Sericin protects against diabetes-induced injuries in sciatic nerve and related nerve cells

Sericin from discarded silkworm cocoons of silk reeling has been used in different fields, such as cosmetology, skin care, nutrition, and oncology. The present study established a rat model of type 2 diabetes by consecutive intraperitoneal injections of low-dose （25 mg/kg） streptozotocin. After intragastrical perfusion of sericin for 35 days, blood glucose levels significantly declined, and the expression of neurofilament protein in the sciatic nerve and nerve growth factor in L4-6 spinal ganglion and anterior horn cells significantly increased. However, the expression of neuropeptide Y in spinal ganglion and anterior horn cells significantly decreased in model rats. These findings indicate that sericin protected the sciatic nerve and related nerve cells against injury in a rat type 2 diabetic model by upregulating the expression of neurofilament protein in the sciatic nerve and nerve growth factor in spinal ganglion and anterior horn cells, and downregulating the expression of neuropeptide Y in spinal ganglion and anterior horn cells.

Effect of ciliary neurotrophic factor on bcl-2 and c-jun gene and protein expression in cultured retinal nerve cells

BACKGROUND：In various retinal neurodegenerative animal models,ciliary neurotrophic factor （CNTF） exhibits prominent neuroprotective effects on retinal nerve cells.Bcl-2 is an anti-apoptotic protein.c-Jun is upregulated and phosphorylated in the activated c-Jun N-terminal kinase pathway,which subsequently mediates apoptosis.However,the effect of CNTF on Bcl-2 and c-Jun expression in retinal nerve cells remains unclear.OBJECTIVE：To determine the dynamic changes in retinal nerve cell apoptosis,as well as bcl-2 and c-jun gene and protein expression,following a single dose of CNTF in a short period of time.DESIGN,TIME AND SETTING：A single-blind,randomized,controlled,in vitro experiment was performed at the Central Laboratory of Beijing Tongren Hospital from May 2008 to April 2009.MATERIALS：Neonatal bovine retinal nerve cells （Chinese Holstein）,recombinant human CNTF （PeproTech,Rocky Hill,NJ,USA）,rabbit polyclonal anti-Bcl-2 and c-Jun antibodies （Abeam,Cambridge,UK）,fluorescein isothiocyanate-conjugated annexin V/propidium iodide kit （BioVision,Mountain View,CA,USA）,real time polymerase chain reaction instrument （ABI,Foster City,CA,USA）,and flow cytometer （BD FACSCalibur,Franklin Lakes,NJ,USA）.METHODS：Neonatal bovine retinal cells from passage 2 were cultured for 3 days and incubated with,or without,50 ng/mL CNTF （control）.MAIN OUTCOME MEASURES：Cell apoptosis was detected via Annexin V-FITC/PI double-staining and flow cytometry.bcl-2 and c-jun mRNA and protein expression were detected by quantitative real time polymerase chain reaction and western blot analysis.RESULTS：The proportion of late-stage apoptotic cells was significantly decreased at 2,4,and 6 days after CNTF treatment compared with the control group （P 〈 0.01）.CNTF did not alter bcl-2 mRNA expression at the three time points,but significantly increased Bcl-2 protein expression at 2 and 4 days （P 〈 0.01）.c-jun mRNA expression was significantly decreased 4 days after CNTF treatment （P〈 0.01）.In addition,c-Jun protein expression was slightly increased at 4 days （P〈 0.01）,but decreased at 6 days,compared with the control group （P〈 0.05）.CONCLUSION：A single dose of CNTF （50 ng/mL） upregulated Bcl-2 protein and downregulated c-jun mRNA expression,followed by a parallel,but lagged,change in c-Jun protein production in cultured neonatal bovine retinal nerve cells.These results suggested that CNTF reduces retinal nerve cell apoptosis by modifying Bcl-2 and c-Jun expression.

Regulation of nerve cells using conductive nanofibrous scaffolds for controlled release of Lycium barbarum polysaccharides and nerve growth factor

Currently,more and more patients suffer from peripheral nerve injury due to trauma,tumor and other causes worldwide.Biomaterial-based nerve conduits are increasingly recognized as a potential alternative to nerve autografts for the treatment of peripheral nerve injury.However,an ideal nerve conduit must offer topological guidance and biochemical and electrical signal transduction mechanisms.In this work,aligned conductive nanofibrous scaffolds comprising polylactic-co-glycolic acid and multiwalled carbon nanotubes(MWCNTs)were fabricated via coaxial electrospinning,and nerve growth factor(NGF)and Lycium barbarum polysaccharides(LBP)purified from the wolfberry were loaded on the core and shell layers of the nanofibers,respectively.LBP were confirmed to accelerate long-distance axon regeneration after severe peripheral nerve injury.In addition,the synergistic promotion of LBP and NGF on nerve cell proliferation and neurite outgrowth was demonstrated.MWCNTs were introduced into the aligned fibers to further increase the electrical conductivity,which promoted the directional growth and neurite extension of neurons in vitro.Further,the combination of conductive fibrous scaffolds with electrical stimulation that mimics endogenous electric fields significantly promoted the differentiation of PC12 cells and the axon outgrowth of neurons.Based on robust cell-induced behaviors,conductive composite fibers with optimized fiber alignment may be used for the promotion of nerve recovery.

Substrate stiffness in nerve cells

Recently, substrate stiffness has been involved in the physiology and pathology of the nervous system. However, the role and function of substrate stiffness remain unclear. Here, we review known effects of substrate stiffness on nerve cell morphology and function in the central and peripheral nervous systems and their involvement in pathology. We hope this review will clarify the research status of substrate stiffness in nerve cells and neurological disorder.

Lamotrigine protects against cognitive deficits,synapse and nerve cell damage,and hallmark neuropathologies in a mouse model of Alzheimer’s disease

Lamotrigine(LTG)is a widely used drug for the treatment of epilepsy.Emerging clinical evidence suggests that LTG may improve cognitive function in patients with Alzheimer’s disease.However,the underlying molecular mechanisms remain unclear.In this study,amyloid precursor protein/presenilin 1(APP/PS1)double transgenic mice were used as a model of Alzheimer’s disease.Five-month-old APP/PS1 mice were intragastrically administered 30 mg/kg LTG or vehicle once per day for 3 successive months.The cognitive functions of animals were assessed using Morris water maze.Hyperphosphorylated tau and markers of synapse and glial cells were detected by western blot assay.The cell damage in the brain was investigated using hematoxylin and eosin staining.The levels of amyloid-βand the concentrations of interleukin-1β,interleukin-6 and tumor necrosis factor-αin the brain were measured using enzyme-linked immunosorbent assay.Differentially expressed genes in the brain after LTG treatment were analyzed by high-throughput RNA sequencing and real-time polymerase chain reaction.We found that LTG substantially improved spatial cognitive deficits of APP/PS1 mice;alleviated damage to synapses and nerve cells in the brain;and reduced amyloid-βlevels,tau protein hyperphosphorylation,and inflammatory responses.High-throughput RNA sequencing revealed that the beneficial effects of LTG on Alzheimer’s disease-related neuropathologies may have been mediated by the regulation of Ptgds,Cd74,Map3k1,Fosb,and Spp1 expression in the brain.These findings revealed potential molecular mechanisms by which LTG treatment improved Alzheimer’s disease.Furthermore,these data indicate that LTG may be a promising therapeutic drug for Alzheimer’s disease.

Bridging long gap peripheral nerve injury using skeletal muscle-derived multipotent stem cells

Long gap peripheral nerve injuries usually reulting in life-changing problems for patients. Skeletal muscle derived-multipotent stem cells （Sk-MSCs） can differentiate into Schwann and perineurial/endoneurial cells, vascular relating pericytes, and endothelial and smooth muscle cells in the damaged peripheral nerve niche. Application of the Sk-MSCs in the bridging conduit for repairing long nerve gap injury resulted favorable axonal regeneration, which showing superior effects than gold standard therapy--healthy nerve autograft. This means that it does not need to sacrifice of healthy nerves or loss of related functions for repairing peripheral nerve injury.

Macrophage polarization in nerve injury： do Schwann cells play a role？

In response to peripheral nerve injury, the inflammatory response is almost entirely comprised of infiltrating macrophages. Macrophages are a highly plastic, heterogenic immune cell, playing an indispensable role in peripheral nerve injury, clearing debris and regulating the microenvironment to allow for efficient regeneration. There are several cells within the microenvironment that likely interact with macrophages to support their function – most notably the Schwann cell, the glial cell of the peripheral nervous system. Schwann cells express several ligands that are known to interact with receptors expressed by macrophages, yet the effects of Schwann cells in regulating macrophage phenotype remains largely unexplored. This review discusses macrophages in peripheral nerve injury and how Schwann cells may regulate their behavior.

Research Status of Astragali Radix on Nerve Cells and Nerve System Diseases

Astragali Radix has a wide application in the nerve system diseases because of its obvious nerve cell protection and recovery effects.Astragali Radix has good clinical effects both in acute and chronic cerebrovascular diseases and neurological degenerative diseases.This paper reviews the experimental and clinical research status of Astragali Radix on nerve system and nerve system diseases,which may promote its experimental research and clinical application.

Nerve growth factor promotes in vitro proliferation of neural stem cells from tree shrews

Neural stem cells promote neuronal regeneration and repair of brain tissue after injury,but have limited resources and proliferative ability in vivo.We hypothesized that nerve growth factor would promote in vitro proliferation of neural stem cells derived from the tree shrews,a primate-like mammal that has been proposed as an alternative to primates in biomedical translational research.We cultured neural stem cells from the hippocampus of tree shrews at embryonic day 38,and added nerve growth factor（100 μg/L） to the culture medium.Neural stem cells from the hippocampus of tree shrews cultured without nerve growth factor were used as controls.After 3 days,fluorescence microscopy after DAPI and nestin staining revealed that the number of neurospheres and DAPI/nestin-positive cells was markedly greater in the nerve growth factor-treated cells than in control cells.These findings demonstrate that nerve growth factor promotes the proliferation of neural stem cells derived from tree shrews.

Substance P combined with epidermal stem cells promotes wound healing and nerve regeneration in diabetes mellitus

Exogenous substance P accelerates wound healing in diabetes,but the mechanism remains poorly understood.Here,we established a rat model by intraperitoneally injecting streptozotocin.Four wounds（1.8 cm diameter） were drilled using a self-made punch onto the back,bilateral to the vertebral column,and then treated using amniotic membrane with epidermal stem cells and/or substance P around and in the middle of the wounds.With the combined treatment the wound-healing rate was 100% at 14 days.With prolonged time,type I collagen content gradually increased,yet type III collagen content gradually diminished.Abundant protein gene product 9.5-and substance P-immunoreactive nerve fibers regenerated.Partial nerve fiber endings extended to the epidermis.The therapeutic effects of combined substance P and epidermal stem cells were better than with amniotic membrane and either factor alone.Our results suggest that the combination of substance P and epidermal stem cells effectively contributes to nerve regeneration and wound healing in diabetic rats.

Human umbilical cord blood stem cells and brainderived neurotrophic factor for optic nerve injury: a biomechanical evaluation

Treatment for optic nerve injury by brain-derived neurotrophic factor or the transplantation of human umbilical cord blood stem cells has gained progress, but analysis by biomechanical indicators is rare. Rabbit models of optic nerve injury were established by a clamp. At 7 days after injury, the vitreous body received a one-time injection of 50 μg brain-derived neurotrophic factor or 1 × 10^6 human umbilical cord blood stem cells. After 30 days, the maximum load, maximum stress, maximum strain, elastic limit load, elastic limit stress, and elastic limit strain had clearly improved in rabbit models of optical nerve injury after treatment with brain-derived neurotrophic factor or human umbilical cord blood stem cells. The damage to the ultrastructure of the optic nerve had also been reduced. These findings suggest that human umbilical cord blood stem cells and brain-derived neurotrophic factor effectively repair the injured optical nerve, improve biomechanical properties, and contribute to the recovery after injury.

Human umbilical cord blood-derived stem cells and brain-derived neurotrophic factor protect injured optic nerve:viscoelasticity characterization

The optic nerve is a viscoelastic solid-like biomaterial.Its normal stress relaxation and creep properties enable the nerve to resist constant strain and protect it from injury.We hypothesized that stress relaxation and creep properties of the optic nerve change after injury.Moreover,human brain-derived neurotrophic factor or umbilical cord blood-derived stem cells may restore these changes to normal.To validate this hypothesis,a rabbit model of optic nerve injury was established using a clamp approach.At 7 days after injury,the vitreous body received a one-time injection of 50 μg human brain-derived neurotrophic factor or 1 × 106 human umbilical cord blood-derived stem cells.At 30 days after injury,stress relaxation and creep properties of the optic nerve that received treatment had recovered greatly,with pathological changes in the injured optic nerve also noticeably improved.These results suggest that human brain-derived neurotrophic factor or umbilical cord blood-derived stem cell intervention promotes viscoelasticity recovery of injured optic nerves,and thereby contributes to nerve recovery.

Microencapsulation improves inhibitory effects of transplanted olfactory ensheathing cells on pain after sciatic nerve injury

Olfactory bulb tissue transplantation inhibits P2X2/3 receptor-mediated neuropathic pain. However, the olfactory bulb has a complex cellular composition, and the mechanism underlying the action of purified transplanted olfactory ensheathing cells（OECs） remains unclear. In the present study, we microencapsulated OECs in alginic acid, and transplanted free and microencapsulated OECs into the region surrounding the injured sciatic nerve in rat models of chronic constriction injury. We assessed mechanical nociception in the rat models 7 and 14 days after surgery by measuring paw withdrawal threshold, and examined P2X2/3 receptor expression in L4–5 dorsal root ganglia using immunohistochemistry. Rats that received free and microencapsulated OEC transplants showed greater withdrawal thresholds than untreated model rats, and weaker P2X2/3 receptor immunoreactivity in dorsal root ganglia. At 14 days, paw withdrawal threshold was much higher in the microencapsulated OEC-treated animals. Our results confirm that microencapsulated OEC transplantation suppresses P2X2/3 receptor expression in L4–5 dorsal root ganglia in rat models of neuropathic pain and reduces allodynia, and also suggest that transplantation of microencapsulated OECs is more effective than transplantation of free OECs for the treatment of neuropathic pain.

Histological observation on acellular nerve grafts co-cultured with Schwann cells for repairing defects of the sciatic nerve

BACKGROUND： Animal experiments and clinical studies about tissue engineering method applied to repair nerve injury mainly focus on seeking ideal artificial nerve grafts, nerve conduit and seed cells. Autologous nerve, allogeneic nerve and xenogeneic nerve are used to bridge nerve defects, it is one of the methods to promote the repair of nerve injury by culturing and growing Schwann cells, which can secrete various neurotrophic factor activities, in the grafts. OBJECTIVE ： To observe the effect of acellular nerve grafts co-cultured with Schwann cells in repairing defects of sciatic nerve. DESIGN： An observational comparative study.SETTING： Tissue Engineering Laboratory of China Medical University.MATERIALS： The experiment was carried out in the Tissue Engineering Laboratory of China Medical University between April 2004 and April 2005. Forty neonatal Sprague-Dawley rats of 5-8 days （either males or females） and 24 male Wistar rats of 180-220 g were provided by the experimental animal center of China Medical University. METHODS： ① Culture of Schwann cells： The bilateral sciatic nerves and branchial plexus were isolated from the 40 neonatal SD rats. The sciatic nerves were enzymatically digested with collagenase and dispase, isolatd, purified and cultured with the method of speed-difference adhersion, and identified with the SABC immunohistochemical method. ② Model establishment： In vitro Schwann cells were microinjected into 10-mm long acellular nerve grafts repairing a surgically created gap in the rat sciatic nerve. According to the different grafted methods, the animals were randomly divided into three groups： autografts （n=8）, acellular nerve grafts （n=8）, or acellular nerve grafts with Schwann cells （n=8）. ③ The regenerated nerve fiber number and average diameter of myeline sheath after culture were statistically anlayzed. MAIN OUTCOME MEASURES： ① The regenerated nerve ultrastructure, total number and density of myelinated nerve fibers, and the thickness of myeline sheath were observed under electron microscope. ② The images were processed with the Mias-1000 imaging analytical system to calculate the number of myelinated nerve fibers, and the thickness of myeline sheath. RESULTS： All the 24 Wistar rats were involved in the analysis of results. ① Results observed under transmission electron microscope： The regenerated myelinated nerve fibers in the group of acellular nerve grafts with Schwann cells were more even than those in the group of acellular nerve grafts, the number of myelinated nerve fibers and thickness of myelin sheath were close to those in the allografts group （P 〉 0.05）, but significantly different from those in the group of acellular nerve grafts （P 〈 0.05）. ② Results observed under scanning electron microscope： A great amount of Schwann cells with two polars were observed in the group of grafts with Schwann cells, the feature of cultured Schwann cells showed shoulder by shoulder, head to head. ③ The number of myelinated nerve fibers and thickness of myelin sheath analyzed by Mias-1000 imaging system in the group of acellular nerve grafts with Schwann cells were close to those in the autografts group （P 〉 0.05）, but significantly different from those in the group of acellular nerve grafts （P 〈 0.05）.CONCLUSION： Host axonal regeneration is significantly increased after implant of acellular nerve grafts. Acellular nerve grafts with Schwann cells offers a novel approach for repairing the gap of nerve defect.

Combined use of Y-tube conduits with human umbilical cord stem cells for repairing nerve bifurcation defects

Given the anatomic complexity at the bifurcation point of a nerve trunk,enforced suturing between stumps can lead to misdirection of nerve axons,thereby resulting in adverse consequences.We assumed that Y-tube conduits injected with human umbilical cord stem cells could be an effective method to solve such problems,but studies focused on the best type of Y-tube conduit remain controversial.Therefore,the present study evaluated the applicability and efficacy of various types of Y-tube conduits containing human umbilical cord stem cells for treating rat femoral nerve defects on their bifurcation points.At 12 weeks after the bridging surgery that included treatment with different types of Y-tube conduits,there were no differences in quadriceps femoris muscle weight or femoral nerve ultrastructure.However,the Y-tube conduit group with longer branches and a short trunk resulted in a better outcome according to retrograde labeling and electrophysiological analysis.It can be concluded from the study that repairing a mixed nerve defect at its bifurcation point with Y-tube conduits,in particular those with long branches and a short trunk,is effective and results in good outcomes.

Neurotrophins and their receptors in satellite glial cells following nerve injury

Peripheral neuropathy is a condition where damage resulting from mechanical or pathological mechanisms is inflicted on nerves within the peripheral nervous system （PNS）. Physical injury is the most common cause and may result in nerves being partially or completely severed, crushed, compressed or stretched. Other causes include metabolic or endocrine disorders, with e.g.,

The active principle region of Buyang Huanwu decoction induced differentiation of bone marrow-derived mesenchymal stem cells into neural-like cells Superior effects over original formula of Buyang Huanwu decoction

The present study induced in vitro-cultured passage 4 bone marrow-derived mesenchymal stem cells to differentiate into neural-like cells with a mixture of alkaloid, polysaccharide, aglycone, glycoside, essential oils, and effective components of Buyang Huanwu decoction （active principle region of decoction for invigorating yang for recuperation）. After 28 days, nestin and neuron-specific enolase were expressed in the cytoplasm. Reverse transcription-PCR and western blot analyses showed that nestin and neuron-specific enolase mRNA and protein expression was greater in the active principle region group compared with the original formula group. Results demonstrated that the active principle region of Buyang Huanwu decoction induced greater differentiation of rat bone marrow-derived mesenchymal stem cells into neural-like cells in vitro than the original Buyang Huanwu decoction formula.

Wharton's jelly mesenchymal stem cells differentiate into retinal progenitor cells

Human Wharton＇s jelly mesenchymal stem cells were isolated from fetal umbilical cord. Cells were cultured in serumfree neural stem cellconditioned medium or neural stem cellconditioned medium supplemented with Dkk1, a Wnt/13 catenin pathway antagonist, and LeftyA, a Nodal signaling pathway antagonist to induce differentiation into retinal progenitor cells. Inverted microscopy showed that after induction, the spindleshaped or fibroblastlike Wharton＇s jelly mesenchymal stem cells changed into bulbous cells with numerous processes. Immunofluorescent cytochemical stain ing and reversetranscription PCR showed positive expression of retinal progenitor cell markers, Pax6 and Rx, as well as weakly downregulated nestin expression. These results demonstrate that Wharton＇s jelly mesenchymal stem cells are capable of differentiating into retinal progenitor cells in vitro.