Efficacy of SMTP-7, a small-molecule anti-inflammatory thrombolytic, in embolic stroke in monkeys.

SMTP-7 (Stachybotrys microspora triprenyl phenol-7) is a small molecule that promotes thrombolysis and suppresses inflammation possibly through plasminogen modulation and soluble epoxide hydrolase (sEH) inhibition, respectively. Here, we demonstrate an efficacy of SMTP-7 in a severe embolic stroke model in monkeys. The middle cerebral artery was embolized by an autologous blood clot. Saline, SMTP-7, or tissue-type plasminogen activator (t-PA) (n = 5 in each group) was given after 3 hours, and neurologic deficit scoring and infarct characterization were performed after 24 hours. Hemorrhagic infarct-accompanied premature death was observed for two animals in t-PA group. SMTP-7 treatment significantly reduced the sizes of infarct by 65%, edema by 37%, and clot by 55% compared to saline treatment. Plasma levels of the products of plasminogen activation (plasmin-α2-antiplasmin complex) and sEH reaction (dihydroxyeicosatrienoic acid) in SMTP-7 group were 794% (P < 0.05) and 60% (P = 0.085) compared to saline group, respectively. No significant changes in the plasma levels of MMP-9, CRP, MCP-1, and S100B were found. There was an inverse correlation between plasmin-α2-antiplasmin complex level and infarct volume (r = 0.93, P < 0.05), suggesting a role of thrombolysis in the SMTP-7 action to limit infarct development. In conclusion, SMTP-7 is effective in treating severe embolic stroke in monkeys under conditions where t-PA treatment tends to cause hemorrhagic infarct-associated premature death.

Diurnal Variation of Melatonin Concentration in the Cerebrospinal Fluid of Unanesthetized Microminipig.

BACKGROUND/AIM: The aim of this study was to develop a method for sequentially collecting cerebrospinal fluid (CSF) from an unanesthetized microminipig, which shares many physiological and anatomical similarities with humans, such as diurnality, and investigate the diurnal variation of melatonin concentration in the CSF. MATERIALS AND METHODS: A catheter was placed percutaneously into the subarachnoid space of an anesthetized animal, and the tip of the catheter was placed into the cisterna magna under X-ray. We then sequentially collected CSF at light-on and -off times from the unanesthetized animal for several weeks. After catheter placement, a period of one week or more was necessary to relieve the contamination of RBCs in the CSF. RESULTS: A higher melatonin level in the CSF was noted during lights-off time, and the level was higher than that in the serum. CONCLUSION: This model of sequential collection of CSF will contribute to research in brain functions.

Biogenetically inspired total syntheses of Lycopodium alkaloids, (+)-flabellidine and (-)-lycodine.

The first asymmetric total synthesis of (+)-flabellidine (2) and the shortest total synthesis of (-)-lycodine (3) were accomplished by a strategy featuring the one-pot construction of a tetracyclic lycodine skeleton from a linear precursor, which was inspired by the biosynthetic consideration of Lycopodium alkaloids.

Effect of estrogen on bone resorption and inflammation in the temporomandibular joint cellular elements.

Several epidemiological studies have reported that temporomandibular disorder is more prevalent in women, which suggests the involvement of sex hormones, such as estrogen, in the pathogenesis of this disease. PCR amplification and Western blotting were employed to target the expression of estrogen receptors (ERs) in human fibroblast-like synovial and ATDC5 cells. The effect of estrogen was investigated through the expression of RANKL, osteoprotegerin (OPG), M-CSF/CSF-1 and c-fms. We showed expression of M-CSF/ CSF-1 and c-fms, with time-dependent increase in both after the addition of estrogen. Based on previous studies reporting that M-CSF/CSF-1 regulates the proliferation and differentiation of hemopoietic progenitor cells into mature macrophages, we put forward a new hypothesis based on the increased inflammation and tendency of females to suffer more from temporomandibular disorder (TMD) in the presence of external exacerbating factors. Detection of RANKL and OPG in ATDC5 and expression of both in HFLS was confirmed with complete disappearance of the RANKL band, and marked increase in the expression of OPG after 1 h from the addition of estrogen.

Ginsenoside Rb1 reduces neurodegeneration in the peri-infarct area of a thromboembolic stroke model in non-human primates.

Ginsenoside Rb1 (GRb1), a major component of the traditional herb ginseng, has been reported to show a neuroprotective effect in a rodent ischemic model. The purpose of this study was to investigate effects of GRb1 on early and delayed brain injuries in a non-human primate thromboembolic stroke model. Thromboembolic stroke was induced by occlusion of the middle cerebral artery by injection of an autologous blood clot into the left internal carotid artery. GRb1 (300 microg/kg per day, i.v.) and vehicle were administered from 7 days before embolization to the day following embolization (total: 8 times). Neurological deficits were observed at 1, 6, and 24 h and at 2, 4, and 7 days after embolization. At 7 days after embolization, neuron damage in the peri-infarct area and core region were assessed by NeuN, TUNEL, and GFAP staining. GRb1 improved the skeletal muscle coordination score of the neurologic deficits (median: GRb1 vs vehicle = 10 vs 12, P<0.05). In the GRb1 group, positive neurons expressed by NeuN staining were noted in the ischemic peri-infarct area, and TUNEL- and GFAP-positive cells significantly decreased, when compared with vehicle. These results demonstrated that GRb1 ameliorated both early and delayed injuries in the thromboembolic stroke model in non-human primates.

Bovine lactoferrin potently inhibits liver mitochondrial 8-OHdG levels and retrieves hepatic OGG1 activities in Long-Evans Cinnamon rats.

BACKGROUND/AIMS: To assess the effect of lactoferrin on oxidative liver damage and its mechanism, we used Long-Evans Cinnamon (LEC) rats that spontaneously develop fulminant-like hepatitis and lethal hepatic failure. METHODS: Four-week-old female LEC rats were divided into the untreated and treated groups. The latter was fed bovine lactoferrin at 2% mixed with conventional diet. RESULTS: The cumulative survival rates were 75.0% vs. 100% at 14 weeks, 37.5% vs. 91.7% at 15 weeks, and 12.5% vs. 91.7% at 16 weeks, respectively, for untreated and treated rats (P=0.0008). The 8-OHdG levels in liver mitochondrial DNA and malondialdehyde in plasma and liver tissues were significantly lower in treated than untreated rats (P<0.001, =0.017 and 0.034, respectively). Mitochondrial DNA mutations were more common in untreated rats. OGG1 mRNA and protein expression levels were significantly lower in untreated than treated rats (P=0.003 and 0.007, respectively). Hypermethylation of the second CpG island located upstream of OGG1 gene was observed in untreated rats. CONCLUSIONS: Our findings indicated that lactoferrin inhibits oxidative liver damage in LEC rats. Lactoferrin could be potentially useful for the treatment of oxidative stress-induced liver diseases.

Early apoptosis and cell death induced by ATX-S10Na (II)-mediated photodynamic therapy are Bax- and p53-dependent in human colon cancer cells.

AIM: To investigate the roles of Bax and p53 proteins in photosensitivity of human colon cancer cells by using lysosome-localizing photosensitizer, ATX-S10Na (II). METHODS: HCT116 human colon cancer cells and Bax-null or p53-null isogenic derivatives were irradiated with a diode laser. Early apoptosis and cell death in response to photodynamic therapy were determined by MTT assays, annexin V assays, transmission electron microscopy assays, caspase assays and western blotting. RESULTS: Induction of early apoptosis and cell death was Bax- and p53-dependent. Bax and p53 were required for caspase-dependent apoptosis. The levels of anti-apoptotic Bcl-2 family proteins, Bcl-2 and Bcl-x(L), were decreased in Bax- and p53-independent manner. CONCLUSION: Our results indicate that early apoptosis and cell death of human colon cancer cells induced by photodynamic therapy with lysosome-localizing photosensitizer ATX-S10Na (II) are mediated by p53-Bax network and low levels of Bcl-2 and Bcl-x(L) proteins. Our results might help in formulating new therapeutic approaches in photodynamic therapy.

Detection of sentinel nodes by a novel red-fluorescent dye, ATX-S10Na (II), in an orthotopic xenograft rat model of human gastric carcinoma.

BACKGROUND AND OBJECTIVE: We developed a new imaging system to detect sentinel nodes (SNs) using a novel fluorescent tracer, ATX-S10Na(II), and investigated its usefulness in an animal model. STUDY DESIGN/MATERIALS AND METHODS: Human gastric carcinoma cells were implanted orthotopically into nude rats. ATX-S10Na(II) was injected subserosally into the primary tumor lesion, and visualized by a fluorescence spectro-laparoscope. Presence of tumor cells in lymph nodes (LNs) was determined by RT-PCR specific for human beta-actin. RESULTS: Injection of ATX-S10Na(II) was successful in 27 tumor-bearing rats. A red fluorescence was incorporated into the left gastric and hepatic LNs in 25 and 2 rats, respectively. Of note, human beta-actin was detected in most of these LNs. Fluorescence was not detected in LNs that did not contain cancer. CONCLUSION: ATX-S10Na(II) is useful for the detection of cancer-containing SNs in an animal model of gastric carcinoma, and may serve as a novel tracer in SN navigation surgery.

Effects of intra-arterial urokinase on a non-human primate thromboembolic stroke model.

One of the most important prognostic factors in the thrombolytic treatment of acute ischemic stroke is to re-canalize. The purpose of this study was to evaluate the effectiveness and safety of urokinase in a primate thromboembolic stroke model. Thromboembolic stroke was accomplished via occlusion of the middle cerebral artery (MCA) obtained by injecting an autologous blood clot into the left internal carotid artery in 21 male cynomolgus monkeys. Animals were randomly assigned to the following treatment groups: Group 1: vehicle (saline), Group 2: urokinase (40,000 IU), Group 3: urokinase (120,000 IU,) over 2 or 6 h via intra-internal carotid catheter starting 1 h after embolization, respectively. In the urokinase-treated groups, neurologic deficits were improved in consciousness and skeletal muscle coordination, but not sensory and motor systems. The infarction size in Group 2 (11.9 +/- 3.9% of the hemisphere) and 3 (7.6 +/- 2.5%) were significantly smaller than that (24.7 +/- 3.5%) in Group 1. However, 2 of 5 animals in Group 3 died. In conclusion, urokinase improved neurologic deficits and reduced cerebral infarction on thromboembolic stroke in the cynomolgus monkey.

Effects of hypothermia and hyperthermia on attentional and spatial learning deficits following neonatal hypoxia-ischemic insult in rats.

We previously reported that rats exposed to neonatal hypoxic-ischemic (HI) insult showed selective and long-lasting learning and memory impairments in the plus maze, 8-arm radial maze, choice reaction time (CRT) task, and water maze, and that they showed severe brain injury to areas such as parietal cortex, hippocampus, striatum and thalamus. In this study, we examined the effects of hypothermia and hyperthermia on learning and memory deficits following neonatal HI insult. Seven-day-old Wistar rats were subjected to left carotid artery ligation followed by 2 h of hypoxia (8% O2/92% N2) under three different temperature conditions: 27 degrees C (hypothermia), 33 degrees C (normothermia) and 37 degrees C (hyperthermia) in temperature-controlled chambers. Hypothermia significantly reduced attentional deficits in the CRT task and spatial learning deficits in the water maze, and protected against severe brain injury in comparison with the control temperature. On the other hand, hyperthermia aggravated the behavioral deficits and brain injury. These outcomes clearly show that temperature regulation during HI insult plays an important role in the induction of behavioral and histological changes following neonatal HI insult in rats.

Can arthrocentesis release intracapsular adhesions? Arthroscopic findings before and after irrigation under sufficient hydraulic pressure.

PURPOSE: To investigate the effect of release of intra-articular adhesions of arthrocentesis, we examined patients with closed lock of the temporomandibular joint by arthroscopy before and after irrigation. PATIENTS AND METHODS: In 6 closed lock cases in which adhesions in the upper joint space were observed by arthroscopy before arthrocentesis, arthroscopic examination was performed again to confirm whether the adhesion was released after the procedure. Range of mouth opening and joint pain were examined to determine the clinical efficiency of the procedure. RESULTS: Adhesion was not released by irrigation under low pressure but could be released by irrigation under high pressure. After irrigation under low and high pressure, the maximum mouth opening of the patients improved from 0 to 1 mm (average, 0.3 mm) and 2 to 6 mm (average, 3.7 mm), respectively. CONCLUSIONS: The results indicate that arthrocentesis with sufficient pressure could be effective for closed lock cases with adhesions in the upper joint compartment.

The pharmacological characterization of attentional processes using a two-lever choice reaction time task in rats.

Activating the noradrenergic and cholinergic systems is known to enhance attentional processes, while stimulating dopaminergic, serotonergic, and GABAergic systems suppresses them. The objective of the present study was to investigate the pharmacological characterization in the attentional processes of a two-lever choice reaction time (CRT) task using different centrally acting drugs. We designed seven parameters in this task: the correct response (CR) rate; error response rate; nonresponse (NR) rate; differential reinforcement of other behavior (DRO) responses; number of incorrect lever pressings during both the intertrial interval and DRO periods; the mean CRT of CR; and activity during 30 trials. The compounds produced different profiles at each dose. 1) Facilitative and disruptive effects on attentional processes occurred with changes in CRT alone. Scopolamine (0.1 mg/kg) and prazosin (0.3-1 mg/kg) prolonged the CRT, whereas methamphetamine (0.3 mg/kg) shortened the CRT. 2) Attentional deficits occurred with abnormal behavior showing premature response or perseverative behavior. Scopolamine (0.2-1 mg/kg), methamphetamine (3 mg/kg), delta(9)-tetrahydrocannabinol (10 mg/kg), and MK-801 (0.1-0.3 mg/kg) produced a marked increase in the number of total lever pressings. 3) Motor function deficits rather than attentional deficits occurred. 8-OH DPAT (1 mg/kg) and muscimol (1 mg/kg) produced a decrease in CR and an increase in NR with a marked decrease in activity and prolonged the CRT. Activating noradrenergic alpha(1) receptors was found to enhance the attentional processes, while blocking muscarinic receptors, alpha(1) receptors, and NMDA receptors, and stimulating cannabinoid receptors and the dopaminergic systems impaired the attentional processes in the two-lever CRT task.

Dexamethasone prevents long-lasting learning impairment following neonatal hypoxic-ischemic brain insult in rats.

We examined for 18 weeks the effect of dexamethasone treatment on learning and memory impairment produced by hypoxic-ischemic stress at postnatal day 7 in rat in addition to brain histological study. Dexamethasone of 0.5 mg/kg was injected i.p. 4 h before hypoxic-ischemic stress, in which the left carotid artery was ligated followed by 2 h hypoxia (8% oxygen). Dexamethasone treatment improved behavior in each learning task: in choice reaction time tasks relating to the attention process, in 8-arm radial maze task examining working and reference memory, and in water maze task relating to reference memory. Improvement to the extent of the sham-control level was observed. Dexamethasone treatment also completely prevented histological brain damage. No adverse effect in learning and memory tests was observed in the animals treated with dexamethasone without hypoxic-ischemic stress. It is concluded that dexamethasone treatment is significantly effective in prevention not only of histological brain damage but also of learning and memory impairment occasioned by subsequent hypoxic-ischemic insult, warranting further clinical investigation.

Development of a novel experimental rat model for neonatal pre-ganglionic upper brachial plexus injury.

A neonatal upper brachial plexus injury, referred to as Erb's palsy, is a serious obstetric problem. Some surgical methods are used to treat this injury, but they are inadequate. To seek new treatments for Erb's palsy, we used a model for cervical preganglionic root transection in neonate rats and evaluated the behavioral and histological compatibility of this model with Erb's palsy. Two groups were used in this study. In the group, receiving the Erb operation, the left anterior and posterior roots of spinal vertebra C5-C7 were transected at the preganglionic level, and the results were compared with those of a group that received a sham operation. In the group, receiving the Erb operation, walking difficulties and behavioral abnormalities were observed. These observations were noted on the side where the transection took place, and the problems were attributed to proximal muscle weakness in the forelimb. Additionally, the forepaw grip was not impaired. Furthermore, in this group, the number of anterior horn cells in the cervical cord on the transected side was significantly lower than that on the contralateral side (P < 0.001). The results of this study indicate that the model fulfills the criteria for the clinical symptoms of Erb's palsy and that it may also serve as a new method for enabling treatment of the condition.