A genetic diagnosis of maturity-onset diabetes of the young (MODY): experiences of patients and family members.

AIMS: Genetic testing for maturity-onset diabetes of the young (MODY) facilitates a correct diagnosis, enabling treatment optimization and allowing monitoring of asymptomatic family members. To date, the majority of people with MODY remain undiagnosed. To identify patients' needs and areas for improving care, this study explores the experiences of patients and family members who have been genetically tested for MODY. METHODS: Fourteen semi-structured interviews with patients and the parents of patients, and symptomatic and asymptomatic family members were conducted. Atlas.ti was used for thematic analysis. RESULTS: Most people with MODY were initially misdiagnosed with Type 1 or Type 2 diabetes; they had been seeking for the correct diagnosis for a long time. Reasons for having a genetic test included reassurance, removing the uncertainty of developing diabetes (in asymptomatic family members) and informing relatives. Reasons against testing were the fear of genetic discrimination and not having symptoms. Often a positive genetic test result did not come as a surprise. Both patients and family members were satisfied with the decision to get tested because it enabled them to adjust their lifestyle and treatment accordingly. All participants experienced a lack of knowledge of MODY among healthcare professionals, in their social environment and in patient organizations. Additionally, problems with the reimbursement of medical expenses were reported. CONCLUSIONS: Patients and family members are generally positive about genetic testing for MODY. More education of healthcare professionals and attention on the part of diabetes organizations is needed to increase awareness and optimize care and support for people with MODY.

Newborn screening for hunter disease: a small-scale feasibility study.

Hunter disease (Mucopolysaccharidosis type II, MPS II) is an X-linked lysosomal storage disorder caused by deficiency of iduronate-2-sulfatase (IDS). Two main therapies have been reported for MPS II patients: enzyme-replacement therapy (ERT) and hematopoietic stem-cell transplantation (HSCT). Both treatment modalities have been shown to improve some symptoms, but the results with regard to cognitive functioning have been poor. Early initiation of therapy, i.e., before neurological symptoms have manifested, may alter cognitive outcome. The need for early identification makes Hunter disease a candidate for newborn screening (NBS). Our objective was to explore the use of a fluorometric assay that could be applicable for high-throughput analysis of IDS activity in dried blood spots (DBS). The median IDS activity in DBS samples from 1,426 newborns was 377 pmol/punch/17 h (range 78-1111). The IDS activity in one sample was repeatedly under the cutoff value (set at 20% of the median value), which would imply a recall rate of 0.07%. A sample from a clinically diagnosed MPS II individual, included in each 96-well test plate, had IDS activities well below the 20% cutoff value. Coefficients of variation in quality control samples with low, medium, and high IDS activities (190, 304, and 430 pmol/punch/17 h, respectively) were 12% to 16%. This small-scale pilot study shows that newborn screening for Hunter disease using a fluorometric assay in DBS is technically feasible with a fairly low recall rate. NBS may allow for identification of infants with Hunter disease before clinical symptoms become evident enabling early intervention.

[Orphanet: a European database for rare diseases]. / Orphanet: een Europese database over zeldzame ziekten.

Orphanet is a European initiative that aims to improve the management and treatment of rare diseases. It comprises a database dedicated to information on rare diseases and orphan drugs, and offers services adapted to the needs of patients and their families, health professionals, and researchers. The database can be accessed through the website (www.orpha.net) and has some interesting options for searching, for example research projects, support groups or searching by clinical signs. Health professionals are encouraged to add activities concerning rare diseases to the database.

Off-Pump versus on-pump coronary artery bypass: can OPCAB reduce neurologic injury?

OBJECTIVE: Coronary artery bypass grafting (CABG) with cardiopulmonary bypass is still the gold standard for surgical myocardial revascularization. Despite advances in techniques and technologies, documented evidence indicates that cardiopulmonary bypass remains the major source of intraoperative brain injury. This study was set up to test whether offpump coronary artery bypass (OPCAB) is superior to CABG regarding postoperative neurologic outcome or neurocognitive function. METHODS: Between January 1999 and June 2001, 251 patients scheduled for coronary revascularization were divided into 2 groups, CABG (control) and OPCAB. All patients underwent an extensive neurologic and neurocognitive battery of tests preoperatively and postoperatively at 48 hours, 7 days, and 3 months following surgery. RESULTS: There were no statistically significant differences between the 2 groups regarding the preoperative or intraoperative data. The means for patient age, number of grafts, and number of central anastomoses were, respectively, 65.4 years (CABG) and 64.6 years (OPCAB), 3.0 (CABG) and 2.2 (OPCAB), and 2.0 (CABG) and 1.2 (OPCAB). The occurrence of stroke was 2.3% (CABG) and 0% (OPCAB). CONCLUSION: Neurologic complications and postoperative neurocognitive dysfunction remain major concerns in coronary artery surgery. Besides the occurrence of stroke, which dramatically reduces the success of the heart operation, the importance of neurocognitive disorders for postoperative quality of life is not yet well defined. OPCAB significantly improves postoperative neurocognitive function, which may in turn improve the postoperative quality of life.

Variable leukocyte composition of red blood cell concentrates prepared in top-bottom systems: possible implications for pre-transplant blood transfusion.

BACKGROUND AND OBJECTIVES: The beneficial effect of blood transfusion on kidney graft survival requires the presence of leukocytes in the transfusate, but a minimal dose has not been defined, nor has the role of individual leukocyte subsets been investigated. In the Netherlands, a standard pre-transplant blood transfusion consists of a buffy coat (BC)-depleted red blood cell concentrate (RBCC) containing a maximum of 1.2x10(9) residual leukocytes per unit. However, leukocyte subset composition is not standardized. MATERIALS AND METHODS: Using FACS analysis, this study compared the residual leukocyte composition of RBCCs produced by Compomat((R)) and Optipress((R)), two currently used top-bottom systems. RESULTS: While the total leukocyte content of the RBCCs was equivalent in both press types (0.5x10(9)), the percentage of mononuclear cells (lymphocytes and monocytes) was significantly higher in the Compomat as compared with the Optipress system (p < 0. 0001), resulting in significantly higher numbers of transfused T cells, B cells, HLA-DR-positive cells, NK cells and stem cells. CONCLUSIONS: The leukocyte composition of a pre-transplant blood transfusion depends on the BC depletion method used; this might differentially affect the tolerizing or immunizing potential of a pre-transplant blood transfusion.

Germ-free mice do not develop ankylosing enthesopathy, a spontaneous joint disease.

Ankylosing enthesopathy (ANKENT) is a naturally occurring joint disease in mice with numerous parallels to human ankylosing spondylitis (AS). Similarities between AS and ANKENT include not only affected tissue (joint entheses) but also association of the disease with genetic background, including MHC genes, gender, and age. Young males with the C57Bl/10 background have been described to suffer from ANKENT and, among H-2 congenic strains, high frequency of afflicted joints has been recorded in B10.BR (H-2(k)) males. Interestingly, the incidence of ANKENT is higher in conventional (CV) males that in their specific-pathogen-free (SPF) counterparts. The latter finding suggests that microbes could play a role as an ANKENT-triggering agent. To further examine this hypothesis we have established a germ-free (GF) colony of B10.BR mice and observed ANKENT incidence in both GF males and their conventionalized (ex-GF) male littermates; 20% of ex-GF males developed ANKENT before 1 year of age. In contrast, no joint disease was observed under GF conditions (p < 0.0001). Our results show that live microflora is required in ANKENT pathogenesis.

Increased risk of developing ankylosing spondylitis among first-born children.

OBJECTIVE: Studies in mice have demonstrated an increased risk of ankylosing enthesopathy in earlier litters compared with later-born offspring. In humans, birth order and maternal age as risk factors for ankylosing spondylitis (AS) have not been investigated previously. This study was undertaken to investigate whether first-born children have a higher risk of AS than later-born children and whether maternal age at delivery is another risk factor. METHODS: The birth order of 162 AS patients was compared with that of their healthy siblings, both for the total group and with stratification for maternal age at first delivery. Maternal age at the time of delivery of AS patients who were first-born children was compared with the mean maternal age at first delivery in the Dutch population. RESULTS: The number of first-born children with AS was significantly higher than would be expected in case of an equal risk between first-born and later-born children (26 versus 20 for families with 2 children [P = 0.029] and 63 versus 47.6 for all families [P = 0.004]). Also, the mean maternal age at first delivery was lower in mothers of AS patients (24.8 years) compared with mothers of healthy controls (26.1 years). CONCLUSION: Low birth order is a risk factor for AS in humans.

Strong association of HLA-B27 heavy chain with beta 2-microglobulin.

Monoclonal antibody TG1 recognizes specifically antigens HLA-B27, B7, B22 and B17 on cell surface in cytotoxicity and cytofluorometry tests. When cell detergent extracts were subjected to SDS PAGE under mild conditions (no heating and no reduction of the sample) followed by Western blotting, TG1 detected exclusively a complex of B27 heavy chains with beta(2)-microglobulin (as a 50 kDa band) whereas the other B-locus antigens (B7, B22, B17) were detected as free 43 kDa heavy chains under the same conditions. When the samples were boiled prior to SDS PAGE, TG1 detected again the 43 kDa free heavy chains of B7, B22 and B17 but no zone corresponding to B27 could be detected indicating that the epitope in free B27 chains is more sensitive to denaturation by SDS. Thus, our main finding is that the interaction of HLA-B27 heavy chain with beta(2)-microglobulin appears to be stronger than that of the other HLA-B chains. The resistance of the HLA-B27/beta(2)-microglobulin complex to the SDS dissociation is strikingly similar to the behavior of MHC class II molecules under similar conditions. Thus, it may be speculated that HLA-B27 complexes can be also more stable than other MHC class I molecules under more physiological dissociative conditions (e.g. in endosomal compartments). This feature might potentially influence antigen presentation by HLA-B27 and contribute to the well known disease linkage of HLA-B27.

Histopathology of murine ankylosing enthesopathy.

Ankylosing enthesopathy is a spontaneously occurring progressive stiffening of the ankle and/or tarsal joints in mice of C57Black background. In C57BL/10 mice and mice of the same genetic background that had been made transgenic for HLA-B27, the start of the disease was detected by weekly testing for decreased mobility in the ankle/tarsus region. Ankylosing enthesopathy was found to begin with a short phase of proliferative inflammation of the joints and adjacent tissues, with some fibrinous exsudation, some leucocytic infiltration and slight bone erosion. This inflammation is soon accompanied and followed by proliferation of cartilaginous cells at the bone insertions of joint capsule ligaments (entheses). Ossification of the cartilage proliferations and some desmal ossification lead to large osteophytes that inhibit mobility. Fusion of osteophytes occasionally leads to marginal ankylosis. The histopathology of the successive stages of murine ankylosing enthesopathy and the preponderance in males and HLA-B27 transgenic mice are reminiscent of ankylosing spondylitis in man. The spine, however, was not affected.

HIV counselling and testing in Zambia: the Kara Counselling experience.

PIP: This paper examines the impact of voluntary HIV counseling and testing (VCT) in Zambia based on the Kara Counseling Experience. The Kara Counseling and Training Trust has offered VCT services since 1992. Overall, it is noted that although counseling by people other than family members or community elders is a new concept in Zambia, it was seen as useful by virtually all attendees who valued the opportunity to share problems with a counselor who was not judgmental. Although the demand has risen, considerable barriers to HIV counseling still exist such as fear of discrimination and abandonment. Poor communication between partners is in part due to traditional beliefs about discussing sensitive subjects and roles of men and women within marriage. This paper recommends the need to teach young people about HIV and gender awareness. Moreover, VCT should also be linked with medical care, and improving medical services for people with HIV will help reduce the barriers to testing. Lastly, improving linkages to palliative and home care services will help alleviate the fears of those who test positive.^ieng

The role of MHC class I heterodimer expression in mouse ankylosing enthesopathy.

Ankylosing enthesopathy (ANKENT) is a spontaneous mouse joint disease with strikingly similar pathology to human HLA-B27-associated enthesopathies such as ankylosing spondylitis. In C57Bl/10 mice, transgenic HLA-B*2702 as well as H2 genes have been shown to be relative risk factors for ANKENT. To investigate the role of major histocompatibility complex (MHC) class I expression in disease pathogenesis, ANKENT occurrence was compared among beta2-microglobulin (beta2m) knockout littermates with or without transgenes for HLA-B*2702 and human beta2m. In the knockout phenotype lacking beta2m, ANKENT occurrence is significantly reduced (P = 0.016). In the absence of beta2m, B*2702 is not detected on the cell membrane, nor does it increase the risk for ANKENT. This means that the previous finding that HLA-B*2702 increases susceptibility to ANKENT in C57Bl/10 mice cannot be ascribed to a transgene insertion effect. Rather, in order to increase disease susceptibility, B*2702 must be coexpressed with mouse beta2m (mo-beta2m). In contrast, when HLA-B*2702 is expressed with beta2m of human origin, disease susceptibility is not affected. Thus, both H2(b)-derived class I heterodimers and HLA-B*2702/mo-beta2m heterodimers contribute to ANKENT susceptibility.

Grouped caging predisposes male mice to ankylosing enthesopathy.

OBJECTIVE: To evaluate the number of males per cage as a possible risk factor for murine ankylosing enthesopathy (ANKENT)--a spontaneous joint disease with parallels to human seronegative spondylarthropathies--since ANKENT shows incomplete penetrance of genetic susceptibility factors among individuals living in a stable environment. METHODS: Frequency of ANKENT was compared among males housed with females, with other males, or alone. RESULTS: In three independent cohorts, a trend was observed that males housed with females rarely develop the disease, in contrast to males housed with other males (P < 0.25, P < 0.05, and P < 0.01). Furthermore, no males caged alone developed ANKENT, whereas disease did occur in males grouped together (P < 0.01). When healthy males (retired breeders) were recaged either alone or with other males, ANKENT developed among the grouped males only (P < 0.005). CONCLUSIONS: Caging males together is a relative risk factor for ANKENT. Grouped caging may perturb the immune system through endocrine pathways or modify microbiological load through behaviour (for example, infection due to biting).

Solitary caging protects mice from ankylosing enthesopathy.

Murine ankylosing enthesopathy (ANKENT) is a spontaneous joint disease with numerous parallels to the human seronegative spondylarthropathies at the level of disease distribution, genetics and pathology. Although several genetic susceptibility factors have been identified previously, there is variation in disease susceptibility among genetically identical individuals, living in a stable environment. Preliminary observations indicate that males housed together with females rarely develop the disease, in contrast to males housed with other males (p < 0.25, p < 0.05, and p < 0.01). Furthermore, males caged under solitary conditions developed no ANKENT, whereas the disease did develop among individuals housed in groups of 4 (p < 0.01). The mechanism through which grouped caging induces a risk for ANKENT requires further study.

Maternal age influences risk for HLA-B27 associated ankylosing enthesopathy in transgenic mice.

OBJECTIVE: To study further the temporal clustering of ankylosing enthesopathy (AE) noted originally during a study of the influence of mouse major histocompatibility complex (MHC) H-2 and transgenic HLA-B27 on the frequency of AE. METHODS: The relationship between maternal age at littering and frequency of AE was analysed. RESULTS: Mice born to mothers aged eight months or older had a significantly lower disease frequency of AE than mice born to mothers younger than eight months of age. This phenomenon was observed in three independent cohorts evaluated to date (p < 0.01, 0.025, and 0.05). CONCLUSION: Maternal age is a novel, non-genetic risk factor as defined in relation to an MHC associated enthesopathy. Its mode of action and relevance to human disease require further investigation.

HLA-B27 as a relative risk factor in ankylosing enthesopathy in transgenic mice.

HLA-B27 is a risk factor for several human diseases through a mechanism that is not yet understood. This article describes a naturally occurring joint disease in laboratory mice, ANKENT. ANKENT begins with mild inflammation and culminates in irreversible stiffening of the ankle and/or tarsal joints in one or both hind paws. The macroscopic and histologic features of ANKENT, its relationship to age, gender, and environment, and some immunologic aspects are considered. With respect to genetics, it is demonstrated that an HLA-B27 transgene is a relative risk factor for ANKENT. Its impact depends on the H-2 haplotype, reaching a relative risk value of 9.4 for C57Bl/10, H-2b males (p < 0.025). Several features of ANKENT are reminiscent of human AS: joint pathology, age and gender distribution, the presence of non-MHC as well as MHC risk factors (including HLA-B27), and the suspicion that environmental factors are involved.

New technique for sigmoid colectomy.

A combination of laparoscopic and rectoscopic procedures has enabled complete resection of the sigmoid colon and eliminated the need for minilaparotomy when constructing the anastomosis or withdrawing the specimen. This combined procedure was performed in different ways in a series of animal models until the definitive technique had been standardized. These techniques were then successfully used in 15 consecutive medium-sized pigs. Histological evaluation of the specimens, including the anastomosis (examined 2-3 weeks after operation) confirmed the efficacy and safety of the method.

Characterization of a human plasmacytoma line.

The TH line was established by bringing tumour cells from a multiple myeloma patient into suspension culture and subsequently cloning them by limiting dilution. The cultured cells show marked heterogeneity; there are ultrastructural differences between small and large TH cells, particularly with respect to the rough endoplasmatic reticulum (RER). Karyotyping revealed chromosome numbers in the triploid range, with many structural abnormalities, at the 14q32 region among others. A t(14;18) could not be demonstrated. TH was shown to have germline and a rearranged allele for kappa light chain, and only a single rearranged gene for heavy chain immunoglobulin. TH expressed PCA-1, CD9, CD28 and CD38 antigens, HLA class II, RER and kappa light chain, but few or no other antigens associated with the B-cell lineage. Light chain kappa and trace amounts of IgG3 were found intracellularly as well as in culture supernatant. The addition of IL-6 to cultures of TH increased proliferation, as well as the secretion of kappa light chain and the membrane expression of CD28 and CD38 antigens. Because TH has relatively few B cell markers on its membrane, it may be useful for the induction of monoclonal antibodies specific for human plasma cells. It also provides a model for the demonstration that IL-6 can act as a paracrine growth and differentiation factor for cells of myelomal origin.

Pharmacodynamic modeling of lymphocytopenia and whole blood lymphocyte cultures in prednisolone-treated individuals.

In this study, we describe the time course of the influence of a single oral dose of prednisolone (10 mg) in man on the proliferative response of peripheral blood lymphocytes in whole blood. The data were fitted to an integrated pharmacokinetic-pharmacodynamic model, relating the plasma concentrations of prednisolone to its effect. The determination of prednisolone-induced lymphocytopenia and monocytopenia in vivo and the assessment of the influence of varying lymphocyte and monocyte numbers on proliferative responses in vitro enabled us to calculate the relative contributions of several prednisolone-induced effects to the diminished lymphoproliferative responses in whole blood after administration of prednisolone in vivo. We demonstrate that the decrease of the proliferative response in whole blood lymphocyte cultures stimulated with a monoclonal antibody directed against CD3 is mainly determined by the time course of the prednisolone-induced lymphocytopenia. The time course of the monocytopenia and the relative changes in the CD4+ and CD8+ T-cell subpopulations induced by prednisolone and the direct inhibitory effect of the changing plasma concentrations of the drug also contribute to the decrease of aCD3-stimulated whole blood lymphocyte cultures to some extent. The decrease of the proliferative response in whole blood lymphocyte cultures stimulated with a horse anti-human lymphocyte serum closely followed the time course of the lymphocytopenia induced by prednisolone. However, this response remained decreased for a longer period of time than could be expected on the basis of the prednisolone-induced lymphocytopenia in vivo. A possible mechanism which might explain this discrepancy between the prednisolone-induced effects in vivo and in vitro will be discussed.

The influence of prednisolone on the recirculation of peripheral blood lymphocytes in vivo.

In humans and in laboratory animals, administration of a single does of glucocorticosteroids induces a rapid and transient decrease in the numbers of peripheral blood lymphocytes through a redistribution of circulating lymphocytes. This lymphocytopenic effect is an important factor in the immunosuppressive capacity of corticosteroids. We have investigated whether the redistribution is due to an increased influx of circulating lymphocytes from the blood into the lymphoid organs or to a decreased efflux from the organs. Fluoresceinated lymphocytes were injected intravenously in unrestrained normal and prednisolone-treated rabbits and the distribution curves in the peripheral blood were analysed on the basis of a two-compartment model (known from pharmacokinetic studies). In this way, it is shown that the rapid fall in peripheral blood lymphocyte numbers induced by prednisolone is due to a decreased efflux of circulating lymphocytes from the lymphoid organs. In addition, it is demonstrated that prolonged infusion of prednisolone leads to an altered distribution pattern of the circulating lymphocytes among the organs.