The relationship between HbA1c and fasting plasma glucose in patients with increased plasma liver enzyme measurements.

BACKGROUND: HbA(1c) is currently being introduced for diagnostic purpose in diabetes. Previous studies have, however, indicated that patients with liver disease have false low HbA(1c) levels. We therefore investigated the correlation between HbA(1c) and plasma glucose in patients with different levels of increased liver enzyme concentrations. METHODS: Data from 10,065 patients with simultaneous measurement of HbA(1c), venous fasting plasma glucose, alanine aminotransferase and γ-glutamyl transferase were extracted from our laboratory database. Correlations were investigated in four patient groups divided according to their liver enzyme concentrations. RESULTS: The correlation between HbA(1c) and plasma glucose was high in all groups, with r = 0.77 for men and r = 0.78 for women (P < 0.001), a correlation confirmed with multiple regression analysis (P < 0.001). However, interaction analysis revealed that linear regression lines were significantly different for men and women, with increase of both liver enzyme measurements and also, for women, with increased alanine aminotransferase. When compared with biological variation for HbA(1c), only men with increased measurements of both liver enzymes had a clinically important decrease in HbA(1c). CONCLUSIONS: Increased liver enzyme concentrations do not bias the correlation between HbA(1c) and fasting plasma glucose. However, men with low plasma glucose and increased concentrations of both liver enzymes do have a slightly decreased HbA(1c) and, if the clinical suspicion is strong enough, one should consider supplement testing.

Functional characteristics of N8, a new recombinant FVIII.

The aim of this study was to evaluate the in vitro function of the new recombinant factor VIII (FVIII) compound, N8. The specific activity of N8 as measured in a FVIII:C one-stage clot assay was 9300±400 IU mg(-1) based on the analysis of seven individual batches. The ratio between the FVIII:C activity measured in clot and chromogenic assays was 1.00 (95% confidence interval 0.97-1.03). N8 bound to von Willebrand factor with Kd values of 0.2 nm when measured by ELISA and by surface plasmon resonance. FVIIIa cofactor activity was determined from the kinetic parameters of factor IXa-catalysed factor X (FX) activation. The rate of activation of N8 by thrombin as well as Km and kcat for FX activation was in the same range as those observed for Advate®. The rate of activated protein C (APC)-catalysed inactivation was similar for activated N8 and Advate®. N8 improved thrombin generation in a dose-dependent manner and induced similar rates of thrombin generation as Advate® and the plasma-derived FVIII product Haemate®. Using thromboelastography (TEG®), N8 was shown to improve the clot formation and clot stability in whole blood from haemophilia A patients. Comparable potency and efficacy of N8 and Advate® was found based on TEG® parameters. Finally, similar binding profiles to immobilized lipoprotein receptor-related protein (LRP) of N8 and Advate® were observed. The study demonstrated that N8 is fully functional in a variety of assays measuring FVIII activity. No functional differences were found between N8 and comparator compounds.

Purification and characterization of a new recombinant factor VIII (N8).

SUMMARY: A new recombinant factor VIII (FVIII), N8, has been produced in Chinese hamster ovary (CHO) cells. The molecule consists of a heavy chain of 88 kDa including a 21 amino acid residue truncated B-domain and a light chain of 79 kDa. The two chains are held together by non-covalent interactions. The four-step purification includes capture, affinity purification using a monoclonal recombinant antibody, anion exchange chromatography and gel filtration. The specific clotting activity of N8 was 8800-9800 IU mg(-1). Sequence and mass spectrometry analysis revealed two variants of the light chain, corresponding to two alternative N-terminal sequences also known from plasma FVIII. Two variants of the heavy chain are present in the purified product, namely with and without the B-domain linker attached. This linker is removed upon thrombin activation of N8 rendering an activated FVIII (FVIIIa) molecule similar to plasma FVIIIa. All six known tyrosine sulphations of FVIII were confirmed in N8. Two N-linked glycosylations are present in the A3 and C1 domain of the light chain and two in the A1 domain of the heavy chain. The majority of the N-linked glycans are sialylated bi-antennary structures. An O-glycosylation site is present in the B-domain linker region. This site was glycosylated with a doubly sialylated GalNAc-Gal structure in approximately 65% of the product. In conclusion, the present data show that N8 is a pure and well-characterized FVIII product with biochemical properties that equal other FVIII products.

Isolation of a spontaneously fusing BC3H1 muscle cell line: fusion alters the response to serum stimulation.

Differentiation of skeletal muscle cells involves two distinct events: exit from the cell cycle and expression of muscle-specific contractile genes and formation of multinucleated myocytes. Although many studies have shown that growth factors regulate the initial step of differentiation, little is known about regulation of fusion. BC3H1 cells are a skeletal muscle cell line characterized by a nonfusing phenotype and an ability to dedifferentiate. When subjected to serum or growth factors, differentiated BC3H1 cells lose muscle-specific gene expression and re-enter the cell cycle. In this study, we describe a spontaneously fusing clone of BC3H1 cells. We demonstrate that this fusion capability is not due to altered muscle regulatory factor or adhesion molecule expression. Furthermore, we show that fusion inhibits dedifferentiation. Multinucleated BC3H1 cells do not lose myosin expression, nor do they re-enter the cell cycle. Fused BC3HI cells react to serum stimulation with a hypertrophic response. Our results suggest that the state of differentiation, mono- or multi-nucleated, is essential to how myocytes react to growth stimulation and may provide a mechanism for how differentiation, fusion, and hypertrophy are regulated in vivo.

Expression of fibroblast growth factor family during postnatal skeletal muscle hypertrophy.

The potential role of the fibroblast growth factor (FGF) family during stretch-induced postnatal skeletal muscle hypertrophy was analyzed by using an avian wing-weighting model. After 2 or 11 days of weighted stretch, anterior latissimus dorsi (ALD) muscles were, on average, 34 (P < 0.01) and 85% (P < 0.01) larger, respectively, than unweighted ALD control muscles. By using quantitative RT-PCR, FGF-1 mRNA expression was found to be significantly decreased in ALD muscles stretched for 2 or 11 days. In contrast, FGF-4 and FGF-10 mRNA expression was significantly increased 2 days after initiation of stretch. FGF-2, FGF-10, fibroblast growth factor receptor 1, and FREK mRNA expression was significantly increased at 11 days poststretch. Increases in FGF-2 and FGF-4 protein could be detected throughout the myofiber periphery after 11 days of stretch. On a cellular level, FGF-2 and FGF-4 proteins were differentially localized. This differential expression pattern and protein localization of the FGF family in response to stretch-induced hypertrophy suggest distinct roles for individual FGFs during the postnatal hypertrophy process.