Gorog Thrombosis Test: analysis of factors influencing occlusive thrombus formation.

We used the Gorog Thrombosis Test to analyze the factors influencing the occlusion time, which represents platelet activation and subsequent occlusive thrombus formation, in 132 healthy Japanese volunteers (116 men, 16 women; mean age, 45.0 +/- 12.0 years). The Gorog Thrombosis Test was designed to evaluate platelet aggregation and thrombolytic activity under a high shear stress condition (175 dynes/cm) in a native blood sample in vitro. The mean +/- SD occlusion time was 154.8 +/- 64.7 s (men, 153.4 +/- 64.2 s and women, 165.4 +/- 56.5 s). The occlusion time was inversely correlated with von Willebrand factor ristocetin cofactor activity (VWF:Rco) (r = -0.242, P = 0.0055) and von Willebrand factor antigen (r = -0.230, P = 0.0080). The mean occlusion time in the group with VWF:Rco of at least 170% (137 s) was significantly shorter than that in the group with VWF:Rco less than 170% (156 s, P < 0.05). Platelet counts, other coagulation markers and smoking showed no significant correlations with occlusion time. Red blood cells (r = -0.177, P = 0.0365), hemoglobin (r = -0.191, P = 0.0245) and hematocrit (r = -0.182, P = 0.0329) also showed inverse correlations with the occlusion time. This report is the first to clearly demonstrate the role of von Willebrand factor in the formation of occlusive thrombi in the Gorog Thrombosis Test.     

Surgical treatment for intrathoracic aneurysm of the innominate artery in an 83-year-old asymptomatic woman

Aneurysm of the innominate artery is uncommon compared with other peripheral aneurysms, and holds the potential for rupture, embolization, or thrombosis as well as various complications caused by compression to the adjacent structures. The most effective treatment for this condition is surgical resection, but the earlier reports described high mortality rates. We report the case of an 83-year-old asymptomatic woman with an aneurysm in the innominate artery, which was successfully resected and repaired with the use of modern surgical techniques of hypothermic circulatory arrest and selective cerebral perfusion. Aggressive surgical intervention should be employed despite the fact that a patients is asymptomatic.     

Analysis of small dense LDL in patients with type 2 diabetic mellitus by the modified Krauss method using an internal standard

In patients with Type 2 diabetes mellitus (Type 2 DM), the relationship between the prevalence rate of small dense LDL (sdLDL) and parameters of lipid metabolism was analyzed using the method devised by modified Krauss method using apoferritin as an internal standard. The prevalence rate of sdLDL was 34% compared with it of normal subjects in this study. When the severity of Type 2 DM was classified into three groups of the HbA1c value, neither the sdLDL size nor its prevalence rate differed significantly depending upon the severity of the Type 2 DM. Also, when the prevalence rate of sdLDL was analyzed in relation to the severity of complications, i.e., of microangiopathy (retinopathy and nephropathy) or macroangiopathy (cerebral infarction), there was no significant difference in the prevalence rate of sdLDL depending on the severity of any of these complications. On the other hand, the prevalence rate of sdLDL was found to be correlated with the serum TG level. The serum level of TG-rich remnants (metabolites of TG) was also high in patients with sdLDL. It should take notice that the assessment of sdLDL should be used the authorized method for the evaluation. Thus it is concluded that the levels of sdLDL were important in evaluation of Type 2 DM. The prevalence rate of sdLDL did not correlate with the severity, nor the modalities for the complications of Type 2 DM.     

Measurement of plasma von Willebrand factor cleaving protease in patients with varied thrombotic microangiopathy

Thrombotic thrombocytopenic purpura(TTP) is a multisystem disorders characterized by thrombocytopenia, microangiopathic hemolytic anemia associated with red cell fragmentation, and neurological and renal symptoms. Plasma of patients with TTP has been shown to contain unusually large von Willebrand factor(vWF) multimers that may cause platelet agglutination in vivo. Recently, a metalloprotease responsible for cleavage of vWF multimers has been isolated from normal human plasma and was found to be deficient in some patients with TTP. We examined the activity of the vWF-cleaving protease(vWF-CP), by modified Furlan's method, in plasma from patients with a familial TTP, 3 acquired TTP, 4 thrombotic microangiopathy(TMA) and 2 veno-occlusive disease(VOD) associated after allo-BMT. Diluted plasma samples of patients were incubated with protease-free vWF purified from normal human plasma, in the presence of urea and barium ions. The extent of vWF degradation was assayed by electrophoresis in SDS-agarose gels and immunoblotting. Activity of vWF-CP from 12 normal plasma have been shown as 77-180%(average 115%), whereas, no vWF-CP(below 5%) was observed in plasma from familial TTP, before and after plasma exchange, although FFP infusion therapy has been effective for this patient to recover thrombocytopenia. In 3 acquired TTP, 2 patients showed lack of vWF-CP activity in plasma, and inhibitors against vWF-CP have been elucidated by plasma cross-mixing test. After extensive plasma exchange and FFP infusion followed by corticosteroid therapy, normal vWF-CP was recovered in plasma from 2 acquired TTP patients. Among BMT patients, plasma from 4 BMT-TMA showed normal vWF-CP activities as 55-111%, whereas plasma from 2 BMT-VOD revealed low vWF-CP activity, as 24% and 37%, respectively. Thus, measurement of vWF-CP is crucial to predict differentiation of primary forms of TMA to establish the pathogenesis in varied endothelial dysfunction.     

Ramp rate of blood pressure changes does not affect aortic afferent sensitivity in anesthetized rats

To investigate whether the rate of change in blood pressure affects the sensitivity of the aortic baroreceptor afferent response, the change in aortic nerve activity (ANA) to two different rates of ramp increase in mean blood pressure (MBP), elicited by phenylephrine administration, was determined in the rat under urethane (1.5 g kg⁻¹) anesthesia. The sensitivity of the increase in ANA following a rapid (average ramp rate, 9.14 ± 0.60 mmHg s⁻¹, n = 11) or gradual (1.78 ± 0.24 mmHg s⁻¹, n = 11) increase in MBP was 2.03 ± 0.14% and 1.81 ± 0.20% of baseline mmHg⁻¹, respectively. These values were not significantly different from each other (P = 0.16). Furthermore, we found no correlation between the rate of ramp increase in MBP and the sensitivity of the increase in ANA (r = 0.24, P = 0.29, n = 22). These results suggest that, at least within the normal physiological range of MBP, the rate of the ramp change in blood pressure does not affect aortic baroreceptor afferent sensitivity in the anesthetized rat.     

Enzymatic kinetics regarding reversible metabolism of CS-0777, a sphingosine 1-phosphate receptor modulator,via phosphorylation and dephosphorylation in humans

1.?CS-0777, a candidate compound for autoimmune diseases, becomes phosphorylated active metabolite, M1, by fructosamine 3-kinase (FN3K), FN3K-related protein (FN3K-RP); and M1 is reverted back to CS-0777 by alkaline phosphatase (ALP) in the body. We performed enzyme kinetic analysis of phosphorylation of CS-0777 by FN3K, FN3K-RP, human erythrocytes and human platelets; and dephosphorylation of M1 by various ALP isozymes and human liver, kidney, lung and small intestine microsomes.

2.?The Michaelis constants of human FN3K, FN3K-RP and erythrocytes for CS-0777 phosphorylation were in the range from 498?μM to 1060?μM. FN3K inhibitor, 1-deoxy-1-morpholinofructose, suppressed only about 20% of CS-0777 phosphorylation activity in human erythrocyte lysate. Immunodepletion of FN3K and FN3K-RP decreased M1 formation activity by about 25% and 50%, respectively, in human erythrocyte lysate.

3.?The Michaelis constants of four human ALPs and microsomes were in the range from 10.9?μM to 32.1?μM. The ALP inhibitor, levamisole, suppressed over 50% of M1 dephosphorylation activity in liver, kidney and lung microsomes.

4.?FN3K-RP is expected to take a prominent role in the phosphorylation of CS-0777 in human erythrocytes; dephosphorylation of M1 was observed in all ALPs and human tissue microsomes examined, with a similar affinity towards M1 among them.     

Reference values for urinary steroids in Japanese newborn infants: gas chromatography/mass spectrometry in selected ion monitoring

Urinary steroid profile analysis using gas chromatography/mass spectrometry (GC/MS) has been reported for the diagnosis of abnormal steroidogenesis in newborn infants with some success. We tried to establish the reference values of 63 urinary steroids in Japanese newborn infant, using GC/MS in selected ion monitoring (SIM) that utilizes two characteristic mass ions for each steroid for definitive identification. We studied 36 healthy full-term newborn infants (1-56 days of age) on spot urine samples to define the reference values (mg/g creatinine, median and 10-90 percentile range) and to investigate the possible difference between daytime and nighttime levels. We also studied 23 healthy adult females (20-24 years of age) on 24-hour-urine for the comparison of the reference values of newborn infants. Fifty metabolites of DHEA, pregnenolone, 17-hydroxypregnenolone, androstenedione, progesterone, 17-hydroxyprogesterone, 21-deoxycortisone, corticosterone, 18-hydroxycorticosterone, aldosterone, 18-hydroxycortisol, 11-deoxycortisol, cortisone, cortisol, and estrogen in each infant were measurable without interference, but 13 metabolites of 11-hydroxyandrostenedione, pregnenolone, 11-deoxycorticosterone, corticosterone, 11-dehydrocorticosterone, 21-deoxycortisol, 11-deoxycortisol and cortisol were unmeasurable in each infant due to the interference of fetal cortex steroids as confirmed by abnormal peak area ratios of two mass ions. All 63 metabolites in each control adult were measurable without interference. 16alpha-, 16beta-, and 15beta-hydroxy metabolites of 3beta-hydroxy-5-en-steroids, and 6beta-, 18-hydroxy and 11-oxo-metabolites of corticosteroids were significantly higher in full-term newborn infants than those in adults as previously reported. Urinary steroids showed little circadian variation in the newborn infants, indicating that spot urine can substitute for 24-hour urine.     

Evaluation of the efficacy of an Helicobacter pylori eradication treatment for idiopathic thrombocytopenic purpura patients]

The relationship between Helicobacter pylori (H. pylori) and gastric diseases (e.g. peptic ulcer, MALT lymphoma, and stomach cancer) has been widely accepted. Recent studies have also suggested an association between H. pylori infection and idiopathic thrombocytopenic purpura (ITP). In this study, an H. pylori eradication treatment was administered to 20 ITP patients and elucidated for its effectiveness. Among those 20 patients, H. pylori infection was confirmed in 17 (85%) through a C14 urea breath test, a rapid urease test, or a culture examination of a biopsied sample obtained by gastrointestinal endoscopy. Although the other 3 were negative to H. pylori, the H. pylori eradication treatment was also attempted because no other effective treatments had been established at the time of this study. In the H. pylori eradication treatment, lansoprazole (LPZ) 60 mg bid, amoxicillin (AMPC) 1500 mg bid, and clarithromycin (CAM) 400 mg bid were given to each patient for 7 days. For 4 cases, CAM was replaced with metronidazole (MNZ) 750 mg bid. The patients whose H. pylori infection was not eradicated after the first treatment received the re-eradication treatment with LPZ 60 mg bid, AMPC 1500 mg bid, and MNZ 750 mg bid for 7 days. After the treatments, the success of eradicating H. pylori was confirmed in all 17 H. pylori positive patients. In addition, platelet recovery was obtained in 11/20 patients (55%), which included 2 H. pylori negative patients and 2 patients whose H. pylori eradication was not successful after the first treatment. No relationship was found between the eradication effectiveness and the following clinical parameters: age, gender, previous therapies, disease duration, presence of anti-nucleus antibody, endoscopic atrophic change in the stomach, or kinds of antibiotics used for the treatment. These results support the efficacy of an H. pylori eradication treatment for ITP patients. A noteworthy result of this study was that an increase of platelet count was observed not only in H. pylori positive ITP patients, but also in 2 out of 3 H. pylori negative ITP patients after H. pylori eradication. Further studies are required to elucidate the efficacy of H. pylori eradication therapy in the patients negative for H. pylori.     

Trends of hepatitis B virus genotype distribution in chronic hepatitis B patients in Japan

Journal of Gastroenterology - Hepatitis B virus (HBV) is one of the most prevalent chronic viral infections that causes chronic hepatitis B (CHB). In Japan, genotypes B and C account for most of...