UVB-induced calmodulin increase in pig epidermis: Analysis of the effect of the calmodulin antagonist,W-13

Calmodulin (CaM) is a multifunctional calcium-binding protein that has been implicated in the control of cell proliferation. In order to determine the role of CaM in keratinocyte proliferation, we investigated the effect of the CaM antagonist, W-13 on thymidine incorporation into pig epidermis. W-13 significantly inhibited thymidine incorporation into pig epidermis, while W-12, a closely related compound with much less anti-CaM activity, had little effect. The effect of W-13 was detected after as little as 2 h of incubation. Using a short-term (2-h) incubation system, the effects of other chemicals affecting various transmembrane signalling systems of keratinocytes were also investigated. None of these chemicals (epinephrine, histamine, forskolin, HA-1004, bradykinin, mezerein, phorbol 12-myristate, 13-acetate, H-7, staurosporin) inhibited thymidine incorporation. The effect of W-13 was reversible; its removal from the incubation medium resulted in the reinitiation of thymidine incorporation. Pig epidermis responded to 2.5 MED UVB irradiation showing an initial (24–48 h after irradiation) decrease and a subsequent (96–120 h after irradiation) increase in thymidine incorporation. The CaM content was not significantly altered during the initial hypoproliferative phase, but was significantly increased during the 72–120 h after UVB irradiation sometimes slightly preceding but mostly coinciding with the increase in thymidine incorporation. The inhibitory effect of W-13 on keratinocyte proliferation was observed at time 0, and at 72 h and 168 h following the UVB irradiation but not at 24–48 h or 96–144 h, i.e. during the hypo- and hyperproliferative phases, respectively. Methotrexate, which inhibits de novo nucleic acid synthesis significantly augmented thymidine incorporation into the UVB-induced hyperproliferative epidermis, but the addition of W-13 together with methotrexate caused no inhibitory effect on thymidine incorporation into UVB-induced hyperproliferative epidermis.Abbreviations CaM calmodulin - W-13 N-(4-aminobutyl)-5-chloro-2-naphthalenesulphonamide hydrochloride - W-12 N-(4-aminobutyl)-2-naphthalenesulphonamide hydrochloride - HA-1004 N-(2-guanidinoethyl)-5-isoquinolinesulphonamide - PMA phorbol 12-myristate, 13-acetate - H-7 1-(5-isoquinoline-sulphonyl)-2-methyl piperazine dihydrochloride - UVB ultraviolet B radiation - MED minimal erythema dose - RB retinoblastoma gene product     

Decoding higher-order motor information from primate non-primary motor cortices.

To investigate the involvement of primate non-primary motor cortices in bimanual sequential movements, we recorded neuronal activity in the supplementary motor area (SMA) and presupplementary motor area (pre-SMA) while an animal was performing bimanual motor tasks that required two sequential arm movements consisting of either pronation or supination of the right or left arms with delay periods. We also recorded electromyograms (EMGs) from the arm while the animal performed the bimanual task to compare muscle and neuronal activity. This paper focuses on the neuronal activity before the onset of sequential movements. We found that the prime-mover forelimb muscles were selectively active when an impending arm movement involved recorded muscles, but was not dependent on whether the arm movements were bimanual or unimanual. In contrast, we found that neurons in the non-primary motor cortices showed different activity depending on whether the forthcoming sequential arm movements were unimanual or bimanual. Our results suggest that neuronal activity in the SMA and pre-SMA reflects higher-order information about arm use before motor execution. By extracting this type of information, we can use it to control prosthetic arms in a more intelligent manner through a brain-machine interface.     

The Influence of a Heparin-Coated Oxygenator During Cardiopulmonary Bypass on Postoperative Lung Oxygenation Capacity in Pediatric Patients with Congenital Heart Anomalies

A bstract Background : Cardiopulmonary bypass (CPB) causes an inflammatory response and remarkably depresses the oxygenation capacity of the lung in pediatric patients with pulmonary hypertension. Although a heparin-coated circuit is more biocompatible than an uncoated circuit, the beneficial effect of a heparin-coated circuit on the postoperative lung function in the pediatric patients remains unknown. Methods : Sixty patients younger than 3-years-old undergoing heart operations for ventricular septal defect were divided into three groups: group I = children (n = 11) without pulmonary hypertension who underwent CPB with an uncoated oxygenator; group II = children (n = 32) with pulmonary hypertension who underwent CPB with an uncoated oxygenator; and group III = children (n = 17) with pulmonary hypertension who underwent CPB with a heparin-coated oxygenator. A respiratory index (RI) was used to assess the oxygenation capacity of the lung. Results : RI in group II was significantly higher than in group I and intubation time in group II was significantly longer than in group I. There was a positive correlation between preoperative pulmonary-systemic blood pressure ratio and RI at 3 hours post-CPB. Three and six hours post-CPB, RI in group III was significantly lower than in group II, but there was no significant difference in RI between both groups at 12 hours post-CPB. Conclusions : Pulmonary hypertensive pediatric patients were vulnerable to postperfusion lung injury. Beneficial effects of a heparin-coated oxygenator in a CPB circuit was limited to the early hours post-CPB and the postoperative clinical course was not modified by the heparin-coating of a membrane oxygenator.     

Rapamycin as an inhibitor of osteogenic differentiation in bone marrow-derived mesenchymal stem cells

Background An autograft of cultured bone marrow-derived mesenchymal stem cells has already been used in clinical practice. In those patients whose bone marrow cannot be used, a cell allograft with the use of immunosuppressant drugs will be an option in the future. However, little is known about the effects of immunosuppressant drugs on mesenchymal stem cells. This study assessed the effects of immunosuppressant drugs on osteogenic differentiation of mesenchymal stem cells and analyzed the manner in which immunosuppressant drugs modulate the osteogenic effect of dexamethasone. Methods Rat bone marrow cells were cultured with or without dexamethasone as an osteogenic supplement. In each experimental group, one of three immunosuppressants (rapamycin, cyclosporine A, or FK506) was added. As a control, cells were cultured without immunosuppressants. Histologically, mineralization was assessed by alizarin red S staining and phase-contrast microscopy. Biochemically, alkaline phosphatase activity, calcium content, and osteocalcin content were assessed. Results On histological analysis, no mineralized nodules were seen on alizarin red S staining or phase-contrast microscopy in the groups not treated with dexamethasone, except in the group that was treated with FK506. Mineralized nodules were seen in the groups treated with dexamethasone, except in the group that was treated with rapamycin. On biochemical analysis, it was found that, compared to the control group, rapamycin reduced alkaline phosphatase activity and the calcium content of mesenchymal stem cells; FK506 increased alkaline phosphatase activity, calcium content, and osteocalcin content; and cyclosporine A had negligible effects. Dexamethasone increased alkaline phosphatase activity, calcium content, and osteocalcin content, but these effects were decreased by rapamycin. Conclusions Rapamycin did not have an osteogenic effect on mesenchymal stem cells, but inhibited the effect of osteogenic differentiation induced by dexamethasone. In contrast, FK506 had an osteogenic effect on mesenchymal stem cells. Therefore, FK506 might be more useful than rapamycin in allogeneic transplantation of mesenchymal stem cells.     

Axonal regeneration of retinal ganglion cells in the cat geniculocortical pathway

The optic nerve of anesthetized cats was completely cut and the autologous sciatic nerve was transplanted. Sixty days later some populations of retinal ganglion cells were shown to regenerate the axon with retrograde HRP labeling. We verified that ganglion cells that had projected to the lateral geniculate nucleus (LGN) were able to regenerate through the transplant with a double-labeling method: diI was injected into the LGN prior to the transplantation, and dextran-fluorescein was injected into the graft after axonal regeneration. Intracellular injection of HRP into regenerating ganglion cells in an in vitro preparation revealed that the two major cell types projecting to the LGN, and β, regenerated axons and showed normal dendritic morphology.     

K1 Antigen, Serotype and Antibiotic Susceptibility of Escherichia coli Isolated from Cerebrospinal Fluid, Blood and Other Specimens from Japanese Infants

K1 antigens, serotypes and antibiotic susceptibilities of Escherichia coli isolates from neonates and infants were investigated. The presence of K1 antigen was tested by the K1-specific phage method. The number of K1 positive strains was 27 (84%) of 32 isolates from cerebrospinal fluid, 11 (25%) of 44 from blood and 4 (22%) of 18 from other specimens. Fourteen (33%) of the K1 positive strains were serotyped as O16:H6, and 8, 7 and 5 were serotyped as O18ac:H7, O1:H7 and O7:H-, respectively. One of 5 of the K1 negative strains were distributed into 30 different combinations of O and H antigens. The ampicillin resistance rates were 19% in K1 positive strains and 45% in K1 negative ones. The incidence of chloramphenicol resistance was the same in K1 positive and negative strains (21%). Ampicillin resistance was not noted in O16: H6 strains, but the incidence of antibiotic resistance was high (65% to ampicillin and 53% to chloramphenicol) in the rough-type strains.     

Creatine kinase release from regenerated muscles after eccentric contractions in rats

The purpose of this study was to test the hypothesis that an increase in plasma creatine kinase (CK) activity after eccentric contractions (ECC) would be attenuated in regenerated muscle fibres. Adult male Wistar rats (aged 12–14 weeks) were randomly assigned to a treatment group (n =14) or a control group (n = 10). In the treatment group, 1.2% barium chloride solution (BaCl₂) was injected into the tibialis anterior (TA) and extensor digitorum longus (EDL) muscles to induce degeneration and subsequent regeneration. The same amount of isotonic saline solution was injected into TA and EDL for the control group. Histological observation showed that approximately 50% of the fibres in the transverse sections of both muscles underwent necrosis 2 days after BaCl₂ injection. The CK activity increased about tenfold at 2–4 h after BaCl₂ injection. At 4 weeks after BaCl₂ injection, when the regeneration process was almost complete, the TA and EDL of anaesthetized rats from both groups were subjected to ECC in which maximal dorsiflexion was caused by nerve electrical stimulation and the flexed foot was forcibly extended by a lever arm connected to a motor. This action was performed in 2 sets of 30 repetitions. Maximal isometric torque of the dorsiflexors decreased to about 15% (P<0.01) of the pre-ECC value immediately after the exercise. Blood samples were collected before and 2, 4, 12, 24, 48 h after ECC. The CK activity increased significantly (P < 0.01) and peaked at 2–4 h after ECC, and there was no significant difference in the amount of CK increase between the treatment [1007 (SEM 120) IU · I^–1] and the control [1064 (SEM 120) IU · 1^–1 group. Contrary to the hypothesis, CK release after ECC was not attenuated in muscle regenerated from BaCl₂-induced myonecrosis.     

Public, Experts, and Acceptance of Advanced Medical Technologies: The Case of Organ Transplant and Gene Therapy in Japan

In 1997, after long social debates, the Japanese government enacted a law on organ transplantation from brain-dead bodies. Since 1993, on gene therapy, administrative agencies have issued a series of guidelines. This study seeks to elucidate when people became aware of the issues and when they formed their opinions on organ transplant and gene therapy. At the same time, it aims to examine at which point in time experts, those in university ethical committees and in academic societies, consider these technologies became accepted among the public. A self-administered questionnaire was sent by mail to a stratified random sampling of 3000 people nationwide in Japan. Another questionnaire was sent both to the member societies of the Japanese Association of Medical Sciences and to the ethical committees of all the medical schools in Japan. Results of the surveys indicated that many of the public remained undecided on the desirability of organ transplant or gene therapy at the time of enactment of official guidelines. A substantial part of them formed their opinions in subsequent periods, especially around the time of first implementation and thereafter. Experts of the academic societies and of the university ethical committees regarded the time of implementation as an important factor in the acceptance of the technologies in society. Since many people formed their opinion during the period of technological implementation, communications efforts to facilitate public understanding of science and technology, as well as to advance practical discussion on policy alternatives in this period can play a key role in determining the fate of technological innovation and ethical debates in medicine.