Yuanming Ouyang

Nerve Guidance Conduits from Aligned Nanofibers: Improvement of Nerve Regeneration through Longitudinal Nanogrooves on a Fiber Surface

A novel fibrous conduit consisting of well-aligned nanofibers with longitudinal nanogrooves on the fiber surface was prepared by electrospinning and was subjected to an in vivo nerve regeneration study on rats using a sciatic nerve injury model. For comparison, a fibrous conduit having a similar fiber alignment structure without surface groove and an autograft were also conducted in the same test. The electrophysiological, walking track, gastrocnemius muscle, triple-immunofluorescence, and immunohistological analyses indicated that grooved fibers effectively improved sciatic nerve regeneration. This is mainly attributed to the highly ordered secondary structure formed by surface grooves and an increase in the specific surface area. Fibrous conduits made of longitudinally aligned nanofibers with longitudinal nanogrooves on the fiber surface may offer a new nerve guidance conduit for peripheral nerve repair and regeneration.

Polymerizing Pyrrole Coated Poly (l-lactic acid-co-ε-caprolactone) (PLCL) Conductive Nanofibrous Conduit Combined with Electric Stimulation for Long-Range Peripheral Nerve Regeneration.

Electrospinning and electric stimulation (ES) are both promising methods to support neuron adhesion and guide extension of neurons for nerve regeneration. Concurrently, all studies focus on either electrospinning for conduits material or ES in vitro study to accelerate nerve regeneration; few work on the combined use of these two strategies or ES in vivo study. Therefore, this study aimed to investigate the abilities of direct current ES through electrospinning conductive polymer composites composed of polypyrrole and Poly (l-lactic acid-co-ε-caprolactone) (PPY/PLCL) in peripheral nerve regeneration. PPY/PLCL composite conduits were synthesized by polymerizing pyrrole coated electrospun PLCL scaffolds. Morphologies and chemical compositions were characterized by scanning electron microscope and attenuated total reflection fourier transform infrared (ATR-FTIR) microscope. Rat pheochromocytoma 12 (PC12) cells and dorsal root ganglia (DRG) cells cultured on PPY/PLCL scaffolds were stimulated with 100 mV/cm for 4 h per day. The median neurite length and cell viability were measured in PC-12 cells. The levels of brain-derived neurotrophic factor (BDNF), glial cell derived neurotrophic factor (GDNF) and neurotrophin-3 (NT-3) were analyzed in DRG cells. In rats, 15 mm gaps of sciatic nerves were bridged using an autograft, non-stimulated PPY/PLCL conduit and PPY/PLCL conduit stimulated with 100 mV potential, respectively. A 100 mV potential direct current ES was applied for 1 h per day at 1, 3, 5 and 7 days post-implantation. The PPY/PLCL conduits with ES showed a similar performance compared with the autograft group, and significantly better than the non-stimulated PPY/PLCL conduit group. These promising results show that the PPY/PLCL conductive conduits’ combined use with ES has great potential for peripheral nerve regeneration.

Potential Value of miR-221/222 as Diagnostic, Prognostic, and Therapeutic Biomarkers for Diseases

microRNAs (miRNAs) are short non-coding RNAs that regulate gene expression by base pairing with their target messenger RNAs. Dysregulation of miRNAs is involved in the pathological initiation and progression of many human diseases. miR-221 and miR-222 (miR-221/222) are two highly homologous miRNAs, and they are significantly overexpressed in several types of human diseases. Silencing miR-221/222 could represent a promising approach for therapeutic studies. In the present review, we will describe the potential value of miR-221/222 as diagnostic, prognostic, and therapeutic biomarkers in various diseases including cancer and inflammatory diseases.

Efficient and Non-Toxic Biological Response Carrier Delivering TNF-α shRNA for Gene Silencing in a Murine Model of Rheumatoid Arthritis

Small interfering RNA (siRNA) is an effective and specific method for silencing genes. However, an efficient and non-toxic carrier is needed to deliver the siRNA into the target cells. Tumor necrosis factor α (TNF-α) plays a central role in the occurrence and progression of rheumatoid arthritis (RA). In this study, we pre-synthetized a degradable cationic polymer (PDAPEI) from 2,6-pyridinedicarboxaldehyde and low-molecular-weight polyethyleneimine (PEI, Mw = 1.8 kDa) as a gene vector for the delivery of TNF-α shRNA. The PDAPEI/pDNA complex showed a suitable particle size and stable zeta potential for transfection. In vitro study of the PDAPEI/pDNA complex revealed a lower cytotoxicity and higher transfection efficiency when transfecting TNF-α shRNA to macrophages by significantly down-regulating the expression of TNF-α. Moreover, the complex was extremely efficient in decreasing the severity of arthritis in mice with collagen-induced arthritis. PDAPEI delivered TNF-α shRNA has great potential in the treatment of RA.

Evaluation of the proliferation and differentiation behaviors of mesenchymal stem cells with partially converted borate glass containing different amounts of strontium in vitro.

The objective of this study was to examine the proliferation and differentiation behaviors of different compositions of strontium-containing (from 0-12 mol%) borate glasses with mesenchymal stem cells (MSCs). The Cell Counting Kit-8 (CCK-8) assay revealed that after three days of culturing, the 6Sr group had the highest cell growth rate. Analysis of cell morphology revealed that cells proliferated well near the particles of the samples in all the groups on day 3. On day 7, cells in the 6Sr group demonstrated a higher proliferation rate than other 4 groups under the microscope. When performing the Live-Dead staining experiment, the 6Sr group had the least number of dead cells. Total DNA qualification indicated that the 6Sr group had a statistically higher concentration compared with the remaining groups. It was found that on day 7, compared with the 0Sr group, the core binding factor α1 (Cbfa1) mRNA expression level was significantly higher in the 6Sr, 9Sr and 12Sr groups. On day 14, compared with the 0Sr group, the bone sialoprotein (BSP) mRNA level was significantly higher in the 6Sr group. Additionally, on day 21, the 6Sr and 9Sr groups demonstrated higher osteocalcin (OCN) mRNA expression levels compared with the 0Sr group. In the alkaline phosphatase (ALP) activity test, on day 21, the 6Sr group presented a higher activity than the 0Sr group. Further, the number of mineralized nodules per unit in MSCs was measured by Alizarin Red S staining. The results showed that the 6Sr and 9Sr groups had the greatest number of mineralized nodules. Therefore, it could be concluded that borate glasses containing strontium oxide of 0, 3, 6, 9 and 12 mol% demonstrate a significant level of proliferation when interacting with MSCs. The borate glass containing 6 mol% strontium oxide had the greatest level of proliferation when cultured with MSCs. The borate glass containing 6 and 9 mol% strontium oxide facilitated an improved bone formation ability compared with the remaining two compositions.

Effect of celecoxib on proliferation, collagen expression, ERK1/2 and SMAD2/3 phosphorylation in NIH/3T3 fibroblasts

In the present study, the effects of celecoxib on proliferation, collagen expression, ERK1/2 and SMAD2/3 phosphorylation in NIH/3T3 fibroblasts were investigated. NIH/3T3 fibroblasts stimulated with fibroblast growth factor-2 (FGF-2) or transforming growth factor-β1 (TGF-β1) were examined in the presence of celecoxib. Proliferation was assessed by MTT assays; ERK1/2 expression and SMAD2/3 expression were assessed by quantitative RT-PCR and western blotting; ERK1/2 phosphorylation and SMAD2/3 phosphorylation were assessed by western blot analysis. The results indicated that celecoxib could suppress cell proliferation stimulated by FGF-2 (IC(50) FGF+group, 75±1.9μmol/l) and TGF-β1 (IC(50) TGF+group, 48±1.4μmol/l), by inhibiting ERK1/2 phosphorylation but not ERK1/2 expression. Celecoxib also suppressed collagen expression (0.35-fold COL3 and 0.43-fold COL1 at 320μmol/l celecoxib relative to the untreated control after stimulation with TGF-β1 for 3h, P<0.01), by inhibiting SMAD2/3 phosphorylation but not SMAD2/3 expression. The suppression of NIH/3T3 fibroblast proliferation and collagen expression upon stimulation by FGF-2 and TGF-β1 is likely a result of the inhibition of ERK1/2 and SMAD2/3 phosphorylation by celecoxib.

Platelet-Rich Plasma Derived Growth Factors Contribute to Stem Cell Differentiation in Musculoskeletal Regeneration

Stem cell treatment and platelet-rich plasma (PRP) therapy are two significant issues in regenerative medicine. Stem cells such as bone marrow mesenchymal stem cells, adipose-derived stem cells and periodontal ligament stem cells can be successfully applied in the field of tissue regeneration. PRP, a natural product isolated from whole blood, can secrete multiple growth factors (GFs) for regulating physiological activities. These GFs can stimulate proliferation and differentiation of different stem cells in injury models. Therefore, combination of both agents receives wide expectations in regenerative medicine, especially in bone, cartilage and tendon repair. In this review, we thoroughly discussed the interaction and underlying mechanisms of PRP derived GFs with stem cells, and assessed their functions in cell differentiation for musculoskeletal regeneration.

Lentivirus-mediated microRNA-124 gene-modified bone marrow mesenchymal stem cell transplantation promotes the repair of spinal cord injury in rats

Our study aims to explore the effects of lentivirus-mediated microRNA-124 (miR-124) gene-modified bone marrow mesenchymal stem cell (BMSC) transplantation on the repair of spinal cord injury (SCI) in rats. BMSCs were isolated from the bone marrow of rats. The target gene miR-124 was identified using a luciferase-reporter gene assay. Seventy-two rats were selected for construction of the SCI model, and the rats were randomly divided into the blank group, sham group, SCI group, negative control (NC) group, overexpressed miR-124 group and si-PDXK group. The mRNA expression of miR-124 and the mRNA and protein expression of pyridoxal kinase (PDXK) were detected by quantitative real-time polymerase chain reaction and western blotting. The locomotor capacity of the rats was evaluated using the Basso, Beattie and Bresnahan (BBB) scale. Brdu, neuron-specific enolase (NSE), neurofilament (NF) and microtubule-associated protein 2 (MAP2) were detected using immunohistochemistry. The expression levels of thyrotropin-releasing hormone (TRH), prostacyclin (PGI2) and gangliosides (GM) were measured using an enzyme-linked immunosorbent assay. PDXK was identified as the target gene of miR-124. The overexpressed miR-124 group exhibited higher miR-124 expression than the SCI, NC and si-PDXK groups. Compared with the SCI and NC groups, the PDXK expression was downregulated in the overexpressed miR-124 and si-PDXK groups, and the BBB scores were significantly increased 7, 21 and 35 days after transplantation. The double-labeled positive cell densities (Brdu+NSE/NF/MAP2) and the expression levels of TRH, PGI2 and GM in the overexpressed miR-124 group were significantly higher than those in the NC and SCI groups. These results indicated that miR-124 targeted PDXK to accelerate the differentiation of BMSCs into neurocytes and promote SCI repair.

Advances in Roles of miR-132 in the Nervous System

miR-132 is an endogenous small RNA and controls post-transcriptional regulation of gene expression via controlled degradation of mRNA or transcription inhibition. In the nervous system, miR-132 is significant for regulating neuronal differentiation, maturation and functioning, and widely participates in axon growth, neural migration, and plasticity. The miR-132 is affected by factors like mRNA expression, functional redundancy, and signaling cascades. It targets multiple downstream molecules to influence physiological and pathological neuronal activities. MiR-132 can influence the pathogenesis of many diseases, especially in the nervous system. The dysregulation of miR-132 results in the occurrence and exacerbation of neural developmental, degenerative diseases, like Alzheimer’s disease, Parkinson’s disease and epilepsy, neural infection and psychiatric disorders including disturbance of consciousness, cognition and memory, depression and schizophrenia. Regulation of miR-132 expression relieves symptoms, alleviates severity and finally effects a cure. This review aims to discuss the clinical potentials of miR-132 in the nervous system.

Celecoxib suppresses fibroblast proliferation and collagen expression by inhibiting ERK1/2 and SMAD2/3 phosphorylation.

This study aimed to investigate whether celecoxib suppresses fibroblast proliferation and collagen expression by inhibiting extracellular signal-regulated kinase 1/2 (ERK1/2) and SMAD2/3 phosphorylation. Celecoxib was added to NIH/3T3 fibroblasts stimulated by fibroblast growth factor-2 (FGF-2) or transforming growth factor-β1 (TGF‑β1). NIH/3T3 fibroblast proliferation and viability were assessed by MTT assays; ERK1/2 expression and SMAD2/3 expression were assessed by quantitative RT-PCR and Western blot analysis. The results indicated that celecoxib suppressed cell proliferation (IC50 FGF+ group, 75 ± 1.9 µmol/l) stimulated by FGF-2, and also inhibited cell viability (IC50 FGF- group, 252 ± 2.3 µmol/l) by inhibiting ERK1/2 phosphorylation but not ERK1/2 expression. In addition, celecoxib treatment led to the apoptosis of NIH/3T3 fibroblasts (IC50 FGF- group, 35 ± 1.4 µmol/l). Celecoxib also suppressed collagen expression (0.35-fold COL3 and 0.43-fold COL1 with 320 µmol/l celecoxib relative to the untreated group following stimulation for 3 h, p<0.01) when stimulated by TGF‑β1, by inhibiting SMAD2/3 phosphorylation but not SMAD2/3 expression. Celecoxib is capable of inhibiting ERK1/2 and SMAD2/3 phosphorylation, which is responsible for NIH/3T3 fibroblast proliferation and collagen expression.