Shu-Hua Yang

Thermosensitive chitosan-gelatin-glycerol phosphate hydrogel as a controlled release system of ferulic acid for nucleus pulposus regeneration.

In the degenerative disc, overproduction of reactive oxygen species (ROS) involves in apoptosis and senescence of nucleus pulposus (NP) cells that could accelerate the degenerative process. Ferulic acid (FA) has been reported to have an excellent antioxidant property. In the study, injectable thermosensitive chitosan/gelatin/glycerol phosphate (C/G/GP) hydrogel was applied as a controlled release system for FA delivery. The study was aimed to evaluate possible therapeutic effects of FA-incorporated C/G/GP hydrogel on hydrogen peroxide (H(2)O(2))-induced oxidative stress NP cells. The results showed that the release of FA from C/G/GP hydrogel could decrease the H(2)O(2)-induced oxidative stress. Post-treatment of FA-incorporated C/G/GP hydrogel on H(2)O(2)-induced oxidative stress NP cells showed up-regulation of Aggrecan and type II collagen and down-regulation of MMP-3 in mRNA level. The results of sulfated-glycosaminoglycans (GAGs) to DNA ratio and alcian blue staining revealed that the GAGs production of H(2)O(2)-induced oxidative stress NP cells could reach to normal level. The results of caspase-3 activity and TUNEL staining indicated that FA-incorporated C/G/GP hydrogel decreased the apoptosis of H(2)O(2)-induced oxidative stress NP cells. The results suggested that the C/G/GP hydrogel was very suitable for sustained delivery of FA. The FA-incorporated C/G/GP hydrogel would be used to treat the degenerative disc in the early stage before it developed into the latter irreversible stages.

In Vitro Study on Interaction Between Human Nucleus Pulposus Cells and Mesenchymal Stem Cells Through Paracrine Stimulation

Study Design. Coculture of human nucleus pulposus (NP) cells and mesenchymal stem cells (MSCs) using a noncontact system. Objective. To investigate the interaction between NP cells and MSCs through paracrine stimulation. Summary of Background Data. Cell-based therapies have a potential role in the treatment of intervertebral disc degeneration. Upregulating the viability of NP cells and differentiating MSCs into NP-like cells are potential alternatives to achieve viable cells. Methods. Culture plates and inserts were used to coculture MSCs and NP cells without direct contact or exchange of cellular components. Cellular proliferation and RNA expression of selected genes were then evaluated after coculture. Results. Coculturing slightly promoted the proliferation of MSCs, and expression of collagen I and Fas-associated death domain protein significantly decreased. MSCs, which initially expressed no collagen II, started to show collagen II expression after coculturing; the expression level was highest when the cells were cultured with a higher number of NP cells. On the converse, proliferation of NP cells significantly rose even after cocultured with a few MSCs. Increasing the number of cocultured MSCs did not further enhance proliferation of NP cells. Expression of aggrecan in the NP cells significantly increased when the cells were cultured with a higher number of MSCs. Conclusion. The results showed a possible mechanism of interaction between MSCs and NP cells mediated by secreted factors. The most significant effect on NP cells was enhancement of cellular proliferation when they were cocultured with even a small number of MSCs. To differentiate MSCs into NP-like cells with heightened collagen II expression, MSCs must be in an environment containing numerous NP cells.

Proximal kyphosis after short posterior fusion for thoracolumbar scoliosis.

Thoracolumbar idiopathic scoliosis usually is treated by anterior spinal fusion. However, short posterior spinal fusion that includes only the structural curve has been tried in a limited number of patients. The fusion may end cranially in the lower thoracic region and cause an increase in sagittal decompensation at the proximal junction. From July 1989 to July 1998, 14 patients were treated with thoracolumbar idiopathic scoliosis by short posterior spinal fusion. The lateral radiographs were evaluated preoperatively, immediately postoperative, and during followup. The focal kyphotic angle was used to examine the changes in focal sagittal alignment. A 10° progression was defined as the radiographic criterion for the development of junctional kyphosis. Proximal junctional kyphosis occurred in six of the 14 patients, in which one patient needed revision surgery. In all six patients, the average preoperative lumbar lordosis was greater than 35°, and decreased more than 10° during surgery. In the five patients with a focal kyphotic angle larger than 10°, four had proximal junctional kyphosis develop. According to the current findings, short posterior spinal fusion can be done only if the focal kyphotic angle proximal to the fusion is less than 10°, and the lumbar lordosis must be preserved carefully during surgery.

Stress analysis of the distal locking screws for femoral interlocking nailing.

In femoral locked nailing, the distal locking screws are vulnerable to mechanical failure. Biomechanical studies have shown that the stress on these screws is substantially affected by the fit of the nail in the medullary canal. In this study, a “closed form” mathematical model based on elastic beam‐column theory was developed to investigate how the nail‐cortical contact, which was simulated by a linear elastic foundation, affected the stress on the distal locking screws. Providing data for the model was a construct of a fractured femur with an intramedullary locked nail loaded by an eccentric vertical load. The stress on the locking screw was analyzed as a function of the distance from the fracture to the locking screw in the distal fragment under two situations: with and without nail‐cortical contact in the distal fragment. With nail‐cortical contact, the screw stress decreased as the length of nail‐cortical contact and the distance between the distal locking screw and the fracture site increased, but this stress contrarily increased when the nail reached the femoral region at which the screw length increased. The screw stress was much higher without nail‐cortical contact than with contact and continued to increase as the nail was inserted further. The mathematical model developed here can be a convenient means of rapid stress evaluation and parametric analysis for locked femoral nailing. It may be used to improve the design of interlocking nails and surgical technique.

In situ forming hydrogels composed of oxidized high molecular weight hyaluronic acid and gelatin for nucleus pulposus regeneration

Encapsulation of nucleus pulposus (NP) cells within in situ forming hydrogels is a novel biological treatment for early stage intervertebral disc degeneration. The procedure aims to prolong the life of the degenerating discs and to regenerate damaged tissue. In this study we developed an injectable oxidized hyaluronic acid-gelatin-adipic acid dihydrazide (oxi-HAG-ADH) hydrogel. High molecular weight (1900 kDa) hyaluronic acid was crosslinked with various concentrations of gelatin to synthesize the hydrogels and their viscoelastic properties were analyzed. Interactions between the hydrogels, NP cells, and the extracellular matrix (ECM) were also evaluated, as were the effects of the hydrogels on NP cell gene expression. The hydrogels possess several clinical advantages, including sterilizability, low viscosity for injection, and ease of use. The viscoelastic properties of the hydrogels were similar to native tissue, as reflected in the complex shear modulus (∼11-14 kPa for hydrogels, 11.3 kPa for native NP). Cultured NP cells not only attached to the hydrogels but also survived, proliferated, and maintained their round morphology. Importantly, we found that hydrogels increased NP cell expression of several crucial ECM-related genes, such as COL2A1, AGN, SOX-9, and HIF-1A.

Postoperative Meralgia Paresthetica After Posterior Spine Surgery: Incidence, Risk Factors, and Clinical Outcomes

Study Design. A prospective study on postoperative meralgia paresthetica after posterior thoracolumbar spine surgery on the Relton-Hall frame. Objectives. To assess the incidence of postoperative lateral femoral cutaneous nerve (LFCN) neuralgia and to investigate its risk factors and clinical outcomes. Summary of Background Data. Postoperative meralgia paresthetica is a common complication of posterior thoracolumbar spine surgery. The injury mechanism is external compression to the LFCN near the anterior superior iliac spine in the prone position. Methods. A total of 252 patients were examined for signs of meralgia paresthetica before and after surgery. Patients with a LFCN injury were followed regularly until sensory impairment resolved. Several possible contributing factors were assessed to evaluate the correlations. Results. Postoperative meralgia paresthetica was experienced by 60 patients (23.8%). Patients with an LFCN injury had a significantly greater body mass index (23.6 vs. 22.4 kg/m2) and a longer surgical time (3.7 vs. 3.2 hours). Overweight/obese patients had a significantly greater incidence (odds ratio, 1.83; 95% confidence interval, 1.02–3.29). Patients operated for degenerative spinal disorders also had a significantly higher incidence of LFCN injury (odds ratio, 2.81; 95% confidence interval, 1.53–5.13). Recovery took 10.5 days on average (range, 2 days to 2 months). Thirty-two patients (53%) recovered completely within the first week and every patient recovered within 2 months. Conclusion. Postoperative meralgia paresthetica is a common but benign complication of posterior thoracolumbar spine surgery. Degenerative spinal disorders, overweight/obesity, and longer surgical time are factors related to a higher incidence of LFCN injury. The clinical outcome is always excellent, and complete recovery can be expected within 2 months.

Small intestinal submucosa as a potential bioscaffold for intervertebral disc regeneration.

Study Design. To evaluate the capacity of porcine small intestine submucosa to support the in vitro proliferation of human disc cells and the synthesis of extracellular matrix that could restore the biochemical properties of the disc. Objective. To evaluate if porcine small intestine submucosa is a potential bioactive scaffold for rescuing degenerative disc cells. Summary of Background Data. Discogenic back pain is associated with alterations of the disc and abnormal turnover of the disc extracellular matrix. We hypothesize that a biodegradable and biocompatible acellular scaffold such as small intestine submucosa, which contains entrapped growth factors, may stimulate disc cells to synthesize extracellular matrix, thereby arresting the degeneration, or even promoting the regeneration, of the disc. Methods. Human degenerative anulus and nucleus cells were seeded onto small intestine submucosa scaffolds, and evaluated over a 3-month period for cell growth (proliferation assay, deoxyribonucleic acid content) and matrix composition (glycosaminoglycan and collagen contents). Results. As hematoxylin and eosin staining revealed, more than 70% of seeded cells attached to the small intestine submucosa surface and invaded throughout the scaffold. The macroscopic appearance of cell-seeded scaffolds was dramatically modified over time. Cell metabolic activity was confirmed for up to 3 months. Seeded scaffolds showed a higher glycosaminoglycan content as compared to control scaffolds. Toluidine blue staining detected large areas of proteoglycans. Positive gene expression for collagens I, II, and X, aggrecan, and Sox-9 confirmed deposition of new extracellular matrix components. Conclusions. This pilot study shows that small intestine submucosa is a promising bioactive material that could potentially serve as a temporary scaffold for intervertebral disc regeneration.

Thermosensitive hydrogel made of ferulic acid-gelatin and chitosan glycerophosphate.

Reactive oxygen species-induced oxidative stress is involved in apoptosis of nucleus pulposus (NP) cells that can alter cellular phenotype and accelerate disc degeneration. Ferulic acid (FA) possesses an excellent antioxidant and anti-inflammatory properties. In the study, we developed the thermosensitive FA-gelatin/chitosan/glycerol phosphate (FA-G/C/GP) hydrogel which was applied as a sustained release system of FA to treat NP cells from the damage caused by oxidative stress. The gelation temperature of the FA-G/C/GP hydrogel was 32.17 °C. NP cells submitted to oxidative stress promoted by H(2)O(2), and post-treated with FA-G/C/GP exhibited down-regulation of MMP-3 and up-regulation aggrecan and type II collagen in mRNA level. The sulfated-glycosaminoglycan production was increased and the apoptosis was inhibited in the post-treatment group. The results suggest that the thermosensitive FA-G/C/GP hydrogel can treat NP cells from the damage caused by oxidative stress and may apply in minimally invasive surgery for NP regeneration.

Three-dimensional Culture of Human Nucleus Pulposus Cells in Fibrin Clot: Comparisons on Cellular Proliferation and Matrix Synthesis With Cells in Alginate

Regeneration of nucleus pulposus (NP) tissue may stop or reverse early intervertebral disk (IVD) degeneration. Cellular proliferation and matrix synthesis can be promoted by incorporation of cells and bioscaffolds. However, insertion of preshaped solid bioscaffolds may damage remaining IVD integrity. Fibrin clots can be introduced in a minimally invasive manner with polymerization in desired three-dimensional shape and retention of cells. In this study, we investigated the cellular proliferation and matrix synthesis of human NP cells in the fibrin clots in vitro. Monolayer-expanded cells were embedded in fibrin clot or alginate and were cultivated in vitro for 2 weeks. Increased DNA content and decreased expression of apoptosis stimulating fragment (Fas)-associated death-domain protein in fibrin scaffolds suggested higher cellular proliferation and reduced apoptosis. Superior proteoglycan synthesis was found in fibrin scaffolds. As expression of collagens I and X increased and SOX9 expression decreased, fibrin scaffolds tended to promote fibrotic transformation and inhibit chondrogenesis. Adjustments of fibrin preparations are needed to make it more suitable for IVD regeneration.

Influence of age-related degeneration on regenerative potential of human nucleus pulposus cells

Nucleus pulposus (NP) cells, sourced from herniation surgeries, may be used as a cell‐based therapy for intervertebral disc (IVD) degeneration. But, both the regenerative potential of these degenerative adult NP cells and how to stimulate optimum matrix synthesis is not yet clear. The purpose of the current study was to understand the different phenotypic behaviors between degenerative adult NP cells and normal adolescent NP cells. Degenerative adult NP cells produced a significantly higher amount of proteoglycans and collagens than adolescent cells. Insulin‐like growth factor‐1 was the only anabolic cytokine with increased endogenous expression in degenerative adult NP cells. TGF‐β1 treatment of degenerative NP cells promoted matrix synthesis but stimulated too much type I collagen and suppressed type II collagen and aggrecan. Adult degenerative NP cells possess upregulated regenerative potential, but stimulation in addition to TGF‐β1 is needed to enhance matrix productivity and optimize the collagen expression profile.