Flavio Habal

Oral 5-aminosalicylic acid for inflammatory bowel disease in pregnancy: safety and clinical course.

BACKGROUND Oral 5-aminosalicylic acid (5-ASA) has proven an effective maintenance therapy of ulcerative colitis and may also be useful in Crohns disease, but its safety in pregnancy has not been established. The present study therefore examined the course and outcome of pregnancies in patients with inflammatory bowel disease who continued to take oral 5-ASA. METHODS Ten patients with ulcerative colitis and 7 patients with Crohns disease with a total of 19 pregnancies were studied while they were receiving 5-ASA. All patients were previously in remission on 5-ASA, at a mean dose of 1.7 g/day (range, 0.8-2.4 g/day). They continued taking the drug without a change in dose and were followed up throughout their pregnancies and postpartum. RESULTS Eighteen pregnancies resulted in full-term delivery. No fetal abnormalities were found at delivery, and there were no clinical or biochemical abnormalities in the neonatal period. Four patients had a relapse. One patient required a colectomy but carried on to a full-term pregnancy. One patient had a miscarriage, but she had miscarried on four previous occasions before taking 5-ASA. She subsequently had a successful pregnancy on 5-ASA. CONCLUSIONS Oral 5-ASA appears to be safe for the management of inflammatory bowel disease during pregnancy.

Isolation of two functionally different kininogens from human plasma—separation from proteinase inhibitors and interaction with plasma kallikrein☆

Abstract A high (HMW) and a low (LMW) molecular weight kininogen were isolated in highly purified form from human plasma, using QAE-Sephadex chromatography, followed by ammonium sulfate precipitation, gel filtration through Sephadex G-200, re-precipitation with ammonium sulfate, CM-Sephadex and SP-Sephadex chromatography. The initial preparative step was done at room temperature and the remaining procedures at 4°. In aqueous media, the apparent molecular size of the HMW-kininogen was about four times the size of the LMW-kininogen (200,000 vs 50,000). During the process of purification, proteinase inhibitors were separated from the two kininogens: α 1 -antitrypsin and α 2 -macroglobulin from the LMW-kininogen preparations: Cl-inactivator and inter-α-trypsin inhibitor from the HMW-kininogen preparations. There was a well defined functional difference between the two kininogens with respect to kinin generation by plasma kallikrein. This enzyme released kinin at a much faster rate from the HMW-kininogen than from the LMW-kininogen. When equipotent preparations of kininogens were incubated for 10 min with kallikrein, 60 times more enzyme was required to release the same amount of kinin from the LMW-kininogen as from the HMW-kininogen.

Review article: a decision‐making algorithm for the management of pregnancy in the inflammatory bowel disease patient

Inflammatory bowel disease affects patients who are in their reproductive years. There are many questions regarding the management of IBD patients who are considering or who are already pregnant. These include the effect of the disease and the medications on fertility and on the pregnancy outcome.

The inhibition of activated factor XII (hageman factor) by antithrombin III: The effect of other plasma proteinase inhibitors

Abstract Human factor XII was activated by adsorption onto kaolin in the presence of high molecular weight kininogen. The washed kaolin-containing precipitates activate prekallikrein to kallikrein. When antithrombin III was added to the reaction mixture, the conversion of prekallikrein to kallikrein was inhibited, the degree of inhibition depending on the concentration of antithrombin and the time of incubation. Heparin had a slight enhancing effect with low concentrations of antithrombin and short incubation times. However, the inhibition of the generated kallikrein by antithrombin III was markedly enhanced by heparin. Antithrombin III inhibited also the effect of activated factor XII on the partial thromboplastin time, using factor XII-deficient plasma. Of other plasma proteinase inhibitors used (α1-antitrypsin, α2-macroglobulin, C l -inactivator) only C l -inactivator inhibited activated factor XII.

Interaction between factor XII (Hageman factor), high molecular weight kininogen and prekallikrein.

Abstract Adsorption of factor XII and plasma kallikrein onto kaolin induces activation of factor XII. Trace amounts of high molecular weight (HMW)-kininogen markedly enhances the rate and the amount of activation of factor XII. Activation of factor XII was measured by its capacity to convert prekallikrein to kallikrein. This potentiating effect is dependent on the amount of HMW-kininogen. Low molecular weight kininogen has no enhancing effect. When the reactants are adsorbed to kaolin and the latter separated by centrifugation the prekallikrein-converting activity is detectable only in the kaolin-containing precipitates. Less activation of factor XII occurs when the HMW-kininogen is added not initially to the kaolin-factor XII mixture, but to the prekallikrein, indicating that the HMW-kininogen probably acts when factor XII is being activated, rather than at a later stage. Treatment of HMW-kininogen with excess trypsin, plasmin or kallikrein abolishes its potentiating effect. When HMW-kininogen is treated with increasing concentrations of kallikrein there is an inverse relationship between the generated kinin and the ability of the kininogen to enhance the activation of factor XII.

Standard Treatment of Ulcerative Colitis

Ulcerative colitis (UC) is an idiopathic, chronic inflammation of the colon which may present with a range of mild to severe symptoms. The disease may be localized to the rectum or can be more extensive and involve the left side of the colon or the whole colon. Treatment in UC is directed towards inducing and maintaining remission of symptoms and mucosal inflammation. The key parameters to be assessed for the most appropriate treatment are the severity and extent of the inflammation. Meta-analyses of published trials have shown that topical treatment with 5-aminosalicylic acid (5-ASA) is the treatment of choice in active distal mild-to-moderate UC. Oral aminosalicylates are effective in both distal and extensive mild-to-moderate disease, but in distal disease, the rates of remission are lower than those obtained with topical 5-ASA. New steroids, such as budesonide and beclomethasone dipropionate (BDP), administered as enemas, constitute an alternative to 5-ASA therapy. In some studies, these have been shown to be as effective as conventional steroids but with significantly lower inhibition of plasma cortisol levels. Patients with unresponsive disease or those with more severe presentation will require oral corticosteroids and sometimes intravenous therapy. Approximately 10% of patients with unresponsive UC have severe attacks requiring hospitalization. Patients with severe disease should be managed jointly by a medical and surgical team, and intensive intravenous treatment should be started with high-dose steroids. Early recognition of failure of therapy will allow the introduction of immunosuppressive therapy with intravenous cyclosporine. Patients who respond are shifted to oral cyclosporine associated with azathioprine/6-mercaptopurine, whereas those who fail will require proctocolectomy. Oral aminosalicylates are the first-line therapy in maintenance of remission. Topical 5-ASA may play a role in distal disease. Patients who are steroid dependent can be started on azathioprine or 6-mercaptopurine although it may take up to 3 months for the treatment to become effective. They may have reversible immediate side effects, such as pancreatitis or bone marrow suppression, which disappear upon discontinuation of therapy. Close monitoring of these hematologic and biochemical parameters will improve safety. The use of biologic therapy with infliximab in more severe disease has not been established.

Neutral Proteases of Human PMN Leukocytes with Kininogenase Activity

A neutral protease with kininogenase activity was isolated from human polymorphonuclear (PMN) leukocytes by cation exchange chromatography and gel filtration. The protease appears heterogeneous by cation exchange chromatography, isoelectric focusing and cationic disc gel electrophoresis, but homogeneous by gel filtration, sucrose density gradient ultracentrifugation, SDS-disc gel electrophoresis and immunoelectrophoresis. By carrying out the electrophoresis of the protease in acrylamide gels of varying concentrations, it was shown that they represent charge isomers. The protease was stable at pH 4-10, but labile to heat, being almost completely inactivated when incubated for 30 min at 70 degrees C. It exhibited proteolytic activity between pH 5 and 9, being maximal at 7.5-8.5. The molecular weight of the PMN protease was estimated to be about 20,000 daltons by gel filtration in aqueous buffer and about 26,000-28,000 daltons by SDS-disc gel electrophoresis and gel filtration in Sepharose 6B in the presence of the dissociating agent guanidine HCl. Its sedimentation coefficient was about 2.7S. Corresponding to the charge heterogeneity, by isoelectric focusing, the kinin-generating and esterolytic activities of the PMN granule lysate focused between pH 6.0 and 11.5, whereas the isolated PMN protease focused between 10.0 and 11.8. With respect to kinin generation, caseinolysis, and alanine esterase activity, the protease was inhibited by DFP and certain chloromethyl ketone inhibitors, as well as the plasma protease inhibitory a1-antitrypsin, a2-macroglobulin and antithrombin III. Both bradykinin and a methionyl-lysyl-bradykinin-like peptide were generated from highly purified kininogens by a lysosomal lysate containing the PMN protease. However, this assay was done with a crude enzyme preparation which contains an aminopeptidase capable of converting lysyl-bradykinin or methionyl-lysyl-bradykinin to bradykinin. When injected intradermally, the protease induced hyperemia, hemorrhage, and moderate enhancement of vascular permeability, but the mixture of the protease and kininogen induced a marked enhancement of vascular permeability.

The inhibition of human plasma Kallikrein by antithrombin III

Abstract Prekallikrein, prekallikrein activator, high molecular weight kininogen and antithrombin III were isolated from human plasma. The prekallikrein was converted to kallikrein by prekallikrein activator. The kallikrein had arginine esterase activity and generated kinin from kininogen. Both activities were inhibited by antithrombin III in concentrations of approximately 1 2 of that of plasma. Physiological concentrations of heparin were required for the rapid and complete inhibition of kallikrein. In the absence of heparin only partial and much slower inhibition was achieved. The proteolytic, i.e. kinincleaving activity of kallikrein was more readily inhibited than the esterolytic activity.

Generation of kinin by plasma kallikrein and plasmin and the effect ofα(in1) antitrypsin and antithrombin III on the kininogenases

low molecular weight Kgn I, which has the bradykinin (BK) sequence at its carboxyl terminus, and Kgn II, with the BK moiety inside the molecule (J.V. Pierce and M.E. Webster, in Hypotensive Peptides, E.G. Erd6s, N. Back, F. Sicuteri, eds., Springer, 1966), were equally good substrates for pepsin. All of the kinin was released in three minutes when 10 mg of Kgn B3.2c~ (containing ca. 15/Jg of BK/mg, as determined with trypsin) was incubated with 50/Jg of 2X crystallized pepsin at pH 2.0 and 37 ~ . The released kinin, equivalent to 18 #g of BK on the isolated guinea pig ileum, was stable in the incubation mixture. It was inactivated by carboxypeptidase B and could be converted to an 8-fold more active peptide upon incubation with human plasma aminopeptidase or dipeptidyl aminopeptidase I (DAP-I). The peptic Kinin was separated from the bulk of protein by gel filtration on Sephadex G-25 and was further purified by SP-Sephadex G-25 and CM-cellulose chromatography. The protein fraction did not generate any additional kinin upon incubation with trypsin or human plasma kallikrein. Several criteria were used to identify the purified peptic kinin as MLBK: a) it has the same retention volume as MLBK on an analytical SP-Sephadex C-25 column which is able to resolve BK, LBK, and MLBK; b) it was converted by treating with DAP-I to BK, identified by the 8-fold activity increase and by its now having the same retention volume as BK on the SP-Sephadex C-25 columnn; and c) it had the same amino acid composition as MLBK. The kinin released from other pure Kgns by pepsin was also identified as MLBK by some of the above criteria. Pepsin, like trypsin and plasmin, cleaves several peptide bonds in Kgns other than the bonds involved in the release of MLBK. The kinin-releasing activity of pepsin extends to pH 5.0 and is inhibited by pepstatin at both pH 2.0 and 5.0 Our highly purified preparations retained full Kgn activity at pH 2.0 after several days at room temperature (ca. 25~ MLBK was first isolated from bovine plasma which had been dialyzed against 0.01 M HCI and then incubated at pH 7.5 (D.F. Elliott and G.P. Lewis, Biochem. J. 95, 435, 1965). Since plasma contains pepsinogen, its activation in the acid dialysis step could explain the release of MBLK observed by Elliott and Lewis. GENERATION OF KININ BY PLASMA KALLIKREIN AND PLASMIN AND THE EFFECT OF oq ANTITRYPSIN AND ANTITHROMBIN III ON THE KININOGENASES M. FLAVIO, HABAL, E. CLEMENT, BURROWES and Z HENRY, M O V A T Division of Experimental Pathology, Department of Pathology, University of Toronto,

Lupus-Like Syndrome Caused by 5-Aminosalicylic Acid in Patients with Inflammatory Bowel Disease

BACKGROUND Although 5-aminosalicylic acid (5-ASA) preparations used to treat inflammatory bowel disease are reported to have fewer side effects than sulphasalazine, increased clinical use of these compounds has resulted in increased reports of significant side effects. OBJECTIVE To report four patients with antinuclear antibody-positive migratory arthralgias and acute inflammation unrelated to the underlying inflammatory bowel disease, fulfilling the criteria of a drug-induced lupus-like syndrome. SETTING A university-affiliated teaching hospital. INTERVENTION Cessation of treatment with 5-ASA compounds. RESULTS The cases described constitute a drug-induced lupus-like syndrome. All patients improved rapidly after discontinuation of 5-ASA compounds. CONCLUSIONS Reversible lupus-like syndrome appears to be a rare but significant side effect of 5-ASA compounds. Patients treated with 5-ASA compounds who experience acute inflammatory symptoms or clinical deterioration not related to their gastrointestinal disease should be screened to rule out a lupus-like reaction.