bluuebunny/crossref_metadata_2025 huggingface dataset:
An Antigenicity of the Platelet-Clumping Substance
Authors: Yamazaki H. , Sano T. , Shimamoto T.
Published: June 2019

We have reported an appearance of platelet-clumping substance in heparinized plasma of rabbit after acidification to pH around 5.5 (Blood 31, 348, 1968). This substance is different from ADP, collagen, thrombin etc. and may play an important role in hemostasis and thrombogenesis. The chemical analyses suggested a close relationship between the substance and acid mucopolysaccharide bound to protein (Plenary session in the 14th ISH, 1972). So we examined an antigenicity of the substance. One ml of acidified heparinized rabbit plasma (pH 5.5), which showed the platelet-clumping activity, was emulsified with 1 ml of Freund’s complete adjuvant (Difco). Six rabbits were injected with the emulsion every week repeatedly during 7 to 10 weeks and the sera were collected. The titer of platelet-clumping activity in the plasma was determined by serial dilution. The titers of acidified plasma collected from the rabbits before the immunization were 32 to 64 and those from the same rabbits after the immunization were 4 to 8. When the acidified plasmas collected before the immunization were mixed with control serum, the titers were 8 to 32 (13.3±3.9, mean ± SE). When the above plasmas were mixed with the immune serum, the titers were 2 to 8 (4.3±0.6). A statistically significant difference (p < 0.05) was observed between both the titers. These results suggest a presence of antigenicity of the platelet-clumping substance.
Binding of Anti-Actin Auto-Antibodies to Platelets
Authors: Bouvier C. A. , Gabbiani G. , Majno G.
Published: June 2019

It was recently shown (Gabbiani & Coll., Am. J. Path. 72, 473-488, 1973) that a natural antibody occurring in patients with chronic hepatitis is specifically directed against smooth and striated muscle actin or, in the case of platelets, Thrombosthenin A. Thus it is proposed to change its name from Smooth Muscle Antibody (SMA) to Anti Actin Antibody (AAA). The possibility that AAA would contain a contaminant directed against cell membranes was ruled out because it does not bind (nor its titer in supernatant decrease) to intact platelets, but only to damaged ones (by freeze-thawing). However AAA binds to a platelet-rich clot, and bright fluorescent staining is obtained both for such clots and bone-marrow megacaryocytes. AAA does not induce platelet aggregation in vitro when added to platelet-rich plasma; it does not inhibit or potentiate ADP, Adrenalin or Collagen induced aggregation, and does not inhibit platelet-rich plasma clot retraction.

With aggregation controlled in an aggregometer, the following observations were made : AAA adsorption (and consequent fluorescent staining) begins immediately after addition of the trigger, and becomes maximal when a plateau is reached (so-called irreversible aggregation). However, when liminary amounts of ADP are added, so as to obtain reversible aggregation, a large number of disaggregated platelets do bind with AAA, suggesting that actin-related antigens have become available to antibody. This implies that a major change in membrane permeability to large molecules has occurred. The same holds true when ADP-induced aggregation is prevented by EDTA.
Interaction of Fibrinogen and Factor VIII with Human Platelets
Authors: Schneider-Trip M. , Wanders R. , Cate J. W. ten , Jenkins C.
Published: June 2019

Binding of 125I fibrinogen to platelets was studied after incubation of 125I fibrinogen with PRP, gel filtered and washed platelets. Thereafter 125I fibrinogen was completely removed from the platelets by four washes by the albumin cushion method. When the 125I fibrinogen preparation was contaminated by factor VIII antigen binding to platelets was observed. This suggested that factor VIII antigen present in some fibrinogen preparations could be responsible for the controversy in the literature concerning binding of fibrinogen to platelets or not. Therefore factor VIII antigen (V0 fraction of sepharose 6 B chromatografied cryoprecipitate, van Mourik) was labeled to study its binding to platelets. Factor VIII (V0) labeled with 125I as described by MacFarlane resulted in decomposition of the factor VIII molecule as measured by its antigenic and procoagulant activity and Von Wille brand factor activity, whereas the labeling; with the Chloramine T method resulted in a different behaviour of the molecule on the polyacrylamide gel-electrophoresis. Using the lactoperoxidase method the factor VIII molecule remained completely intact. Incubation studies were performed with PRP and this 125I labeled factor VIII preparation. Subsequently the platelets were washed four times by the albumin cushion method and radioactivity was measured of the washing buffer and of the washed platelet suspensions. Binding was observed of radioactivity to the platelets. A dose-relationship was demonstrated.
Human Prothrombin Activation Products
Authors: Downing Michael R. , Butkowski Ralph J. , Mann K. G.
Published: June 2019

SDS gel electrophoretic analysis of the products of human prothrombin (IIH) activation reveal a similar product distribution to that observed for bovine prothrombin (IIB). However, some subtle differences are noted when the amino-terminal sequences of the activation intermediates are determined. The amino-terminal sequences of IIH and HH-intermediate 3 are identical (ALA, ASN, PRO, PHE, LEU, GLU, GLU, VAL, ARG, LYS) and homologous to the sequence for IIB. Similarly the amino-terminal sequences of IIH-mtermediate 1 and IIH-intermediate 4 (SER, GLU, GLY, SER, SER, VAL, ASN, LEU, SER, PRO) are entirely homologous with their bovine counterparts. On the other hand IIH-intermediate 2 and α-thrombin A chain have the sequence THR, PHE, GLY, SER, GLY, GLU, ALA, ASN and this sequence is homologous to the IIB intermediate 2, α IIa A chain beginning with residue 14. Studies of the cleavage of IIH-intermediate 1 with factor Xa in the presence of DFP and Hirrudin and with thrombin in the presence of soybean trypsin inhibitor indicate that the IIH intermediate 2 originally produced by factor Xa is secondarily cleaved by thrombin. Thrombin cleavage of the factor X- produced IIH-intermediate 2 removes the amino-terminal 13 residues of this fragment. The amino-terminal sequence of the factor Xa produced intermediate 2 is THR, SER, THR, -, GLU. Supported by NIH grant HL 16150 and the American Heart Association.
The Role of Membrane Receptors in Pathways of Human Platelet Activation by Endotoxin and other Microbial Products
Authors: Hawiger A. , Hawiger J. , Timmons S. , Steckley S. , Cheng C.
Published: June 2019

Endotoxin-producing gram-negative rods cause thrombocytopenia and contribute to shock and death of 25% of bacteremic patients. Cellular pathways of clot-promoting activity of endotoxin compared with other microbial products were studied. Membrane receptor for endotoxin was demonstrated using human platelets as target cells separated from plasma proteins by albumin gradient-Sepharose 2B gel filtration. Endotoxin receptor was distinct from thrombin-sensitive protein and from IgG Fc receptor. It was resistant to neuraminidase and trypsin. It was related to membrane hydrophobic region(s) as demonstrated with fluorescent probe. Endotoxin-receptor interaction triggered unmasking of platelet procoagulant (Platelet Factor 3) and selective release of 3H-serotonin. A similar receptor was found for histoplasmin (yeast antigen). In contrast, staphylococcal protein A (cell wall antigen) required IgG Fc fragment and complement for its interaction with platelet membrane.

Thus in addition to the well known indirect pathway of activation of human platelets by microbial antigens through receptors for IgG Fc fragment and complement there is a direct pathway of activation due to the existence of specific antigen receptors on human platelets.

(Supported by NIH Grants HL 16646 and HL 08399 and VA.)
Immunohistological Studies of Coagulation Factors in Normal Blood Vessels and Platelets
Authors: Bloom A. L. , Giddings J. C. , Shearn S. A. M.
Published: June 2019

Rabbit antisera to factors II, V, VIII (related protein), X, XI, fibrinogen and fragment D have been used to localise these factors in normal blood vessels and platelets by an indirect fluorescent antiglobulin technique. Localisation of factors V and VIII (RP) confined, to the endothelium of normal blood vessels was confirmed but the presence of fibrinogen and fragment D at this site was variable and these latter antigens were also demonstrated in the sub-intima and media. There was no evidence for the presence of prothrombin and factor X in normal blood vessels. Platelets, separated by albumingradient centrifugation were washed up to 12 times in buffer with and without Ca++. Factor VIII (RP) and factor V were present in platelets and resisted removal by repeated washings. Initial studies indicated that factor XI is not present in platelets or is easily removed. Weakly positive reactions were obtained for prothrombin with four-times washed platelets but the reaction was enhanced by repeated washings. Factor X (or Xa) was removed from platelets by Ca-free buffer but not by Ca - containing buffer. The results indicate the selective presence of coagulation factors in vascular endothelial cells and platelets and are consistent with the finding of a Ca-dependent link between phospholipid and factor Xa which was also demonstrated in conventional chromatographic studies.
Current Knowledge of the AHF-Like Antigen
Authors: Meyer Dominique
Published: June 2019

Factor VIII and von Willebrand Factor activities are associated with a high molecular weight protein which can be isolated from plasma and may be studied by immunological methods. Homologous antibodies to Factor VIII are directed towards the active site of the Factor VIII molecule; they do not neutralize Willebrand Factor activity and do not precipitate with normal plasma. The use of such antibodies has allowed the distinction between Hemophilia A+ and A-. Specific precipitation of Factor VIII antibodies using polyethylene glycol will be reported, allowing typing of heavy and light chains of purified antibodies. Heterologous antisera prepared in rabbits against purified human Factor VIII complex neutralize Factor VIII and von Willebrand Factor activities and precipitate with AHF-like antigen. Estimation of this antigen in plasma has allowed (1) the differenciation of the molecular abnormalities in Hemophilia A and classical von Willebrand’s disease; (2) the comparison between normal and Hemophilic AHF-like antigen; (3) the detection of carriers of Hemophilia A; (4) the study of variants of von Willebrand’s disease; (5) the demonstration of this antigen in platelets and in endothelial cells. Factor VIII activity and AHF-like antigen are probably separate entities, circulating as a complex in normal plasma, as suggested by the following experiments: transfusion studies in von Willebrand’s disease; immuno-adsorption studies; comparison of Factor VIII complex in cryoprecipitate and supernatant; and dissociation in high salt buffer, demonstrating that Factor VIII includes two biologically linked but distinct fragments, of high (HMW) and low (LMW) molecular weight. The non functional HMW subunit, controlled by an autosomal locus, is identified by the presence of AHF-like antigen and Willebrand Factor activity. The LMW subunit, product of an X-chromosome locus, does not contain AHF-like antigen, but it carries Factor VIII activity, as demonstrated by the following facts: inactivation by both human and rabbit antibodies to Factor VIII; transient activation by thrombin; obtention of antisera which specifically inactivate Factor VIII.
Hemophilia: Modified by a Post-Exercise Plasminogen Activator
Authors: Hampton J. W. , Weidenbach A. , Skye G. E. , Rubenstein C. , Taylor F. B.
Published: June 2019

A family with 9 members of both sexes in 4 generations was demonstrated to have enhanced fibrinolysis associated with excessive bleeding. Three with the worst bleeding, all males, had 10% Factor VIII activity (high antigen, bleeding time normal and ristocetin platelet aggregation normal) with no male to male transmission. Antiplasmin proteins, α-I antitrypsin and α-2 macroglobulin were present and functionally normal. Inconsistently elevated fibrin degradation products, low fibrinogen and short euglobulin lysis times first suggested that a plasminogen activator might be present and enhanced by physical activity. Epsilon amino caproic acid (4 gms/day) given to one member with low Factor VIII inhibited the plasma activator but his Factor VIII activity only rose from 10 to 30%. In 2 affected members tested after exercise the activator activity increased up to 10-fold and required exogenous plasminogen for its demonstration. Ammonium sulfate precipitation (35-45% ) of the post-exercise plasma was pH, temperature and concentration dependent and again required exogenous plasminogen. At optimal conditions for plasminogen activation plasmin was obtained with the ammonium sulfate precipitation. Chromatography of the precipitate on Sephadex G-200 showed separation of the activator from both plasminogen and fibrinogen. The protein showed no reaction with an anti-urokinase on double-immunodiffusion and did not require Hageman fragments for its activation. We conclude that the Factor VIII deficiency in this family more closely resembles classic hemophilia and that exercise-induced activator excess modifies the expression of the genetic disorder. The presence of the activator is not Factor VIII dependent since the addition of Factor VIII in vitro did not inhibit the activation of plasminogen.
On the Origin of the Aα Chain Heterogeneity of Human Fibrinogen
Authors: Semeraro N. , Collen D. , Verstraete M.
Published: June 2019

Human fibrinogen was highly purified with over 95 per cent yield by affinity chromatography on insolubilized fibrin. The chains were separated after reduction and S-carboxy-methylation by CM-cellulose chromatography. The extent of Aα chain degradation was quantitated by SDS-polyacrylamide gel electrophoresis in combination with densitometric scanning. Calibration curves with mixtures of intact Aα chains and Aα chains degraded to variable extents revealed that the reproducibility of the method was better than 10 per cent.

Fresh frozen plasma as obtained from the blood bank contained 11.3±3.7 per cent degraded Aα chains (n = 5). Fresh plasma collected on aprotinin (Trasylol) and p-nitrophenyl-p’-guanidinobenzoate, (p-NPGB) and immediately processed at room temperature contained 6.4±2.5 per cent degraded Aα chains (n = 4). Plasma obtained from blood stored for 3 to 6 weeks at 4° C contained between 38 and 61 per cent degraded Aα chains (n = 3).

These results indicate that the major portion of the fibrinogen Aα chain heterogeneity demonstrated in outdated human plasma is the result of in vitro degradation which occurs during storage and that the more protective measures one takes the lower the residual Aα chain degradation. The extent to which the residual 6.4 per cent of Aα chain degradation results from incomplete prevention of in vitro proteolysis or from in vivo fibrinogenolysis remains to be established.
The Presence and Significance of Electrophoreticaily Normal and Abnormal Factor VIII Related Protein in von Willebrand’s Disease
Authors: Peake I. R. , Bloom A. L. , Giddings J. C.
Published: June 2019

The presence of normal amounts of a factor-VII related protein (FVIIIRP) which has abnormal electrophoretic mobility at pH 9.2 has been demonstrated on two dimensional crossed immunoelectrophoresis in some patiente with von Willebrand’s disease (vWd) and these investigations have now been extended to a study of over 14 patients. In those with reduced levels of FVIIIRP, concentrates were prepared by precipitation with 3% ethanol at -3°C. The results showed that in three of the patients with “typical vWd” in whom levels of procoagulant factor VIII and FVIIIRP were parallel and both always reduced the latter had abnormal electrophoretic mobility indistinguishable from that of the “atypical” cases. In all patients in whom abnormal FVIIIRP was demonstrated the bleeding time was grossly prolonged and ristocetin platelet aggregation grossly impaired. In patients in whom the mobility of FVTIIRP was indistinguishable from normal the bleeding times and ristocetin platelet aggregation were sometimes less severely impaired despite gross reduction of the circulating level of FVIIIRP. It is suggested that in vWd synthesis of abnormal variants cf FVIIIRP is relatively common but that in some patients there is reduced syntehsis of normal FVIIIRP.
Tissue Factor Activity of Cultured Macrophages. Enhancement by Specific Antigenic Stimulation
Authors: Beguin S. , Goutner A. , Josso F.
Published: June 2019

Peritoneal macrophage suspensions (95 p. cent pure) were obtained in rabbits either intact or previously immunized against human serum. Cell suspensions exhibited a clear tissue factor activity, closely related to the cell concentration.

Macrophages (10000/mm3) were cultured in the presence of PO4)2 Ca3 adsorbed and decomplemented serum of either rabbit or human origin. Clotting activity of the supernatant medium was tested after 24 hour culture. In immunized animals, tissue factor activity was much higher when the cells were cultured in the presence of the antigens (human serum) instead of rabbit serum; no difference was observed in intact rabbits.

Contamination of the cell suspensions by lymphocytes seems too low to play a role in this stimulation of macrophage activity. The reported data suggest that antibodies bound to macrophage membrane react with the medium antigens with subsequent increase of the cell activity.

Such a phenomenon could initiate the activation of the clotting system observed in some immune reactions.
Modifications of Platelets that Affect Platelet Adherence to Collagen or Damaged Aorta aud Platelet Survival
Authors: Cazenave J.-P. , Reimers H.-J. , Greenberg J. , Niewiarowski S. , Packham M. A. , Mustard J. F.
Published: June 2019

Platelet changes which govern their removal from the circulation are poorly understood We have modified platelets by thrombin degranulation, treatment with plasmin, removing surface sialic acid with neuraminidase, or exposure to sodium periodate. Washed rabbit platelets were prelabeled with 51Cr so that their adherence to collagen-coated glass surfaces or to intima of damaged aortas could be quantitated and their survival in vivo studied (Cazenave et al., J. Lab. Clin. Med. 82, 978, 1973; Reimers et al., Br. J. Haemat, 25, 675, 1973). The thrombin-treated platelets from which more than 70% of the serotonin had been released, showed decreased adherence to collagen (by 43±5%) and decreased adherence to damaged aorta (by 74 ±2%). Survival time for thrombin-treated platelets was 82±2 hr and for control platelets, 82±2 hr. Platelets treated with plasmin were insensitive to ADP unless fibrinogen were added. Plasmin treatment diminished platelet adherence to collagen by 40±5% and to damaged aorta by 68±2%. The survival time was 93±2 hr for plasmin-treated platelets (controls, 88±3 hr). Treatment with purified neuraminidase removed 8-35% of total platelet sialic acid with no release of granule contents. This did not inhibit platelet adherence to collagen or damaged aorta. If more than 15% of the sialic acid were removed, the platelets were cleared from the circulation within 1 hr. Exposure of platelets to NaIO4 (0.1-1 mM) for up to 10 min diminished adherence to collagen by 47 ±4%. When platelets were treated with 0.5 to 1 mM NaIO4 more than 70% of them were cleared from the circulation within 1 hr. Thus, treatments which alter platelet interaction with surfaces do not necessarily affect platelet survival, whereas removal of surface sialic acid shortens platelet- survival but does not influence platelet adhesion to surfaces. NaIO4 affects both adherence and survival but it may affect several platelet components.
Radioiodinated Human Fibrinogen. In Vitro Effects of Increasing Iodination
Authors: Ly B. E. , Kierulf P.
Published: June 2019

Fibrinogen preparations with increasing contents of iodine, ranging from 0.2 to 20 atoms of iodine per molecule fibrinogen, were obtained with the ICl method. Aggregation and shortening of the thrombin clotting time occurred when the content of iodine exceeded 3 atoms per molecule.

Upon the action of thrombin, the increase in N-terminal glycine, reflecting fibrin formation, was almost identical in native and iodinated fibrinogen. At visible gelation, however, decreased amounts of N-terminal glycine were found in heavily iodinated fibrinogen, thus indicating enhanced fibrin polymerization. N-terminal analysis of heavily iodinated fibrinogen demonstrated a deficiency in N-terminal tyrosine concomitantly with the apparance of a new N-terminal amino acid, identified as mono-iodo-tyrosine.

Polyacrylamide gel electrophoresis at pH 8.9 revealed an increase in mobility following extensive iodination, but no shift in the isoelectric point was observed upon isofocusing.

Neither clottability nor the behaviour of fibrinogen and its subunit polypeptide chains on SDS-gel electrophoresis was affected by iodination.
Disseminated Intravascular Coagulation: Experience in a Major Cancer Center
Authors: Al-Mondhiry Hamid
Published: June 2019

Disseminated intravascular coagulation (DIC) developed in 89 patients seen at Memorial Sloan-Kettering Cancer Center between 1971 and 1974. Criteria for the diagnosis of DIC wore: 1. Hypofibrinogenemia or > 50% reduction in plasma fibrinogen on serial determinations. 2. Prolonged thrombin time and/or elevated titer of fibrinogen-related material in the serum. 3. Thrombocytopenia not due to drugs or disease. Patients with severe liver disease or uremia were excluded. The patients included 19 with leukemia (17 acute), 3 with multiple myeloma, 15 with lymphoma, 46 with metastatic solid tumors (12 genitourinary, 10 lung, 9 breast, 8 gastrointestinal, 7 miscellaneous), 4 with vascular tumors, and 3 without tumor. Other conditions which might have precipitated or initiated DIC such as gramnegative sepsis (20 patients), mild liver impairment (31 patients), and mucin secreting tumors (4 patients), were also noted. Four patients with vascular tumors, two with leukemia, and one with vasculitis had markedly shortened fibrinogen survival. Bleeding occurred in 75% of the patients and was fatal in 36%. Thromboembolism occurred in 22.5%. Thirteen percent were asymptomatic. Treatment with heparin was helpful in only three of twenty patients. Eighty percent of the patients died within one to over 30 days of the onset of DIC. Post mortem evidence of DIC was present in 18 of 43 autopsies.

Results of this study indicate that DIC is a frequent complication of a wide variety of tumors and that its occurrence causes morbidity and mortality in a significant number of patients. Treatment with heparin is of little help unless remission is induced and the precipitating factor(s) are reversed.
Dosage of i.v. Brinase in Man Based on Determination of the Brinase Inhibitor Capacity and Coagulation Studies
Authors: Frisch E. P. , Blombäck M. , Kiessling H.
Published: June 2019

Brinase, a proteolytic enzyme from Aspergillus oryzae, is in plasma inhibited by two endogenous inhibitors. In vitro addition of the enzyme to plasma lowers the brinase inhibitor capacity (BTC); the calculated regression lino was used to predict lowering of BIC resulting from i,v. infusion of orinase in man.

A total of 355 i.v. brinase infusions (single doses of 50 200 mg) were given to 36 patients suffering from advanced peripheral arterial disease. Anticoagulant treatment (dicoumarolor heparin) was given concomitantly to 34/36 patients. BIC was determined by the azocollagen method before and after infusion of the enzyme. Good correlation was found between calculated lowering of BIC and determined post-infusion values. Individual dose requirements could be calculated and free proteolytic activity by depletion of inhibitors was avoided.

Changes of α1-antitrypsin and α2-macroglobulm induced by brinase infusion showed the same trends as those recorded for BIC; there was no quantitative correlation. Fibrinogen was lowered by infusion of the enzyme, subnormal values were determined at 4 occasions. There was a moderate increase of fibrinogen degradation products and thrombin time was prolonged. Positive ethanol gelation test following brinase infusion was a constant finding. Brinase induced slight lowering of Thrombotest values and prolongation of the activated partial thromboplastin time; dosage of anticoagulants was not influenced. Other coagulation tests showed insignificant changes only.

In two patients, not on anticoagulants, with a. history of renal disease, transient renal failure occurred subsequent to brinase infusion. Laboratory data indicated intravascular coagulation as a possible cause. Bleeding complications were not observed.
Effects of Inhibitors of ADP-induced Aggregation on Platelet Nucleo-sidediphosphate Kinase (NDK) Activity
Authors: Packham M. A. , Mustard J. F. , Guccione M. A. , Perry D. W.
Published: June 2019

We showed previously (Am. J. Physiol. 227, 1143, 1974) that ADP reacts with NDK on the surface of platelets; when 14C-ADP is added to suspensions of washed rabbit platelets, it is partially converted to 14C-ATP in the suspending fluid. This is more difficult to show with human platelets because of their greater ATPase activity. With both species, -external ATP (10 μM), and other NTP’s at higher concentrations, enhance formation of “C-ATP form “C-ADP. All the NTP’s inhibit ADP-induced aggregation, possibly by acting as external donors of a phosphoryl group which, in their absence, is normally donated to ADP by platelets. Studies of other inhibitors have been done with suspensions -of washed rabbit platelets. The other NDP’s and AMP inhibit, ADP-induced shape change, aggregation, and conversion of 14C-ADP to 14C-ATP, probably by competition for the same site on NDK. AMP inhibits NDK both in the presence and absence of external ATP. Parachloromercuribenzene sulfonic acid has a similar effect. In contrast, prostaglandin E1 and hydroxylamine inhibit ADP-induced shape change and aggregation, but inhibit NDK only in the absence of external ATP, not in its presence. ADP-induced shape change without aggregation occurs in the presence of EGTA or EDTA but only half as much 14C-ADP is converted to 14C-ATP in the first minute. EGTA does not inhibit NDK in the presence of external ATP but EDTA does. Inhibition of NDK by adenosine was observed in some experiments but the adenosine derivatives 2-n-amylthioadenosino, 2-cyclopentylthioadenosine and 2-cyclophexylthioadenosine (Kohjin Co., Tokyo) inhibited strongly both in the presence and absence of external ATP; these compounds also inhibit ADP-induced aggregation. Thus all compounds tested which inhibit NDK also inhibit ADP-induced aggregation, indicating that NDK may be the ADP receptor on the platelet surface. Donation of a phosphoryl group from the platelets to ADP, catalyzed by this enzyme, may be responsible for initiating ADP-induced platelet aggregation.
Specificity of an Acquired Antiplatelet Antibody Occuring in a Polytransfused Thrombasthenic Patient
Authors: Levy-Toledano S. , Bredoux R. , Rendu F. , Tobelem G. , Degos L. , Caen J. P.
Published: June 2019

An IgG antibody which developed in a polytransfused thrombasthenic patient reacted in complement fixation with platelets from 350 normal individuals but not with platelets from 8 other thrombasthenic patients. It agglutinates human platelets in PRP but not thrombasthenic platelets. This agglutination increased if the PRP was preincubated at 37°C or if the platelets were isolated before the addition of the antibody. Dog but not rabbit platelets are agglutinated by the patient plasma. Neither adenosine, nor PGE1 inhibit this agglutination which is slightly reduced by acetylsalicilic acid and disappears with EDTA and EGTA (3,8 mM).

Its activity is reduced or abolished after incubation with control platelet membranes but not with those obtained fron) thrombasthenic or rabbit platelets.

It does not inhibit the ADP-induced shape change of normal platelets, and it prevents all the ADP mediated platelet aggregations but, not those induced by bovine factor VIII and ristocetin.

This antibody seemed to be directed against a molecule absent or structurally modified in thrombasthenic platelets which would be involved in platelet aggregation and more especially in ADP mediated platelet aggregation.
Low-Dose Heparin for Prevention of Deep-Vein Thrombosis in Patients Undergoing Hip-Arthroplasty
Authors: Citterio L. E. , Mannucci P. M.
Published: June 2019

The efficacy of low-dose heparin in patients undergoing hip surgery is not established. 95 patients over the age of 40 undergoing an elective operation of hip-replacement arthroplasty have been randomly allocated to two groups on the basis of numbered sealed envelopes. 45 patients received 5000 units of subcutaneous calcium heparin preoperatively and 8-hourly after this until they were fully mobile (days of treatment: mean 14.1, SD ±5.0), 50 patients received no treatment and acted as controls. The two groups were comparable in respect to age, weight, blood-group, previous disease, findings on admission, type of operation and other drugs used. The 125I-fibrinogen-uptake test was used to diagnose DVT by injecting in all patients 100 μCi postoperatively on the day of operation and scanning the legs for seven days. The frequency of DVT was 42% (21/50) in the control group and 20% (9/45) in the treated group. The difference is statistically significant (P < 0.05) using a χ2 test with Yates’ correction. Since Hampson et al. (Lancet ii, 795, 1974) have recently suggested that low-dose heparin has no significant effect on the frequency of DVT after total-hip replacement arthroplasty but appears to delay its onset, a new trial is presently being carried out in which 100 μCi of 125I fibrinogen are injected twice and the legs scanned for 14 days.
Studies on the Clottable Protein of Limulus Polyphemus
Authors: Fumarola D. , Pasquetto N. , Telesforo P. , Donati M. B.
Published: June 2019

The circulating blood cells of Limulus Polyphemus contain material (coagulogen) which can be clotted by bacterial endotoxin or by trypsin. In this study, several substances interacting with the clotting mechanism of mammalians have been tested on lysates from Limulus cells. Gelation of the amebocyte lysate was observed in the presence of sterile preparations of thrombin, reptilase, botropase, thrombin-coagulase and rabbit or human thromboplastin. No gelation occurred when calcium chloride, ethanol or protamine sulphate were added to the lysate. Gelation was not inhibited by heparin even in the presence of its cofactor. Plasmin neither affected the ability of coagulogen to gel nor caused visible dissolution of preformed gels. No staphylococcal dumping occurred in the presence of the coagulogen. Acidification experiments (method of Solum, Thrombos. Res., 2 55, 1973) suggested that thromboplastin, thrombin and the thrombin-like enzymes tested in this study are able to activate, like endotoxin, an inactive proenzyme in the lysate to form an enzyme responsible for gelation of the coagulogen. In contrast, trypsin would be able to transform directly the coagulcgen into a gel.
Disseminated Intravascular Coagulation in Dengue Haemorrhagic Fever
Authors: Himawan T. , Kho L. K. , Melani S.
Published: June 2019

The most dreaded complications of dengue fever are haemorrhagic manifestations (Dengue Haemorrhagic Fever) and shock (Dengue Shock Syndrome). We have the opportunity to carry out clinical and haematological studies on 68 selected cases of disseminated intravascular coagulation (DIC) occuring in DHF. Shock was encountered in the majority of the cases (70.6%), while haemorrhagic manifestations were observed in all the children. Other laboratory findings revealed the presence of anemia in 38.1%. hemo-concentration in 72.6% ; the W. B.C. varied between 1,600 and 39,400, thrombocytopenia 83.8%. Fragmented red cells were found in the periferal blood smear. Prolonged bleeding time 69.3%; prolonged clotting time 48.7%; prolonged prothrombine time 66.6%; prolonged partial thromboplastin time 56% ; fibrinogenopenia 80% and positive tourniquet test 85.7%.

The management was directed to the improvement of the general condition with intravenous fluid replacement therapy, electrolytes, plasma expanders or blood. Heparin was administered to 36 children with severe DIC. The mortality rate was high (33.8%).
Sex Hormones and Phytomenadione Evidence for their Relationship in an Induced Coagulation Defect
Authors: Sanderson J. H. , Chart I. S. , Shingles C.
Published: June 2019

Rats dosed with benzyl 3,5-dichloro-2,6-difluoro-2-pyridyl ether show a coagulation defect which is much more severe in males than females, and dose levels can be chosen at which males only are affected. The defect is produced by a lowering of Factors VII X and II.

This sex differentiation can be abolished if male rats are castrated or if they are treated with oestrogens, and normal male rats can be protected by treatment with Phytomenadione.

It is suggested that the use of this compound may provide a tool for understanding mechanisms of anti-coagulation.
Factor XI Deficiency, Juvenile Rheumatoid Arthritis (JRA), and Systemic Lupus Erythematosus (SLE) : Report of the First Case
Authors: Blatt P. M. , Yount W. J. , Roberts H. R. , Hadler N. M. , Utsinger P. D. , Korn J. H.
Published: June 2019

A number of inborn errors have recently been associated with a diathesis for collagen vascular disorders. For example, hereditary deficiencies of Clr or C2 components of complement have been associated with certain features of SLE. We wish to record the syndrome of JRA with evolution to SLE associated with a familial deficiency of Factor XI (DTA). The propositus, a 26 year old Sephardic female, presented at age 4 with symmetrical destructive polyarthritis, fever and rash. Progressive renal disease began in her early teens, and she developed multiple diagnostic criteria for SLE beginning at age 25. Three maternal relatives manifest arthritis and two showed mild bleeding after surgery. Physical findings included a deforming arthritis and rash. Low levels of C3 and C4, high titers of antibody to native DNA, positive LE cell preparations and Clq precipitins were present. Skin biopsy showed deposits of IgG, IgM, Clq, C3 and C4 at the dermal-epidermal junction.

Factor XI levels were 5 per cent in the propositus and mother, with normal levels in a sister. There was no evidence for a circulating anticoagulant and other clotting factors were normal.

We believe the present case represents the first association of an inborn error in the clotting system (Factor XI deficiency) and collagen vascular disease with immune complex deposition.
Membrane Glycoproteins of Human and Rabbit Platelets: Studies In Vitro and after Circulation in Vivo
Authors: George J. N. , Lewis P. C. , Sears D. A.
Published: June 2019

The initial events of hemostasis and thrombosis involve platelet contact interactions and may be mediated by surface glycoproteins. Human and rabbit platelets were labeled with 125I-diazotized diiodosulfanilic acid (I), which reacts covalently with proteins, and proteins were separated by SDS-polyacrylamide gel electrophoresis. Only exposed membrane proteins were labeled because: 1) protein specific activity of membranes was 4-7 times that of whole platelets, 2) different proteins were labeled when I was reacted with isolated membranes, and 3) trypsin-hydrolysis of labeled intact platelets altered the radioactive peaks. Like Phillips (Biochem. 11, 4582, 72) and Nachman et al. (JBC 248, 2928, 73) we found that lactoperoxidase iodinated the 93,000 dalton glycoprotein (GP) of human platelets. In contrast, I labeled both the 93,000 and 118,000 dalton membrane GP of human platelets, and all 3 membrane GP of rabbit platelets.

Rabbit platelets labeled simultaneously with I and 51Cr had identical density and therefore age distribution of the 2 labels. After infusion into rabbits, initial recovery of I was 23% of the Cr recovery. After 3 hrs, I disappearance was exponential and more rapid (T/2 = 17 hrs) than the linear Cr disappearance (T/2 = 30 hrs, p < .01). This was due to in vivo removal of I from circulating platelets since 1 did not elute more rapidly from platelets harvested after 3 hrs circulation and incubated in plasma at 37° (T/2 of I elution = 43 hrs, Cr = 33 hrs). Platelets harvested after 14-20 hrs circulation had the same distribution of I on the membrane GP as before circulation. We postulate that this symmetrical label loss indicates uniform loss of membrane GP, suggesting that platelets lose pieces of their plasma membrane during circulation. This could occur during contact interaction in the process of hemostasis.
Inhibition of Formation and “Protection” of the Factor X-Activator by Hirudin
Authors: Lechler E.
Published: June 2019

The essential role of thrombin in the formation of the factor X-activator from F. IXa, F. VIII and phospholipids has been documented in the literature. We have sedimented an “active” factor X-activator by high speed centrifugation when the preparations of factors used in the formation of the factor X-activator were contaminated with traces of (pro-)thrombin. In the presence of hirudin only a weak factor X-activator activity could be sedimented.

In “plasmas” from blood incubated with hirudin no gross changes of coagulation factors were observed after gel filtration (which removes hirudin). However, following kaolin contact activation of blood containing hirudin prothrombin and F. X peaks were no more detectable in gel filtration. It is suggested that 1. trace amounts of thrombin for the factor X-activator formation still have become available, and 2. the consumption of prothrombin and of factor X is due to the protection of the factor X-activator against thrombin by hirudin.
Retraction of Thrombin-Induced Fibrin by Rhabdomyosarcoma BA 1112 Cultured Cells
Authors: Donati M. B. , Dolfini E. , Cavenaghi A. , Morasen L. , Gaetano G. de
Published: June 2019

The capacity to induce fibrin retraction has been considered a specific property of platelets until recently, when Niewiarowski et al. (Proc. Soc. Exp, Biol. Med. 140, 1199, 1972) observed fibrin retraction induced by human fibroblasts. As a part of a larger study on the interactions of cultured cells with fibrin, we have investigated the ability of the following cell lines to retract fibrin: KB (human oral epidermoid carcinoma); HeLa (human cervix carcinoma); Chang Liver (human, normal epithelium; Chang Conjunctiva (human normal epithelium); NCTC clone 929 (L) (fibroblasts from C3H/AN mice) and BA 1112 (rhabdomyosarcoma developed on WAG/Rji inbred rats). Cells were cultured in Eagle’s MEM, in Hank’s blanced salt solution plus 10% calf serum, removed from the flasks by trypsin treatment and resuspended at a concentration of 2 × 106/ml in Tyrode-albumin solution, containing Ca++ and Mg++. Human citrated platelet-poor plasma was clotted in a test tube at 37° C by thrombin in the presence of either the cell suspensions or buffer. Only BA 1112 cells were able to retract fibrin; the presence of Ca++, cellular integrity and random distribution in the sample were required for this activity. BA 1112 cells were able to modify the structuration of thrombin-induced fibrin as indicated by the marked increase of the maximal amplitude of thrombelastogram. BA 1112-induced fibrin retraction was inhibited by PGE1 and by some pyrimido-pyrimidine derivatives, not by apsirin. No retraction occurred when reptilase instead of thrombin was used as the clotting agent, even if the cells were preincubated with ADP. These results suggest that BA 1112 cells have a susceptibility to thrombin similar to that of platelets; this hypothesis is interesting in view of the muscular origin of these cells.

(Supported by Grant NIH-PHRB-10RO1 CA 12764–01.)
The Puzzle of Immunologic Reactivity to Factor VIII in Haemophilia A.
Authors: Frommel D. , Gaillandre A. , Meunier L. , Allain J. P.
Published: June 2019

Factor VIII neutralizing antibodies occur in 6–21% of transfused haemophiliacs A. Two types of specific immune response can be observed. In high responding patients, anti-factor VIII antibodies develop after relatively short exposure to the antigen, and anamnestic responses are characterized by antibodies of high affinity and titer. In low responding haemophiliacs, occurrence of anti-factor VIII antibodies is delayed and secondary response is either inconstant or requires strong antigenic challenge. Both types of immune response can be observed in kindreds. Low responsiveness to human factor VIII can switch to high responsiveness as a result of stimulation with heterologous factor VIII.

In experimental situations, it has been found that immunologic reactivity to weak native isologous antigens is dose-dependent and shows extreme variations from one individual to another. The state of high, low or non responder is, in many instances, controlled by well defined genetic mechanisms. Considering possible analogies between immune response to factor VIII in man and specific responses controlled by Ir genes in animal, it seems useful to determine whether or not the capacity of immune response to factor VIII is linked to loci coding for HL-A specificities or is expressed on other cellular receptors amenable to analysis.
Comparative Study of Blood Fibrinolytic Activity and its Inhibitors in U.S. Blacks and Whites
Authors: Szczeklik A. , Dischinger P. , Kueppers F. , Tyroler H.A. , Hames C. G. , Cassel J. , Creagan S.
Published: June 2019

In 182 black and 155 white men aged 20 to 50 years, all presently employed and living in southern Georgia, the following parameters related to fibrinolysis and its inhibitors were determined: Plasma fibrinolytic activity, fibrinolytic activity of euglobulins, plasminogen in euglobulins, antiplasmin, alpha1 antitrypsin and alpha2 macroglobulin. The first 4 components were determined by means of a fibrin plate technique while the last 2 were assayed using immunodiffusion and electro immunodiffusion, respectively. Whites were similar to blacks with respect to age, height, weight, physical activity and serum cholesterol. Blacks, however, had lower serum triglyceride level and a greater proportion of blacks than whites were cigarette smokers. In addition, the blacks in the sample had a lower socio-economic status as determined by data on education and occupation. Resting fibrinolytic activity was detected in the plasma of 26.6% of blacks and in only 9.3% of whites, this difference being highly significant by the K-S test (P = .02). Antiplasmin levels were significantly lower in blacks than in whites (P < 0.001). While mean values of both alpha1 antitrypsin and alpha2 maeroglobulin were lower in blacks than whites, these differences were not statistically significant. There was no correlation between cigarette smoking and the fibrinolytic components studied. From regression analysis, it is apparent that differences in fibrinolysis between blacks and whites in the sample were not merely secondary to differences in triglyceride levels. Blacks, but not whites, demonstrated the decrease in fibrinolytic activity and alpha1 antitiypsin with rise in social class. The results obtained indicate that in a community where the incidence of coronary heart disease has been previously shown to be markedly higher in white than in black men. whites also exhibit a significant impairment of fibrinolysis in comparison with blacks.
Computational analysis of composition-structure-property-relationships in NZP-type materials for Li-ion batteries
Authors: Mutter Daniel , Urban Daniel F. , Elsässer Christian
Published: June 2019

Compounds crystallizing in the structure of NaZr2(PO4)3 (NZP) are considered as promising materials for solid state electrolytes in Li-ion batteries. Using density functional theory (DFT), a systematic computational screening of 18 NZP compounds, namely, LiX2(LO4)3 with X=Ti, V, Fe, Zr, Nb, Ru, Hf, Ta, Os, and L=P, Mn, is performed with respect to their activation energies for vacancy-mediated Li migration. It is shown how the different ionic radii of the cationic substitutions influence structural characteristics such as the octahedron volumes around Li ions on the initial-state and transition-state sites, which affect the activation energies (“composition–structure–property” relationships). The prevalent assumption that structural bottlenecks formed by triangularly arranged oxygen atoms at a certain location along the migration path determine the energy barriers for Li migration is not supported by the DFT results. Instead, the ionic neighborhood of the migrating ion in the initial and in the transition state needs to be taken into account to relate the structure to the activation energies. This conclusion applies to Na-containing NZP compounds as well.
Nonlinear discovery of slow molecular modes using state-free reversible VAMPnets
Authors: Chen Wei , Sidky Hythem , Ferguson Andrew L.
Published: June 2019

The success of enhanced sampling molecular simulations that accelerate along collective variables (CVs) is predicated on the availability of variables coincident with the slow collective motions governing the long-time conformational dynamics of a system. It is challenging to intuit these slow CVs for all but the simplest molecular systems, and their data-driven discovery directly from molecular simulation trajectories has been a central focus of the molecular simulation community to both unveil the important physical mechanisms and drive enhanced sampling. In this work, we introduce state-free reversible VAMPnets (SRV) as a deep learning architecture that learns nonlinear CV approximants to the leading slow eigenfunctions of the spectral decomposition of the transfer operator that evolves equilibrium-scaled probability distributions through time. Orthogonality of the learned CVs is naturally imposed within network training without added regularization. The CVs are inherently explicit and differentiable functions of the input coordinates making them well-suited to use in enhanced sampling calculations. We demonstrate the utility of SRVs in capturing parsimonious nonlinear representations of complex system dynamics in applications to 1D and 2D toy systems where the true eigenfunctions are exactly calculable and to molecular dynamics simulations of alanine dipeptide and the WW domain protein.
Direct numerical simulation of transitional and turbulent round jets: Evolution of vortical structures and turbulence budget
Authors: Anghan Chetankumar , Dave Sagar , Saincher Shaswat , Banerjee Jyotirmay
Published: June 2019

Direct numerical simulation of transitional and turbulent round jets is reported in a comparative framework. Such a comparison is central toward revealing the roles that molecular viscosity and vorticity intensification play in the evolution of jets. The initial and intermediate evolution is differentiated based on the assessment of the starting jet, roll-up frequency, dynamics of vortex rings, and emergence of the secondary instability. Long-term behavior is differentiated based on the assessment of preferred mode frequency, time averaged vortical structures, half jet-width, and volume flow rate obtained from the time-averaged velocity field. The present study demonstrates that viscous damping of cross-stream vorticity plays a key role in establishing helical instability as the dominant mode in long-term evolution of the transitional jet. On the contrary, varicose mode is dominant in the turbulent jet, despite preferred mode frequency being the same in both cases. Finally, a novel attempt is made toward comparing individual terms constituting turbulence budget between both regimes. Through such a comparison, relative dominance of various transport mechanisms governing the evolution of turbulence kinetic energy (K) is revealed. It is observed that terms accounting for a forward cascade of K from inertial to smallest scales are comparatively larger for the turbulent jet, while those accounting for the backscatter of K are comparatively larger for the transitional jet. It is also established that turbulence dissipation is evidently the same for both jets. Thus, the property of turbulence dissipation being independent of Reynolds number for turbulent jets can also be extrapolated to transitional jets.