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Biomedical subjects

K W Lam

Publications and source records attributed to K W Lam.

At least 55 records · Page 3Linked to original sources
PubMed1990

Comparison of tartrate resistant acid phosphatase in a giant cell bone tumor and a spleen infiltrated with hairy cells.

Acid phosphatase (E.C.3.1.3.2) in a giant cell bone tumor and a spleen infiltrated with hairy cells was extracted by citrate buffer and then by 0.3 mol/L NaCl. The cationic acid phosphatase in the crude extract was isolated by CM-cellulose chromatography, and further separated by high pressure liquid chromatography. The majority of the tartrate resistant acid phosphatase in the hairy cell spleen was unabsorbed on CM-cellulose and was insensitive to iron. A much larger portion of the acid phosphatase in the bone tumor, than in the spleen, was cationic and was eluted from the column by 0.8 mol/L NaCl. The cationic acid phosphatase was further separated into consecutive peaks of acid phosphatases with different sensitivity to iron. A major portion of acid phosphatase in the giant cell bone tumor was enhanced by iron, while the amounts of iron-enhanced and iron-insensitive acid phosphatase were about the same in the spleen. The differences of the phosphatases in these two types of pathologic specimens indicate the occurrence of two types of enzymes with different biological significance.

Acid Phosphatase↗
PubMed1990

Ascorbate-enhanced copper toxicity on bovine corneal endothelial cells in vitro.

The present results indicate that 30 micrograms/ml copper was toxic to bovine corneal endothelial cells (BCEC) cultured in a serum-free medium (SFM) when the duration of treatment was 72 hours or more. Copper at 10 micrograms/ml, if mixed with 50 micrograms/ml ascorbate 2 to 3 hours before treatment, caused a transient decrease in the number of nuclei/mm2 at 72 hour, whereas 10 micrograms/ml copper alone was apparently non-toxic. When 10 micrograms/ml copper was added to 50 micrograms/ml ascorbate at the time of treatment, the toxicity was increased. All the treated cells failed to survive beyond 24 hours, and copper at a lower concentration of 1 microgram/ml could inhibit the proliferation of BCEC. We propose that copper toxicity on BCEC is augmented by ascorbate possibly through the increased replenishment of Cu+ and the subsequent enhanced production of free radicals by copper auto-oxidation.

Analysis of Variance↗
PubMed1989

Improved immunohistochemical detection of prostatic acid phosphatase by a monoclonal antibody.

A monoclonal antibody with high affinity to acid phosphatase isoenzyme 2 (Ab-AcP2) was selected to examine its binding to different normal and tumor tissues using the indirect immunohistochemical method. Both mature prostatic epithelial cells in the prostate and the highly dedifferentiated prostatic cancer cells in the bone marrow showed strong binding to the antibody. Among nonprostatic tissues, only bone marrow, breast, and kidney showed trace staining in some specimens. The specificity of Ab-AcP2 was much better than that of the polyclonal antibody to acid phospatase previously reported. When the antibody to the prostate-specific antigen (Ab-PSA) was used, weak background staining was often encountered, and weak to moderate stains were seen in the prostatic stroma, bone marrow, lung, skin, and melanoma.

Acid Phosphatase↗
PubMed1989

Generation of monoclonal antibodies to prostatic acid phosphatase isoenzyme 2 and application in solid-phase enzyme immunoassay.

Monoclonal antibodies specific to prostatic acid phosphatase (PAP) isoenzyme 2 were generated by using an improved hybridoma technique. After three subcutaneous immunizations and three intravenous boosters, cell fusion experiments were performed. The hybrid cells were first cultured in a semisolid medium containing methylcellulose and later transferred to a liquid medium for further subculture. Out of a total of 600 colonies recovered after two cell fusion experiments, 13 were shown to exhibit affinity to PAP isoenzyme 2 by radioimmunoassay. Nine hybrid cell lines which showed high affinity and specificity were established for further evaluation. Their immunoglobulin subclass was determined to be immunoglobulin G. The association constants between PAP isoenzyme 2 and each monoclonal antibody were determined by titration curve in radioimmunoassay (RIA). Three of them (PAP 1, PAP 03, and PAP 019) were shown to be over 1 X 10(9) M-1. From the results of a matrix cross-matching procedure, a pair of antibodies (PAP 03 and PAP 1) reacting with discrete antigenic determinants were identified for preparing a solid phase sandwich enzyme immunoassay (EIA) kit. The designed EIA procedure could be performed within 40 min in a one-stage incubation protocol. The assay time was shorter than that of other commercial RIA or EIA kits, and the sensitivity was 0.4 ng/ml which was comparable to that of RIA kits. The EIA kit was shown not to cross-react with human thyroid stimulating hormone, alpha-fetoprotein, carcinoembryonic antigen, and acid phosphatases derived from tissues other than prostate. Therefore, this design was a simple and rapid method with high sensitivity and specificity for determining PAP isoenzyme 2 in human serum.

Acid Phosphatase↗
PubMed1989

Degradation of bovine corneal collagen by alkali.

This study was undertaken to demonstrate the effect of alkali on the molecular size of collagen. Type I and V collagen from bovine cornea were incubated with different concentrations of NaOH at room temperature and 37 degrees C for various times. The samples were then neutralized and analyzed by high-pressure liquid chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Degradation of collagen molecules to a heterogeneous mixture of peptides with molecular weights less than 20,000 daltons occurred at concentrations greater than 0.25N, and the effect of alkali was much faster at 37 degrees C than at room temperature. In contrast, the molecular size of collagen was not affected by similar concentrations of HCl under similar conditions of time and temperature.

Alkalies↗
PubMed1988

Ascorbate peroxidase in bovine retinal pigment epithelium and choroid.

Peroxidase, catalyzing hydrogen peroxide reduction concurrent with ascorbate oxidation, was demonstrated in the extract of retinal pigment epithelium and choroid. The peroxidase in the choroid, RPE, and retina are 236.1, 25.1, and 0.5 units/mg protein respectively. Ammonium sulfate fractionation and high pressure liquid chromatography showed that the peroxidase in the RPE-choroid is associated with a group of heme proteins with absorption maxima at 410 nm, and optimal activity at pH 4.5. The high peroxidase activity in the RPE-choroid explains the observation of dehydroascorbate in these tissues and indicates a possible role of this enzyme in the removal of H2O2.

Ammonium Sulfate↗
PubMed1988

Study of prostaglandins and intraocular pressure in rabbits after Nd:YAG laser anterior segment procedures.

Radioimmunoassay of rabbit aqueous humor one hour after Nd:YAG laser anterior capsulotomies revealed increased concentrations of 6-keto-prostaglandin F1 alpha (PGF1 alpha), and thromboxane B2 (TBB2). Coincidental rises in intraocular pressure (IOP) and aqueous humor protein were observed. These changes correlated with the laser power used. Pretreatment with topical 1% prednisolone acetate or 1% indomethacin prior to anterior capsulotomies inhibited the post laser rise in PGF1 alpha and TBB2, but did not eliminate the post laser rise in IOP. Small rises in IOP and the concentrations of aqueous PGF1 alpha, TBB2, and protein were observed after Nd:YAG laser iridectomies. These changes were not affected by prior treatment with 1% topical prednisolone acetate. Although PGF1 alpha and TBB2 are elevated after Nd:YAG laser anterior segment procedures, a causal relationship between post treatment IOP rise and PGF1 alpha or TBB2 release could not be determined.

Animals↗
PubMed1988

Elevated tear IgG and conjunctival plasma cell infiltrate in a graft versus host disease patient.

Two patients developed graft versus host disease (GvHD) following allogeneic bone marrow transplantation for leukemia. One patient developed acute GvHD 12 days after transplantation, and the second developed chronic GvHD 100 days after transplantation. Tear analysis and conjunctival impression cytology were performed. The results were compared to a normal control and to another patient who was 18 months status post bone marrow transplantation with successfully treated GvHD. Tear sampling on the patient with chronic GvHD revealed greatly elevated IgG levels with an inverse IgG to IgA ratio, as compared with normals. In addition, two of three leukemia patients demonstrated decreased mucin levels in tears. Conjunctival impression cytology from the patient with chronic GvHD revealed an abundance of plasma cells.

Adult↗
PubMed1988

Preliminary characterization of a polymorphonuclear leukocyte stimulant isolated from alkali-treated collagen.

This study reports the preliminary characterization of a stimulant released from alkali-treated collagen which activates the respiratory burst of PMNs. The supernatant fraction from alkali-treated collagen (SATC) was precipitated with ammonium sulfate, resuspended, and centrifuged through a sucrose gradient (10-30%, W/V). Proteins were detected throughout the gradient but PMN stimulatory activity was found mainly in fractions 1 and 2 (bottom of gradient), indicating a very high molecular weight. When SATC was layered on a denser sucrose gradient (20-85%, W/V), protein was spread over the upper half of the gradient. The peak of PMN stimulatory activity was found in the middle of the gradient (fractions 8 and 9), indicating a molecular weight similar to the 200,000 dalton standard. High pressure liquid chromatography (HPLC) of fractions from this heavier sucrose gradient showed a small peak, followed by a much higher main peak. The small peak, with the highest molecular weight peptide, is responsible for stimulation of the respiratory burst of PMNs (fraction 8 and 9). The source of this stimulant is yet to be determined.

Alkalies↗
PubMed1988

Integrity of the blood-aqueous barrier in hyperuricemia.

Neither the origin nor the role of high levels of uric acid, found in the aqueous humor of some human glaucomatous eyes, is known. A model of hyperuricemia in the rat was evaluated for its contribution to urate levels in the aqueous humor. Epinephrine-induced hyperuricemia in male rats was shown to increase mean serum urate levels from 1.2 mg/dL to 6.7 mg/dL. Female rats were shown to have a lower uric acid baseline and a lower absolute hyperuricemic level than males. The mean aqueous-humor urate levels were increased from 0.06 mg/dL to 0.33 mg/dL for male rats. In our studies, the blood-aqueous barrier effectively prevented equilibration between the systemic and ocular compartments. The increased aqueous-humor uric acid levels in glaucoma may arise from intraocular mechanisms.

Animals↗
PubMed1988

The effect of YAG laser anterior capsulotomy on prostaglandin concentration in aqueous humor.

The YAG (yttrium aluminum garnet) laser has been recommended for anterior capsulotomies. One major complication is elevated intraocular pressure. We report a study of the biochemical content of the aqueous humor after a YAG laser anterior capsulotomy. We analyzed 6-keto-prostaglandin F1 alpha, thromboxane B2 and protein concentrations in the aqueous humor of human eyes. The average values of protein, 6-keto-prostaglandin F1 alpha, and thromboxane B2 in the control eyes were 81.3 +/- 14.0 mg/dL, 17 +/- 30 pg/mL, and 10 +/- 10 pg/mL, respectively. These values were elevated to 182.4 +/- 81.3 mg/dL, 401 +/- 55 pg/mL, and 576 +/- 148 pg/mL, respectively, after YAG laser anterior capsulotomy. The samples with a moderate level of 6-keto-prostaglandin F1 alpha (less than 300 pg/mL) had negligible changes of thromboxane. The elevation of thromboxane was obvious only when prostaglandin levels rose above 300 pg/mL.

6-Ketoprostaglandin F1 alpha↗
PubMed1988

Effect of Yag-laser anterior capsulotomy on the biochemical content of aqueous humor.

Elevation of prostaglandins and protein in the aqueous humor is an immediate response to Yag-laser anterior capsulotomy. There was a significant correlation between prostaglandins and thromboxane to the elevation of intraocular pressure. The elevation of protein concentration was proportional to the elevation of prostaglandin F1, but not to thromboxane. The biochemical changes in the aqueous humor were more severe as laser power was increased above 3 mj. While the role of prostaglandins in the elevation of intraocular pressure is uncertain, the present data recommend the selection of laser power below 3 mj to avoid pressure rise instead of inhibition of all prostaglandin synthesis by medication.

Aqueous Humor↗
PubMed1987

Molecular cloning of cDNA for human prostatic acid phosphatase.

A human liver cDNA library in lambda gt11 was screened with polyclonal antiserum to human acid phosphatase isoenzyme 2a/4. About eleven positive clones have been obtained. Two clones, lambda Hap21 and lambda Hap22 were further characterized: clone lambda Hap21 contained a 0.8-kb cDNA insert and clone lambda Hap22 a 1.8-2.0-kb insert. XbaI digestion of lambda Hap22 generated two fragments of 1.0 and 0.9 kb. BglII digestion resulted in a 1.2-kb fragment and several smaller fragments of undetermined size. Clone lambda Hap22 contained all the genes carried by lambda gt11(lac5cI857nin5Sam100) and the 2-kb insert. An Escherichia coli(lambda Hap22) lysogen was generated, and its acid phosphatase activity was approximately ten-fold higher than that in the control nonlysogenic lysate. Western-blot analysis of total proteins present in this E. coli(lambda Hap22) lysate revealed that the non-induced lambda Hap22 prophage directed the synthesis of an approx. 175-kDa protein. This protein was recognized by antibody to the human acid phosphatase isoenzyme 2a/4 and anti-beta-galactosidase and was produced only upon induction with IPTG. These results indicated that lambda Hap22 carried a major portion of the gene coding for the human acid phosphatase isoenzyme 2a and/or 4 and this protein fragment of acid phosphatase was sufficient to manifest enzymatic activity.

Acid Phosphatase↗
PubMed1987

Factors affecting ascorbate oxidation in aqueous humor.

Aqueous humor from human glaucomatous eyes was divided into two groups. Group I aqueous humor contains a negligible amount of ascorbate, while group II contains ascorbate. Group I aqueous humor stimulates ascorbate oxidation. This oxidative effect is inhibited by the addition of EDTA, indicating the presence of metallic ions. The amount of copper determined by atomic spectrophotometry in these samples was 1.54 micrograms/ml. On the contrary, group II aqueous humor protects against ascorbate oxidation. The small amount of albumin present in these samples accounts for part of this protective effect.

Aqueous Humor↗
PubMed1986

Retinyl palmitate hydrolase activity in the bovine retina.

Retinyl palmitate hydrolase (RPH) activity of bovine tissues was estimated from retinol formation following incubation of tissue homogenates with all-trans retinyl palmitate. The quantity of retinol produced in the incubation mixture was analyzed by high-performance liquid chromatography. RPH activities of retinal pigment epithelium (RPE), liver, retina, muscle and brain were 194.2, 138.0, 72.5, 25.0 and 5.1 units/gm protein respectively. The RPH activity in the retina was far above that attributable solely to RPE contaminations. The presence of RPH in the retina suggests that retina can utilize retinyl esters for the formation of visual pigments and/or cellular metabolism.

Animals↗
PubMed1986

Monoclonal antibody specific to acid phosphatase isoenzyme 4.

Prostatic acid phosphatase isoenzyme 4 was purified by ion exchange column chromatography, followed by high pressure liquid chromatography. The highly purified enzyme was used to produce monoclonal antibody from immunized BALB/c mice. The antibody was specific to isoenzyme 4, with negligible affinity to isoenzyme 2. The specificity of the monoclonal antibody was evaluated by Western blot analysis and by inhibition of radioimmunoassay. Immunohistochemistry method using the antibody to isoenzyme 2 showed heavy staining on the cell surface in contrast to the even staining throughout the cytoplasm when monoclonal anti-isoenzyme 4 was used. These results reflect the secretory nature of isoenzyme 2 and the non-secretory nature of isoenzyme 4.

Acid Phosphatase↗
PubMed1986

Immunoultrastructural demonstration of prostatic acid phosphatase isoenzyme 2 in prostatic carcinoma.

Human prostatic acid phosphatase isoenzyme 2 (HPAcP-2) was isolated from semen. This purified enzyme was immunized to rabbit to produce polyclonal antibodies. The specificity of the antibodies was tested by Western blot transfer method. Rabbit IgG-peroxidase conjugate was prepared from the antiserum and used to localize HPAcP-2 in prostatic carcinoma. It was found that in the tumor glandular acinus the normal basal cells were replaced by tumor cells containing reaction product. In the tumor cells, the reaction product was seen in the cisternae of rough endoplasmic reticulum (ER) and Golgi apparatus. The secretory vesicles which contained reaction product-stained granules and some amorphous material were seen to fuse with the apical plasma membrane and discharged their content into the glandular lumen. On the other hand, some secretory vesicles in the tumor cells facing to the basement membrane also discharged their similar content into the extracellular spaces. Reaction product-stained granules were found in the interstitial spaces surrounding the tumor cells. These findings suggest that HPAcP-2 is synthesized on the bound ribosomes and discharged into the cisternae of rough ER. The molecules are transported to the Golgi cisternae. After concentration and packaging, HPAcP-2 molecules are then transferred to the secretory vesicles, and discharged into the glandular lumen and to the extracellular spaces. The isoenzyme released in the extracellular space may reach the blood stream through the interstitial spaces or the lymphatic system, resulting in the elevation of serum HPAcPase level in some prostatic cancer patients.

Acid Phosphatase↗