Prediction of progression to AIDS by analysis of CD4 lymphocyte counts in a haemophilic cohort.

Serial CD4 lymphocyte counts were recorded in 112 anti-HIV-positive haemophiliacs who were followed for up to 8 years after seroconversion. The patients remained at low risk of developing AIDS until their CD4 lymphocyte count fell to 0.25 X 10(9)/l. From this point, the risk increased as their count approached zero. Using this result and on the assumption (which is evaluated) that the underlying trend over time in CD4 lymphocyte counts is linear, the predicted rate of progression to AIDS was calculated for the cohort. It was estimated that 73% (95% confidence limits 60-86%) of the cohort will develop AIDS within 15 years of HIV-seropositivity. During 8 years of follow-up, this cohort had shown similar rates of progression to AIDS to other cohorts--haemophilic and otherwise--suggesting that this estimate may well have general applicability. The method described could be used to plan the provision of health-care resources for groups of anti-HIV-positive patients as it allows the number of new cases of AIDS to be predicted year by year, even when the patients' dates of seroconversion are unknown.

p24 antigenaemia, CD4 lymphocyte counts and the development of AIDS.

A cohort of 111 HIV-infected haemophiliacs has been followed for up to 11 years, during which time 33 patients have been diagnosed with AIDS. Twenty-seven of the cohort developed detectable p24 antigenaemia while remaining free of AIDS. These patients experienced an increased risk of progression to AIDS compared with those patients who were persistently p24-negative (relative risk 7.24; P less than 0.0001, Cox proportional hazards model). The relative risk was reduced to 5.42 (P less than 0.0001) after adjustment for age and cytomegalovirus seropositivity. After adjustment for the patients' declining CD4 lymphocyte count during follow-up, the relative risk fell dramatically to 1.97 and became non-significant (P = 0.2). p24-antigenaemic patients tended to develop AIDS at levels of similar CD4 lymphocyte counts to those who were persistently p24-antigen-negative (median CD4 lymphocyte counts, 70 and 50 x 10(6)/l, respectively). These results suggests that the association between p24 antigenaemia and the rate of progression to AIDS can be explained largely by a more rapid decline in CD4 lymphocyte count among patients with p24 antigenaemia than in those without. The major pathological effects of increased plasma viral load, as detected by the presence or absence of p24 antigenaemia, appear to act via progressive CD4 lymphocyte depletion.

Impaired carboxylation of osteocalcin in warfarin-treated patients.

The total circulating osteocalcin and ratio of inactive (noncarboxylated; GLU) to active (carboxylated; GLA) form of circulating osteocalcin were measured in patients receiving long term warfarin treatment (n = 20), age-matched control patients not receiving warfarin treatment (n = 10), and normal subjects before and after the administration of 30 mg warfarin (n = 7). There was no significant difference in the total osteocalcin concentrations between the control patients and the patients receiving long term warfarin treatment, and it did not significantly change after warfarin ingestion in the normal subjects. The GLU/GLA ratio was significantly increased (P less than 0.002) in the patients receiving long term warfarin treatment compared with that in the control patients. There was a significant increase (P less than 0.01) in the GLU/GLA ratio after warfarin ingestion in the normal subjects. This study demonstrates that osteocalcin carboxylation in humans is a vitamin K-dependent process and that circulating osteocalcin is structurally altered by warfarin administration. This finding has pathophysiological implications for the fetal warfarin embryopathy syndrome, bone disease associated with chronic liver diseases, and possibly for osteoporosis, in which vitamin K deficiency has been implicated.

The cumulative risk of AIDS as the CD4 lymphocyte count declines.

A method is proposed for assessing the cumulative risk of various AIDS-defining conditions as the CD4 lymphocyte count declines in HIV-infected individuals. The method is analogous to survival analysis but is based on the CD4 lymphocyte count rather than on time. Thus, the level to which the CD4 lymphocyte count has declined, rather than the length of time since seroconversion, is considered as an individual's survival interval. The survival interval may be censored (due to lack of follow-up) or treated as an interval to failure (if the individual develops AIDS). The Kaplan-Meier (product-limit) estimates, of the proportion of individuals developing AIDS before reaching a given low CD4 lymphocyte count, may be useful for determining when prophylactic treatment should begin.

More rapid progression to AIDS in older HIV-infected people: the role of CD4+ T-cell counts.

The tendency for older people with HIV infection to progress more rapidly to AIDS than younger people was studied in a group of 111 anti-HIV-positive haemophiliacs followed for up to 10 years from seroconversion. After 7 years of seropositivity, those aged over 30 years at the time of the first positive anti-HIV test had a cumulative progression rate to AIDS of 50%, compared with only 12% for those aged 10-19 years (Kaplan-Meier estimates). Overall, the relative risk of developing AIDS by any given time after seroconversion was 1.45 for each 10 year increase in age (p = 0.002; 95% confidence limits of 1.15, 1.85; Cox proportional hazards model). After adjustment for the CD4+ T-cell count (median of 10 count measurements per patient, fitted as a time-dependent covariate), the relative risk fell to 1.31 but remained statistically significant (p less than 0.05; 95% confidence limits of 1.03, 1.67). This implies that older people may be at higher risk of progression than their younger counterparts, even if their CD4+ T-cell counts are the same. Hence, prophylaxis against opportunistic infections may be indicated at higher CD4+ T-cell counts in older people than in younger people.

Rationale and evolution of therapy with porcine factor VIII:C.

Highly purified porcine factor VIII:C (FVIII:C) concentrate prepared by polyelectrolyte fractionation has been available for therapeutic use since 1980. Over the last decade substantial international experience has confirmed the value of porcine FVIII:C in management of hemophilia with inhibitors, and recent studies have underlined its particular effectiveness in treating patients with the acquired form of the disease. The rationale for use of porcine FVIII:C is based on a twofold premise. First, most inhibitors interact less strongly with porcine FVIII:C than they do with the human factor; cross-reactivity is especially low, and often negligible, among patients with acquired disease. Second, when measurable levels of circulating FVIII:C can be achieved, the likelihood of clinical hemostasis is maximized. In a variable proportion of patients with the congenital disease, anamnestic rises in titers of the inhibitor against human FVIII:C may follow treatment with the porcine factor, and this phenomenon may constrain therapy. These events seem to occur rarely in persons with acquired inhibitors, however, thus broadening therapeutic application of porcine FVIII:C to these patients. Although anamnesis often is perceived as a limitation, significant untoward transfusion reactions are highly unusual after porcine FVIII:C therapy. Although early experience with this form of treatment centered on management of major bleeding episodes and hemostatic crises, use of porcine FVIII:C has more recently been extended to more routine bleeding problems, immune tolerance induction regimens, prophylaxis, and home therapy. These and other advances are discussed.

Studies on the neutralization of rabbit antibodies to human factor VIII.

The capacity of normal and haemophilic cryoprecipitates to neutralize the anticoagulant effect of rabbit antibodies to human factor VIII (anti-VIII) was assessed using a quantitative estimation of antibody. About 4 times as much anti-VIII could be neutralized by normal factor VIII as was required to neutralize clotting activity. This suggests that there are probably several antigenic sites intimately associated with factor VIII clotting activity, quite apart from any antigenic sites which may be detected using antibodies directed against other components of the factor VIII complex. The neutralizing capacity of factor VIII was only slightly greater for the rabbit antibodies employed in this study than has been previously reported for antibodies of human origin, thus indicating only minor differences in specificities. Additional evidence in support of this concept was the finding that cryoprecipitates prepared from haemophilic plasmas previously recognized as either lacking or possessing the capacity to neutralize antibodies of human origin neutralized least or most quantities of rabbit antibodies, respectively.

A study of the effect of ethamsylate (Dicynene) on the bleeding time, von Willebrand factor level and fibrinolysis in patients with von Willebrand's disease.

Nine patients with clinically moderate or severe Type I von Willebrand's disease were treated for 2 weeks with ethamsylate (2 g/day in four equal doses) and with a matched placebo in a randomised double-blind trial. Template bleeding time, von Willebrand factor activity (ristocetin co-factor) and antigen, euglobulin lysis time and type I tissue plasminogen activator inhibitor were determined before and at the end of each treatment period. None of these parameters showed any significant change attributable to ethamsylate. Thus, despite the fact that five patients thought subjectively that their bleeding symptoms improved during ethamsylate treatment compared to only one while on placebo, we obtained no evidence that the drug was of benefit to patients with von Willebrand's disease.

A comparison of coagulation factor replacement with and without prednisolone in the treatment of haematuria in haemophilia and Christmas disease.

A double-blind controlled study was carried out to investigate the effectiveness of treatment with factor VIII or factor IX concentrate and a reducing dose of prednisolone in contolling haematuria in patients with haemophilia and Christmas disease. 41 episodes of haematuria were studied in 30 different patients. No appreciable benefit was observed in the treated, as compared with the control group and this is at variance with the results of the few studies reported elsewhere.

Production and therapeutic use of a factor XI concentrate from plasma.

Factor XI deficiency is an uncommon bleeding disorder usually manifested by excessive bleeding after surgery or trauma. Until recently the only effective therapy has been fresh-frozen plasma (FFP) infusion. We describe the efficacy and safety of a new factor XI concentrate produced from human donor plasma by a modification of the method used for antithrombin III concentrate. The mean recovery of factor XI in the circulation measured on 62 occasions was approximately 91% of the injected dose, and the mean half-disappearance-time was 52 h. The concentrate was used for 31 invasive procedures in 30 patients, including 16 patients who had a definite bleeding tendency on previous occasions, with normal haemostasis being achieved in all but 1. Only 1 patient (previously experiencing allergy to FFP) experienced adverse effects during infusion. Monitoring of liver function tests and viral antibody status in suitable patients has shown no evidence of transmission of hepatitis viruses, HIV-1 or parvovirus B19. We conclude that this concentrate provides effective treatment for patients with factor XI deficiency. Preliminary results suggest safety from virus transmission, but this needs to be established in further studies of previously untreated patients.

Disappearance of antibodies to Factor VIII in a patient with acquired haemophilia and carcinoma of the pancreas during cytotoxic therapy with fluorouracil and CCNU.

An inhibitor to clotting factor VIII (anti-VIII:C) developed in a 70 year old woman with carcinoma of the pancreas three months after palliative by-pass surgery. A life-threatening sublingual haemorrhage was controlled by infusion of human factor VIII concentrate in high dosage. With the objective of reducing pancreatic tumour size, combination cytotoxic therapy with fluorouracil and CCNU was given. Reduction in the size of the tumour was associated with disappearance of anti-VIII:C, reappearance of normal quantities of clotting factor VIII (VIII:C) in the plasma and resolution of the bleeding tendency. The anti-VIII:C was characterised as being predominantly of the IgG4 sub-class with k light chains. In vitro and in vivo studies showed the inactivation of VIII:C by anti-VIII:C was markedly non-linear. Normal quantities of factor VIII coagulant antigen (VIII:CAg) were detected in the patient's plasma when VIII:C levels were negligible.

Fluconazole for oropharyngeal candidiasis in anti-HIV positive haemophiliacs.

Candidiasis of the oropharynx and oesophagus is one of the most common problems encountered in patients with HIV disease. Fluconazole is a bis-triazole antifungal agent with a long serum half-life. Sixteen anti-HIV positive patients (15 haemophiliacs and one blood transfusion recipient) with a clinical diagnosis of oropharyngeal candidiasis were treated with 50 mg fluconazole daily for 14-28 days and then either 150 mg fluconazole or placebo weekly for 6 months in a prophylactic phase. Clinical cure occurred in all patients, and mycological cure occurred in 13/16 (81%) patients. In the prophylactic phase, there were 2/5 (40%) relapses in the placebo arm compared with 1/8 (12.5%) in the fluconazole arm, but this was not statistically significant by Fisher's one-sided exact test (P = 0.31). It is concluded that fluconazole is an effective treatment of oropharyngeal candidiasis and has potential for prophylactic use.

Hemostatic evaluation in bleeding disorders from native blood. Clinical experience with the hemostatometer.

Primary hemostasis (PH), i.e., hemostatic platelet plug formation, and the subsequent coagulation were recorded and quantified from the same nonanticoagulated venous blood sample with the use of the Haemostatometer. In addition, platelet thrombus formation induced by interaction of flowing native blood with a collagen fiber under low shear rates (450 s-1) was simultaneously analyzed by this device. The effect of monoclonal antibodies (MoAbs) directed against von Willebrand's factor antigen (vWF:Ag), platelet glycoprotein Ib (GPIb) and the GPIIb/IIIa complex, and fibrinogen were studied. PH was significantly inhibited by MoAbs against vWF:Ag, GPIIb/IIIa, and fibrinogen but was unaffected by antibody against GPIb. Collagen-induced thrombosis was prevented by MoAbs against vWF:Ag and GPIb, slightly inhibited by antifibrinogen, and unaffected by blockage of platelet membrane GPIIb/IIIa. The effect of a single 600-mg dose of aspirin was monitored, and abnormal PH was still detectable five days later. From the 13 hemophiliacs tested, 7 showed significantly prolonged PH. In von Willebrand's disease, a characteristic defect of PH with significant inhibition or absence of collagen-platelet interaction was observed in all the 11 patients. PH was greatly prolonged in both of the two patients with storage pool deficiency. The technique detected improvement of platelet function, i.e., PH in all of six patients with bleeding disorders after replacement therapy or DDAVP infusion. The authors conclude that the Haemostatometer technique is a sensitive test for determining platelet dysfunction and monitoring efficacy of factor-replacement or DDAVP therapy.

Haemophilia A: carrier detection and prenatal diagnosis by linkage analysis using DNA polymorphism.

Restriction fragment length polymorphisms (RFLPs) within or close to the factor VIII locus are very useful for genetic linkage analysis. Such RFLPs allow a mutant allele to be tracked in a family, segregating haemophilia A even when, as is usually the case, the precise mutation causing failure to synthesise factor VIII is unknown. To date two markers tightly linked to the factor VIII locus have been described, one of which is highly polymorphic and therefore informative in most kindreds. A significant crossover rate, however, does not make diagnosis absolute. Three intragenic RFLPs have been defined, which, taken together, are informative in about 70% of women, providing virtually deterministic genetic diagnosis.

Acidosis and severe megaloblastic anaemia.

Ten patients with severe megaloblastic anaemia were studied to investigate whether the causative metabolic defects might predispose them to lactic or other acidosis. One patient had compensated acidosis with hyperlactataemia before treatment but there were obvious causes other than anaemia. No other patient developed an acidosis. Neither anaemia per se nor the metabolic defects of vitamin B(12) or folic acid deficiency are likely to cause clinically significant lactic acidosis or hyperlactataemia.

Non-invasive investigation of liver disease in haemophilic patients.

Liver biopsy specimens previously taken from 16 haemophilic patients with chronic non-A, non-B hepatitis were reviewed. The degree of fibrosis correlated with serum procollagen III peptide (sPIIIP) concentrations, measured both at the time of biopsy and 4.25 years later. Two patients with extremely high sPIIIP concentrations had collateral veins on computed tomography, suggesting portal hypertension. Twenty eight of 47 patients (60%) had splenomegaly on computed tomography, and of 28 patients in whom intravenous contrast medium was used, seven (25%) had collateral oesophageal veins. Serum procollagen III peptide estimations and computed tomography, both non-invasive investigations, indicated that hepatic fibrosis and portal hypertension had developed in a proportion of haemophilic patients with non-A, non-B hepatitis. Infection with the human immunodeficiency virus (HIV) may modify the course of this presumably cytopathic virus infection of the liver.

The application of polyethylene glycol to radioimmunoassays used in haemostasis.

Use of polyethylene glycol 6000 in the second stages of double antibody radioimmunoassays for fibrinopeptide A, B-thromboglobulin and platelet factor 4 facilitates the more rapid separation of free from bound antigen, and allows precipitating antiserum to be used in greater dilution. At a final PEG 6000 concentration of 5%, separation of free from bound antigen was complete within 1 hour, and antisera could be used in dilutions 3-9 times greater than those recommended by the manufacturers. PEG 6000 had a negligible effect on the affinity of first stage antibodies for their respective antigens. Radioimmunoassays using PEG 6000 were sensitive to protein concentration, failure to adjust standards to similar protein concentrations as those of test samples causing artefactually low results.

Changes in coagulation and fibrinolysis during pregnancy: evidence of activation of coagulation preceding spontaneous abortion.

In order to monitor physiological changes in coagulation and fibrinolysis that occur during normal pregnancy, blood samples were collected in each trimester of pregnancy from 17 volunteers. Control samples were collected from 12 non-pregnant female volunteers. As pregnancy advanced there was a rise in the basal levels of fibrinopeptide A, cross linked D-dimer fragment and the B beta 15-42 fragment and an increase in the in vitro rate of fibrinopeptide A generation. These results were consistent with an increased activation of coagulation during normal pregnancy, compensated for by a concomitant rise in fibrinolytic activity. In two patients who spontaneously aborted, evidence of uncompensated activation of coagulation could be detected before the manifestation of any clinical signs. In a second pregnancy in one of these patients similar changes were observed, but were reversed by heparin treatment and the pregnancy progressed to full-term delivery of a normal infant.

Homozygous protein C deficiency with delayed onset of symptoms at 7 to 10 months.

We report an inbred family with two cases of homozygous protein C deficiency and review 11 other such cases. Both patients presented in the second half of their first year of life with recurrent rapidly disappearing ecchymotic skin lesions, disseminated intravascular coagulation, and venous thrombosis. Successful treatment has been achieved by frequent infusions of plasma or prothrombin complex then maintained with Warfarin. Homozygous recessive protein C deficiency usually presents in the neonatal period with purpura fulminans. Two cases have been described elsewhere which presented in the second decade of life with milder symptoms. The present cases appear to be intermediate in time of presentation and severity of symptoms. We also review the distinction that is now evident between recessive and dominant protein C deficiency.

The metabolism of dihomo-gamma-linolenic acid in man.

Orally administered dihomo-gamma-linolenic acid (DHLA) is well absorbed in man; it appears in blood after ca. 4 hr first as triglyceride ester and later as phospholipid. After sustained-dosing, DHLA penetrated membrane pools and all phospholipid components but, depending on the dosage, reached a metabolic equilibrium in 4-16 days. Intact platelets do not accumulate arachidonate following DHLA administration, and species differences occur in the capacity of animals to metabolize DHLA to arachidonic acid (AA). The rat appears to be unusual in having a very active hepatic delta5-desaturase enzyme system. Potentially antithrombotic changes in platelet function which followed the administration of DHLA to man were accompanied by a significant increase in the capacity of platelets to synthesize PGE1. Concomitant increases in PGE2 synthesis do not apparently result from an increased production of AA and suggest that DHLA, or a DHLA metabolite, interferes with the metabolism of AA. Effects on thromboxane and prostacyclin synthesis are being studied.