Development and validation of an immunoluminometric assay (ILMA) for fetuin-A-glycoprotein.

BACKGROUND: To identify dialysis patients with low fetuin-A levels, a sensitive immunoluminometric assay (ILMA) was developed. METHODS: For the two-site ILMA, one monoclonal antibody was coated to polystyrene beads and one polyclonal antibody was labelled with acridinium ester. RESULTS: The lower detection threshold was 0.013 g/L, with the normal range 0.20-0.87 g/L (arithmetic mean 0.437 +/- 0.118 g/L). Serum fetuin-A levels in the dialysis patients were significantly lower (arithmetic mean: 0.352 +/- 0.099 g/L, p < 0.0001). CONCLUSION: This ILMA has been proved to be a reliable method for the determination of serum fetuin-A concentrations in dialysis patients.

Myoglobin plasma level related to muscle mass and fiber composition: a clinical marker of muscle wasting?

Progressive muscle wasting is a central feature of cancer-related cachexia and has been recognized as a determinant of poor prognosis and quality of life. However, until now, no easily assessable clinical marker exists that allows to predict or to track muscle wasting. The present study evaluated the potential of myoglobin (MG) plasma levels to indicate wasting of large locomotor muscles and, moreover, to reflect the loss of MG-rich fiber types, which are most relevant for daily performance. In 17 cancer-cachectic patients (weight loss 22%) and 27 age- and gender-matched healthy controls, we determined plasma levels of MG and creatine kinase (CK), maximal quadriceps muscle cross-sectional area (CSA) by magnetic resonance imaging, muscle morphology and fiber composition in biopsies from the vastus lateralis muscle, body cell mass (BCM) by impedance technique as well as maximal oxygen uptake (VO(2)max). In cachectic patients, plasma MG, muscle CSA, BCM, and VO(2)max were 30-35% below control levels. MG showed a significant positive correlation to total muscle CSA (r = 0.65, p < 0.001) and to the CSA fraction formed by type 1 and 2a fibers (r = 0.80, p < 0.001). However, when adjusted for body height and age by multiple regression, MG yielded a largely improved prediction of total CSA (multiple r = 0.83, p < 0.001) and of fiber type 1 and 2a CSA (multiple r = 0.89, p < 0.001). The correlations between CK and these muscle parameters were weaker, and elevated CK values were observed in 20% of control subjects despite a prior abstinence from exercise for 5 days. In conclusion, plasma MG, when adjusted for anthropometric parameters unaffected by weight, may be considered as a novel marker of muscle mass (CSA) indicating best the mass of MG-rich type 1 and 2a fibers as well as VO(2)max as an important functional readout. CK plasma levels appear to be less reliable because prolonged increases are observed in even subclinical myopathies or after exercise. Notably, cancer-related muscle wasting was not associated with increases in plasma MG or CK in this study.

Transgenic mice overexpressing erythropoietin adapt to excessive erythrocytosis by regulating blood viscosity.

Severe elevation of red blood cell number is often associated with hypertension and thromboembolism resulting in severe cardiovascular complications. However, some individuals such as high altitude dwellers cope well with an increased hematocrit level. We analyzed adaptive mechanisms to excessive erythrocytosis in our transgenic (tg) mice that, due to hypoxia-independent erythropoietin (Epo) overexpression, reached hematocrit values of 0.8 to 0.9 without alteration of blood pressure, heart rate, or cardiac output. Extramedullar erythropoiesis occurred in the tg spleen, leading to splenomegaly. Upon splenectomy, hematocrit values in tg mice decreased from 0.89 to 0.62. Tg mice showed doubled reticulocyte counts and an increased mean corpuscular volume. In tg mice, plasma volume was not elevated whereas blood volume was up to 25% of the body weight compared with 8% in wild-type (wt) siblings. Although plasma viscosity did not differ between tg and wt mice, tg whole-blood viscosity increased to a lower degree (4-fold) than expected from corresponding hemoconcentrated wt blood (8-fold). This moderate increase in viscosity is explicable by the up to 3-fold higher elongation of tg erythrocytes at physiologic shear rates. Apart from the nitric oxide-mediated vasodilation we reported earlier, adaptation to high hematocrit levels in tg mice involves regulated elevation of blood viscosity by increasing erythrocyte flexibility.

Prevention of chemotherapy-related toxic side effects by infection with adeno-associated virus type 2.

Drug resistance and toxic side effects are major limiting factors in the clinical use of antineoplastic chemotherapy. Patients with pancreatic cancer generally do not benefit from chemotherapy. The nonpathogenic adeno-associated virus type 2 (AAV-2) has been shown to sensitize human tumor cells to gamma irradiation and chemotherapeutic drugs. In the present study, we characterized the therapeutic role of AAV-2 infection in combination with 5-fluorouracil (5-FU)-based chemotherapy on pancreatic cancer cells in an animal model. In Lewis rats bearing s.c. implants of syngeneic DSL6A pancreatic cancer cells, intratumoral infection with AAV-2 (MOI 10E8 i.u.) in combination with 5-FU (5 or 50 mg/kg body weight) resulted in significantly reduced tumor growth and prolonged survival time compared with 5-FU single therapy. Most surprisingly, AAV-2-infected rats remained in a much better physical condition compared to their noninfected counterparts. While rats treated with 5-FU single therapy lost weight, were sluggish and died within 4 months after tumor implantation, animals infected with AAV showed much better vigilance, with body weight, leukocyte number and hemoglobin levels similar to healthy rats. In particular, 5-FU-related side effects like thrombocytopenia and leukopenia were significantly reduced in animals treated with the combination regimen. By in vitro analysis, human (Capan-1 and DANG) pancreatic cancer cell lines were shown to be sensitized to 5-FU chemotherapy to an extent similar to DSL6A cells. AAV-2 infection enhanced 5-FU-induced apoptosis by a factor of 8 to 14 in both human and rat pancreatic cancer cell lines. The data suggest that infection with the nonpathogenic AAV-2 significantly improves both chemotherapy efficacy and physical appearance and offers a novel strategy in cancer treatment.