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Histochemical patterns of human T lymphocyte subpopulations with nonspecific esterase staining.

Alpha-naphthyl-acetate-esterase (ANAE) activity was histochemically demonstrated in human T lymphocytes and their subpopulations. Examination of T lymphocytes bearing Fc receptors for IgM showed the presence of circumscribed deposits of esterase activity, while T cells with FC receptors for IgG showed of predominantly diffuse granular pattern. The difference in the topography of the enzymatic activity was shown to be related to cellular activation resulting from exposure to IgG-sensitized bovine erythrocytes or to strong polyclonal activation by lectin mitogens. The ANAE stain also characterized monocytes, which exhibited an intense nongranular cytoplasmic stain. Non-T cells showed no detectable ANAE activity when tested by this method.

Animals↗

Hydrolysis of prednisolone succinate by esterase in rabbit ocular tissue.

Prednisolone concentrations in serum after both subconjunctival and intravenous injections of prednisolone sodium succinate at a 1 mg/kg equivalent dose of prednisolone in rabbits were analyzed by regularly phased high-performance liquid chromatography. The systemic availability after subconjunctival injection was determined to be almost 1.0 by comparing areas under the serum concentration-time curves for both administration routes. The initial hydrolytic rate of prednisolone succinate ester was dependent on the amount of total protein in ocular tissue homogenates and fluids as well as that in serum. The rate followed Michaelis-Menten kinetics, with an almost consistent Michaelis constant. Although the maximum hydrolytic rate per unit amount of tissue protein was the highest in conjunctival tissue, followed by corneal tissue and serum, the esterase activity corrected by total protein in the whole rabbit body should be the highest in serum, followed by conjunctival tissue. These results indicate that prednisolone sodium succinate administered subconjunctivally is hydrolyzed rapidly by esterase and absorbed almost completely into the systemic circulation.

Animals↗

Developmental changes in hepatic esterase activity towards chloramphenicol succinate and its Michaelis-Menten constant of liver, kidney and lung in human.

The hepatic esterase activities towards chloramphenicol succinate were determined in the tissues from 45 human subjects, including 18 electively aborted fetuses, 5 premature and 8 full-term newborn babies, 8 infants and 6 adults. The enzyme activities in the tissues from the fetuses and neonates were significantly lower compared with those obtained from the infants and adults. This suggests that the activity showed postnatal development. Kinetic studies of the esterase activity revealed that hepatic Km values were similar to those of the lung, but renal Km values were about 3 times higher than those of the liver and lung. In each organ, no age-related changes in Km values were observed. However, all the Vmax values in each organ showed developmental increases. Therefore, the liver plays the most important role in hydrolysis of chloramphenicol succinate when the weight of the organ is taken into consideration.

Adult↗

Beta-VLDL-induced cholesterol ester deposition in macrophages may be regulated by neutral cholesterol esterase activity.

We examined the role of enzyme activities involved in cholesterol ester metabolism in the accumulation of cholesterol ester in macrophages. When [3H]cholesterol linoleate-beta-very low density lipoproteins (beta-VLDLs) were incubated with rat resident peritoneal macrophages (RPMs), thioglycolate-elicited macrophages (TEMs), or alveolar macrophages (AMs), cholesterol ester accumulation was the greatest in TEMs and the least in AMs. The uptake of [3H]cholesterol linoleate-beta-VLDL into AMs was four times that into RPMs, and the uptake into TEMs was twice that into RPMs. The [3H]cholesterol released from [3H]cholesterol linoleate-beta-VLDL-loaded AMs was five times higher than from TEMs and RPMs, whereas those from RPMs and TEMs were almost the same. In studies using cell homogenates, the acid cholesterol esterase activity in AMs was 1.7 times that in TEMs and six times that in RPMs. The acyl-coenzyme A:cholesterol acyltransferase (ACAT) activities in AMs and TEMs were similar but were higher than that in RPMs. The neutral cholesterol esterase activity was seven times higher in AMs than in RPMs and TEMs. In the study using intact cells, the hydrolysis of loaded [3H]cholesterol linoleate-beta-VLDL in the presence of the ACAT inhibitor CL277,082 and the cholesterol [14C]oleate synthesis from [14C]oleate were enhanced in AMs about twofold compared with RPMs and TEMs. The reduction of de novo synthesized cholesterol [14C]oleate in the presence of CL277,082 was enhanced in AMs four times compared with TEMs or RPMs.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

In vitro tissue potencies of converting enzyme inhibitors. Prodrug activation by kidney esterase.

The inhibition of angiotensin converting enzyme by ramipril, ramiprilat, enalapril, enalaprilat, and captopril was studied in the plasma and various tissues (lung, heart, renal cortex, renal medulla) of normotensive rats and spontaneously hypertensive rats. Displacement curves for [3H]ramiprilat were established on each tissue with the converting enzyme inhibitors, and their potencies were expressed as the concentration that inhibited 50% of the specific [3H]ramiprilat binding. In the plasma, lung, and heart, the order of activities was: ramiprilat greater than enalaprilat greater than captopril greater than ramipril greater than enalapril. This order was different in the kidney (cortex and medulla): ramiprilat greater than enalaprilat greater than ramipril greater than captopril greater than enalapril. For ramiprilat, enalaprilat, and captopril, there were no differences in their respective potencies between tissues or between rat strains. However, the two prodrugs ramipril and enalapril were 10-30 times more active in the kidney than in the other tissues in both groups of rats. This was due to the deesterification of the prodrugs: in the presence of an esterase inhibitor (diethyl nitrophenyl phosphate, 10 microM), the potencies of ramipril in the kidney were not different from that obtained in the lung, which was not affected by the presence of the esterase inhibitor. These results suggest that the variations in the tissue activities of an angiotensin converting enzyme inhibitor are probably not due to differences in tissue affinities of the angiotensin converting enzyme inhibitor but depend on the concentration of this angiotensin converting enzyme inhibitor in each tissue.

Angiotensin-Converting Enzyme Inhibitors↗

Biodegradation of a dental composite by esterases: dependence on enzyme concentration and specificity.

Studies have shown that inflammatory (cholesterol esterase, CE) and salivary (pseudo-cholinesterase, PCE) enzymes can cause the breakdown of bisphenol-A diglycidyl dimethacrylate (bisGMA) and triethylene glycol dimethacrylate (TEGDMA) components from composite resins. Based on the above consideration, it was desired to show how CE- and PCE-catalyzed hydrolysis of resin components was dependent on the enzymes' concentration and to determine their distinct specificities (if any) towards resin components. Photopolymerized model composite resin samples (60% weight fraction silanated barium glass filler) based on bisGMA and TEGDMA monomers (55/45 weight ratio of the matrix, respectively) were incubated with PBS and either 0.01, 0.05, 0.1 or 1 unit/ml of CE or PCE for 16 days (pH 7.0, 37 degrees C). Incubation solutions were analyzed by high-performance liquid chromatography (HPLC), UV spectroscopy and mass spectrometry. The composite samples were characterized by scanning electron microscopy (SEM). Degradation rates of bisGMA and TEGDMA monomers were assessed. The results showed that CE had a greater specificity towards cleaving bisGMA while PCE showed a greater specificity towards TEGDMA. A strong enzyme concentration dependence was observed which suggests that the level of degradation products generated for a material will depend on the esterase make-up of an individual's saliva in combination with the specific formulation of monomer components used.

Biodegradation, Environmental↗

Development of gamma G, gamma A, gamma M, beta IC-beta IA, C 1 esterase inhibitor, ceruloplasmin, transferrin, hemopexin, haptoglobin, fibrinogen, plasminogen, alpha 1-antitrypsin, orosomucoid, beta-lipoprotein, alpha 2-macroglobulin, and prealbumin in the human conceptus.

The synthesis of gammaG, gammaA, gammaM, beta(1C)/beta(1A), C'1 esterase inhibitor, ceruloplasmin, transferrin, hemopexin, haptoglobin, fibrinogen, alpha(1)-antitrypsin, orosomucoid, beta-lipoprotein, alpha(2)-macroglobulin, and prealbumin was studied in 15 normal human embryos and fetuses of 29 days to 18 wk gestation and in the yolk sacs of four embryos from 5.5 to 11.5 wk gestation using tissue culture in (14)C-labeled amino acids followed by radioimmunoelectrophoresis. The human embryo as early as 29 day gestation synthesized beta(1C)/beta(1A), C'1 esterase inhibitor, transferrin, hemopexin, alpha(1)-antitrypsin, beta-lipoprotein, alpha(2)-macroglobulin, and prealbumin in culture. At 32 days gestation ceruloplasmin and orosomucoid were also synthesized, but synthesis of fibrinogen was not observed before 5.5 wk. Synthesis of gammaM occurred as early as 10.5 wk gestation, and gammaG synthesis was found in cultures as early as 12 wk gestation; gammaA synthesis was not detected in any of the tissue cultures. With the exception of the gamma-globulins, each of the proteins studied was synthesized by the liver, but additional sites of synthesis for some of these proteins were also found. Synthesis of gammaG and gammaM occurred primarily in the spleen, but other sites of synthesis were noted as well. Changes in the concentrations of most of these proteins and plasminogen in embryonic and fetal serum from 5.5 to 41 wk gestation, in amniotic fluid from 6.5 to 38 wk gestation, and in the sera of neonates during the 1st 3 wk postpartum are described. Although gammaA, gammaM, ceruloplasmin, or haptoglobin were not detectable in some of the embryonic and fetal sera, gammaA and ceruloplasmin were both present as early as 6.5 wk gestation, haptoglobin by 9.5 wk gestation, and gammaM by 17 wk gestation. Each of the other proteins were present in all of the sera examined.

Abortion, Therapeutic↗

Esterase activity in various Candida species.

The aim of the present study is to ascertain esterase activities of various species of Candida. A total of 125 strains isolated and identified by conventional methods were tested for esterase activity using the Tween 80 opacity test. Our results showed that 58 of 59 strains of Candida albicans, all of the Candida tropicalis strains (n = 38) and all of the Candida guilliermondii strains (n = 3) produced positive results, whereas the remaining Candida species did not. The Tween 80 opacity test is a useful method because it is simple, economical and easy to prepare and interpret.

Candida↗

Salivary esterase activity and its association with the biodegradation of dental composites.

Pseudocholinesterase (PCE) and cholesterol esterase (CE) can hydrolyze bisphenylglycidyl dimethacrylate (bisGMA) and triethylene glycol dimethacrylate (TEGDMA) monomers. This study will test the hypothesis that enzyme activities showing CE and PCE character are found in human saliva at levels sufficient to hydrolyze ester-containing composites important to restorative denstistry. The study also seeks to ask if the active sites of CE and PCE with respect to composite could be inhibited. Photo-polymerized model composite resin was incubated in PCE and CE solutions, in the presence and absence of a specific esterase inhibitor, phenylmethylsulfonyl fluoride (PMSF). Incubation solutions were analyzed for resin degradation products by high-performance liquid chromatography (HPLC), UV spectroscopy, and mass spectrometry. Saliva was found to contain both hydrolase activities at levels that could degrade composite resins. PMSF inhibited the composite degradation, indicating a material hydrolysis mechanism similar to the enzymes' common function.

Analysis of Variance↗

Comparisons of alveolar and peritoneal macrophages: soluble protein, esterase, dipeptidyl aminopeptidase II, and proteinase inhibitor.

The content of soluble protein, nonspecific esterase, dipeptidyl aminopeptidase II (DAP II), and proteinase inhibitors was compared for alveolar (AM) and peritoneal (PM) wash cells of rats. The cells present in the wash fluids were 85-90% macrophages in the peritoneal wash and 95% in the alveolar wash. Macromolecular components were resolved from whole cell homogenates by polyacrylamide gel isoelectric focusing (PAGIF) on horizontal gels and were identified cytochemically. Banding patterns clearly indicated a larger number of esterase zones in peritoneal compared to alveolar macrophages and ten previously unrecognized isozymes of DAP II in peritoneal macrophages with only three evident in their alveolar counterparts. (On whole blood smears, these cytochemically demonstratable enzymes were limited to macrophages, although DAP II was seen also in some mast cell granules). A protein band similar to the M6 band of alpha-1-antitrypsin in human serum was seen both in alveolar and peritoneal wash preparations. In addition, nine other major trypsin-binding protein bands were observed in the peritoneal macrophages, including two bands not observed in the alveolar macrophage extracts.

Animals↗

Fractionation and further characterization of granulocytic and monocytic alpha-naphthyl acetate (ANAE) esterases.

Following characterization of myeloid nonspecific esterases by isoelectric focusing (IEF), two main groups of alpha-naphthyl acetate (ANAE) esterase isoenzymes were defined and fractionated from cytoplasmic extracts by chromato focusing techniques according to differences in their isoelectric points (pI). The first of these ANAE enzyme groups was common to leukocytes of both granulocytic and monocytic lineage, while the other, which characteristically comprised a group of isoenzymes within the pI range 5.5-6.1, was specifically associated with monocytic differentiation. The properties of the two purified ANAE enzyme fractions were compared by inhibition (heat and sodium fluoride) and further electrophoretic studies, and the results discussed in relation to the cytochemical characterization of these enzymes as markers of specific myeloid cell differentiation.

Carboxylic Ester Hydrolases↗

Immunohistochemical detection of monocytes by the antiserum specific to monocytic esterase.

An antiserum specific to esterase Ib was produced in a rabbit. The antigen-antibody reaction was visualized by the strong esterase activity in the precipitin band in Ouchterlony double diffusion and immunoelectrophoresis. Immunohistochemical procedure demonstrated strong staining in the monocyte-infiltrated splenic sections and tissue sections of true histiocytic lymphoma. Negative results were observed in T- and B-cell lymphomas, granulocytic sarcoma, and chronic granulocytic leukemia. This antibody may be useful for the identification of monocytes and histocytes in paraffin-embedded tissue sections.

Animals↗

Nonspecific esterase isozymes that focus at identical isoelectric points are kinetically analogous.

Isoelectric focusing (IEF) of soluble nonspecific esterases of rat kidney and testis exhibits an identical array of organophosphate-resistant cathodal isozymes. To ascertain whether such isozymes that focus at the same pI are also kinetically analogous, two isozymes, both focused at pI 7.2, were isolated, one from each organ, by elution from cut-out, unstained gel segments. Although the esterases of the whole soluble fraction of kidney and testis exhibited different kinetic properties and organophosphate susceptibility, no differences were observed with regard to the isozymes. Therefore, because of similar electrophoretic and kinetic behavior, the two isozymes can be regarded as phenotypic expressions of similar genetic products.

Animals↗

Deficiency of arginine esterase in cystic fibrosis of the pancreas: demonstration of the proteolytic nature of the activity.

Proteolytic activity, defined as the hydrolysis of peptide bonds involving the carboxyl group of L-arginine, in plasma of patients with cystic fibrosis, heterozygotes, and control subjects has been assayed using a fluorometric method with protamine as the substrate and fluorescamine as the reagent. The mean total proteolytic activity in plasma of patients with cystic fibrosis was approximately one-half the mean total activity in control subjects and heterozygotes. The mean proteolytic activity inhibited by soybean trypsin inhibitor in plasma of patients with cystic fibrosis was approximately one-third that of control subjects and heterozygotes. The relationship of arginine esterase activity to proteolytic activity was investigated. The pH optimum and action of reversible and irreversible inhibitors were similar for both activites, suggesting that the arginine esterase activity and proteolytic activity represent similar catalytic entities. These findings are consistent with our hypothesis that the basic defect in cystic fibrosis may reside in the deficieny of a proteolytic enzyme which results in the accumulation of the various cationic macromolecular "factors" described by other investigators in serum of patients with cystic fibrosis. The demonstration of a deficiency of proteolytic activity as assayed by the hydrolysis of protamine, a cationic polypeptide, could explain the presence of ciliotoxic cationic protein or polypeptide factors in serum of patients with cystic fibrosis and may, in some unknown manner, be related to the clinical manifestations of the disease.

Adolescent↗

Conversion of high molecular weight human epidermal growth factor (hEGF)/urogastrone (UG) to small molecular weight hEGF/UG by mouse EGF-associated arginine esterase.

Human epidermal growth factor (hEGF) has previously been isolated from urine and appears to be identical to beta-urogastrone (UG), an inhibitor of stimulated gastric acid secretion. A high molecular weight (HMW) form of hEGF/UG has recently been found in human urine which is fully immunoreactive but is less bioactive as measured by receptor binding activity. A specific arginine esterase, the EGF-binding protein from mouse submandibular glands, was capable of cleaving HMW-hEGF to yield a small molecular weight (SMW)-hEGF with full immunoreactivity and bioactivity, whereas trypsin produced a SMW-hEGF with much less bioactivity. SMW-hEGF produced by the arginine esterase appeared to be immunologically, biologically (both by receptor binding and mitogenic activity) and chromatographically similar to highly purified hEGF. These data suggest that HMW-hEGF may play a precursor role in the biosynthesis of hEGF/UG in man.

Animals↗

Drosophila neurotactin, a surface glycoprotein with homology to serine esterases, is dynamically expressed during embryogenesis.

Drosophila neurotactin is a transmembrane glycoprotein with an apparent molecular mass of 135 x 10(3). Neurotactin is regionally expressed at the cellular blastoderm stage; later in embryogenesis the expression of the protein becomes restricted to cells of the peripheral and central nervous system. Immunocytochemical localization shows neurotactin protein at points of cell-cell contact. Using the anti-neurotactin monoclonal antibody BP-106, a neurotactin cDNA was isolated that encodes a 846 residue polypeptide. The chromosomal location of the neurotactin gene is 73C. The extracellular domain at the carboxyterminal end of the neurotactin protein shows a strong structural and sequence homology to serine esterases without retaining the amino acids forming the active center. Neurotactin therefore belongs to a growing group of proteins including Drosophila glutactin and thyroglobulins that are known to share this serine esterase protein domain motif without retaining the active center of the enzyme.

Amino Acid Sequence↗

Purification, properties and reactivity of the esterase from Micrococcus sp.

The esterase from Micrococcus sp., which hydrolyzes n-propyl-2-fluorocyclopropanecarboxylate (3) enantioselectively, was highly purified by three types of chromatography. The purified enzyme was inactivated by Hg and diisopropyl fluorophosphate (DFP). It was a monomer with a molecular weight of about 35000. The enzyme exhibits esterase activity towards many aliphatic propyl esters. The enantioselectivity for substrate (3) using purified enzyme did not differ from that of crude enzyme.

Esterases↗

Total synthesis of (+/-)-acetomycin and design of esterase-resistant analogs.

The synthesis of acetomycin and related analogs was investigated. Acetomycin was synthesized from diethyl allyl(methyl)malonate in 6.5% yield over 18 steps. The total number of steps was improved compared to our previous synthesis; i.e., four steps shorter, and the total yield was 4.5% greater than the previous synthesis. Acetomycin analogs with benzoyloxy and pivaloyloxy groups, instead of an acetoxy group at the 5-position of the gamma-butyrolactone ring were designed as esterase-resistant models and prepared similarly. Although they showed a similar level of cytotoxicity as acetomycin in vitro, they were not resistant to porcine liver esterase, and lost cytotoxicity in vivo.

Animals↗