Analysis of plasmin generation and clot lysis of plasma fibrinogen purified from a heterozygous dysfibrinogenemia, B beta Gly15Cys (Hamamatsu II)

This is a non-final version of an article published in final form in Blood Coagulation & Fibrinolysis. 20(8):726-732, December 2009.We found a heterozygous dysfibrinogenemia caused by the substitution of B beta Gly15Cys and designated it fibrinogen Hamamatsu II (H-II). Although the propositus suffered an infarction of the medulla oblongata, other thrombotic risk factors, paradoxical cerebral infarction, and arterial dissection were not found. To determine whether the delayed lysis of fibrin clots or not in the context of the B beta Gly15Cys substitution, we examined the clot lysis and plasmin generation of propositus' fibrinogen. Fibrinogen was purified from the propositus' and normal control plasma by immunoaffinity chromatography and was used for the following experiments: sodium dodecyl sulfate-polyacrylamide gel electrophoresis, fibrin polymerization, scanning electron microscopic observation of fibrin clot and fibers, clot lysis, and tissue-type plasminogen activator-mediated plasminogen activation. The H-II plasma fibrinogen showed the presence of albumin-binding variant forms, a dimeric molecule of variant fibrinogen, and impairment of lateral aggregation during fibrin polymerization. The H-II fibrin clot showed lower density of bundles and thinner diameters of fibers than in the normal fibrin clot. In the clot lysis experiments with overlaid plasmin, H-II fibrin showed a similar lysis period and lysis rate to the normal control. Moreover, plasmin generation from a mixture of thrombin, tissue-type plasminogen activator, plasminogen, and H-II fibrinogen also showed a similar rate to normal fibrinogen. Although the propositus suffered an infarction, the present study did not observe delayed clot lysis, that is, the clot was not resistant to plasmin degradation. Therefore, we did not clarify an association between the B beta Gly15Cys dysfibrinogenemia and arterial thrombosis.ArticleBLOOD COAGULATION & FIBRINOLYSIS. 20(8):726-732 (2009)journal articl

Heterozygous B beta-chain C-terminal 12 amino acid elongation variant, B beta X462W (Kyoto VI), showed dysfibrinogenemia

A heterozygous patient with dysfibrinogenemia with slight bleeding and no thrombotic complications was diagnosed with fibrinogen Kyoto VI (K-VI). To elucidate the genetic mutation(s) and characterize the variant protein, we performed the following experiments and compared with identical and similar variants that have already been reported. The proposita's PCR-amplified DNA was analyzed by sequencing and her purified plasma fibrinogen underwent SDS-PAGE followed by immunoblotting, fibrin polymerization, and scanning electron microscopic observation of fibrin clot and fibers. Sequence analyses showed that K-VI fibrinogen substituted W (TGG) for terminal codon (TAG), resulting in 12 amino acid elongation 462-473 (WSPIRRFLLFCM) in the B beta-chain. Protein analyses indicated that the presence of some albumin-binding variant fibrinogens and a dimeric molecule of variant fibrinogens reduced fibrin polymerization, with a thinner fiber and aberrant fibrin network. These results are almost the same as for the identical variant of Magdeburg, however, different from the similar variant of Osaka VI [ 12 amino acid elongation 462-473 (KSPIRRFLLFCM) in the B beta-chain] in the presence of variant forms and clot structure. We speculate the side-chain difference at 462 residues, W in K-VI, K in Osaka VI, and/or the difference in the presence of disulfide bridged forms of variant fibrinogens, led to the notable difference in the fibrin bundle network. Although a strong evolutional and structural association between B beta-chain and gamma-chain molecules is established, the corresponding recombinant 15 residue elongation variants of the fibrinogen gamma-chain showed reduced assembly and secretion.ArticleBLOOD COAGULATION & FIBRINOLYSIS. 23(1):87-90 (2012)journal articl

Notoamides A-D: prenylated indole alkaloids isolated from a marine-derived fungus, Aspergillus sp.

金沢大学大学院自然科学研究科生理活性物質科学金沢大学薬学

シシツ テイカ リョウホウ ニヨル ケイドウミャク プラーク アンテイカ ノ ヒョウカ : チョウオンパ integrated backscatter オ モチイタ カラー マッピング システム ノ カイハツ ト リンショウ オウヨウ

Background : The carotid plaque vulnerability is related to myocardial and cerebral infarction. We intended to develop an imaging system which enables to visualize tissue characteristics in the carotid plaques based on ultrasound integrated backscatter（IB）. And to test its clinical efficacy, effect of the statin therapy on the plaques was evaluated with our software. Methods and Results : Carotid ultrasound examination was performed and ultrasonographic RAW data of the plaques were obtained from８patients undergoing carotid artery endarterectomy. Tissue characteristics in the plaques of resected examples were compared with preoperative ultrasonic images and the tissue IB values corresponding to the specimens were determined for developing our imaging system. Using this system, Color-coded maps of plaques in the three patients were constructed before and after lipid lowing therapy. We could demonstrate that lipid fraction in each plaque decreased and fibrous or calcification fraction increased in the follow-up study. Conclusions : Changes in histology of carotid plaques by statin could visualized with our imaging system. This technique may become a useful tool for the management of atherosclerosis

Prevention of hypoglycemia by intermittent-scanning continuous glucose monitoring device combined with structured education in patients with type 1 diabetes mellitus : A randomized, crossover trial

Aims: We conducted a randomized, crossover trial to compare intermittent-scanning continuous glucose monitoring (isCGM) device with structured education (Intervention) to self-monitoring of blood glucose (SMBG) (Control) in the reduction of time below range. Methods: This crossover trial involved 104 adults with type 1 diabetes mellitus (T1DM) using multiple daily injections. Participants were randomly allocated to either sequence Intervention/Control or sequence Control/Intervention. During the Intervention period which lasted 84 days, participants used the first-generation FreeStyle Libre (Abbott Diabetes Care, Alameda, CA, USA) and received structured education on how to prevent hypoglycemia based on the trend arrow and by frequent sensor scanning (≥10 times a day). Confirmatory SMBG was conducted before dosing insulin. The Control period lasted 84 days. The primary endpoint was the decrease in the time below range (TBR; <70 mg/dL). Results: The time below range was significantly reduced in the Intervention arm compared to the Control arm (2.42 ± 1.68 h/day [10.1 %±7.0 %] vs 3.10 ± 2.28 h/day [12.9 %±9.5 %], P = 0.012). The ratio of high-risk participants with low blood glucose index >5 was significantly reduced (8.6 % vs 23.7 %, P < 0.001). Conclusions: The use of isCGM combined with structured education significantly reduced the time below range in patients with T1DM

Effects of back massage on psychological status and salivary biomarkers

Massage therapy promotes psychosocial relaxation and reduce stress. In addition, this therapy has been reported to improve immune function. Although evaluation of psychosocial status has been performed with subjective psychological tests such as State-Trait Anxiety Inventory （STAI）, subjective psychological tests are of limited value if the subjects fail to report reliably. Salivary biomarkers have been recently suggested as useful objective indicators for assessing psychosocial status. To determine whether salivary biomarkers are useful objective indices for assessing eff ects of back massage on psychological status in 25 young healthy female volunteers, we measured heart rate and salivary biomarkers（ α-amylase, cortisol and chromogranin-A） and assessed STAI score before and after back massage. Back massage significantly reduced heart rate from 73.4±11.8 to 69.8±11.2 and STAI from 41.0±6.0 to 32.3±4.9. In contrast salivary chromogranin-A signifi cantly increased from 2.93±2.21 to 5.29±5.46 pmol/mg protein whilst salivary α-amylase and cortisol did not change. Therefore, salivary biomarkers tested may not indicate changes in psychological relaxation following back massage. Massage therapy has been reported to not only reduce psychosocial stress but also enhance immune functions in cancer patients. In the present study, massage therapy significantly increased chromogranin-A release. As several reports clearly show that chromogranin-A has antibacterial and antifungal activities, back massage may increase host defense with salivary chromogranin-A release against oral microbial invasion

Robustness of Clonogenic Assays as a Biomarker for Cancer Cell Radiosensitivity

Photon radiation therapy is a major curative treatment for cancer. However, the lack of robust predictive biomarkers for radiosensitivity precludes personalized radiation therapy. Clonogenic assays are the gold standard method for measuring the radiosensitivity of cancer cells. Although a large number of publications describe the use of clonogenic assays to measure cancer cell radiosensitivity, the robustness of results from different studies is unclear. To address this, we conducted a comprehensive detailed literature search of 256 common cancer cell lines and identified the eight cell lines most-frequently examined for photon sensitivity using clonogenic assays. Survival endpoints and experimental parameters from all 620 relevant experiments were compiled and analyzed. We found that the coefficients of variation for SF2 (surviving fraction after 2 Gy irradiation) and for D10 (dose that yields a surviving fraction of 10%) were below 30% for all cell lines, indicating that SF2 and D10 have acceptable inter-assay precision. These data support further analysis of published data on clonogenic assays using SF2 and D10 as survival endpoints, which facilitates robust identification of biological profiles representative of cancer cell sensitivity to photons

Functional Temporomandibular Joint Reconstruction in Costochondral Grafting of Micrognathia

Summary:. Rib bone and costochondral complex grafting has been used to treat micrognathia classified as Pruzansky type III. To acquire more physiological joint movement, we reconstructed a temporomandibular joint with the glenoid fossa in addition to the mandibular ramus. The patient underwent a tracheostomy to correct her airway obstruction at 2 months of age. After that, no further surgical treatments were performed on the micrognathia. When she was 6 years of age and during consultation at our department, micrognathia caused by Goldenhar syndrome was confirmed. A head and neck computed tomography scan showed hypoplasia and deficit of the mandible, severe glossoptosis and airway constriction. Initially, a bilateral mandibular body distraction was performed at 6 years of age, and 15 mm of elongation was obtained. Subsequently, reconstruction of the right ramus and right temporomandibular joint fossa was performed at 8 years of age to achieve extubation. Part of her sixth rib and costochondral complex graft was used for the ramus, and costochondral graft was used for the joint fossa. Some new ideas for temporomandibular joint reconstruction were added. Postoperatively, the open mouth range was increased and improvement of the airway space narrowing was observed in a computed tomography scan. The main points of this new method are prevention of ankylosis, skull cortex thinning, and reconstructed ramus’ dislocation. This method may become an effective new treatment for cases of micrognathia with a ramus classified as Pruzansky type III