Protection of ischemic hippocampal neurons by ginsenoside Rb1, a main ingredient of ginseng root.

Our previous study showed that the oral administration of red ginseng powder before but not after transient forebrain ischemia prevented delayed neuronal death in gerbils, and that a neuroprotective molecule within red ginseng powder was ginsenoside Rb1. However, it remains to be clarified whether or not ginsenoside Rb1 acts directly on the ischemic brain, and the mechanism by which ginsenoside Rb1 protects the ischemic CA1 neurons is not determined. Without elucidation of the pharmacological property of ginsenoside Rb1, the drug would not be accepted as a neuroprotective agent. The present study demonstrated that the intracerebroventricular infusion of ginsenoside Rb1 after 3.5 min or 3 min forebrain ischemia, precluded significantly the ischemia-induced shortening of response latency in a step-down passive avoidance task and rescued a significant number of hippocampal CA1 neurons from lethal ischemic damage. The intracerebroventricular infusion of ginsenoside Rb1 did not affect hippocampal blood flow or hippocampal temperature except that it caused a slight increase in hippocampal blood flow at 5 min after transient forebrain ischemia. Furthermore, ginsenoside Rb1 at concentrations of 0.1-100 fg/ml (0.09-90 fM) rescued hippocampal neurons from lethal damage caused by the hydroxyl radical-promoting agent FeSO4 in vitro, and the Fenton reaction system containing p-nitrosodimethylaniline confirmed the hydroxyl radical-scavenging activity of ginsenoside Rb1. These findings suggest that the central infusion of ginsenoside Rb1 after forebrain ischemia protects hippocampal CA1 neurons against lethal ischemic damage possibly by scavenging free radicals which are overproduced in situ after brain ischemia and reperfusion. The present study may validate the empirical usage of ginseng root over thousands of years for the prevention of cerebrovascular diseases.

Suppression by platelet factor 4 of the myogenic activity of basic fibroblast growth factor.

The effect of platelet factor 4 (PF4) on myoblast cultures with or without basic fibroblast growth factor (bFGF) or other growth factors was investigated in the present in vitro experiments, with reference to bFGF binding to myoblast membrane fraction. When PF4 was added to the culture medium 1 day after myoblast cultivation, the nuclei of both myoblasts and myotubes were markedly reduced in number in a dose-dependent manner, whereas the inhibitory effect of PF4 on myoblast development was not observed when PF4 was added to the culture medium 3, 7, or 14 days after myoblast cultivation. In contrast, bFGF significantly increased the numbers of myoblast and myotube nuclei. When bFGF and PF4 were simultaneously added to the culture medium, PF4 abolished the facilitatory effects of bFGF on myogenesis. The real-time biospecific interaction analysis (BLA) core system showed that the myoblast membrane fraction at 1 day after cultivation contains bFGF-binding elements which are blocked by PF4 in a dose-dependent manner. Moreover, [126I]-bFGF binding experiments indicated the existence of both high and low affinity binding sites on myoblast membranes, although the high affinity binding sites decreased in number and the dissociation constant increased in value as the culture period was prolonged. Among the six other growth factors examined, acidic fibroblast growth factor and platelet-derived growth factor-BB stimulated myogenesis, and their effects were blocked by PF4 treatment. These findings suggest that: 1) PF4 inhibits myoblast proliferation and myotube formation only for a limited initial period of cultivation, possibly because of the time-dependent down-regulation of high affinity bFGF receptors: and 2) PF4 may be used as a tool to investigate the function of endogenous heparin-binding growth factors upregulated transiently at a certain developmental stage or in case of tissue damage and repair, even though it is not monospecific to bFGF.

Expression of basic fibroblast growth factor-like immunoreactivity in the nuclei of regenerating hepatocytes.

This study, utilizing rats subjected to two-thirds partial hepatectomy or sham operation, was designed (1) to investigate the content of basic fibroblast growth factor (bFGF) in the subcellular fractions of regenerating and sham-operated rat livers by immunoblot experiments and enzyme-linked immunosorbent assay (ELISA), (2) to show that bFGF immunoreactivity and proliferating cell nuclear antigen (PCNA) immunoreactivity are markers for hepatocellular mitosis before and after partial hepatectomy, and (3) to observe the location and fine structure of the bFGF immunoreaction within the regenerating liver with special attention to bFGF immunoreactivity in the nuclei of regenerating hepatocytes. Immunoblot experiments and ELISA showed a transient increase in high-molecular-weight forms of bFGF in the nuclear subcellular fraction of regenerating liver 48 h after partial hepatectomy. By light microscopy, bFGF and PCNA immunoreactivities were detected in the nuclei of regenerating hepatocytes. Electron microscopy demonstrated bFGF-like immunoreactivity mainly in the nuclear euchromatin and rarely in the heterochromatin or nucleoli of regenerating hepatocytes. The transient increase in high-molecular-weight forms of bFGF in the nuclear euchromatin of regenerating hepatocytes, together with the concomitant expression of PCNA in the regenerating liver, suggests an important role of the high-molecular-weight forms of bFGF in hepatocyte proliferation and/or mitosis, although authentic bFGF with a molecular form of 18 kDa is not considered to be involved in hepatic regeneration.

Perikaryal projections of developing spinal ganglion neurons in the chick demonstrated by scanning electron microscopy.

The perikaryal projections of sensory ganglion neurons in chick embryos were observed by scanning electron microscopy after removal of the connective tissues and satellite cells by enzymatic digestion treatment. The perikaryal projections were seen not only on the surface of the perikarya but also on the surface of the stem processes. The projections were up to 3 microm in length, and their transverse diameters ranged between 0.12 and 0.24 microm from incubation (embryonic) day 10 to posthatching day 2. On days 6 and 8 of incubation, thicker projections with transverse diameters of 0.24-0.9 microm were observed transiently in addition to those described above, and some of them looked like vestiges of neuronal processes during development. The thin projections emerging mainly in the later developmental stages increased in number as spindle-shaped bipolar neurons differentiated into (pseudo)unipolar cells. Morphometric analysis revealed that the density of perikaryal projections correlated well with the shape and size of each neuron; thin perikaryal projections were more numerous on those of mature pseudounipolar neurons than on the surface of premature ganglion neurons, and they increased in number as the individual ganglion cell bodies grew larger. The neuronal shape- and size-dependent increase in perikaryal projections during development may support the hypothesis that perikaryal projections are structural devices for increasing neuronal surface areas and possibly the efficiency of metabolic activities.

Protective effect of basic fibroblast growth factor-heparin and neurotoxic effect of platelet factor 4 on ischemic neuronal loss and learning disability in gerbils.

Platelet factor 4, which has a potent affinity for heparin, has been shown to inhibit the binding of basic fibroblast growth factor to the cell surface receptor and to counteract the biological activities of basic fibroblast growth factor in certain peripheral tissues. In the present in vitro [125I]basic fibroblast growth factor binding experiments, platelet factor 4 consistently inhibited the binding of iodinated basic fibroblast growth factor to cell membranes of the gerbil hippocampus. To investigate the in vivo function of endogenous basic fibroblast growth factor and/or basic fibroblast growth factor receptor possibly activated in the ischemic gerbil brain, we infused platelet factor 4 continuously into the left lateral ventricle with an osmotic minipump. When platelet factor 4 infusion was started within three days after a 3-min ischemic insult, it significantly enhanced ischemia-induced learning disability and ischemic neuronal loss in the CA1 region of the hippocampus, as demonstrated by the results of the step-down passive avoidance task and by subsequent histological examinations. Infusion of platelet factor 4 into the cerebral ventricle of intact gerbils did not affect learning ability or CA1 neuron number. Basic fibroblast growth factor-neutralizing antibody, when infused continuously in the cerebral ventricle, also exhibited a neurotoxic effect in ischemic but not intact gerbils. Basic fibroblast growth factor co-infused with heparin, but not basic fibroblast growth factor alone, rescued a significant number of ischemic neurons which were destined to degenerate without the infusion of heparinized basic fibroblast growth factor, and it prevented ischemia-induced learning disability.(ABSTRACT TRUNCATED AT 250 WORDS)

[Lymphangial infusion of carboplatin and iodized-oil emulsion: phase I trial].

We examined lymphography as an interventional radiological technique for suppressing microscopic metastasis to the pelvic and paraaortic lymph nodes. There were no reports on this method in our survey of the literature. We performed a dose-escalation study as a phase I trial to determine the maximum dose that could be given without intolerable complications. From September 1991 to April 1992, carboplatin and iodized-oil emulsion was injected into both feet of 10 patients by the Kinmonth method. In the first 5 patients 5 mg of carboplatin was injected into each foot, and 10 mg was injected in the next 5 patients. When the injection of 15 mg was attempted, the injection could not be completed because carboplatin powder was deposited in the syringe. The amount of carboplatin was limited by the instability of the carboplatin-lipiodol emulsion at 15 mg in the present study. There were not intolerable complications. In one case in which 10 mg was injected into each foot, the average platinum concentration in resected pelvic lymph nodes was 0.83 microgram/gWet (maximum: 3.51 micrograms/gWet) even a week after treatment. Serum platinum was undetectable (< 50 ng/ml). These results suggest that a high concentration of carboplatin can be preserved for a long time by this novel interventional technique.

Relation between preoperative CT findings in rectal cancer and local recurrence rate. A pilot study.

To identify predictive factors of local recurrence possible to use as criteria for preoperative radiotherapy, we reviewed the CT scans of 51 patients undergoing curative resection for rectal cancer. Seven patients developed local recurrence. The presence of the CT images of spicular structures or a fibrous soft tissue layer around the rectum was related to extrarectal spread with a positive predictive value of 88% (30/34) when compared with the pathology. Six out of 34 cases with these CT findings developed recurrence, compared to only one out of 17 cases without such findings. The recurrence rate was especially high, (4/14), in patients where the abnormal tissue as judged by CT was attached to the perirectal fascia or extended beyond it. CT may be a useful tool for predicting local recurrence by using the perirectal fascia as a diagnostic marker.

Molecular forms of islet amyloid polypeptide (IAPP/amylin) in four mammals.

Using reverse-phase high performance liquid chromatography combined with radioimmunoassays for human and rat/mouse islet amyloid polypeptide (IAPP), we identified molecular forms of IAPPs in pancreata of four mammals including species in which islet amyloid deposition occurs (human and cat) and those in which amyloid deposition does not occur (rat and mouse). In human pancreas, IAPP (1-37) was the major molecular form, and IAPP (17-37), IAPP (24-37) and four IAPP-immunoreactive peptides were detected as minor components. In rat, mouse and cat pancreata, IAPP (1-37) and IAPP (19-37) were identified with the latter being the major molecular form. Major processing takes place at a single arginine residue at position 18 of rat/mouse and cat IAPPs, but not at the histidine at position 18 of human IAPP, indicating that arginine could yield different processing of IAPP between the 3 species and human. Different processing of IAPP by species suggests that processing of IAPP in pancreas is not responsible for islet amyloid formation. Identification of molecular forms of IAPP is helpful in elucidating the physiological function of the IAPP molecule and in determining the type of system regulating biosynthesis and catabolism of the peptide.

Plasma islet amyloid polypeptide levels in obesity, impaired glucose tolerance and non-insulin-dependent diabetes mellitus.

We examined the response of plasma islet amyloid polypeptide (IAPP) to an oral glucose load in non-obese and obese subjects with normal glucose tolerance or impaired glucose tolerance (IGT), and in non-obese patients with non-insulin-dependent diabetes mellitus (NIDDM). Plasma IAPP response to intravenous glucagon injection in NIDDM patients was also studied. Plasma IAPP concentration was determined by a sensitive and specific radioimmunoassay. Basal levels of plasma IAPP in non-obese subjects with normal glucose tolerance, IGT and NIDDM were not significantly different from each other. Non-obese subjects with IGT showed delayed and higher plasma IAPP response to oral glucose load compared to normal non-obese subjects. In NIDDM patients, IAPP response to glucose was delayed and lower when compared to normal non-obese subjects. Basal levels of plasma IAPP in normal obese subjects and obese subjects with IGT were significantly higher than those in normal non-obese subjects. Plasma IAPP response to glucose load in these obese subjects was higher than that in normal non-obese subjects. Plasma IAPP response was decreased in diabetic patients treated with diet, oral hypoglycemic agents and insulin in that order. We conclude that the secretion of IAPP is reduced with progression of NIDDM, although it appears to be rather augmented in IGT compared to normal non-obese subjects.

Identification and characterization of islet amyloid polypeptide in mammalian gastrointestinal tract.

We identified and determined the content and molecular form of islet amyloid polypeptide (IAPP/amylin) in the gastrointestinal (GI) tract of human, rat, mouse and cat. IAPP was isolated by anti- IAPP- IgG immunoaffinity chromatography and reverse-phase high performance liquid chromatography coupled with radioimmunoassays for human and rat/mouse IAPPs. Human IAPP [1-37], [17-37] and [24-37] were identified in human stomach with IAPP [1-37] being the major molecular form. In the GI tract of rat, mouse and cat, IAPP [1-37] and IAPP [19-37] were identified with the latter being the major molecular form. IAPP is present from stomach to colon with the highest concentration being observed in pyloric antrum of stomach. IAPP content in rat antrum fell to 69% of control after 4 days of fasting, with the molar ratio of IAPP [19-37] to IAPP [1-37] increasing from 1.4 in controls to 2.9 in fasted rats. Identification of IAPP and characteristic morphology of IAPP- cells in the GI tract indicate a possible biological function of IAPP as a gastrointestinal peptide.