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Biomedical subjects

A Dubin

Publications and source records attributed to A Dubin.

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

Isolation of nine human plasma proteinase inhibitors by sequential affinity chromatography.

Purification of nine plasma proteinase inhibitors and one zymogen from a single batch of human plasma, using affinity chromatography has been accomplished. Those isolated were plasminogen (lysine-Sepharose), alpha-2-antiplasmin (plasminogen-Sepharose), high and low molecular weight kininogens (CM-papain-Sepharose), alpha-2-macroglobulin (Zn++ chelate-Sepharose), alpha-1-proteinase inhibitor, alpha-1-antichymotrypsin, Cl-inhibitor, inter-alpha-trypsin inhibitor (Blue-Sepharose) and antithrombin III (heparin-Sepharose). Alpha-2-macroglobulin and alpha-1-proteinase inhibitor required gel filtration as additional purification steps. Each protein was recovered in both high yield and purity.

Alpha-Globulins↗
PubMed1990

Efficacy of immunopriming prior to isolation of tumor infiltrating lymphocytes for use in adoptive immunotherapy.

Adoptive immunotherapy is dependent upon the leukocytic subsets isolated as tumor infiltrating lymphocytes (TIL) prior to in vitro expansion with interleukin-2. To favorably influence T-cell subset representation in TIL the efficacy of bacillus Calmette Guerin (BCG) and cyclophosphamide (CTX) priming was evaluated in rats bearing Dunning R3327-AT prostatic tumors. When assessed by immunohistochemistry, both agents significantly (p less than 0.001) increased helper-T representation and decreased that by suppressor-T cells. As a result helper/suppressor (H/S) T cell ratios of TIL from untreated tumors (0.73 +/- 0.11) were significantly (p less than 0.001) elevated by both BCG (1.93 +/- 0.39) and CTX (1.40 +/- 0.25). Immunopriming might enhance adoptive immunotherapy by increasing the H/S ratio of TIL prior to their culture.

Animals↗
PubMed1989

Application of immunohistologic staining to develop a malignant index to aid in distinguishing benign from malignant prostatic tissue.

The development of antisera with reactivities against intermediate filaments, differentiation antigens, and secretory products has aided in identification and characterization of tissue specimens. Such evaluations may assist pathologists in distinguishing between benign (BPH) and malignant (CAP) tissue of prostatic origin. However, attempts to employ this technique are thwarted by 1) the use of frequently incompletely characterized antisera, 2) the use of both paraffin- and frozen-sectioned materials, and 3) a lack of quantitation in the degree of antisera immunoreactivity. To overcome these shortcomings, a mathematical approach was evaluated using eight BPH and 23 CAP specimens. These were sectioned and stained using commercially prepared antisera against cytokeratin (Cyto P, Cyto M), epithelial membrane antigen (EMA), NK cells (Leu-7), prostatic acid phosphatase (PAP), and prostate specific antigen (PSA). Reactivity was quantitated on a scale of 0-5. Mean values for markers elevated in CAP (relative to BPH) were placed in the numerator; those elevated in BPH (relative to CAP) were placed in the denominator: Malignant index = PSA + EMA + Leu-7 + Cyto M/PAP + Cyto P. This malignant index was significantly greater (P less than .001) in CAP tissues than in BPH regardless of Gleason grade (3.2 +/- 0.9 vs 1.6 +/- 0.9). It was also significantly elevated (3.0 +/- 0.8; P less than .01) in nine specimens representing prostatic atypical hyperplasia. These data suggest that immunohistologic staining may be applied as an aid in distinguishing between BPH and CAP.

Diagnosis, Differential↗
PubMed1988

An elastase inhibitor from equine leukocyte cytosol belongs to the serpin superfamily. Further characterization and amino acid sequence of the reactive center.

Horse leukocyte elastase inhibitor rapidly forms stable, equimolar complexes with both human leukocyte elastase and cathepsin G, porcine pancreatic elastase, and bovine alpha-chymotrypsin. Formation of the inhibitor-pancreatic elastase complex results in peptide bond cleavage at the reactive site of the inhibitor so that a small peptide fragment representing the carboxyl-terminal sequence of the inhibitor is released. Sequence analysis of both this peptide, as well as that of an overlapping peptide obtained by enzymatic inactivation of native inhibitor with either Staphylococcus aureus metalloproteinase, Pseudomonas aeruginosa elastase, or cathepsin B, yields data which indicate that the reactive site encompasses a P1-P1' Ala-Met sequence. However, unlike the human endothelial plasminogen activator inhibitor, which also has a Met residue in the P1' position, oxidation of the horse inhibitor only slightly reduces its association rate constant with either of the elastolytic enzymes tested or with chymotrypsin. Comparison of the amino acid sequence at or near the reactive site of the horse inhibitor (P2-P18') with members of the serpin superfamily of proteinase inhibitors indicates that it not only belongs in this class but also represents the first example of a functionally active intracellular serpin.

Amino Acid Sequence↗
PubMed1988

Degradation of elastin by a cysteine proteinase from Staphylococcus aureus.

Staphylococcus aureus is known to produce three very active extracellular proteinases. One of these enzymes, a cysteine proteinase, after purification to homogeneity was found to degrade insoluble bovine lung elastin at a rate comparable to human neutrophil elastase. This enzyme had no detectable activity against a range of synthetic substrates normally utilized by elastase, chymotrypsin, or trypsin-like proteinases. However, it did hydrolyze the synthetic substrate carbobenzoxy-phenylalanyl-leucyl-glutamyl-p-nitroanilide (Km = 0.5 mM, kcat = 0.16 s-1). The proteolytic activity of the cysteine proteinase was rapidly and efficiently inhibited by alpha 2-macroglobulin and also by the cysteine-specific inhibitor rat T-kininogen (Ki = 5.2 X 10(-7) M). Human kininogens, however, did not inhibit. Human plasma apparently contains other inhibitors of this enzyme, since plasma depleted of alpha 2-macroglobulin retained significant inhibitory capacity. The elastolytic activity of this S. aureus proteinase and its lack of control by human kininogens or cystatin C may explain some of the connective tissue destruction seen in bacterial infections due to this and related organisms such as may occur in septicemia, septic arthritis, and otitis.

Animals↗
PubMed1988

Noncatalytic protein component of elastase from horse leucocytes. A protein with regulatory function.

1. Noncatalytic protein component (NPC), a strongly acidic protein (pH = 4.5) was separated from native horse leucocyte elastase 1. 2. This protein reduces elastinolytic properties of elastases: 1 and 2A probably by decreasing their isoelectric points. 3. A possible regulatory role of this protein may be inferred from a higher affinity of elastase 1 to NPC rather than to elastin.

Animals↗
PubMed1988

Effect of BCG upon functional and phenotypic immune markers in rats bearing the Dunning R3327 MAT-LyLu prostatic adenocarcinoma.

Rats bearing (or not bearing) the Dunning R3327 MAT-LyLu prostatic adenocarcinoma were treated with Bacillus Calmette-Guérin (BCG) and evaluated for immune competence using functional and phenotypic markers. Tumor presence significantly depressed total T and helper T cell representation along with the helper/suppressor T cell ratio. Functional immunity, measured by phytohemmagglutinin (PHA) induced blastogenesis, was also significantly depressed. When BCG was administered to non-tumor bearing animals, it had no effect upon T cell subset distributions but significantly reduced PHA induced blastogenesis. BCG similarly administered to tumor bearing animals did not alter the depressed helper/suppressor T cell ratio found in tumor bearing rats, but did significantly elevate PHA induced blastogenesis. However, these elevated levels of functional immunity in BCG treated tumor-bearing rats remained significantly below normal. These data demonstrate a poor correlation between functional and phenotypic assessments of immune capability.

Adenocarcinoma↗
PubMed1988

Review of acid phosphatase in the diagnosis and prognosis of prostatic cancers.

Acid phosphatase is a secretory product frequently utilized as a tumor marker for disseminated, late stage (D2) prostatic cancer. In the 40 years since this association has been recognized, this enzyme has been subjected to extensive biochemical and immunological characterizations. These techniques have also been adapted for rapid and specific determinations of the prostatic isoenzyme levels using a variety of techniques. Since acid phosphatase levels do not become significantly elevated until late stage cancer, newer markers such as prostate-specific antigen have been sought which appear earlier and may be more useful for the screening and monitoring of high risk populations. At this time it is appropriate to review the current and future status of acid phosphatase as a diagnostic aid.

Acid Phosphatase↗
PubMed1987

Combination therapy using polyamine synthesis inhibitor alpha-difluoromethylornithine and adriamycin in treatment of rats carrying the Dunning R3327 MAT-LyLu prostatic adenocarcinoma.

Prostate cells of human and rat origin produce polyamines in high content, whose apparent functions relate to cellular proliferation and secretory activities. Formation is dependent on the enzyme ornithine decarboxylase (ODC) which is irreversibly inhibited by alpha-difluoromethylornithine (DFMO). It has been postulated that pretreatment with DFMO may render cells more susceptible to subsequent chemotherapy. Copenhagen X Fischer F1 rats bearing the Dunning R3327 MAT-LyLu prostatic adenocarcinoma were given DFMO or adriamycin (ADR), alone or in combination. Those receiving DFMO were continuously provided the drug ad libitum, in water (2.5%), for the duration of the experiment, beginning 2 days prior to ADR administration. At intervals, tumor sizes were measured, animal survivals noted and comparisons made to nontreated, tumor-bearing controls. The results indicate that ADR alone or in combination with DFMO significantly reduced tumor progression, but that only combination therapy significantly prolonged survivals. Decreased tumor progression produced by DFMO alone was not statistically significant. Differences produced with combined use were additive and suggest that DFMO may augment ADR chemotherapy.

Adenocarcinoma↗
PubMed1987

Effect of cyclophosphamide on leukocytic subset distributions in rats carrying the Dunning R3327-MAT-LyLu prostatic adenocarcinoma.

The Dunning R3327 adenocarcinoma represents a model for studying prostate cancer in rats; early studies have indicated its utility for studying relationships between tumor growth, immunologic markers, and chemotherapy. Normal animals and those bearing the metastatic Dunning R3327 MAT-LyLu tumor were treated with 10, 30, and 100 mg/kg doses of cyclophosphamide (CTX) and their spleens assayed for leukocytic subset distributions using monoclonal antibodies. Tumor-bearing animals had significant reductions in helper T cell content as well as reduced helper/suppressor T cell ratios, compared to controls. These effects occurred rapidly following implantation and were not reversed by chemotherapy. When administered to both tumor- and non-tumor-bearing animals, CTX also depleted T cell populations. Despite reductions produced in all subsets, two administrations of CTX (30 mg/kg) were capable of retaining (in non-tumor-bearing animals) or restoring (in tumor-bearing) normal helper/suppressor T cell ratios. Such studies aid in identifying therapeutically effective dosages of cytotoxic drugs that minimize their deleterious effects on the immune system.

Adenocarcinoma↗
PubMed1987

Amino acid composition of uremic middle and low molecular weight retention products.

In order to characterize the spectrum of small peptides retained in chronic renal failure, we carried out high pressure liquid chromatography (HPLC) of serum ultrafiltrates from patients with chronic renal failure (CRF), acute renal failure (ARF), and normal subjects. HPLC patterns in CRF resolved into more than twenty peaks; those in ARF contained fewer peaks and resembled that of normals. We carried out amino acid analysis of HPLC fractions after hydrolysis with 6N HC1 of four patients with CRF, one patient with ARF, and one normal subject. Following hydrolysis each HPLC fraction yielded several amino acids. Glycine, leucine, serine, phosphoserine, glutamic acid, and phenylalanine were found in greatest frequency in the four CRF patients.

Amino Acids↗
PubMed1987

Biochemical elucidation and HPLC fractionation of fluorescent peptides in patients with chronic renal failure.

We evaluated the biochemical characteristics of endogenous fluorescent substances, Ex 380 nm/Em 440 nm and Ex 400 nm/Em 460 nm, present in sera of patients with chronic renal failure (Clin. Chem. 31:1988, 1985). Sera from 23 patients with chronic renal failure (CRF) and from 10 normal subjects were filtered through ultrafiltration membranes (cutoff limit of 500 Da). Fluorescence intensity of the aforementioned substances was significantly elevated as compared to normals (p less than 0.001). Fluorescence characteristics of these substances remained unaltered after ultrafiltration and treatment with beta-glucuronidase. Extraction of these fluorescent compounds with organic solvents (dichloromethane, ethyl acetate, chloroform:methanol) could not be achieved after ultrafiltrates were subjected to 6N hydrochloric acid (HC1) hydrolysis. In addition, treatment with 6N HC1 enhanced fluorescence intensity without altering fluorescence excitation/emission maxima. Removal of fluorescence could be accomplished in toto by adsorption onto activated charcoal with subsequent recovery from charcoal by treatment with sodium hydroxide, pH 12 (Ex 380 nm: 51.1%, Ex 400 nm: 91.8%). Analysis of alkali-treated specimens by high performance liquid chromatography demonstrated that peptides associated with these fluorescent substances were denatured, although fluorescence at these previously described excitation/emission maxima persisted. Our studies indicate that the unique fluorescence observed in the sera of patients with CRF is not an intrinsic characteristic of a specific peptide or its amino acids, but rather an inherent property of fluorescent molecules which may bind to these peptides.

Chromatography, High Pressure Liquid↗
PubMed1987

Horse alpha 2-macroglobulin. Circular dichroism studies of conformational changes upon reaction with proteinases and methylamine.

The interaction of horse alpha 2-macroglobulin with methylamine, trypsin and cathepsin D was studied by circular dichroism in the far and near UV region, by polyacrylamide gel electrophoresis and by determination of its inhibitory activity. The CD spectra of horse alpha 2-macroglobulin resemble those of bovine und human alpha 2-macroglobulin. The CD spectra were changed in a different manner after the interaction of alpha 2-macroglobulin with methylamine, trypsin and inactive or active cathepsin D, indicating that more than one conformational change occurs. Cathepsin D activity was not affected by complex formation with horse alpha 2-macroglobulin. In contrast to the action of trypsin, treatment with methylamine did not increase the electrophoretic mobility of alpha 2-macroglobulin.

Animals↗
PubMed1987

Administration of recombinant tumor necrosis factor to rats bearing the Dunning R3327 MAT-LyLu prostatic adenocarcinoma.

Copenhagen X Fischer F1 rats bearing palpable Dunning R3327 MAT-LyLu prostatic adenocarcinomas were treated by intraperitoneal (i.p.) or intratumor (i.t.) injection with either human serum albumin alone or in combination with recombinant tumor necrosis factor (rTNF). At intervals tumors were measured and survivals noted. A maximum tolerable dose and least toxic route of administration was then determined. Those treated i.t. with rTNF survived significantly longer and ultimately developed significantly smaller tumors than untreated controls. Those administered rTNF by the i.p. route had less significant increases in survival with intermediate final tumor sizes.

Animals↗
PubMed1986

Comparison of antiproteolytic activities of alpha-1-proteinase inhibitors from the plasma of some mammalian species.

Alpha-1-proteinase inhibitors isolated from plasmas of horse, ox, pig, rabbit and man were used for determination of some kinetic parameters of interaction with three horse leucocyte proteinases and bovine pancreatic trypsin and chymotrypsin. Effective molar ratio of enzyme-to-inhibitor, inactivation rate constant and inhibition constant were measured. In horse, ox, pig and rabbit two principal electrophoretic forms of alpha 1-PI could be distinguished. Both forms effectively inhibited trypsin but usually only one form reacted promptly and stoichiometrically with chymotrypsin and leucocyte elastases. It appears that genetic variability and functional heterogeneity of multiple forms of alpha 1-PI as well as lack of other tissue inhibitors of proteinases may be responsible for lung emphysema occurring in man and horse.

Animals↗
PubMed1986

Adherent spleen cell production of E series prostaglandins in rats bearing variants of the R3327 Dunning prostatic adenocarcinoma: effect of cyclophosphamide.

Prostaglandins of the E series (PGE) have been implicated in many facets of immunoregulation, as well as having a possible role in metastatic dissemination. Variant sublines of the Dunning R3327 rat prostatic adenocarcinoma, differing in growth rate, hormonal responsiveness and in propensity for metastasis, were carried in Fisher X Copenhagen F1 animals. Adherent spleen cells were assayed in vitro for their ability to convert arachidonic acid to prostaglandins of the E series. These glass adherent cells presumably include the monocytic and T cell populations which have been implicated as being immunoregulatory. The results indicated that those spleen cells obtained from animals carrying the metastatic R3327-MAT-LyLu subline tumor converted more arachidonic acid to PGE's than cells derived from animals bearing non-metastatic sublines. Cyclophosphamide therapy did not alter such conversion. Multiple regulatory mechanisms for prostaglandin metabolism are suggested.

Adenocarcinoma↗
PubMed1986

Alpha-1-acute phase globulin in the blood of tunicamycin-injected rats. Isolation of the non-glycosylated form, its inhibitory properties and synthesis in liver slices.

Plasma of rats injected with tunicamycin (2 micrograms/g body wt) shows on crossed immunoelectrophoresis the presence of an additional, slowly migrating component of alpha 1-acute-phase globulin (alpha 1-AP-globulin). The native and modified forms of alpha 1-AP-globulin were jointly isolated on the column of immobilized antibodies and then separated by chromatofocusing on polybuffer exchanger. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate demonstrated that tunicamycin-induced form of rat alpha 1-AP-globulin has Mr of 50,000 and is devoid of carbohydrates as inferred from the lack of staining with Schiff reagent. However, during incubation with papain in vitro it is only slightly less effective than mature glycosylated alpha 1-AP-globulin (Mr 68 000) in inhibiting hydrolysis of CBZ-Lys-ONp. Incubation of liver slices from control and tunicamycin-injected rats with 14C-leucine demonstrated that tunicamycin reduces synthesis and release to the medium of alpha 1-AP-globulin and some other plasma proteins, but the proportion of aglyco-alpha 1-AP-globulin is higher than in plasma.

Acute-Phase Proteins↗