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Biomedical subjects

H E Fuchs

Publications and source records attributed to H E Fuchs.

34 records · Page 2Linked to original sources
PubMed1986

Specificity of alpha 2-macroglobulin covalent cross-linking for the active domain of proteinases.

The reaction of alpha 2-macroglobulin (alpha 2M) with the two-chain enzyme plasma kallikrein results in covalent bond formation between the catalytic subunit and the inhibitor. We have recently published a model of alpha 2M which suggests that this phenomenon may be a general mechanism when multisubunit proteinases are inactivated by alpha 2M. In order to test this hypothesis, we studied the reactions of factor Xa, plasmin, streptokinase-plasmin and alpha-thrombin with alpha 2M. In the case of factor Xa the catalytic heavy chain demonstrated greater than 99% covalent incorporation while over 97% of the light chain failed to crosslink to the inhibitor. Preferential binding of the catalytic light chains of plasmin (70% covalent incorporation) and plasmin in complex with streptokinase (79% covalent incorporation) was also observed. Finally, 82% covalent incorporation of the catalytic heavy chain of alpha-thrombin was found. These studies demonstrate that in the case of multisubunit proteinases, the chain containing the active site demonstrates preferential binding as predicted by the model supporting placement of the site of covalent binding close to the "bait region" of alpha 2M.

Binding Sites↗
PubMed1986

Release of tissue plasminogen activator and its fast-acting inhibitor in defective fibrinolysis.

Releasable tissue plasminogen activator (t-PA) and the fast inhibitor of t-PA were measured in 18 controls and a pedigree with venous thrombosis. The functional assay was performed by a technique that destroys the t-PA inhibitor when blood is drawn. It was found that activator and inhibitor levels varied widely in the control group. One patient demonstrated inhibitor levels, on two different occasions, of 2.82 and 3.54 IU of t-PA per milliliter of plasma, as compared with a releasable activator level of 1.87 IU/mL. The t-PA antigen levels of this patient and the remainder of the pedigree were essentially normal for all seven subjects. Thus, it is suggested that the previously reported fibrinolytic disorder in this pedigree represents an imbalance between activator and inhibitor levels rather than an actual deficiency of t-PA.

Antigens↗
PubMed1985

Releasable vascular plasminogen activator and thrombotic strokes.

Releasable vascular plasminogen activator was measured in 28 patients (14 males and 14 females) with a history of thrombotic strokes documented by computed tomographic scanning. Levels were compared with those in a control population of 126 healthy subjects with no history of thromboembolic disease. The patient population tended to have higher levels of activator than the control population, 0.53 Committee on Thrombolytic Agents (CTA) units/ml of plasma for patients versus 0.21 CTA units/ml for control subjects; however, there was a wide distribution of values, as reported in all previous populations. Since plasminogen activator levels distribute in a non-Gaussian manner, patient values and control values were stratified into deciles. By this approach, the distribution among the patient was not significantly different from that among the control subjects except in females, who demonstrated skewing to higher deciles (p = 0.019). It is concluded that thrombotic strokes are not associated with low levels of releasable vascular plasminogen activator, and in fact, these patients may present with levels considerably above the mean for normal subjects.

Adolescent↗
PubMed1985

Catabolism of human tissue plasminogen activator in mice.

The catabolism of human tissue plasminogen activator (t-PA) was studied in mice. The clearance of t-PA labeled with iodine 125 was rapid (t1/2). The clearance of phenylmethylsulfonyl-125I-t-PA, which is active site-inhibited, was identical to the active enzyme. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) demonstrated that the vast majority of 125I-t-PA injected into the circulation was present as free enzyme and not in a complex with inhibitors. The clearance of 125I-t-PA was unaltered by large molar excesses of several ligands of known clearance specificities, including macroalbumin, asialoorosomucoid, and diisopropylphosphorylthrombin and was also not altered in the presence of a 1,000-fold molar excess of unlabeled t-PA. Organ distribution studies demonstrated that the early rapid clearance of 125I-t-PA occurred in hepatocytes, followed by a later renal phase of clearance. The clearance of 125I-urokinase (UK) also was studied and was very similar in all aspects to the clearance of 125I-t-PA. These results suggest that both t-PA and UK are cleared from the circulation by unique nonsaturable processes localized in the liver that are independent of the proteinase active site.

Albumins↗
PubMed1984

Hepatocyte receptors for antithrombin III-proteinase complexes.

The in vivo clearance of antithrombin III-proteinase complexes occurs via a specific and saturable pathway located on hepatocytes. We now report studies of the catabolism of antithrombin III-proteinase complexes in vitro using rat hepatocytes in primary culture. Antithrombin III-thrombin and trypsin complexes were prepared and purified to homogeneity. Ligand uptake by hepatocytes was concentration, temperature, and time dependent. Initial rate studies were performed to characterize the maximum rate of uptake, V, and apparent Michaelis constant Kapp. These studies yielded a V of 12.8 fmol/mg cell protein/min and a Kapp of 144 nM for antithrombin-trypsin complexes. Competition experiments with antithrombin III, antithrombin III-proteinase complexes, alpha 2-macroglobulin-methylamine, asialoorosomucoid and the neoglycoproteins, fucosyl-bovine serum albumin (BSA), N-acetylglucosaminyl-BSA, and mannosyl-BSA indicated that only antithrombin III-proteinase complexes were recognized by the hepatocyte receptor. Uptake studies were performed at 37 degrees C with 125I-antithrombin III-trypsin and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in conjunction with autoradiography. These studies demonstrate time-dependent uptake and degradation of the ligand to low molecular weight peptides. In addition, there was a time-dependent accumulation of a high molecular weight complex of ligand and a cellular protein. This complex disappeared when gels were performed under reducing conditions.

Animals↗
PubMed1984

Hepatocyte uptake of alpha 1-proteinase inhibitor-trypsin complexes in vitro: evidence for a shared uptake mechanism for proteinase complexes of alpha 1-proteinase inhibitor and antithrombin III.

In vivo clearance studies have indicated that the clearance of proteinase complexes of the homologous serine proteinase inhibitors alpha 1-proteinase inhibitor and antithrombin III occurs via a specific and saturable pathway located on hepatocytes. In vitro hepatocyte-uptake studies with antithrombin III-proteinase complexes confirmed the hepatocyte uptake and degradation of these complexes, and demonstrated the formation of a disulfide interchange product between the ligand and a cellular protein. We now report the results of in vitro hepatocyte uptake studies with alpha 1-proteinase inhibitor-trypsin complexes. Trypsin complexes of alpha 1-proteinase inhibitor were prepared and purified to homogeneity. Uptake of these complexes by hepatocytes was time and concentration-dependent. Competition experiments with alpha 1-proteinase inhibitor, alpha 1-proteinase inhibitor-trypsin, and antithrombin III-thrombin indicated that the proteinase complexes of these two inhibitors are recognized by the same uptake mechanism, whereas the native inhibitor is not. Uptake studies were performed at 37 degrees C with 125I-alpha 1-proteinase inhibitor-trypsin and analyzed by sodium dodecyl sulfate-gel electrophoresis in conjunction with autoradiography. These studies demonstrated time-dependent uptake and degradation of the ligand to low molecular weight peptides. In addition, there was a time-dependent accumulation of a high molecular weight complex of ligand and a cellular protein. This complex disappeared when gels were performed under reducing conditions. The sole cysteine residue in alpha 1-proteinase inhibitor was reduced and alkylated with iodoacetamide. Trypsin complexes of the modified inhibitor were prepared and purified to homogeneity. Uptake and degradation studies demonstrated no differences in the results obtained with this modified complex as compared to unmodified alpha 1-proteinase inhibitor-trypsin complex. In addition, the high molecular weight disulfide interchange product was still present on sodium dodecyl sulfate-polyacrylamide gel electrophoresis of solubilized cells. Clearance and clearance competition studies with alpha 1-proteinase inhibitor-trypsin, alkylated alpha 1-proteinase inhibitor-trypsin, antithrombin III-thrombin, and anti-thrombin III-factor IXa further demonstrated the shared hepatocyte uptake mechanism for all these complexes.

Alkylation↗
PubMed1984

Regulation of factor IXa in vitro in human and mouse plasma and in vivo in the mouse. Role of the endothelium and the plasma proteinase inhibitors.

The regulation of human Factor IXa was studied in vitro in human and mouse plasma and in vivo in the mouse. In human plasma, approximately 60% of the 125I-Factor IXa was bound to antithrombin III (ATIII) by 2 h, with no binding to alpha 2-macroglobulin or alpha 1-proteinase inhibitor, as assessed by gel electrophoresis and IgG- antiproteinase inhibitor-Sepharose beads. In the presence of heparin, virtually 100% of the 125I-Factor IXa was bound to ATIII by 1 min. The distribution of 125I-Factor IXa in mouse plasma was similar. The clearance of 125I-Factor IXa was rapid (50% clearance in 2 min) and biphasic and was inhibited by large molar excesses of ATIII-thrombin and alpha 1-proteinase inhibitor-trypsin, but not alpha 2-macro-globulin-trypsin; it was also inhibited by large molar excesses of diisopropylphosphoryl - (DIP-) Factor Xa, DIP-thrombin, and Factor IX, but not by prothrombin or Factor X. The clearance of Factor IX was also rapid (50% clearance in 2.5 min) and was inhibited by a large molar excess of Factor IX, but not by large molar excesses of Factor X, prothrombin, DIP-Factor Xa, or DIP-thrombin. Electrophoresis and IgG- antiproteinase inhibitor-Sepharose bead studies confirmed that by 2 min after injection into the murine circulation, 60% of the 125I-Factor IXa was bound to ATIII. Organ distribution studies with 125I-Factor IXa demonstrated that most of the radioactivity was in the liver. These studies suggest that Factor IXa binds to at least two classes of binding sites on endothelial cells. One site apparently recognizes both Factors IX and IXa, but not Factor X, Factor Xa, prothrombin, or thrombin. The other site recognizes thrombin, Factor Xa, and Factor IXa, but not the zymogen forms of these clotting factors. After this binding, Factor IXa is bound to ATIII and the complex is cleared from the circulation by hepatocytes.

Animals↗
PubMed1983

Regulation of factor Xa in vitro in human and mouse plasma and in vivo in mouse. Role of the endothelium and plasma proteinase inhibitors.

The regulation of human Factor Xa was studied in vitro in human and mouse plasma, and in vivo in mouse. In human plasma, 125I-Factor Xa bound to alpha 1-proteinase inhibitor, antithrombin III, and alpha 2-macroglobulin in a ratio of 4.9:1.9:1 as determined by gel electrophoresis and by adsorption to IgG-(antiproteinase inhibitor)-Sepharose beads. The distribution of Factor Xa in mouse plasma was similar. The clearance of Factor Xa in mice was rapid (50% clearance in 3 min) and biphasic. alpha 1-Proteinase inhibitor-trypsin, even at a 2,000-fold molar excess, failed to inhibit the clearance of Factor Xa, while alpha 2-macroglobulin-trypsin inhibited only the later phase of clearance. The plasma clearance of diisopropylphosphoryl-Factor Xa was more rapid than native Factor Xa (50% clearance in 2.5 min), and the clearance was blocked by diisopropylphosphoryl-thrombin. Electrophoresis experiments confirmed that by 2 min after injection into the murine circulation, 90% of the bound Factor Xa was on alpha 2-macroglobulin, in marked contrast to the in vitro results. Organ distribution studies at 3 and 15 min with 125I-Factor Xa demonstrated that the majority of radioactivity was in the liver, with significant radioactivity also present in lung and kidney. Autopsies performed 30 s after injection of 125I-Factor Xa also demonstrated significant binding to the aorta and vena cava. These studies indicate that Factor Xa binds to specific thrombin-binding sites on endothelial cells, and that this binding alters its proteinase inhibitor specificity. Factor Xa binds to alpha 2-macroglobulin in vivo, whereas the predominant in vitro inhibitor of Factor Xa is alpha 1-proteinase inhibitor.

Animals↗
PubMed1982

A unique pathway for the plasma elimination of alpha 2-antiplasmin-protease complexes in mice.

Radiolabeled alpha 2-antiplasmin cleared slowly from the circulation of mice. Complex formation with either plasmin or trypsin resulted in a significant increase in the plasma elimination rate of the protease inhibitor. Approximately 20 min and 14 min were required for 50% of the injected alpha 2-antiplasmin-plasmin and alpha 2-antiplasmin-trypsin to clear from the circulation, respectively. Significant competition was observed when radiolabeled alpha 2-antiplasmin-plasmin was cleared in the presence of a large molar excess of unlabeled alpha 2-antiplasmin-plasmin. alpha 1-Antitrypsin-trypsin failed to complete with radiolabeled alpha 2-antiplasmin-plasmin even when present at 2000 fold molar excess. Organ distribution studies localized the major site of alpha 2-antiplasmin-plasmin clearance in the liver. Microscopic autoradiography data suggested that the cell responsible for the clearance pathway was the hepatocyte.

Animals↗
PubMed1982

In vivo catabolism of alpha 1-proteinase inhibitor-trypsin, antithrombin III-thrombin and alpha 2-macroglobulin-methylamine.

The clearances of 125I-labeled alpha 1-proteinase inhibitor-trypsin, antithrombin III-thrombin and alpha 2-macroglobulin-methylamine (CH3NH2) were compared in our previously described mouse model. alpha 1-Proteinase inhibitor-trypsin cleared with a t 1/2 of 20 min, antithrombin III-thrombin of 7 min and 125I-labeled alpha 2-macroglobulin-methylamine of 2 min. Competition studies were performed to determine whether one or several pathways clear these three ligands. The clearance of 125I-labeled alpha 1-proteinase inhibitor-trypsin and 125I-labeled antithrombin III-thrombin was blocked by large molar excesses of either ligand, but not by alpha 2-macroglobulin-methylamine. The clearance of 125I-labeled alpha 2-macroglobulin-methylamine can be blocked by a large molar excesses of unlabeled alpha 2-macroglobulin-methylamine but not by alpha 1-proteinase inhibitor-trypsin. These studies demonstrate that the clearance of alpha 1-proteinase inhibitor-trypsin complexes is independent of alpha 2-macroglobulin-methylamine and utilizes the same pathway which is involved in the clearance of antithrombin III-thrombin complexes.

Animals↗
PubMed1980

Three-dimensional structures of C-phycocyanin and B-phycoerythrin at 5-A resolution.

Single crystal x-ray diffraction studies are proceeding on two phycobiliproteins: C-Phycocyanin from Anabaena variabilis and B-Phycoerythrin from Porphyridium cruentum. C-Phycocyanin consists of six alpha and six beta subunits. A three-dimensional x-ray diffraction study of this light-harvesting protein (P6(3); a = b = 154.1 A, c = 40.1 A) at 5-A resolution shows that the molecule is 110 A in diameter, 40 A thick, with a 20-A diameter central channel. It can be divided into three separate domains related by the crystallographic 3 and contains long, columnar regions of density, presumed to represent alpha helix. B-Phycoerythrin is composed of 6 alpha, 6 beta, and a gamma subunit. A 5.25-A resolution study of this protein (R3, a = b = 189 A, c = 60 A) indicates that the molecule is 107 A in diameter and 55 A thick, has 32-D3 symmetry, and can be divided into six regions. The gamma chain lies on a crystallographic 3 and undergoes symmetric disordering. A region of low and unstructured density in the center of the molecule is presumably occupied by this gamma chains. These two low resolution structures are compared with regard to differences in quaternary structure.

Cyanobacteria↗
PubMed1977

Preliminary crystallographic investigations of two phycobiliproteins.

Single crystal x-ray diffraction investigations are in progress on two phycobiliproteins. C-phycocyanin from Anabaena variabilis crystallizes in space group P63 with a = b = 154 A and c = 40 A. The crystallographic asymmetric unit is (alphabeta)2 with a total molecular mass of 7.0-10(4) daltons. B-phycoerythrin from Porphyridium cruentum crystallizes in space group R3 with a = b = 189 A and c = 60 A. This molecule has the unusual molecular stoichiometry (alphabeta)6gamma and the crystallographic asymmetric unit is (alphabeta)2gamma1/3. This requires that the gamma chain undergo a perfect 3-fold disordering about the crystallographic 3 axis, i.e. the gamma chain must occupy three symmetry-equivalent positions, each with an occupancy of one-third.

Cyanobacteria↗
PubMedDate not supplied

Post-traumatic epidermoid cyst: CT appearance.

We report development of an intracranial epidermoid cyst 2 years after a depressed skull fracture. The epidermoid cyst is presumed to be the result of introduction of epidermal elements at the time of trauma. Post-traumatic intracranial epidermoid cysts are rare and appear to be less common than those occurring in the spine.

Adolescent↗
PubMedDate not supplied

The pathology of extracranial scalp and skull masses in young children.

OBJECTIVE: Extracranial subcutaneous masses involving the scalp and/or skull in young children are uncommon lesions that get excised by the neurosurgeon. Although the most common reported lesion is the dermoid cyst, our experience suggests that the spectrum of pathology in these lesions can present diagnostic challenges to the pathologist. MATERIAL: We reviewed 30 consecutive extracranial masses from 29 patients between July 1998 and June 2003. METHOD: Hematoxylin and eosin-stained sections were reviewed in all cases, and immunohistochemistry was performed in select cases. RESULTS: Twenty-three were within the scalp, 5 involved the scalp and skull and 2 were within the limits of the inner and outer tables of the skull. There were 8 dermoid cysts, 2 epidermoid cysts, 6 post-traumatic lesions including 3 calcified cephalhematomas and 3 pseudocysts, 5 vascular lesions including 3 capillary hemangiomas, 1 venous angioma and 1 lymphangioma, 2 cases of cranial fasciitis and 1 case each of benign teratoma, deep granuloma annulare, benign fibrous histiocytoma, congenital melanocytic nevus, hamartoma with ectopic meningothelial elements, cutaneous hyalinised ectopic meningioma and a meningocele with a fibrohistiocytic reaction. No lesions have recurred or exhibited malignant features. CONCLUSIONS: Surgical pathologists and neuropathologists should be aware that the differential diagnosis of "lumps and bumps on babie's heads" is quite varied and can be histologically challenging.

Child, Preschool↗
PubMedDate not supplied

Solitary fibrous tumor of the meninges occurring after irradiation of a mixed germ cell tumor of the pineal gland.

Twenty-nine months after surgery, irradiation, and systemic chemotherapy for a pineal mixed germ cell tumor, an 11-year-old Caucasian male developed a 3 cm dural based nodule in the occipital lobe that proved to be a solitary fibrous tumor by immunohistochemical and ultrastructural examination. Differential diagnosis included fibrous meningioma, neurofibroma, Schwannoma, cranial fasciitis of infancy, and solitary fibrous tumor. A Masson trichrome stain revealed a prominent collagenous stroma and reticulin staining exhibited strong pericellular positivity. Immunohistochemical staining demonstrated diffuse vimentin and focal CD34 positivity of tumor cells. Ultrastructural examination revealed fibroblastic differentiation. These features are consistent with solitary fibrous tumor. Although we favor a radiation-induced origin for the neoplasm, alternative explanations for the tumor's origin include cerebrospinal fluid spread from the original germ cell tumor or a de novo neoplasm.

Antineoplastic Agents↗