Arg60 to Leu mutation of the human thromboxane A2 receptor in a dominantly inherited bleeding disorder.

Recent advances in molecular genetics have revealed the mechanisms underlying a variety of inherited human disorders. Among them, mutations in G protein-coupled receptors have clearly demonstrated two types of abnormalities, namely loss of function and constitutive activation of the receptors. Thromboxane A2 (TXA2) receptor is a member of the family of G protein-coupled receptors and performs an essential role in hemostasis by interacting with TXA2 to induce platelet aggregation. Here we identify a single amino acid substitution (Arg60-->Leu) in the first cytoplasmic loop of the TXA2 receptor in a dominantly inherited bleeding disorder characterized by defective platelet response to TXA2. This mutation was found exclusively in affected members of two unrelated families with the disorder. The mutant receptor expressed in Chinese hamster ovary cells showed decreased agonist-induced second messenger formation despite its normal ligand binding affinities. These results suggest that the Arg60 to Leu mutation is responsible for the disorder. Moreover, dominant inheritance of the disorder suggests the possibility that the mutation produces a dominant negative TXA2 receptor.     

Characterization of adenosine deaminase (ADA)-negative B-lymphoblastoid cells cocultured with ADA-positive fibroblasts

Abstract: A cell line, BAD05, derived from B lymphocytes of an adenosine deaminase (ADA; EC 3,5,4,4)-deficient patient could not proliferate in a serum-free medium containing 100 μmol/l deoxyadenosine. When BAD05 was cultured with ADA-positive fibroblasts, the proliferation of BAD05 was improved. BAD05 cell density increased when the initially mixed ratio of fibroblasts/BAD05 was 1/10 or higher, but decreased when the ratio was 1/20 or lower. Deoxyadenosine concentrations in the medium and ATP and deoxyATP (dATP) levels in the BAD05 were measured after 4 hours of coculture at initial BAD05 cell densities of 1 × 10⁵and 1 × 10⁶cells/ml. Deoxyadenosine concentrations in the medium decreased as the density of fibroblasts increased. The dATP level decreased as the mixed ratio rose. The ratio of fibroblasts/BAD05 rather than the cell density of fibroblasts had a larger effect on the dATP levels in BAD05. Under our experimental conditions, ADA-negative cells proliferated well when the ratio of ADA-positive cells/ADA-negative cells was over 1/10.     

Disorder in reciprocal innervation upon initiation of voluntary movement in patients with Parkinson's disease

Summary Reciprocal innervation of the soleus motoneurones upon initiation of voluntary ankle dorsiflexion was investigated in eight patients with Parkinson's disease. H-reflex and visually guided step tracking methods were used for testing moto-neurone excitability and for controlling the timing of movement initiation, respectively. While reciprocal inhibition appeared almost simultaneously with the agonist electromyographic (EMG) onset in normal subjects (Kagamihara and Tanaka 1985), facilitation appeared in the majority of patients under the same onset condition. It increased slowly, reaching a maximum at about 100 ms after the EMG onset. It then subsided slowly at around 200–300 ms, and was replaced thereafter by an inhibitory effect. No coactivation of the soleus muscle was detected electromyographically. The facilitation between the EMG onset and the onset of mechanical contraction was attributed to the direct effect of the descending command from the brain, suggesting a certain disorder in controlling the system for reciprocal innervation.     

Treatment results of radiotherapy to both the prostate and metastatic sites in patients with bone metastatic prostate cancer

Although systemic therapy is the standard treatment for metastatic prostate cancer, a randomized controlled trial showed radiotherapy to the prostate improved overall survival of metastatic prostate cancer patients with the low metastatic burden. Additionally, a randomized phase II trial showed that metastasis-directed therapy for oligo-recurrent prostate cancer improved androgen-deprivation therapy (ADT)-free survival. Therefore, administering radiotherapy to both prostate and metastatic regions might result in better outcomes. Thus, we report the treatment results of radiotherapy to both prostate and metastatic regions. Our institutional database was searched for patients who received radiotherapy to the prostate and metastatic regions. We summarized patient characteristics and treatment efficacy and performed statistical analysis to find possible prognostic factors. A total of 35 patients were included in this study. The median age was 66 years, and the median initial prostate-specific antigen (PSA) level was 32 ng/ml. The Gleason score was 7 in 10 patients, 8 in 13 patients, and 9 in 12 patients. The median radiotherapy dose was 72 Gy to the prostate and 50 Gy to the metastatic bone region. The 8-year overall survival, cause-specific survival, progression-free survival, and freedom from biochemical failure rate were 81, 85, 53, and 57%. Among the 35 patients, 12 were disease-free even after ADT was discontinued. In selected patients with metastatic prostate cancer, ADT and radiotherapy to the prostate and metastatic sites were effective. Patients with good response to ADT may benefit from radiotherapy to both prostate and metastatic regions.     

Melatonin binding sites in estrogen receptor-positive cells derived from human endometrial cancer

Our previous work showed that melatonin (N-acetyl-5-methoxytryptamine) inhibits proliferation of the human endometrial cancer cell line, Ishikawa, which is estrogen receptor-positive. The aim of the present study was to determine whether Ishikawa cells possess membrane melatonin receptors. Binding of the radioligand 2-[125I]-iodomelatonin to membrane preparations obtained from Ishikawa cells was detectable, saturable and stable. Scatchard analysis revealed that the dissociation constant (Kd) of the binding sites was 179.0 pm (similar to that of the MT2 [Mel1b] melatonin receptor subtype), and that the concentration (Bmax) of the binding sites was 12.9 fmol/mg protein. Luzindole, a selective MT2 melatonin receptor antagonist, significantly suppressed binding of 2-[125I]-iodomelatonin at all concentrations tested (10(-8) to 10(-4) m). These results suggest that the MT2 melatonin receptor subtype is present in the membranes of Ishikawa cells, and that the antiproliferative effect of melatonin on Ishikawa cells is mediated via the MT2 receptor. This may have implications for the use of melatonin in endometrial cancer therapy.     

Effect of dobutamine on the arterial ketone body ratio and portal blood flow velocity in cirrhosis

ABSTRACT— We studied the relationship between the portal blood flow velocity and the arterial ketone body ratio in patients with chronic liver disease receiving a dobutamine infusion. We used an ultrasonic Doppler duplex system to evaluate the portal blood flow velocity. Dobutamine was given intravenously at 5 μg/kg/min for 20 min. Dobutamine infusion induced smaller changes in the portal blood flow velocity and ketone body ratio in liver cirrhosis than in chronic hepatitis. The existence of shunts and the poor increase of the cardiac index in response to dobutamine explained the limited improvement of portal blood flow velocity in cirrhosis patients. The ketone body ratio was improved by dobutamine in cirrhosis patients whose portal blood flow velocity was increased by more than 10%, while this ratio decreased when the increase of it was less than 10%. There was no change in portal oxygen extraction in the cirrhosis group, and portal oxygen uptake only increased when the portal blood flow velocity rose by more than 10%. Dobutamine should only be used to treat liver failure if the portal blood flow velocity is increased by more than 10% or the arterial ketone body ratio is improved by a test infusion.