a lectin-like protein is the major protein of vacuolar structures called lutoids in the latex of rubber trees. We have shown both by in planta and ex planta studies that hevein is involved in the coagulation of latex by bringing together rubber particles. This polyvalent bridging between hevein and rubber particles is mediated by N-acetyl-D-glucosamine and involves a receptor glycoprotein of 22 kDa, which is localized on the surface of the rubber particles. The proposed role of hevein helps us to understand the mechanism of coagulation of latex and assigns a physiological intracellular function to one of the smallest lectins.

Acetylglucosamine↗
PubMed1994

Interleukin 4 increases type 5 acid phosphatase mRNA expression in murine bone marrow macrophages.

Type 5 acid phosphatase is a lysosomal enzyme expressed in cells of monocyte/macrophage lineage frequently used as a marker of osteoclastic differentiation. Oligonucleotide primers for DNA amplification were designed following sequence alignment of rat bone and human macrophage type 5 acid phosphatases. DNA (330 bp in length) obtained using these primers and reverse transcribed total cell RNA from in vitro generated murine osteoclastic cells was cloned and sequenced. DNA sequence analysis of two clones demonstrates that the amplified material was 91% and 96% identical to rat bone type 5 acid phosphatase at the nucleotide and amino acid level, respectively. Northern blots of murine tissue RNA show the presence of 1.5-kb transcripts that are most highly expressed in the long bones. Total cell RNA from the osteoclastic cells contain a marked level of type 5 acid phosphatase mRNA when compared to the levels seen in the tissue samples. Additionally, osteoclastic cell RNA contains two additional transcripts of 2.5 and 5 kb. Bone marrow macrophages grown in the presence of M-CSF express low levels of the 1.5-kb transcript with no signal observed for either of the two larger transcripts that were seen in the osteoclastic RNA samples. Importantly, bone marrow macrophage 1.5-kb type 5 acid phosphatase transcript levels are increased by interleukin 4 treatment in both a time and concentration-dependent manner. These findings indicate that type 5 acid phosphatase, while a cytochemical marker for osteoclasts, can be induced in macrophages by agents that block in vitro osteoclastic differentiation. Increased type 5 acid phosphatase may play a role in interleukin 4-stimulated monocyte activities.

Acid Phosphatase↗
PubMed1994

Contribution of mechanical activity and electrical activity to cellular electrical uncoupling in ischemic rabbit papillary muscle.

We separated mechanical and electrical activity during ischemia and we assessed the contribution of each of these variables to ischemia induced cellular electrical uncoupling. In two groups of rabbit papillary muscles, mechanical and electrical activity at the onset of ischemia was abolished. We determined how this affected the time of onset of uncoupling. Also, action potential duration (APD80), conduction velocity and extracellular pH were recorded. In the first group, the BDM group, mechanical arrest was achieved with 10 mM BDM (2,3-butanedione monoxime). This had no effect on APD80 and conduction velocity. In the second group, the quiescent group, electrical as well as mechanical arrest was obtained by not stimulating the muscles. The effect of electrical arrest was estimated as the difference between this group and the BDM group. Mechanical arrest delayed uncoupling significantly (control: 14.6 +/- 0.9 (min), mean +/- S.E.M., n = 10; BDM: 19.3 +/- 1.1, n = 10; P < 0.025). Electrical arrest caused an even greater delay (quiescent: 41.6 +/- 4.6, n = 7; P < 0.001 v control and BDM). In the BDM group, these effects were not associated with a difference of APD80 during ischemia, but with a smaller decrease of conduction velocity. Delay of uncoupling correlated with delay of extracellular acidosis. We conclude: (1) mechanical and electrical activity both are significant determinants of ischemia induced uncoupling; (2) delay of uncoupling is associated with delay of extracellular acidosis, indicating a reduced rate of metabolism.

Action Potentials↗
PubMed1994

A review of pediatric cases of urinary tract calculi.

A retrospective review was performed of the records of 93 children with urinary tract calculi treated during a 10-year period (1983 to 1992). Two distinct groups of patients were identified. One group comprised 63 Aboriginal children, mainly from the northern region of Western Australia, who presented at a young age (average, 2.8 years) with urinary tract infections and failure to thrive. The other group comprised of 30 non-Aboriginal children, mainly from the metropolitan region, who presented at an older age (average, 6.7 years) with abdominal and flank pain. Calculi were located in the upper urinary tract in 85 children (91.4%) and in the bladder in eight (8.6%). Most patients underwent open surgical procedures for removal of their calculi, and the overall success rate was 86%. In recent years, percutaneous nephrolithotomy has been used successfully in selected patients with renal pelvic calculi. This method seems to be the trend for the future.

Adolescent↗
PubMed1994

Thoracoscopic biopsy of a pathological vertebral body.

When computed tomography (CT) guided biopsy of a pathological vertebral body fails, the accepted alternative has been open thoracotomy and biopsy. The authors wish to report the use of a thoracoscopic approach to biopsy of a pathological thoracic vertebra in a child, after an unsuccessful attempt at CT guided percutaneous biopsy.

Acute Disease↗
PubMed1994

The role of laparoscopic surgery in children.

Although laparoscopy was widely evaluated by Drs Stephen Gans and George Berci in the 1970's, it did not become a useful clinical modality in paediatric surgery until relatively recently. It was reported by these workers to be a useful diagnostic modality, but as a therapeutic tool, its role was limited to simple biopsies. The introduction of computed tomography (CT) scan abdominal ultrasound imaging also limited its clinical application, as these modalities allowed very accurate diagnosis of clinical pathology without resorting to laparoscopy for diagnosis. Recently, the widespread acceptance of laparoscopic surgery in adult surgical practice has lead many centres including ours to evaluate the role of laparoscopic surgery in paediatrics. Although it will be some time before its benefits or otherwise in paediatrics are established, there is no doubt that laparoscopic surgery is safe in children in the hands of the skilled laparoscopist. There is also no doubt that the paediatric surgeon in future will have to be skilled in laparoscopic surgery as increasing demands are made on us to treat common surgical conditions without causing the patient physical harm. This paper reviews some of the common conditions being treated by the laparoscopic route in our institution and attempts to evaluate the early results.

Abdominal Pain↗
PubMed1993

Extracellular-matrix degradation at acid pH. Avian osteoclast acid collagenase isolation and characterization.

Osteoclasts degrade bone matrix, which is mainly type I collagen and hydroxyapatite, in an acidic extracellular compartment. Thus we reasoned that osteoclasts must produce an acid collagenase. We purified this enzyme, a 31 kDa protein, from avian osteoclast lysates (in 100 mM acetate/1 mM CHAPS/1 mM dithiothreitol, pH 4.4), fractionated by (NH2)2SO4 precipitation, gelatin-affinity, cation exchange, and gel filtration. Fraction activity was measured using diazotized collagen or 3H-labelled cross-linked collagen (decalcified and trypsin-treated metabolically L-[4,5-3H]proline-labelled bone) as substrates. Iodoacetate, leupeptin, antipain, pepstatin and mercurials inhibited collagenolysis by the isolated proteinase; mercurial derivatives could not be re-activated by dithiothreitol. Collagen degradation was maximal at pH 4.4; purified proteinase reproduced the collagenolytic activity of cell lysates. The N-terminal amino acid sequence from the isolated protein and its CNBr degradation fragments showed sequence similarity to mammalian cathepsin Bs, and near-identity with avian liver cathepsin B. Peptide substrate specificity of the osteoclastic enzyme resembled those of mammalian cathepsin B and its avian liver counterpart, but degradation of low-molecular-mass substrates by the osteoclastic enzyme was slower, reflecting generally lower kcat. values. Further, kcat/Km varied less between arginine-containing substrates than for previously reported cathepsin Bs, indicating different substrate specificity of the osteoclast enzyme. Polyclonal antibody raised to a 25 kDa fragment of the enzyme recognized a single 31 kDa band in SDS/PAGE of osteoclast lysates blotted to poly(vinylidene difluoride), adsorbed collagenolytic activity of osteoclast lysates, and stained avian osteoclasts in tissue sections. Degenerate sense- and antisense-oligonucleotide primers, predicted from segments of primary amino acid sequence, amplified a 486 bp DNA fragment; this was cloned and sequenced. Of 162 amino acids encoded, 77% are identical with those of human cathepsin B; hybridization identified a 2.4 kb RNA in osteoclast lysates. We conclude that the major avian osteoclast collagenolytic enzyme is a cathepsin B, whose activity varies from other enzymes of its class.

Amino Acid Sequence↗
PubMed1993

1,25-Dihydroxyvitamin D3 increases type 1 interleukin-1 receptor expression in a murine T cell line.

The biologically active metabolite of vitamin D3, 1,25 (OH)2 D3, exerts important immunoregulatory effects in addition to being a central mediator of calcium/phosphate metabolism. Utilizing an interleukin 1 responsive murine T cell line and 125I-interleukin 1 alpha, we show that 1,25 (OH)2 D3 (5,50 nM) enhanced 125I-interleukin 1 alpha binding up to almost 2-fold over control. This 1,25 (OH)2 D3 effect occurred in a dose-dependent manner and was detectable after 24 h but not before 7 h of culture. Scatchard analysis of 125I-interleukin 1 alpha binding data demonstrated that 1,25 (OH)2 D3 enhanced interleukin 1 receptor number without a significant change in affinity. The biologically less potent metabolite of vitamin D3, 25 (OH) D3, also augmented 125I-interleukin 1 alpha binding but at steroid levels 2-3 log orders greater than 1,25 (OH)2 D3. This observation, combined with the presence of high-affinity 3H-1,25 (OH)2 D3 receptors (88 sites/cell, K = 0.45 nM) in cytosolic extracts, strongly suggests that the nuclear vitamin D receptor mediates this steroid's effect on interleukin 1 receptor expression. Based on the capacity of an anti-type 1 interleukin 1 receptor monoclonal antibody (35F5) to block 1,25 (OH)2 D3-enhanced 125I-interleukin 1 alpha binding, we conclude that this steroid augments type 1 interleukin 1 receptor expression. When combined with interleukin 1, a cytokine that also impacts MD10 interleukin 1 receptor expression, 1,25 (OH)2 D3 enhanced interleukin 1 receptor expression. Northern blots hybridized with a 32P-type 1 interleukin 1 receptor cDNA probe show that 1,25 (OH)2 D3 enhanced type 1 interleukin 1 receptor steady state mRNA levels. Functionally, 1,25 (OH)2 D3 pretreatment augmented the MD10 proliferative response to suboptimal levels of interleukin 1 (< 100 fM interleukin 1 alpha). These findings further support 1,25 (OH)2 D3's role as an immunoregulatory molecule and provides a possible mechanism by which this steroid could potentiate certain immune activities.

Animals↗
PubMed1993

Murine osteoblast interleukin 4 receptor expression: upregulation by 1,25 dihydroxyvitamin D3.

The immune cytokine interleukin 4 has newly recognized effects on skeletal metabolism. While the interaction of many cells ultimately determines bone mass, we have examined the possibility that the osteoblast may be an IL-4 target in bone by characterizing IL-4 receptor (IL-4R) expression by MC3T3-E1 (MC3T3) murine osteoblastic cells. Based on 125I-IL-4 binding, MC3T3 cells express large numbers of IL-4 receptors (125I-IL-4 Bmax = 3,000-7,500 sites/cell, 125I-IL-4 K = 13-40 pM) with an affinity similar to the IL-4 receptor expressed by an IL-4-responsive T cell line. Monoclonal anti-IL-4R antibodies (M1) blocked specific MC3T3 125I-IL-4 binding and MC3T3 total cell RNA contained full-length IL-4R mRNA as detected by reverse transcription DNA amplification utilizing IL-4R primers and Northern blot analysis. Functionally, IL-4 treatment of MC3T3 cells resulted in increased cellular proliferation (10-20%) and inhibition of alkaline phosphatase levels (20-40%). While parathyroid hormone (PTH) exposure did not influence IL-4R levels, vitamin D3 treatment augmented MC3T3 125I-IL-4 binding, in a time-dependent manner, up to threefold after a 24 h exposure with a metabolite specificity indicating the involvement of the vitamin D receptor. Equilibrium binding studies showed that the impact of 1,25 (OH)2 D3 on MC3T3 125I-IL-4 binding was due to an increased IL-4R Bmax. Cycloheximide treatment inhibited 1,25 (OH)2 D3-induced IL-4R upregulation, suggesting that protein synthesis was required. Furthermore, the steroid increased steady-state IL-4R mRNA levels in both a time- and concentration-dependent manner. The IL-4R message half-life was not altered by 1,25 (OH)2 D3, suggesting that increased IL-4R mRNA expression resulted from increased IL-4R gene transcription. Taken together, these findings raise the possibility that IL-4's influence on mineral metabolism could be mediated by osteoblasts and that the effectiveness of this cytokine may be influenced by vitamin D3's impact on IL-4R expression.

Antibodies, Monoclonal↗
PubMed1993

R 56865 delays cellular electrical uncoupling in ischemic rabbit papillary muscle.

The compound R56865 protects the heart from irreversible ischemic damage. The proposed mechanism of its action is a reduction of Ca2+ overload secondary to a reduction of intracellular Na+, caused by blockade of the Na(+)-channel. In addition, cardioprotection is ascribed to blockade of the Na(+)-sensitive K(+)-channel (IK-Na). We tested whether R 56865 delays cellular electrical uncoupling, one aspect of irreversible ischemic damage that is due to Ca2+ overload. Also, we studied whether the Na(+)-channel and IK-Na are involved in cardioprotection by relating delay of the onset of cellular electrical uncoupling to changes of conduction velocity and action potential duration (APD80), respectively. Experiments were performed with isolated perfused rabbit papillary muscles that were treated with 1 microM R 56865 for 45 min prior to ischemia. Uncoupling started at 15.0 +/- 0.8 min (mean +/- S.E.M., n = 12) of ischemia in the control group and at 23.4 +/- 1.7 min in the R 56865 group (n = 9, P < 0.005 vs control). R 56865 tended to decrease conduction velocity and to increase APD80 during pre-treatment, but these changes were not statistically significant. During ischemia, conduction velocity was statistically not different between the R 56865 group and the control group. APD80 was significantly longer in the R 56865 than in the control group during the first 7 min of ischemia and similar after that. We conclude that R 56865 delays the onset of cellular uncoupling during ischemia and that this effect is not related to changes of conduction velocity and at most in part to changes of APD80.

Action Potentials↗
PubMed1993

Ischaemic preconditioning delays ischaemia induced cellular electrical uncoupling in rabbit myocardium by activation of ATP sensitive potassium channels.

OBJECTIVE: The aim was to examine whether ischaemic preconditioning delays the onset of cellular electrical uncoupling during ischaemia, and whether the effect of preconditioning is mediated by the activation of ATP sensitive K+ channels (IK-ATP). METHODS: Onset of uncoupling, action potential duration (APD80), and conduction velocity were measured in an isolated perfused rabbit papillary muscle. Preconditioning consisted of 10 min occlusion and 10 min reperfusion prior to 40 min sustained ischaemia. Five groups were studied: (1) control (sustained ischaemia only); (2) preconditioning; (3) preconditioning with 20 microM glibenclamide, a blocker of IK-ATP, added for 10 min during the reperfusion period; (4) sustained ischaemia after 15 min perfusion with 20 microM cromakalim (BRL 34915), an opener of IK-ATP; (5) sustained ischaemia after 10 min perfusion with 20 microM glibenclamide without preconditioning. RESULTS: Uncoupling started at 15.0(SEM 0.7) min of ischaemia in the control group and at 22.8(1.5) min after preconditioning (p < 0.001 v control group). Blocking IK-ATP during the preconditioning protocol with glibenclamide abolished the delay of uncoupling: onset was at 14.7(1.2) min. Activation of IK-ATP with cromakalim resulted in uncoupling at 23.3(1.9) min (p < 0.002 v control). Glibenclamide without preconditioning had no effect on uncoupling: onset was at 15.6(1.0) min. APD80 during ischaemia was significantly shorter in the preconditioning and cromakalim groups than in the control group from 5 min of ischaemia onward. In the preconditioning+glibenclamide group and the glibenclamide group APD80 was at no point significantly different from the control group. Conduction velocity during ischaemia decreased to about 70% of baseline after 10 min and was not different between the five groups. CONCLUSIONS: (1) Preconditioning delays the onset of electrical uncoupling; (2) the protective effect of preconditioning may be caused by activation of the IK-ATP channel; (3) the protective effect is associated with reduction of action potential duration, but not with changes of conduction velocity.

Action Potentials↗
PubMed1993

Endopyelotomy for pelvi-ureteric junction obstruction in children.

Endopyelotomy or percutaneous pyeloplasty has become an attractive alternative to open pyeloplasty in adult patients in recent years. However, the experience and reports of this procedure in the paediatric population have been limited to date. We report our experience in paediatric patients undergoing this procedure. Seventeen children with primary (14) or secondary (3) pelvi-ureteric junction (PUJ) obstruction were consented for endopyelotomy over a 3-year period. Patient age ranged from 4 months to 16 years, and the male to female ratio was 11:6. Percutaneous nephrostomy was established in 5 patients prior to endopyelotomy either for diagnosis or for drainage of an acutely obstructed upper tract. The remaining 12 had endopyelotomy as a one-stage procedure. The average duration of the procedure was 103 min (range 60-150 min) and the average hospital stay was 5 days (range 3-9 days). Follow-up averaged 15 months (range 3-36 months). Endopyelotomy was successful in 10/13 patients (77%), unchanged in 1, and failed in the remaining 2. We believe that percutaneous pyeloplasty can be safely and successfully performed in children with primary and secondary PUJ obstruction.

Adolescent↗
PubMed1993

IL-1-induced murine osteoblast IL-6 production is mediated by the type 1 IL-1 receptor and is increased by 1,25 dihydroxyvitamin D3.

IL-1-induced osteoblast IL-6 production represents one possible mechanism by which IL-1 augments bone resorption. In this report, we show that the murine osteoblastic cell line (MC3T3-E1) expresses type 1 IL-1 receptors based on 125I-HrIL1 alpha binding, blocked by type 1 IL-1R antibodies (35F5), and analysis of MC3T3 RNA by reverse transcription (RT)-DNA amplification and Northern analysis. MC3T3 cells do not express detectable type 2 IL-1R mRNA by RT-DNA amplification. IL-1 induces (IL-1 ED50, 0.1 pM) IL-6 production through the type 1 IL-1R as 35F5 antibodies block IL-1-stimulated IL-6 production. Vitamin D3 increases IL-1R expression dose- and metabolite-dependently, with 1,25-(OH)2D3 having the greatest potency, and also enhances IL-1's capacity to stimulate IL-6 production at low IL-1 levels. Both IL-1 and 1,25-(OH)2D3 induce type 1 IL-1R and not type 2 IL-1R upregulation based on ligand binding and RT-DNA amplification. Increased IL-1R expression requires a 5-7-h treatment and is protein/RNA synthesis dependent. These observations imply that IL-1-induced IL-6 production in osteoblasts is mediated by type 1 IL-1Rs and that increased IL-1R expression could play a role in mediating IL-1-induced skeletal responses.

3T3 Cells↗
PubMed1992

Reversible inhibition of osteoclastic activity by bone-bound gallium (III).

Gallium(III) is a new therapeutic agent for hypercalcemia. Ga3+ reduces osteoclast action, but how it inhibits the cell's physiology is unknown. In vivo, 7-12 microM Ga(III) reduces calcium release from bone, but surprisingly, 10-100 microM Ga3+ added to isolated avian osteoclasts did not reduce their degradation of L-(5-3H)-proline bone. 3H-proline labels bone collagen specifically, and collagenolysis is an excellent indicator of bone dissolution because collagen is the least soluble component of bone. Ga(III) greater than 100 microM inhibited osteoclasts in vitro, but also killed the cells. To resolve this apparent conflict, we measured 67Ga distribution between bone, cells, and media. Gallium binds avidly but slowly to bone fragments. One hundred micrograms of bone clears 60% of 1 microM gallium from 500 microliters of tissue culture medium, with steady state at greater than 24 h. Osteoclasts on bone inhibited gallium binding capacity approximately 40%, indicating a difference in available binding area and suggesting that osteoclasts protect their substrate from Ga binding. Less gallium binds to bone in serum-containing medium than in phosphate-buffered saline; 30% reduction of the affinity constant suggests that the serum containing medium competes with bone binding. Consequently, the effect of [Ga] on bone degradation was studied using accurately controlled amounts of Ga(III) pre-bound to the bone. Under these conditions, gallium sensitivity of osteoclasts is striking. At 2 days, 100 micrograms of bone pre-incubated with 1 ml of 1 microM Ga3+, with 10 pmoles Ga3+/micrograms bone, was degraded at 50% the rate of control bone; over 50 pM Ga3+/micrograms bone, resorption was essentially zero. In contrast, pre-treatment of bone with [Ga3+] as high as 15 microM had no significant effect on bone resorption rate beyond 3 days, indicating that gallium below approximately 150 pg/micrograms bone acts for a limited time and does not permanently damage the cells. We conclude that bone-bound Ga(III) from medium concentrations less than 15 microM inhibits osteoclasts reversibly, while irreversible toxicity occurs at solution [Ga3+] greater than 50 microM.

Animals↗
PubMed1991

Single-increment dilatation for percutaneous renal surgery: an experimental study.

Renal nephrostomy track dilatation is an integral part of percutaneous renal surgery. The traditional method has employed sequential dilators of increasing size, or balloon dilatation. This study used a canine model to investigate the effects of single-increment renal track dilatation to 24 F and compares it with the conventional techniques. Single-increment dilatation proved to be a safe technique with minimal haemorrhage or parenchymal damage and healing at 6 weeks by a fine linear scar. It was as safe as conventional techniques. We believe this study proves that single-stage dilatation is a safe technique for use in humans.

Animals↗
PubMed1991

Passive chloride permeability charge coupled to H(+)-ATPase of avian osteoclast ruffled membrane.

We prepared proton-transporting membrane vesicles from the avian osteoclast's ruffled membrane, a specialized region of the cell surface that acidifies the bone resorption space. We demonstrated a unique conductive Cl- permeability that is charge coupled to the vesicle H(+)-ATPase and is required for acidification. Ion replacement indicated an anion selectivity of Br- approximately Cl- greater than SO4(2-) greater than NO3- approximately SCN- in supporting acidification. The anion channel blocker 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (10 microM) was a competitive inhibitor of acidification and raised the Michaelis constant for ATP of the proton pump approximately 11-fold in 120 mM KCl. Inhibition was reversed by valinomycin, which provides an alternate path for charge neutralization. The Cl- dependence of acidification was nonlinear and yielded a Hill coefficient of 3-4, showing that it is distinct from a linear Cl- dependence reported for acidification of renal cortical endosomes. The K+ ionophore valinomycin augmented H+ transport in K2SO4, and not in KCl. Dependence of Cl- transport on membrane potential was confirmed by direct measurement of 36Cl- transport. We uncoupled charge transport from proton transport with a large excess of ammonia, which had no effect on 36Cl- accumulation in vesicles, and by measuring 36Cl- accumulation in response to a membrane diffusion potential, produced with a [K+] gradient and valinomycin in the absence of ATP. These experiments demonstrate that the electrogenic proton pump of the osteoclast ruffled membrane is charge coupled to a passive Cl- permeability in the same membrane.

Acridine Orange↗