Cellular mechanism of the conduction abnormalities induced by serum from anti-Ro/SSA-positive patients in rabbit hearts.

In this study, IgG fractions from sera of SLE patients with anti-Ro/SSA or anti-Ro/SSA and anti-La/SSB activity were tested in Langendorff preparations of adult rabbit hearts, aiming to reproduce the cardiac manifestations observed in neonatal lupus in an experimental model. The hearts were perfused with normal Tyrode's solution for 30 min, followed by perfusion with Tyrode's containing 0.3 mg/ml of anti-Ro/SSA- (or anti-Ro/La-) positive IgG (nine sera), anti-ribonucleoprotein (RNP)-positive IgG (five sera), or IgG fractions from normal donors (five sera). In one third of the experiments done with anti-Ro/La-positive IgG, heart block was observed. With the remaining fractions, a decrease in heart rate of 17.1% was observed, but normal sinus rhythm was maintained. The IgG fractions with anti-RNP activity (five experiments) and from normal sera (six experiments) reduced heart rates by 12.9 and 3.3%, respectively, but heart block was not observed. To further characterize the cellular mechanisms involved in the conduction disturbances observed in the whole rabbit hearts, we conducted experiments with ventricular myocytes isolated from young rabbit hearts, studied by whole cell patch-clamp technique. In these experiments, the slow inward currents were analyzed during the superfusion of the cell with normal Tyrode's solution and 5 min after superfusion with Tyrode's solution containing 0.3 mg/ml of anti-Ro/SSA- (or anti-Ro/La-) positive IgG (five sera), anti-RNP-positive IgG (three sera), or IgG from normal donors (four sera). Resting and action potential amplitudes were not affected by any of the sera used. The anti-Ro/SSA IgG fraction induced a mean reduction in the peak slow inward current of 31.6%. IgG fractions with anti-RNP activity reduced slow inward current by 4.4%, whereas IgG fractions from normal donors increased this current by 3.3%. IgG-free fractions from sera of patients with anti-Ro/SSA activity did not alter the peak slow inward current. These results show, for the first time, that the presence of anti-Ro/SSA or anti-Ro/SSA and anti-La/SSB antibody activity in IgG fractions from lupus patients' sera can induce cardiac conduction disorders similar to those observed in neonatal lupus.     

Differential Stimulation of Noradrenaline Release by Reversal of the Na+/Ca2+ Exchanger and Depolarization in Chromaffin Cells

We compared the effectiveness of Ca²⁺ entering by Na⁺/Ca²⁺ exchange with that of Ca²⁺ entering by channels produced by membrane depolarization with K⁺ in inducing catecholamine release from bovine adrenal chromaffin cells. The Ca²⁺ influx through the Na⁺/Ca²⁺ exchanger was promoted by reversing the normal inward gradient of Na⁺ by preincubating the cells with ouabain to increase the intracellular Na⁺ and then removing Na⁺ from the external medium. In this way we were able to increase the cytosolic free Ca²⁺ concentration ([Ca²⁺]_c) by Na⁺/Ca²⁺ exchange to 325 ± 14 nM, which was similar to the rise in [Ca²⁺]_c observed upon depolarization with 35 mM K⁺ of cells not treated with ouabain. After incubating the cells with ouabain, K⁺ depolarization raised the [Ca²⁺]_c to 398 ± 31 nM, and the recovery of [Ca²⁺]_c to resting levels was significantly slower. Reversal of the Na⁺ gradient caused an −6-fold increase in the release of noradrenaline or adrenaline, whereas K⁺ depolarization induced a 12-fold increase in noradrenaline release but only a 9-fold increase in adrenaline release. The ratio of noradrenaline to adrenaline release was 1.24 ± 0.23 upon reversal of the Na⁺/Ca²⁺ exchange, whereas it was 1.83 ± 0.19 for K⁺ depolarization. Reversal of the Na⁺/Ca²⁺ exchange appeared to be as efficient as membrane depolarization in inducing adrenaline release, in that the relation of [Ca²⁺]_c to adrenaline release was the same in both cases. In contrast, we found that for the same average [Ca²⁺]_c, the Ca²⁺ influx through voltage-gated channels was much more efficient than the Ca²⁺ entering through the Na⁺/Ca²⁺ exchanger in inducing noradrenaline release from chromaffin ceils. This greater effectiveness of membrane depolarization in stimulating noradrenaline release suggests that there is a pool of noradrenaline vesicles which is more accessible to Ca²⁺ entering through voltage-gated Ca²⁺ channels than to Ca²⁺ entering through the Na⁺/Ca²⁺ exchanger, whereas the adrenaline vesicles do not distinguish between the source of Ca²⁺.     

Action of antiestrogens on the (Ca2+ + Mg2+)-ATPase and Na+/Ca2+ exchange of brain cortex membranes

The effect of tamoxifen (TAM) and other antiestrogens on the Ca2+ transport activity of synaptic plasma membranes (SPM) and microsomal membranes isolated from sheep brain cortex was investigated. The maximal (Ca2+ + Mg2+)-ATPase activity of SPM, which is reached at a pCa of about 6.0-6.5, is decreased by about 30% in the presence of 50 microM TAM, whereas the (Ca2+ + Mg2+)-ATPase activity of microsomes, which is maximal at a pCa of about 5.0, is decreased by about 90% by 50 microM TAM. In parallel experiments, we observed that the ATP-dependent Ca2+ uptake is also affected differently by TAM in the two membrane preparations. We found that 50 microM TAM inhibits SPM Ca2+ uptake by about 25-30%, whereas the ATP-dependent Ca2+ uptake by the microsomal fraction is inhibited by about 60%. No significant effect of TAM was observed on the Na+/Ca2+ exchange of either membrane system. The results indicate that TAM is a more potent inhibitor of the active, calmodulin-independent Ca2+ transport system of the intracellular membranes than of that of the plasma membranes, which is calmodulin-dependent. It appears that TAM inhibits calmodulin-mediated reactions, probably through its binding to calmodulin, as we showed previously. However, the Ca2+ transport system of microsomes, which does not depend on calmodulin, is also particularly sensitive to TAM.     

Acute Polymyositis Following Renal Transplantation

Myositis is a rare complication following renal transplantation and is most commonly the result of drug-mediated myotoxicity. Other causative disorders include viral infection, electrolyte imbalance and myositis of autoimmune origin. We describe a 60-year-old patient who developed acute polymyositis 4 weeks after a 000 human leukocyte antigen (HLA) mismatch cadaveric renal transplant. Following an uncomplicated transplant course with maintenance triple immunosuppression (prednisolone, mycophenolate mofetil and cyclosporine), the patient presented with severe symmetrical proximal muscle weakness associated with a rise in serum creatine kinase to 46800 U/L. Electromyography confirmed myopathic changes and muscle biopsy demonstrated extensive muscle-fiber necrosis with an inflammatory infiltrate. There were no obviously culpable drugs and viral studies were negative. Prompt initiation of high-dose steroid therapy led to clinical and biochemical recovery. Acute polymyositis may occur following renal transplantation. Potential mechanisms include viral antigen transmission or a localized form of graft vs. host disease.     

Telomerase activity in prognostic histopathologic features of melanoma.

BACKGROUND: Telomerase activity (TA) is believed to play a role in the regulation of senescence and to limit the number of cell divisions. The deregulation of telomerase appears to contribute to oncogenesis and the formation of immortal cell lines. As a result, it is believed that it could be used as a prognostic marker in melanoma. METHODS: TA was assayed by the polymerase chain reaction PCR-ELISA-based telomeric repeat amplification protocol (TRAP assay). One hundred and eight samples were distributed in four histological groups: 30 samples from primary cutaneous melanomas, 24 from peritumoural skin sites, 28 from benign melanocytic lesions, and 26 from normal skin sites as a control. RESULTS: TA was different among the four tested groups (Kruskall-Wallis test p<0.001), and increasing values of TA were observed progressing from normal skin to benign and then to malignant lesions. Among melanoma samples, there was a significant association between TA and ulceration (p=0.025), TA and vascular invasion (p=0.018) and TA and mitotic rate (p=0.029) (Mann-Whitney test). A linear regression analysis showed significant associations between the increase of TA with Breslow thickness (p=0.004) and the presence of satellites (p=0.002). CONCLUSIONS: We observed that TA had increased from control skin to peritumoural skin, and then to benign melanocytic lesions and finally to melanoma, suggesting tumour progression. TA showed higher values in the presence of some important histopathologic parameters related to poor prognosis in cutaneous melanoma such as ulceration, vascular invasion, satellites, high rates of mitosis, and in thicker tumours.     

Blood pressure on arising, morning blood pressure surge and blood pressure variability in white coat hypertensives and in matched normotensives and sustained hypertensives.

INTRODUCTION: It is still controversial whether subjects with white-coat hypertension (WCHT) exhibit higher cardiovascular risk compared to normotensive subjects (NT). In subjects with WCHT it is not known whether the abnormal blood pressure (BP) reaction in the office also occurs at other times of day, particularly on arising and immediately after waking, i.e. the times at which the majority of cardiovascular events are reported to occur. OBJECTIVE AND METHODS: To evaluate with 24h ambulatory BP measurement the values of morning BP surge, BP on arising and BP variability in subjects with WCHT in comparison with age-, gender- and weight-matched normotensives (BP) and untreated sustained hypertensives (BP). RESULTS: Groups of BP, WCHT and BP were matched for age, gender and body weight: BP: n=69, age 49 +/- 7 years, 54 % female, BMI 26 +/- 1, casual BP 126/79 +/- 5/4 mmHg, daytime BP 124/80 +/- 6/6 mmHg; WCHT: n=74, age 52 +/- 8 years, 57% female, BMI 26 +/- 2, casual BP 152/95 +/- 7/7 mmHg, daytime BP 126/80 +/- 5/6 mmHg; HT: n=79, age 53 +/- 7 years, 56% female, BMI 27 +/- 2, casual BP 154/97 +/- 9/8 mmHg, daytime BP 143/89 +/- 12/10 mmHg. Of the three groups, subjects with WCHT exhibited BP on arising (121/81 +/- 13/8 mmHg) similar to that of NTs (120/80 +/- 13/9 mmHg, NS), both significantly lower than that of HTs (137/92 +/- 17/10 mmHg, p < 0.01), suggesting the absence of an alerting BP reaction in WCHT at that time. By contrast, subjects with WCHT showed higher values of systolic morning BP surge vs. NTs (25 +/- 10 vs. 22 +/- 11 mmHg, p < 0.05), both lower than that observed in hypertensives (33 +/- 11 mmHg, p < 0.01 vs. NT and WCHT) and greater daytime variability (systolic BP standard variation), i.e. 12 2 vs. 10 +/- 2 mmHg, p < 0.05, both lower than that observed in hypertensives (14 +/- 3 mmHg, p < 0.01 vs. NT and WCHT). CONCLUSIONS: Although subjects with WCHT did not show any alerting blood pressure reaction on arising, morning BP surge and BP variability were greater in these subjects than in control normotensives, although lower than sustained hypertensives. Although this is still speculative, we cannot exclude the possibility that even a slight increase in morning BP surge might in the long term constitute an additional load on the circulation that could increase cardiovascular risk in subjects with WCHT compared to matched normotensives.     

The ED50 and ED95 of Intrathecal Isobaric Bupivacaine with Opioids for Cesarean Delivery

Background: The ideal intrathecal isobaric bupivacaine dose for cesarean delivery anesthesia is uncertain. While small doses (5-9 mg) of bupivacaine may reduce side effects such as hypotension, they potentially increase spinal anesthetic failures. This study determined the ED50 and ED95 of intrathecal isobaric bupivacaine (with adjuvant opioids) for cesarean delivery.

Methods: After institutional review board approval and written informed consent were obtained, 48 parturients undergoing elective cesarean delivery under combined spinal-epidural anesthesia were enrolled in this double-blind, randomized, dose-ranging study. Patients received a 5-, 6-, 7-, 8-, 9-, 10-, 11-, or 12-mg intrathecal isobaric bupivacaine dose with 10 [mu]g fentanyl and 200 [mu]g morphine. Overall anesthetic success was recorded when no intraoperative epidural supplement was required during the cesarean delivery. ED50 and ED95 values for overall anesthetic success were determined using a logistic regression model.

Results: ED50 and ED95 values for overall anesthetic success were 7.25 and 13.0 mg, respectively. No advantages for low doses could be demonstrated with regard to hypotension, nausea, vomiting, pruritus, or maternal satisfaction, although this study was underpowered to detect significant differences in secondary outcome variables.     

Type 2 diabetes: assessment of endothelial lesion and fibrinolytic system markers.

This study aimed to investigate whether endothelial cells are damaged and to evaluate fibrinolytic system function in patients with type 2 diabetes. For this proposal, plasma levels of von Willebrand factor (an endothelial marker of injury), homocysteine (an inductor of endothelial injury), D-dimer (a marker of coagulation cascade activation) and plasminogen activator inhibitor-1 (a fibrinolysis marker) were measured in individuals with both type 2 diabetes and high blood pressure, with type 2 diabetes, with high blood pressure and in healthy control individuals. No significant differences among groups were observed for von Willebrand factor and homocysteine plasma levels. The type 2 diabetes and high blood pressure group presented a significant difference to the other groups for D-dimer and also presented high values for plasminogen activator inhibitor-1. The high blood pressure group and type 2 diabetes group presented separately higher values of plasminogen activator inhibitor-1 compared with the control group. High levels of D-dimer and plasminogen activator inhibitor-1 in patients with type 2 diabetes and high blood pressure with normoalbuminuria therefore indicate a state of hypercoagulability and hypofibrinolysis, despite no evident microvascular injury supported by normal levels of von Willebrand factor and homocysteine.