Biomedical subjects

R Thorpe

Publications and source records attributed to R Thorpe.

At least 19 recordsLinked to original sources

PubMed2005

Kinetics of development and characteristics of antibodies induced in cancer patients against yeast expressed rDNA derived granulocyte macrophage colony stimulating factor (GM-CSF).

We have determined the presence and kinetics of granulocyte macrophage colony stimulating factor (GM-CSF) antibodies induced after repeated administration of a yeast expressed GM-CSF product in prostate cancer patients with minimal recurrent disease using a panel of assays for detection and characterization of antibodies. Results showed that all 15 prostate cancer patients treated with GM-CSF developed GM-CSF reactive antibodies during the course of therapy. Most patients (87%) developed GM-CSF reactive antibodies within 3 months while in other patients (13%), these antibodies were induced after additional cycles of GM-CSF treatment. For most patients, the timing of occurrence of these antibodies was the same regardless of whether the ELISA or surface plasmon resonance (SPR) assays were used for detection. However, in two patients, the recognition of GM-CSF reactive antibodies by SPR assays preceded their detection by ELISA. A significant number of patients (n=9, 60%) developed GM-CSF antibodies which neutralized the biological activity of GM-CSF in vitro in a cell-line based bioassay. These antibodies also recognized GM-CSF protein from different expression systems including the non-glycosylated protein from E. coli indicating that the antibody response is directed towards the amino acid backbone of the protein. A significant effect of GM-CSF antibodies on PSA modulation was not observed in this small cohort of patients despite an alteration in PSA levels in some treated patients. The study design used here did not allow conclusions regarding the relationship between neutralizing antibodies and the PSA levels which were used as a marker for clinical outcome. Implementation of a clinical strategy which permits monitoring for antibody development and for levels of a relevant pre-determined clinical marker at appropriate time-points is necessary for assessing the impact of antibody development on the therapeutic efficacy of the protein.

Aged↗

PubMed2005

Comparative studies on cellular gene regulation by HIV-1 based vectors: implications for quality control of vector production.

Among the recent gene therapy protocols, vectors that can stably express transgenes, for example, HIV-1-based vectors, are particularly desirable. There have been no direct reports on insertional mutagenesis by lentiviral vectors; however, the severe pathogenic nature of their parental virus (HIV-1) is still a major safety concern surrounding these vectors and prevents the progress with their clinical application. We reason that by investigating the host response we shall be able to assess the safety and potential effects of the vectors on targeted cells and understand the interaction between vectors and the host. For this, two major sets of experiments were conducted. Initially, we used cDNA microarray methodology to examine cellular gene profile in human primary umbilical cord endothelial cells (HUVECs) after HIV-1-based VSV-G/GFP vector transduction and observed a modest effect of HIV-1-based vectors on HUVECs. The represented functional categories include transcription and translation factors, tumour antigens, complement factors and signal transduction factors. Some of the differentially expressed genes, for example, Clusterin, CD151, Ku antigen and eIF4gamma, could have oncogenic potential. In the second approach, we systematically compared five different viral vectors, that is, HIV-1-based VSV-G/Empty, VSV-G/GFP, VSV-G/puro, Amph/GFP and MLV-based Amph/Laz, for the effects of individual viral components on cellular gene regulation. Our comparative results demonstrated a regulatory function of Gag/Pol proteins on cellular gene expression. The significance of our findings in relation to the safety of HIV-1 vectors and the importance of quality control of vector production will be presented and discussed.

Blotting, Western↗

PubMed2005

Engineering of the gut commensal bacterium Bacteroides ovatus to produce and secrete biologically active murine interleukin-2 in response to xylan.

AIMS: The aim of this work was to engineer a gut commensal bacterium, Bacteroidesovatus, to produce and secrete a biologically active cytokine in a regulated manner as a basis for novel immunotherapies for chronic gut disorders. METHODS AND RESULTS: Bacteroides ovatus was engineered to produce murine interleukin-2 (MuIL2) intracellularly in response to xylan in culture media by inserting the MuIL2 gene into the xylanase operon of the organism. A second strain was engineered to secrete MuIL2 by adding Bacteroides fragilis enterotoxin secretion signal sequence to the protein. The recombinant strains produced MuIL2 only in the presence of xylan as determined by ELISA of cell lysates and culture supernatants. The IL2-dependent cell line CTLL-2 was used to demonstrate that MuIL2 produced by both B. ovatus strains was biologically active. This activity could be blocked by an anti-IL2 neutralizing antibody. The xylan-inducible nature of this system was demonstrated by RT-PCR. CONCLUSIONS: Bacteroides ovatus was successfully engineered to produce and secrete biologically active MuIL2 in a xylan-inducible manner. SIGNIFICANCE AND IMPACT OF THE STUDY: The production and secretion of a biologically active mammalian protein by a member of the gut microflora could lead to the development of new long-term immunotherapies for inflammatory gut diseases.

Animals↗

PubMed2005

International collaborative study to evaluate a candidate reference preparation to define an appropriate specified limit of anti-D in intravenous immunoglobulin products.

BACKGROUND AND OBJECTIVES: The aim of the study was to evaluate a lyophilized intravenous immunoglobulin (IVIG) preparation containing anti-D (02/228; nominal reciprocal titre of 8) for its suitability to define the maximum limit of anti-D in IVIG products when used in a proposed reference method of direct haemagglutination of papain-treated erythrocytes, in an international collaborative study. MATERIALS AND METHODS: Twenty laboratories tested 02/228 along with a negative control IVIG preparation and four IVIG samples containing different levels of anti-D. Nineteen laboratories performed direct haemagglutination methodology using papain-treated erythrocytes; five of these laboratories and one additional laboratory performed their in-house haemagglutination methodology (all indirect antiglobulin tests). RESULTS: The mode titre of 02/228, obtained by using the proposed reference method, was 8 (62.5% of tests). However, there was wide variation in haemagglutination titres between laboratories for three of the four samples. Correcting the titres of the samples relative to those of the proposed reference preparation reduced the interlaboratory variability and increased the frequency of the mode titres in three out of four samples. The indirect antiglobulin tests also showed wide interlaboratory variability and were less sensitive than the direct method in four laboratories. Eleven of the 14 laboratories that expressed an opinion considered that the level of anti-D in 02/228 was appropriate to define a specified limit. CONCLUSIONS: Our results demonstrate the necessity of using a reference preparation to define the maximum level of anti-D in IVIG products and ensure sufficient sensitivity in haemagglutination testing methodology. On the basis of these results, members of the European Pharmacopoeia Expert Group 6B recommended revision of the appropriate monograph to include this new specification and test. The Food and Drug Administration in the USA intends to adopt the same maximal specification defined by the reference preparation and to recommend the same test for the safety of IVIG products. Preparations 02/228 and 02/226 were also established by the World Health Organization as International Reference Reagents to standardize haemagglutination testing for anti-D in normal IVIG products.

Cooperative Behavior↗

PubMed2005

Detection, measurement and characterization of unwanted antibodies induced by therapeutic biologicals.

Assessment of the unwanted immunogenicity of therapeutic biologicals in recipients is an important consideration in the evaluation of these medicinal products. Proper planning of immunogenicity studies with appropriately devised strategies is critical if valid and meaningful conclusions concerning the unwanted immunogenicity are to be derived. An essential requisite for such studies is the need for conducting carefully selected and validated procedures. Several techniques are available for detection, characterization and measurement of antibodies elicited in an immune response. These include various formats of immunoassays, surface plasmon resonance and biological assays. None of these assays alone can provide sufficient information on the characteristics of the induced antibodies. A combination of methods is therefore usually necessary for a detailed understanding of the quantity and type(s) of antibodies generated against a therapeutic product. This manuscript considers the benefits and limitations of the various techniques available for antibody detection and outlines a brief strategy for the assessment of unwanted immunogenicity of therapeutic products.

Animals↗

PubMed2004

Cytokine profile and insulin antibody IgG subclasses in patients with recent onset type 1 diabetes treated with oral insulin.

AIMS/HYPOTHESIS: Tolerance to orally administered antigens may be generated through the induction of T helper cell type 2 and 3 (Th2/Th3) regulatory cells. We previously reported that treatment of recent onset type 1 diabetes with oral insulin had no effect on residual beta cell function. The aim of this study was to evaluate whether this treatment produces a deviation in the immune response, with polarisation of the cytokine pattern and the induction of a Th2-like antibody response. METHODS: Mononuclear cells were collected from a total of 20 patients with type 1 diabetes before and after 12 months of treatment with oral insulin (n=11) or placebo (n=9). Following stimulation of the cells with insulin or phytohaemagglutinin, levels of Th2 and Th3 cytokines (including TGF-beta, IFN-gamma, IL-4 and IL-5) in the culture supernatants were assessed by ELISA. In addition, levels of total and specific insulin antibody IgG subclasses were measured by radioimmunoassay in serum samples drawn from 33 patients with type 1 diabetes before and after 3, 6 and 12 months of therapy with oral insulin (n=18) or placebo (n=15). RESULTS: After 12 months of treatment, the release of TGF-beta was significantly higher in patients who received oral insulin compared with those who received placebo (p=0.025 and p=0.006 for lymphocytes challenged with insulin and phytohaemagglutinin respectively). The two groups had similar levels of IL-4 and IL-5 both at baseline and after 12 months of treatment. The release of IFN-gamma was markedly reduced in patients treated with oral insulin compared with those who received placebo at the 12-month follow-up. Circulating levels of IgG1 and IgG3 subclasses directed against insulin were significantly lower in the oral insulin group than in the placebo group after 12 months of treatment (p=0.05 for IgG1 and p=0.014 for IgG3). CONCLUSIONS/INTERPRETATION: The increased TGF-beta release observed in patients treated with oral insulin suggests that a regulatory response can be induced in vivo by this treatment. The lower levels of insulin antibody IgG1 and IgG3 subclasses present in patients exposed to oral insulin are consistent with a Th2 deviation of the immune response. The failure of oral insulin treatment to provide any measurable clinical benefit may be due to the timing of treatment initiation.

Administration, Oral↗

PubMed2004

Hospital and after: experience of patients and carers in rural and remote north Queensland, Australia.

INTRODUCTION: This article reports a study of patients and carers discharged from the Townsville General Hospital into rural and remote communities in north Queensland, Australia. The findings indicate the importance of focusing on the experiences of patients and carers in attempting to understand the impact of discharge procedures. The four stories and their implications exist within a particular healthcare context that impacts disproportionately on rural patients and their carers. Economic rationalism has shaped contemporary healthcare policy in Australia, creating a system that is encouraged to conform to market principles. The costs borne by individuals, groups and communities have been increasingly privatised. Later admission and earlier discharge from hospital is now the norm. Concern about the impact of this policy context on the lives of rural and remote patients and carers prompted the study, which aimed to: (1) examine, from the patient and carer perspective, the social, economic, cultural and emotional cost of hospitalisation away from home communities; (2) identify the needs of rural and remote patients and their carers before, during and after hospitalisation a long way from home; and (3) make recommendations for improved policies and practices concerning the continuum of care: from admission planning, through hospitalisation, discharge-planning, and post-discharge support, in the context of rural and remote location. METHODS: The experiences presented highlight the depth of the challenges faced by patients and their carers who live in rural and remote communities. Both quantitative and qualitative methodologies were used to obtain insight into the complexity of patients and carers' lives. The four vignettes presented in this paper are taken from in-depth, qualitative interviews with 12 patients and 12 carers. RESULTS: The four stories described reveal the high financial and emotional costs, for patients and carers, of negotiating a healthcare system a long way from home. Challenges faced included inadequate admission planning, excessive accommodation and transport costs, and lack of post-discharge support services in home communities, as well as business failure, marital and family strain. There was an over-reliance on carers who lacked medical caring expertise, had other major commitments (family and work) and who may have had a tenuous and uncertain relationship with the patient. CONCLUSIONS: Recognition of these complex circumstances, exacerbated by rural location, during the discharge planning process should mean that attention to ensuring patients and their carers are linked to adequate support services in their communities is of the highest priority. The communication and dissemination of information to patients and carers is also vital. Information on hospital admission, travel benefits, accommodation options, care requirements post-discharge are particular recommendations. To summarise, the experiences highlighted in this study suggest that patients and carers in rural and remote communities have not benefited from adequate discharge planning, and are struggling to cope in a policy context that encourages later admission, earlier discharge and over reliance on family and friends as carers.

Journal Article↗

PubMed2004

WHO Expert Committee on Biological Standardization.

This report presents the recommendations of a WHO Expert Committee commissioned to coordinate activities leading to the adoption of international requirements for the production and control of vaccines and other biologicals and the establishment of international biological reference materials. The report starts with a discussion of general issues brought to the Committee's attention and provides information on the status and development of reference materials for various antibodies, antigens, blood products and related substances, cytokines, growth factors, and endocrinological substances. The second part of the report, of particular relevance to manufacturers and national regulatory authorities, contains recommendations for the production and quality control of meningococcal group C conjugate vaccines, guidelines for regulatory expectations for clinical evaluation of vaccines, guidelines for the production and quality control of inactivated oral cholera vaccines and guidelines on viral inactivation and removal procedures intended to assure the viral safety of human blood plasma products.

Advisory Committees↗

PubMed2003

Strategies for detection, measurement and characterization of unwanted antibodies induced by therapeutic biologicals.

An important aspect of evaluating the safety of therapeutic biologicals is the assessment of the unwanted immunogenicity of such biologicals in recipients. Properly planned immunogenicity studies with appropriately devised strategies are critical if valid conclusions concerning the unwanted immunogenicity are to be derived. Such studies need to be conducted using carefully selected and validated procedures. Several techniques are available for detection and measurement of immunogenicity including immunoassays, radioimmunoprecipitation assays (RIPAs), surface plasmon resonance (SPR) and bioassays. A combination of methods for characterization of the induced antibodies is usually necessary for a detailed understanding of the type(s) of antibodies generated against a therapeutic product. This review considers the benefits and limitations of the various techniques available for antibody detection and outlines a strategy for the assessment of unwanted immunogenicity of therapeutic products.

Animals↗

PubMed2003

Anti-cytokine autoantibodies in autoimmunity: preponderance of neutralizing autoantibodies against interferon-alpha, interferon-omega and interleukin-12 in patients with thymoma and/or myasthenia gravis.

We have screened for spontaneous anticytokine autoantibodies in patients with infections, neoplasms and autoimmune diseases, because of their increasingly reported co-occurrence. We tested for both binding and neutralizing autoantibodies to a range of human cytokines, including interleukin-1alpha (IL-1alpha), IL-1beta, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12, IL-18, interferon-alpha2 (IFN-alpha2), IFN-omega, IFN-beta, IFN-gamma, tumour necrosis factor alpha (TNF-alpha), transforming growth factor beta-1 (TGF-beta1) and granulocyte-macrophage colony stimulating factor (GM-CSF), in plasmas or sera. With two notable exceptions described below, we found only occasional, mostly low-titre, non-neutralizing antibodies, mainly to GM-CSF; also to IL-10 in pemphigoid. Strikingly, however, high-titre, mainly IgG, autoantibodies to IFN-alpha2, IFN-omega and IL-12 were common at diagnosis in patients with late-onset myasthenia gravis (LOMG+), thymoma (T) but no MG (TMG-) and especially with both thymoma and MG together (TMG+). The antibodies recognized other closely related type I IFN-alpha subtypes, but rarely the distantly related type I IFN-beta, and never (detectably) the unrelated type II IFN-gamma. Antibodies to IL-12 showed a similar distribution to those against IFN-alpha2, although prevalences were slightly lower; correlations between individual titres against each were so modest that they appear to be entirely different specificities. Neither showed any obvious correlations with clinical parameters including thymoma histology and HLA type, but they did increase sharply if the tumours recurred. These antibodies neutralized their respective cytokine in bioassays in vitro; although they persisted for years severe infections were surprisingly uncommon, despite the immunosuppressive therapy also used in most cases. These findings must hold valuable clues to autoimmunizing mechanisms in paraneoplastic autoimmunity.

Adolescent↗

PubMed2003

The influence of platelet additive solutions on cytokine levels and complement activation in platelet concentrates during storage.

BACKGROUND AND OBJECTIVES: The accumulation of platelet-derived cytokines in platelet concentrates (PC) during storage may contribute towards non-haemolytic transfusion reactions (NHTR). We investigated the effect of platelet storage medium on platelet activation, complement activation and cytokine levels in leucocyte-reduced PC. MATERIALS AND METHODS: Hyperconcentrated platelets (3000-6000 x 109/l) were collected by apheresis and diluted in 100% plasma, 70% PASIII, or 70% or 80% PASIII supplemented with magnesium and potassium (PAS IIIM). RESULTS: Levels of transforming growth factor-beta (TGF-beta) and regulated on activation, normal, T-cell expressed, and secreted (RANTES) increased during storage, as did the expression of P-selectin (CD62P), and were highest in PC stored in PASIII. In PC stored in PASIIIM, however, levels of TGF-beta and RANTES were not significantly different from PC stored in plasma. Levels of CD62P expression, however, remained higher in PASIIIM PC than in those stored in plasma by day 5, but were lower than PC stored in PASIII. C3a des arg levels increased during storage in all media with the exception of PASIII and, on day 7, were higher in PC stored in plasma compared to PC stored in the other media. CONCLUSIONS: Our results indicate that replacing plasma in PC with unmodified PASIII for storage results in higher levels of platelet-derived cytokines in PC. Furthermore, it appears that the nature of the medium used for storage of PC has a significant impact on platelet activation and cytokine levels of the PC. These implications should be taken into account when considering replacement of plasma with PAS.

Blood Platelets↗

PubMed2003

Batches of intravenous immunoglobulin associated with adverse reactions in recipients contain atypically high anti-Rh D activity.

BACKGROUND AND OBJECTIVES: The presence of anti-Rh D in intravenous immunoglobulin (IVIG) products has been claimed to be associated with adverse reactions in recipients. There is currently no regulatory specification to control the level of anti-D in IVIG products and it is unclear what this should be. Two reports of haemolysis occurring in recipients of IVIG manufactured from US plasma provided a rare opportunity to investigate whether high anti-D levels could have induced the haemolysis. MATERIALS AND METHODS: We developed a direct microtitre plate haemagglutination method suitable for screening IVIG products and starting plasma pools for haemagglutinating activity. RESULTS: Of 101 batches of IVIG tested, six were found to contain specific anti-D. Four of these batches had anti-D titres ranging from 64 to 256 (including the two batches each associated with a report of haemolysis) and could be linked, in each case, to a starting plasma pool also positive for anti-D. CONCLUSIONS: Our results show that IVIG products can contain appreciable anti-D levels. To avoid potential problems in recipients, we propose an anti-D titre of 8 as the maximum permissible limit of anti-D in IVIG products for batch acceptance and release. The availability of a reference preparation is essential for control of this proposed requirement.

Blood Cells↗

PubMed2003

Effects of rheumatoid arthritis on sexual activity and relationships.

OBJECTIVES: The primary aim of this research was to assess patients' perceptions of the effects of rheumatoid arthritis (RA) on their sexual relationship and sexual activity, the causes of any difficulties and who they would turn to for help. METHODS: A self-report questionnaire was distributed to 74 patients attending a regional rheumatology clinic and returned by prepaid post. Fifty-nine questionnaires were returned (response rate 80%) and 57 of them were usable. RESULTS: Thirty-five per cent of the cohort thought that their disease strained their relationship with their partner, and reasons for this included curtailment of daily and social activities, changes in the balance of the relationship, emotional changes and changed financial circumstances. Over half (56%) of the patients found that their arthritis placed limitations on sexual intercourse and the principal reasons cited were fatigue and pain. Sexual ability was important to 58% of the sample and ageing had a negative impact on its importance (P<0.005). Poor communication was a major problem, patients being reluctant to approach health professionals and vice versa. The topic of sexual relationships was broached with patients attending one nurse practitioner's clinic, but only one other patient had ever been offered any discussion. Thirty-nine per cent of the cohort would consider talking to someone if they had a problem and the nurse and the doctor were the professionals most often chosen. CONCLUSION: RA impacts on the sexual lives of a large minority of patients and this is a problem that patients and health professionals are reluctant to discuss face to face. However, patients do appear to be willing to complete questionnaires and this may be an acceptable format for assessment. Further training and support is needed by nurses and other health professionals in this sadly neglected area.

Adult↗

PubMed2003

Establishment of T cell lines to bovine beta-casein and beta-casein-derived epitopes in patients with type 1 diabetes.

Enhanced cellular immune response to bovine beta-casein has been reported in patients with type 1 diabetes. In this study we aimed to establish beta-casein-specific T cell lines from newly diagnosed type 1 diabetic patients and to characterise these cell lines in terms of phenotype and epitope specificity. Furthermore, since sequence homologies exist between beta-casein and putative beta-cell autoantigens, reactivity to the latter was also investigated. T cell lines were generated from the peripheral blood of nine recent onset type 1 diabetic patients with different HLA-DQ and -DR genotypes, after stimulation with antigen pulsed autologous irradiated antigen presenting cells (APCs) and recombinant human interleukin-2 (rhIL-2). T cell line reactivity was evaluated in response to bovine beta-casein, to 18 overlapping peptides encompassing the whole sequence of beta-casein and to beta-cell antigens, including the human insulinoma cell line, CM, and a peptide from the beta-cell glucose transporter, GLUT-2. T cell lines specific to beta-casein could not be isolated from HLA-matched and -unmatched control subjects. beta-Casein T cell lines reacted to different sequences of the protein, however a higher frequency of T cell reactivity was observed towards the C-terminal portion (peptides B05-14, and B05-17 in 5/9 and 4/9 T cell lines respectively). Furthermore, we found that 1 out of 9 beta-casein-specific T cell lines reacted also to the homologous peptide from GLUT-2, and that 3 out of 4 of tested cell lines reacted also to extracts of the human insulinoma cell line, CM. We conclude that T cell lines specific to bovine beta-casein can be isolated from the peripheral blood of patients with type 1 diabetes; these cell lines react with multiple and different sequences of the protein particularly towards the C-terminal portion. In addition, reactivity of beta-casein T cell lines to human insulinoma extracts and GLUT-2 peptide was detected, suggesting that the potential cross-reactivity with beta-cell antigens deserves further investigation.

Adolescent↗

PubMed2003

Immunogenicity of GM-CSF products in cancer patients following immunostimulatory therapy with GM-CSF.

There is a high risk of developing neutralising and non-neutralising antibodies when GM-CSF is used as an immunomodulatory agent in non-immunocompromised patients. The presence of neutralising antibodies may seriously hamper the clinical response of the patients. This must be taken into account when designing protocols if the biological activity of the exogenously administered GM-CSF is not to be impaired and the endogenous production of GM-CSF is not to be inactivated. Assessment of production of neutralising antibodies during cytokine therapy is important for predicting the clinical response to progressive therapy. Use of validated assays is imperative for evaluation of antibodies generated following therapy with a particular protein.

Autoantibodies↗

PubMed2002

Cytokines as quality indicators of leucoreduced red cell concentrates.

Different types of filters are currently used for leucodepletion of red cell concentrates. These filters meet the specification for leucoreduction (<5 x 10(6) leucocytes/ATD) but the quality of the final product may differ depending on the performance of the filters for effective removal of both leucocytes, platelets and possibly cytokines which are associated with transfusion reactions. We measured the levels of three representative cytokines: IL-8, RANTES and TGF-beta1 in red cell concentrates prior to and subsequent to the filtration procedure on day 1 and after a storage period of 35 days. Low levels of IL-8 (10-24 pg/ml) in the control unfiltered concentrates on day 1 which increased by approximately twofold on storage. Filtration reduced the levels of IL-8 on day 1 and day 35, in filtered concentrates in comparison with their control unfiltered counterparts. Leucoreduced concentrates produced by three different filters showed similar IL-8 levels on day 1 and day 35. However, concentrates prepared using another type of process showed a twofold increase in IL-8 levels on storage in comparison with day 1. None of the concentrates tested contained any detectable RANTES and TGF-beta1 suggesting a minimal platelet content. These results indicate that a combination of IL-8, RANTES and TGF-beta1 are useful quality indicators for validation of leucoreduced red cell preparations.

Biomarkers↗

PubMed2002

Cytokine levels as performance indicators for white blood cell reduction of platelet concentrates.

BACKGROUND AND OBJECTIVES: With the implementation of universal white blood cell (WBC) reduction in the UK, in-process WBC-reduction filters for pooled buffy coat (BC)-derived platelet concentrates (PCs) and apheresis methods are used routinely for the production of WBC-reduced PCs. While these strategies meet the specification for WBC reduction (< 5 x 10(6) WBCs/unit), the products from these processes may differ depending on the process employed and its performance. The aim of this study was therefore to investigate whether PCs prepared using various WBC-reduction processes are sufficiently depleted of WBCs to limit cytokine accumulation during storage and to assess if cytokine levels detected in platelet products can serve as indicators of acceptable platelet activation as a result of the WBC-reduction process. MATERIALS AND METHODS: We measured the levels of cytokines predominantly derived from WBCs [e.g. interleukin-8 (IL-8)] and platelets [e.g. regulated on activation, normal, T-cell expressed, and secreted (RANTES) and transforming growth factor-beta(1) (TGF-beta(1))] under the present experimental conditions in different WBC-reduced PCs, i.e. PCs prepared from three different WBC-reduction filters and control non-filtered PCs using pooled BCs from the same donors and three apheresis types. Supernatant plasma was collected at the beginning (day 1) and end (day 5) of the shelf life of each PC, and the cytokine content was determined using appropriate enzyme-linked immunosorbent assays (ELISAs). Process efficiency was assessed by platelet yield and residual WBC count. RESULTS: We found that products from the apheresis process involving a filtration step (Haemonetics MCS+) showed a lower cytokine content on both day 1 and day 5 in comparison with the fluidized bed (COBE Spectra) or elutriation (Amicus) processes. WBC reduction of BC-PCs of the same origin using three different filters showed comparable levels of cytokines on day 1 in all units. After storage for 5 days, the levels of IL-8 remained essentially unchanged in filtered BC-PCs but increased by more than threefold in control non-filtered BC-PCs, suggesting IL-8 release by residual WBCs present in the control PCs. The concentration of platelet-derived cytokines such as RANTES and TGF-beta(1), however, increased significantly in all filtered and control non-filtered PCs during the storage period. CONCLUSION: These results show that markers of cytokine release from both WBCs and platelets are useful indicators of the performance and efficacy of the WBC-reduction process and of platelet quality.

Biomarkers↗

PubMed2002

Antibodies to bovine beta-casein in diabetes and other autoimmune diseases.

Cow's milk is thought to be an environmental trigger for autoimmune response in Type 1 diabetes. In the present study, our aim was to investigate the antibody response to bovine beta-casein in different immune- and non-immune-mediated diseases and to establish whether such an antibody response is specific to Type 1 diabetes. We measured antibodies to bovine beta-casein using an enzyme-linked immunosorbent assay in a total of 519 sera from subjects as follows: 71 patients with Type 1 diabetes, 33 patients with coeliac disease, 100 patients with latent autoimmune diabetes in adults (LADA), 50 patients with autoimmune thyroid disease (ATD), 50 patients with Type 2 diabetes, 24 patients with multiple sclerosis (MS), and 3 different groups of controls (n = 191). Significantly increased levels of antibodies to beta-casein were found in patients with Type 1 diabetes, coeliac disease and in LADA compared to age-matched controls (p = 0.01, p = 0.02 and p = 0.01, respectively). No differences were observed in beta-casein antibody titres between patients with other disease conditions (MS, and ATD) and age-matched controls. The highest antibody response to beta-casein in Type 1 diabetic patients and in patients with coeliac disease could reflect the gut mucosal immune disorders common to Type 1 diabetes and coeliac disease. Furthermore, the elevated beta-casein antibody levels found in LADA patients suggest that the antibody response to this protein may be relevant in autoimmune diabetes.

Adolescent↗