Refractory hyperhaemolysis in a patient with beta-thalassaemia major

We report the case of a 1-year-old girl with newly diagnosed beta-thalassaemia major. Following an initial blood transfusion with phenotypically matched blood, she developed a haemolytic anaemia which progressed with subsequent transfusions. The Direct Antiglobulin test (DAT) was strongly positive with C3d and weakly with IgG. The only free antibodies detected were a weak anti-H and a weak cold auto-antibody, which did not exhibit a wide thermal range. The indirect Donath-Landsteiner and Ham's tests were negative. There was no sustained clinical response to steroids, immunoglobulin infusions or splenectomy. An HLA identical sibling donor was available for allogeneic bone marrow transplantation (BMT) and the haemolysis resolved during the immunosuppressive transplant conditioning. Such hyperhaemolysis without significant red cell alloantibodies has previously been reported in patients with sickle cell anaemia, but only rarely in patients with beta-thalassaemia major.     

Studies on the mechanism of bacterial resistance to complement-mediated killing. II. C8 and C9 release C5b67 from the surface of salmonella minnesota S218 because the terminal complex does not insert into the bacterial outer membrane

The mechanism for consumption of terminal complement components and release of bound components from the surface of serum-resistant salmonella minnesota S218 was studied. Consumption of C8 and C9 by S218 occurred through interaction with C5b67 on the bacterial surface because C8 and C9 were consumed when added to S218 organisms previously incubated in C8-deficient serum and washed to remove all C5b67 on the bacterial surface because C8 and C9 were consumed when added to S218 organisms previously incubated in C8- deficient serum and washed to remove al but cell bound C5b67. Rapid release of (125)I C5 and (125)I C7 from the membrane of S218 was dependent on binding of C8 because (125)I C5 and (125)I C7 deposition in C8D serum was stable and was twofold higher in C8D than in PNHA, and addition of purified C8 or C8 and C9 to S218 previously incubated in C8D serum caused rapid release of (125)I C5 and (125)I C7 from the organism. Analysis by sucrose density gradient ultracentrifugation of the fluid phase from the reaction of S218 and 10 percent PNHS revealed a peak consistent with SC5b-9, in which the C9:C7 ratio was 3.3:1, but the NaDOC extracted bound C5b-9 complex sedimented as a broad peak with C9:C7 of less than 1.2:1. Progressive elution of C5b67 and C5b-9 from S218 but not serum-sensitive S. minnesota Re595 was observed with incubation in buffers of increasing ionic strength. Greater than 90 percent of the bound counts of (125)I C5 or (125)I C9 were released from S218 by incubation in 0.1 percent trypsin, but only 57 percent of (125)I C9 were released by treatment of Re595 with trypsin. These results are consistent with the concept that C5b-9 forms on the surface of the serum-sensitive S. minnesota S218 in normal human serum, but the formed complex is released and is not bactericidal for S218 because it fails to insert into hydrophobic outer membrane domains.     

Complement activation by artificial blood substitute Fluosol: in vitro and in vivo studies

A perfluorocarbon blood substitute, Fluosol, is undergoing clinical trials as an adjunct to chemotherapy. The adverse effects associated with its administration have been postulated to result from complement activation. When gel electrophoresis and Western blotting of Fluosol are used after its incubation with serum, activated C3 and factors Bb and H are bound to the Fluosol particles in a time-dependent fashion, which suggests that complement activation with Fluosol, as does that with zymosan, occurs on the surface of the particles. Paradoxically, it is found, both by the measurement of Fluosol-bound C3d and by fluid-phase C5a, that lower concentrations of Fluosol cause greater amounts of complement activation, which suggests a complex interaction of activators and inhibitors that changes as the available surface area is decreased. Studies performed with bystander red cell-bound C3d demonstrated in vivo complement activation occurring in six patients receiving Fluosol as an adjunct to chemotherapy for colon cancer. In two patients, there was a marked increase in red cell-bound C3d after Fluosol infusion; these two patients also developed adverse reactions during Fluosol infusion. These studies suggest that the Fluosol surface plays a major role in the initiation and regulation of complement activation that is seen during Fluosol infusion.     

Renal aquaporin-4 associated pathology in TG-26 mice

Human immunodeficiency virus-associated nephropathy (HIVAN) is a leading cause of end-stage renal disease in HIV patients, which is characterized by glomerulosclerosis and renal tubular dysfunction. Aquaporin-4 (AQP-4) is a membrane bound water channel protein that plays a distinct role in water reabsorption from renal tubular fluid. It has been proven that failure of AQP-4 insertion into the renal tubular membrane leads to renal dysfunction. However, the role of AQP-4 in HIVAN is unclear. We hypothesize that impaired water reabsorption leads to renal injury in HIVAN, where AQP-4 plays a crucial role. Renal function is assessed by urinary protein and serum blood urea nitrogen (BUN). Kidneys from HIV Transgenic (TG26) mice (HIVAN animal model) were compared to wild type mice by immunostaining, immunoblotting and quantitative RT-PCR. TG26 mice had increased proteinuria and BUN. We found decreased AQP-4 levels in the renal medulla, increased endothelin-1, endothelin receptor A and reduced Sirtuin1 (SIRT-1) levels in TG26 mice. Also, oxidative and endoplasmic reticulum stress was enhanced in kidneys of TG26 mice. We provide the first evidence that AQP-4 is inhibited due to induction of HIV associated stress in the kidneys of TG26 mice which limits water reabsorption in the kidney which may be one of the cause associated with HIVAN, impairing kidney physiology. AQP-4 dysregulation in TG26 mice suggests that similar changes may occur in HIVAN patients. This work may identify new therapeutic targets to be evaluated in HIVAN.     

EASE OF HANDLING AND CLINICAL EFFICACY OF FLUTICASONE PROPIONATE ACCUHALER/DISKUS™ INHALER COMPARED WITH THE TURBOHALER™ INHALER IN PAEDIATRIC PATIENTS

A total of 323 children aged 4-11 years who were receiving, or had symptoms indicating a clinical requirement for, inhaled corticosteroid at a daily dose of 400 μg budesonide (BUD) or beclomethasone dipropionate (BDP), or 200 μg fluticasone propionate (FP), were randomised into this multicentre, open-label, parallel group study. Patients received either FP 100 μg b.d. administered via the Accuhaler/Diskus inhaler (n=159) or BUD 200 μg b.d. administered via a Turbohaler inhaler (n=164) for four weeks and recorded daily their morning and evening peak expiratory flow (PEF), asthma symptoms and use of relief medication. Device handling was assessed by a questionnaire, with responses recorded on three- or five-point ordinal scales. The primary efficacy parameter was mean percent predicted morning PEF. The device handling results showed the Accuhaler/Diskus inhaler was rated more favourably than the Turbohaler inhaler in terms of ease of correct inhaler use, ease of telling how many doses were left, ease of knowing whether a dose had been inhaled and overall liking of the device. More patients in the Accuhaler/Diskus group (85%) than in the Turbohaler group (58%) said they would be happy to receive the same device again, while 8% and 25% respectively said they would not be happy to be given it again. In addition, the change from baseline to week 4 of treatment in mean percent predicted morning PEF was greater in the FP Accuhaler/Diskus group, indicating that FP 200 μg daily via Accuhaler/Diskus inhaler is at least as clinically effective as BUD 400 μg daily via the Turbohaler inhaler.