Peripheral nerve impairment is a common complication in surgery, clinical researchers always do nerve sutrure using microsurgical technique and adjuvant treatment to improve peripheral nerve regeneration. Western medicine used usually adjuvant drugs, such as neurotrophic factors,are limited by their defects in clinical application. Traditional Chinese medicines (TCMs) classifies peripheral nerve impair as flaccidity Zheng and arthromyodynia, and considers that it is the result of stagnant blood block in the meridians and vessels, deficient of Qi and blood and disuse of bones and muscles. So, drugs usually have the function of invigorating vital energy, activating blood circulation and dredging collaterals. Mono-drugs include astragalus, Salvia miltiorrhiza, Astragali Radix, Epimedii Folium and so on. Extracts of TCMs have Ginkgo Folium, Cervi Cornu Pantotrichum, Achyranthis Bidentatae Radix, and so on. To be ready for further study and development, TCMs which can promote the peripheral nerve regeneration were reviewed by the literatures of the latest years.
Animals ; Drugs, Chinese Herbal ; pharmacology ; Humans ; Nerve Growth Factors ; pharmacology ; Nerve Regeneration ; drug effects ; Peripheral Nerves ; drug effects ; physiology

A scaffold fabricated with lysine/nerve growth factor (NGF)/poly (lactic acid coglycolic acid) copolymer (PLGA) and acellular pigskin was evaluated in vitro as a potential artificial nerve scaffold. Properties of the scaffold such as microstructure, mechanical property, degradation behavior in PBS and water, Schwann cell adhesion property, and release of NGF were investigated. Results showed PLGA had permeated into the porous structure of acellular pigskin; its breaking strength was 8.308 MPa, breaking extensibility was 38.98%, elastic modulus was 97.27 MPa. The porosities of the scaffold ranged from 68.3% to 81.2% with densities from 0.62 g/cm3 to 0.68 g/cm3. At 4 weeks of degradation in vitro, maximum mass loss ratio was 43.3%. The release of NGF could still be detected on the 30th day, and its accumulative release rate was 38%. Lysine added into the scaffold neutralized the acidoid preventing degradation of PLGA to maintain a solution pH value. Schwann cells had grown across the scaffold after co-cultivation for 15 days. These in vitro properties of the pigskin based composite might indicate its potentiality to be an artificial nerve scaffold.
Acellular Dermis ; Animals ; Biocompatible Materials ; Guided Tissue Regeneration ; Lactic Acid ; pharmacology ; Lysine ; pharmacology ; Nerve Growth Factors ; chemistry ; pharmacology ; Nerve Regeneration ; Polyglycolic Acid ; pharmacology ; Swine ; Tissue Engineering ; Tissue Scaffolds

AIMThe protecting effect of human-derived neurotrophin-6 (NT-6) on injured neurons was investigated to test the neurobiological characteristics of human-derived NT-6.

METHODSAdult SD rats were used and divided into two groups, normal control group and experiment group. The right facial nerve of the experimental rats was sectioned. The experimental animals were subdivided into three groups, blank control group, NT-6 group and saline control group. The rats were raised for two weeks. Brain stem of the rats was removed and transversely sliced. Nissle stain and ChAT immunohistochemical stain of the slices were carried out to observe the role of NT-6 in protecting facial motoneurons.

RESULTSThe number of the ChAT positive neurons and the intensity of Nissle stain in the facial nucleus of NT-6 group were significantly augmented compared with that of the blank and saline control groups.

CONCLUSIONThe human-derived NT-6 could partially protect the facial motoneurons from retrograde degeneration induced by their axon damage.

Animals ; Disease Models, Animal ; Facial Nerve ; cytology ; Humans ; Motor Neurons ; drug effects ; pathology ; Nerve Growth Factors ; pharmacology ; Neuroprotective Agents ; pharmacology ; Rats ; Rats, Sprague-Dawley

This study aims to explore the neuroprotective effect of bilobalide(BB) and the mechanisms such as inhibiting inflammatory response in macrophage/microglia, promoting neurotrophic factor secretion, and interfering with the activation and differentiation of peripheral CD4~+ T cells. BB of different concentration(12.5, 25, 50, 100 μg·mL~(-1)) was used to treat the RAW264.7 and BV2 cells for 24 h. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT) assay and cell counting kit-8(CCK-8) were employed to detect the cytotoxicity of BB and appropriate concentration was selected for further experiment. Lipopolysaccharide(LPS) was applied to elicit inflammation in RAW264.7 and BV2 cells, mouse bone marrow-derived macrophages(BMDMs), and primary microglia, respectively. The effect of BB on cell proliferation and secretion of inflammatory cytokines and neurotrophic factors was detected by enzyme-linked immunosorbent assay(ELISA). Spleen monocytes of C57BL/6 female mice(7-8 weeks old) were isolated, and CD4~+ T cells were separated by magnetic beads under sterile conditions. Th17 cells were induced by CD3/CD28 and the conditioned medium for eliciting the inflammation in BMDMs. The content of IL-17 cytokines in the supernatant was detected by ELISA to determine the effect on the activation and differentiation of CD4~+ T cells. In addition, PC12 cells were incubated with the conditioned medium for eliciting inflammation in BMDMs and primary microglia and the count and morphology of cells were observed. The cytoto-xicity was determined by lactate dehydrogenase(LDH) assay. The result showed that BB with the concentration of 12.5-100 μg·mL~(-1) had no toxicity to RAW264.7 and BV2 cells, and had no significant effect on the activity of cell model with low inflammation. The 50 μg·mL~(-1) BB was selected for further experiment, and the results indicated that BB inhibited LPS-induced secretion of inflammatory cytokines. The experiment on CD4~+ T cells showed that the conditioned medium for LPS-induced inflammation in BMDMs promoted the activation and differentiation of CD4~+ T cells, while the conditioned medium of the experimental group with BB intervention reduced the activation and differentiation of CD4~+ T cells. In addition, BB also enhanced the release of neurotrophic factors from BMDMs and primary microglia. The conditioned medium after BB intervention can significantly reduce the death of PC12 neurons, inhibit neuronal damage, and protect neurons. To sum up, BB plays a neuroprotective role by inhibiting macrophage and microglia-mediated inflammatory response and promoting neurotrophic factors.
Female ; Rats ; Mice ; Animals ; Bilobalides/pharmacology* ; Neuroprotection ; Lipopolysaccharides/toxicity* ; Culture Media, Conditioned/pharmacology* ; Mice, Inbred C57BL ; Macrophages/metabolism* ; Microglia ; Cytokines/metabolism* ; Nerve Growth Factors/pharmacology* ; Inflammation/metabolism*

BACKGROUNDPigment epithelium-derived factor (PEDF) is expressed in several normal organs and identified as an inhibitor of neovascularization. In the present study, we investigated the effect of PEDF in an in vitro model of ocular choroidal neovascularization.

METHODSMicrodissection was used to isolate the human choroidal endothelial cells (CECs), followed by the use of superparamagnetic beads (Dynabeads) coated with the CD31 antibody, which selectively binds to the endothelial cell surface. The mitogenic and motogenic effects of vascular endothelial growth factor (VEGF) on cultured choroidal capillary endothelial cells were examined in the presence or absence of PEDF (1, 10, 100, and 1000 ng/ml) using cell counts and migration assays.

RESULTSCells bound to the beads were isolated using a magnetic particle concentrator and they were successfully cultured and characterized to be endothelial cells that possessed greater than 95% immunoreactivity to von Willebrand factor. PEDF suppressed the proliferation and migration of VEGF-induced choroidal capillary endothelial cells. However, the concentration of PEDF which we used has little effect on normal CECs.

CONCLUSIONSPEDF played an important role on the growth and migration of VEGF-stimulated choroidal endothelial cell. These findings suggest that PEDF may be an effective approach to the treatment of choroidal neovascular disorders.

Cell Movement ; drug effects ; Cell Proliferation ; drug effects ; Cells, Cultured ; Choroid ; blood supply ; drug effects ; Choroidal Neovascularization ; drug therapy ; Endothelial Cells ; cytology ; drug effects ; Eye Proteins ; pharmacology ; Humans ; Nerve Growth Factors ; pharmacology ; Serpins ; pharmacology ; Vascular Endothelial Growth Factor A ; pharmacology

Fractal dimension has been widely used in medical images processing and analysis. The neurite growth of cultured dorsal root ganglion (DRG) was detected by fluorescent immunocytochemistry treated with nerve regeneration factor (0.1, 0.5, 2.0 mg/L). A novel method based on triangular prism surface area (TPSA) was introduced and adopted to calculate the fractal dimension of the two-dimensional immunofluorescent images. Experimental results demonstrate that this method is easy to understand and convenient to operate, and the quantititve results are concordant with the observational findings under microscope. This method can be guidelines for analyzing and deciding experimental results.
Cells, Cultured ; Fluorescent Antibody Technique ; methods ; Fractals ; Ganglia, Spinal ; cytology ; Humans ; Image Processing, Computer-Assisted ; methods ; Nerve Growth Factors ; pharmacology ; Neurites ; physiology

As a severe threat to human health, ischemic brain injury has a very complex pathological mechanism involving excitotoxic amino acids, oxygen free radical formation, nitric oxide (NO), Ca2+ overload and inflammation. Traditional Chinese medicine Qingkailing injection have shown good clinical efficacy in the treatment of cerebrovascular disease, and thus it is very significant to studies on its pharmacological mechanism. This essay summarizes relevant studies on pharmacological mechanism of a new compound traditional Chinese medicine Jingzhiqiangkailing (JZQKL) injection in treatment on cerebral ischemia, and explains the pharmacological mechanism of its single effective compounds and their compatibility in treatment of schemic brain injury in the aspects of regulating inflammatory response, neurotrophic factors, vascular protection, blood-brain barrier (BBB) protection and others, and thus providing information for further studies.
Animals ; Blood-Brain Barrier ; drug effects ; Brain Ischemia ; drug therapy ; Cell Adhesion Molecules ; physiology ; Cytokines ; biosynthesis ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Humans ; Injections ; Nerve Growth Factors ; physiology

To get the mature peptide of human-derived neurotrophin-6 (NT-6), NT-6 gene encoding mature peptide was amplified by PCR, using the NT-6 cDNA that had been cloned as templet. The gene encoding mature peptide of NT-6 gene was cloned into pGEX1-lambdaT plasmid to construct the fusion expression vector. Expression of fusion protein in Escherichia coli was defected after induction by isopropyl beta-D-thiogalactoside(IPTG). The mature peptide of NT-6 was collected with GST fusion protein purifying kit. It was shown that a fragment of 460bp was gained by PCR. With the techniques of double-cleave and electrophoresis, the recombinant vector was identified as pGEX1-NT-6. The recombinant vector pGEX1-NT-6 transformed Escherichia coli expressed fusion protein of 41KD after induction by IPTG. Cleaved by thrombin, the mature peptide of NT-6 was obtained; its molecular weight was about 15KD. The cloning and expression of human-derived NT-6 gene encoding mature protein has provided a basis for further studies on the function and clinical application of NT-6.
Escherichia coli ; genetics ; metabolism ; Genetic Vectors ; genetics ; Humans ; Isopropyl Thiogalactoside ; pharmacology ; Nerve Growth Factors ; biosynthesis ; genetics ; Peptides ; genetics ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; isolation & purification

BACKGROUNDThe common pathological characteristics of corneal injury include inflammatory factors activation, vascular endothelial cells or inflammatory cells infiltration into lesions, corneal edema, corneal neovascularization (CNV), and scar formation. PEDF-34 is the functional fragment of pigment epithelium-derived factor (PEDF) that has anti-angiogenic and anti-inflammatory properties and contains an N-terminal 34-amino acid peptide. This study was to investigate the anti-inflammatory effects of PEDF-34 on H2O2-induced corneal injury in vitro.

METHODSAfter cultured in H2O2 (0.1 mmol/L) for 2 hours, human corneal fibroblasts (HCFs) and human umbilical vein endothelial cells (HUVECs) were treated with PEDF-34-nanoparticles (NPs) at different concentrations (0.1, 0.5, 1.0, 2.0 µg/ml) or 2.0 µg/ml control-NPs for 24 hours. The viable cells were quantified using the MTT assay. Western blotting or ELISA analysis was performed for measuring the human vascular endothelial growth factor (VEGF) and intercellular adhesion molecule-1 (ICAM-1) expression of both HCFs and HUVECs. VEGF and nuclear factor κB (NF-κB) mRNA levels of HCFs were semi-quantified by RT-PCR.

RESULTSThe survival rates of HCFs or HUVECs stimulated by H2O2 did not decrease significantly (P > 0.05) compared to those in the normal conditions. As compared to control-NP group, PEDF-34-NPs had dose-dependent inhibitive effect on HUVECs with the MTT assay, but not HCFs. Western blotting analysis showed that the VEGF and ICAM-1 levels in the HCFs and HUVECs stimulated by H2O2 were significantly higher than those in the normal conditions, which were decreased dramatically in those treated with PEDF-34-NPs. RT-PCR analysis revealed that the VEGF mRNA and NF-κB mRNA levels increased in H2O2-stimulated HCFs, while both of them decreased in PEDF-34-NP groups dose dependently.

CONCLUSIONSPEDF-34-NPs may play an important role in regulating the NF-κB pathway, inhibiting inflammatory activity. PEDF-34-NPs may be a potential new drug for treating corneal injury in the future.

Anti-Inflammatory Agents ; chemistry ; pharmacology ; Cells, Cultured ; Corneal Injuries ; chemically induced ; metabolism ; Eye Proteins ; chemistry ; Human Umbilical Vein Endothelial Cells ; drug effects ; metabolism ; Humans ; Hydrogen Peroxide ; pharmacology ; Nerve Growth Factors ; chemistry ; Peptides ; chemistry ; pharmacology ; Serpins ; chemistry

Animals ; Diabetic Neuropathies ; drug therapy ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Humans ; Nerve Growth Factors ; drug effects ; Nerve Regeneration ; drug effects ; Peripheral Nervous System Diseases ; drug therapy ; Phytotherapy ; Schwann Cells ; drug effects