Tanshinone IIA Regulates Synaptic Plasticity in Mg2+-Free-Induced Epileptic Hippocampal Neurons via the PI3K/Akt Signaling Pathway.

BACKGROUND: Tanshinone IIA (TSIIA) is an element of the effective ingredients of Salvia miltiorrhiza Bunge (Labiatae), exhibits a significant therapeutic effect in brain neuroprotection. The focus of this study was the examination of synaptic plasticity of in Mg2+-free-induced epileptic hippocampus neurons and how TSIIA protects against it. METHODS: The purity of the primary hippocampal neurons extracted from Sprague Dawley rats was assessed within 24 hours by microtubule-associated protein (MAP2) immunofluorescence staining. A hippocampal neuron model for Mg2+-free-induced spontaneous recurrent epileptiform discharge was developed, five experimental groups were then randomized: blank (Blank), model (Model), TSIIA (TSIIA, 20 µM), LY294002 (LY294002, 25 µM), and TSIIA+LY294002 (TSIIA+LY294002, 20 µM+25 µM). FIJI software was used to examine variations of neurite complexity, total length of hippocampal neurons, number of primary dendrites and density of dendritic spines. Developmental regulation brain protein (Drebrin) and brain-derived neurotrophic factor (BDNF) expression was evaluated using immunofluorescence staining and the relative expression of phospho-protein kinase B (p-Akt)/Akt, BDNF, synaptophysin (SYN) and postsynaptic density 95 (PSD-95) determined by Western blot. RESULTS: In contrast to the model group, TSIIA drastically reduced damage to synaptic plasticity of hippocampal neurons caused by epilepsy (p < 0.05). The TSIIA group showed a significant increase in the relative expression of PSD-95, SYN, BDNF, and p-Akt/Akt (p < 0.01). CONCLUSIONS: TSIIA was effective in reducing harm to the synaptic plasticity of hippocampal neurons induced by persistent status epilepticus, with the possible mechanism being regulation of the phosphatidylinositol 3-kinase 56 (PI3K)/Akt signaling pathway.

Investigation of the mechanism of tanshinone IIA to improve cognitive function via synaptic plasticity in epileptic rats.

CONTEXT: Tanshinone IIA is an extract of Salvia miltiorrhiza Bunge (Labiatae) used to treat cardiovascular disorders. It shows potential anticonvulsant and cognition-protective properties. OBJECTIVE: We investigated the mechanism of tanshinone IIA on antiepileptic and cognition-protective effects in the model of epileptic rats. MATERIALS AND METHODS: Lithium chloride (LiCl)-pilocarpine-induced epileptic Wistar rats were randomly assigned to the following groups (n = 12): control (blank), model, sodium valproate (VPA, 189 mg/kg/d, positive control), tanshinone IIA low dose (TS IIA-L, 10 mg/kg/d), medium dose (TS IIA-M, 20 mg/kg/d) and high dose (TS IIA-H, 30 mg/kg/d). Then, epileptic behavioural observations, Morris water maze test, Timm staining, transmission electron microscopy, immunofluorescence staining, western blotting and RT-qPCR were measured. RESULTS: Compared with the model group, tanshinone IIA reduced the frequency and severity of seizures, improved cognitive impairment, and inhibited hippocampal mossy fibre sprouting score (TS IIA-M 1.50 ± 0.22, TS IIA-H 1.17 ± 0.31 vs. model 2.83 ± 0.31), as well as improved the ultrastructural disorder. Tanshinone IIA increased levels of synapse-associated proteins synaptophysin (SYN) and postsynaptic dense substance 95 (PSD-95) (SYN: TS IIA 28.82 ± 2.51, 33.18 ± 2.89, 37.29 ± 1.69 vs. model 20.23 ± 3.96; PSD-95: TS IIA 23.10 ± 0.91, 26.82 ± 1.41, 27.00 ± 0.80 vs. model 18.28 ± 1.01). DISCUSSION AND CONCLUSIONS: Tanshinone IIA shows antiepileptic and cognitive function-improving effects, primarily via regulating synaptic plasticity. This research generates a theoretical foundation for future research on potential clinical applications for tanshinone IIA.

Neuronal protective effect of Songling Xuemaikang capsules alone and in combination with carbamazepine on epilepsy in kainic acid-kindled rats.

CONTEXT: Epilepsy is a common life-threatening neurological disorder that is often drug-resistant and associated with cognitive impairment. The traditional Chinese patent medicine Songling Xuemaikang capsules (SXC) is clinically used for epilepsy therapy and alleviation of cognitive impairment. OBJECTIVE: This study investigates the neuronal protective effect of SXC combined with carbamazepine (CBZ) on epilepsy and cognitive impairment in kainic acid-kindled SD rats. MATERIALS AND METHODS: Kainic acid-kindled rats were established by injection of 0.45 µg kainic acid and randomly divided into 5 groups (n = 14): saline (sham-operated), control, CBZ, SXC and CBZ + SXC combined group. Rats in the treatment groups received CBZ (50 mg/kg/d), SXC (600 mg/kg/d) or combined CBZ (50 mg/kg/d) + SXC (600 mg/kg/d) via intragastric injection for 60 days. Epileptic behaviours, cognitive impairment, neuronal apoptosis and expression of p-Akt, Akt and caspase-9 were measured, and the alleviation of cognitive damage and neuronal apoptosis was analyzed. RESULTS: The combined administration of SXC and CBZ significantly decreased the frequency of seizures (1.2 ± 0.3) and the number of episodes (1.3 ± 0.5) above stage III (p < 0.05). Neuronal apoptosis was improved (p < 0.01), and cognitive damage was ameliorated (p < 0.05).The level of p-Akt was enhanced (p < 0.01) whereas the expression of caspase-9 was evidently inhibited (p < 0.01) in the combined group. CONCLUSIONS: The present findings confirm that the combined use of SXC with CBZ can effectively control epileptic seizures, alleviate damage to hippocampal neurons and protect against cognitive impairment. The mechanism of action might be related to the upregulation of p-Akt and inhibition of caspase-9 expression.

Protective effect of compound Danshen (Salvia miltiorrhiza) dripping pills alone and in combination with carbamazepine on kainic acid-induced temporal lobe epilepsy and cognitive impairment in rats.

CONTEXT: Temporal lobe epilepsy (TLE) is resistant to antiepileptic drugs (AEDs) and is associated with cognitive impairment. The modern Chinese medicine, compound Danshen dripping pills (CDDP), is clinically effective in treating epilepsy and improving cognitive impairment. OBJECTIVE: This study evaluated the protective effects of CDDP alone and in combination with carbamazepine (CBZ) on kainic acid-induced TLE and cognitive impairment in rats. MATERIALS AND METHODS: Sprague-Dawley rats were randomly divided into five groups: control (sham operated), model, CDDP, CBZ and combined. A TLE model was then created via bilateral intrahippocampal injection of 0.35 µg kainic acid (KA). Rats received CDDP (85 mg/kg), CBZ (100 mg/kg) or combined (85 mg/kg CDDP +100 mg/kg CBZ) via intragastric administration for 90 d, respectively. Seizure intensity, apoptosis and glial cell line-derived neurotrophic factor (GDNF) were measured. Furthermore, the improvement in cognitive impairment and hippocampal neuronal damage was evaluated. RESULTS: CDDP combined with CBZ significantly decreased seizure severity and frequency (p < 0.05) and ameliorated cognitive impairment (p < 0.05). The model group showed a significant reduction of neurons and Bcl-2/Bax expression in the hippocampus CA3 area (p < 0.01), the combined groups significantly reversed these change (p < 0.01). GDNF expression in the combined groups showed a clear increase over the model group (p < 0.05). CONCLUSION: These findings support the use of CDDP as an adjuvant drug for the treatment of TLE and cognitive deficit. Its mechanism might be related to an anti-apoptosis effect and up-regulation of GDNF.

ß-methasone-containing biodegradable poly(lactide-co-glycolide) acid microspheres for intraarticular injection: effect of formulation parameters on characteristics and in vitro release.

A sustained drug release system based on the injectable poly(lactic-co-glycolic acid) (PLGA) microspheres loaded with ß-methasone was prepared for localized treatment of rheumatic arthritis. The microscopy and structure of microspheres were characterized by scanning electron microscope (SEM) and Fourier transform infrared (FTIR). The effects of various formulation parameters on the properties of microspheres and in vitro release pattern of ß-methasone were also investigated. The results demonstrated that increase in drug/polymer ratio led to increased particle size as well as drug release rate. Increase in PLGA concentration led to increased particle size, but decreased burst release. The drug encapsulation efficiency increased sharply by increasing polyvinyl alcohol (PVA) concentration in the aqueous phase from 1.5 to 2.0%. ß-methasone release rate decreased considerately with decreasing OP (organic phase)/AP (aqueous phase) volume ratio. Stirring rate had significantly influence on the particle size and encapsulation efficiency. Independent of formulation parameters, ß-methasone was slowly released from the PLGA microspheres over 11 days. The drug release profile of high drug loaded microspheres agree with Higuchi equation with a release mechanism of diffusion and erosion, that of middle drug loaded microspheres best agreed with Hixcon-Crowell equation and controlled by diffusion and erosion as well. The low drug loaded microspheres well fitted to logarithm normal distribution equation with mechanism of purely Fickian diffusion.

[Preparation and quality evaluation of fufangxiaoyepipa dispersible tablets].

OBJECTIVE: To study the preparation of fufangxiaoyepipa dispersible tablets and evaluate its quality. METHODS: Using the disintegration time as index to screen out the best prescription of the dispersible tablets by orthogonal design. RESULTS: The prescription assembly was 40% of extraction, 15% of MCC, 18% of CCMC-Na, 25% of Calcium sulfate, 2% of Magnesium stearate, the disintegration time met the provision of Pharmacopoeia. CONCLUSION: The dispersible tablets dissolve faster and disperse uniformly and the dissolution percent in vitro is obviously superior to the conventional tablets, improving the bioavailability of the preparation.

Ginaton improves neurological function in ischemic stroke rats via inducing autophagy and maintaining mitochondrial homeostasis.

PURPOSE: The present study was carried out to confirm the protective effect of extract of Ginkgo biloba (Ginaton) against ischemic neuronal damage post-treatment at 24 h after reperfusion in rats with middle cerebral artery occlusion (MCAO) and further reveal its possible mechanisms. METHODS: Adult male Sprague-Dawley rats were modeled by MCAO for 2 h. The rats were divided into three groups: sham, model, and Ginaton (50 mg/kg). All animals received treatment once a day for 14 days from 24 h after reperfusion. Modified neurological severity score test was performed in 1, 7 and 14 days after MCAO, and beam walking test was performed only 14 days after MCAO. Hematoxylin-eosin straining was implemented to measure infarct volume and immunohistochemical analysis was performed to calculate the number of neurons in ischemic cortex penumbra. Western blot was used to evaluate the expression of autophagy (Beclin1, LC3, AMPK, mTOR, ULK), mitochondrial dynamic protein (Parkin, DRP1, OPA1) and apoptosis (Bcl-2, Bax). RESULTS: Post-treatment with Ginaton for 14 days decreased neurological deficit score, promoted the recovery of motor function, and noticeably reduced infarct size. Besides, Ginaton also alleviated the loss of NeuN-positive cells in ischemic cortex penumbra. In ischemic cortex, Ginaton increased the expression of Beclin1 and LC3-Ⅱ, elevated the AMPK, mTOR and ULK1, and induced autophagy. Moreover, Ginaton treatment upregulated Parkin, DRP1, and OPA1, and elevated the ratio of Bcl-2/Bax in 14 days after MCAO reperfusion injury. CONCLUSION: Ginaton exhibited obvious neuroprotective effects in MCAO rats with initial administered 24 h after MCAO. The mechanism of Ginaton included induction of autophagy via activation of the AMPK pathway, maintenance of mitochondrial homeostasis and inhibition of apoptosis.

Euphornin reduces proliferation of human cervical adenocarcinoma HeLa cells through induction of apoptosis and G2/M cell cycle arrest.

BACKGROUND: The plant Euphorbia helioscopia L. has been used in traditional Chinese medicine for treating various disorders such as tuberculosis and edema. The aim of this study was to investigate the effect of euphornin, a bioactive compound isolated from E. helioscopia, on proliferation of human cervical adenocarcinoma HeLa cells by analyzing cell viability, rate of apoptosis, and cell cycle progression. MATERIALS AND METHODS: The sulforhodamine B assay was used to study the effect of euphornin on the proliferation of HeLa cells. Morphological changes to cell nuclei were identified after Hoechst 33342 staining. Mitochondrial membrane depolarization (MMP) was analyzed after staining with JC-1 dye. The influence of euphornin on the apoptosis rate was analyzed by Annexin V/propidium iodide double staining. Fluorescence-activated cell sorting was applied to investigate the influence of euphornin on cell cycle progression. Proteins were obtained from HeLa cells and analyzed by Western blots. RESULTS: A cell viability assay showed that euphornin inhibited proliferation of HeLa cells in a dose-dependent and time-dependent manner. Euphornin also induced apoptosis in a concentration-dependent manner, with the rates of apoptosis ranging from 25.3% to 52.6%. A high concentration of euphornin was found to block HeLa cells at the G2/M stage. A Western blot analysis suggested that euphornin might exhibit antitumor activity by inducing apoptosis. Euphornin treatment altered the ratio of Bax/Bcl-2 in HeLa cells, which led to the release of cytochrome complex. The levels of cleaved caspase-3, caspase-8, caspase-9, and caspase-10 were also markedly increased by euphornin treatment. Analysis of cell cycles indicated that euphornin induced cell cycle arrest by increasing the level of the phospho-CDK1 (Tyr15) protein. The various assays demonstrated that euphornin treatment resulted in a significant suppression of cell growth accompanied by G2/M cell cycle arrest and increased rate of apoptosis via mitochondrial and caspase pathways. CONCLUSION: Our findings suggest that euphornin has the potential to be used as a cancer therapeutic agent against human cervical adenocarcinoma.

Transdermal application of azithromycin-amlodipine-heparin gel enhances survival of infected random ischaemic flap.

BACKGROUND: Flap necrosis is generally regarded as the result of vasospasm, thrombosis, and infection. METHODS: To improve skin flap survival and lower the risk of side effects due to systemic drug delivery, we formulated and evaluated compound gels for transdermal application. The transdermal delivery of 1% azithromycin (AZM), 0.5% amlodipine besylate (AB), and 300 IU/g low molecular weight heparin (LMWH) in compound gels, singly or in combinations, was measured across rat skin in vitro. The effects of AB and LMWH on flap blood circulation was investigated using fluorescein angiography, by transdermally applying the gel onto the surface of an in vivo ischaemic flap rat model; concentrations of the drugs were detected in both blood plasma and flap tissue at assigned timepoints. Finally, infected ischaemic flaps were treated to evaluate their anti-inflammatory effects and sizes of flap survival area. RESULTS: Each drug efficiently penetrated the in vitro skin in a time-dependent manner. In the in vivo ischaemic flaps, AB or LMWH increased the blood supply. All gel formulations that included AZM were associated with less flap inflammation. The surviving areas after treatment with AZM+LMWH or AZM+AB were significantly larger than that treated with the AZM-only gel, and the largest surviving area was that treated with AZM+AB+LMWH. Gels containing no AZM could not decrease flap inflammation or increase flap survival. CONCLUSION: Transdermal application of a compound gel with AZM, AB, and LMWH combined is a promising method to prevent and treat flap infection, improve blood circulation, and increase the survival of infected ischaemic flaps.