Matrix metalloproteinase 12 is an indicator of intervertebral disc degeneration co-expressed with fibrotic markers.

OBJECTIVE: Recent evidence suggests a role of fibrogenesis in intervertebral disc (IVD) degeneration. We aim to explore if fibrotic genes may serve as IVD degeneration indicators, and if their expression is associated with myofibroblast activity. DESIGN: Transcriptional expression of fibrosis markers (COL1A1, COL3A1, FN1, HSP47, MMP12, RASAL1) were analyzed in degenerated (D) and non-degenerated (ND) human nucleus pulposus (NP) and annulus fibrosus (AF) cells, along with traditional (SOX9, ACAN) and newly established degeneration markers (CDH2, KRT19, KRT18, FBLN1, MGP, and COMP). Protein expression was investigated by immunohistochemistry in human IVDs, and in rodent IVDs undergoing natural ageing or puncture-induced degeneration. Co-expression with myofibroblast markers was examined by double staining on human and rat specimens. Disc degeneration severity and extent of fibrosis were determined by histological scoring and picrosirius red staining respectively. RESULTS: Human D-NP showed more intensive staining for picrosirius red than ND-NP. Among the genes examined, D-NP showed significantly higher MMP12 expression along with lower KRT19 expression. Protein expression analysis revealed increased MMP12(+) cells in human D-IVD. Histological scoring indicated mild degeneration in the punctured rat discs and discs of ageing mouse. Higher MMP12 positivity was found in peripheral NP and AF of the degenerative rat discs and in NP of the aged mice. In addition, human D-NP and D-AF showed increased α-SMA(+) cells, indicating enhanced myofibroblast activity. MMP12 was found co-expressed with α-SMA, FSP1 and FAP-α in human and rat degenerative IVDs. CONCLUSIONS: Our study suggests that in addition to a reduced KRT19 expression, an increased expression of MMP12, a profibrotic mediator, is characteristic of disc degenerative changes. Co-expression study indicates an association of the increased MMP12 positivity with myofibroblast activity in degenerated IVDs. Overall, our findings implicate an impact of MMP12 in disc cell homeostasis. The precise role of MMP12 in IVD degeneration warrants further investigation.

Enrichment of committed human nucleus pulposus cells expressing chondroitin sulfate proteoglycans under alginate encapsulation.

OBJECTIVE: Intervertebral disc (IVD) degeneration is associated with a malfunction of the nucleus pulposus (NP). Alginate culturing provides a favorable microenvironment for the phenotypic maintenance of chondrocyte-like NP cells. However, NP cells are recently evidenced to present heterogeneous populations, including progenitors, fibroblastic cells and primitive NP cells. The aim of this study is to profile the phenotypic changes of distinct human NP cells populations and describe the dynamic expression of chondroitin sulfate glycosaminoglycans (CS-GAGs) in extended alginate encapsulation. METHOD: Non-degenerated (ND-NPC) and degenerated (D-NPC) NP cells were expanded in monolayers, and subject to 28-day culture in alginate after serial passaging. CS-GAG compositional expression in monolayer-/alginate-cultured NP cells was evaluated by carbohydrate electrophoresis. Cellular phenotypic changes were assessed by immunologic detection and gene expression analysis. RESULTS: Relative to D-NPC, ND-NPC displayed remarkably higher expression levels of chondroitin-4-sulfate GAGs over the 28-day culture. Compared with monolayer culture, ND-NPC showed increased NP marker expression of KRT18, KRT19, and CDH2, as well as chondrocyte markers SOX9 and MIA in alginate culture. In contrast, expression of fibroblastic marker COL1A1, COL3A1, and FN1 were reduced. Interestingly, ND-NPC showed a loss of Tie2+ but gain in KRT19+/CD24+ population during alginate culture. In contrast, D-NPC showed more consistent expression levels of NP surface markers during culture. CONCLUSION: We demonstrate for the first time that extended alginate culture selectively enriches the committed NP cells and favors chondroitin-4-sulfate proteoglycan production. These findings suggest its validity as a model to investigate IVD cell function.

Magnetic-film atom chip with 10 µm period lattices of microtraps for quantum information science with Rydberg atoms.

We describe the fabrication and construction of a setup for creating lattices of magnetic microtraps for ultracold atoms on an atom chip. The lattice is defined by lithographic patterning of a permanent magnetic film. Patterned magnetic-film atom chips enable a large variety of trapping geometries over a wide range of length scales. We demonstrate an atom chip with a lattice constant of 10 µm, suitable for experiments in quantum information science employing the interaction between atoms in highly excited Rydberg energy levels. The active trapping region contains lattice regions with square and hexagonal symmetry, with the two regions joined at an interface. A structure of macroscopic wires, cutout of a silver foil, was mounted under the atom chip in order to load ultracold (87)Rb atoms into the microtraps. We demonstrate loading of atoms into the square and hexagonal lattice sections simultaneously and show resolved imaging of individual lattice sites. Magnetic-film lattices on atom chips provide a versatile platform for experiments with ultracold atoms, in particular for quantum information science and quantum simulation.

Interplay between multiple scattering, emission, and absorption of light in the phosphor of a white light-emitting diode.

We study light transport in phosphor plates of white light-emitting diodes (LEDs). We measure the broadband diffuse transmission through phosphor plates of varying YAG:Ce(3+) density. We distinguish the spectral ranges where absorption, scattering, and re-emission dominate. Using diffusion theory, we derive the transport and absorption mean free paths from first principles. We find that both transport and absorption mean free paths are on the order of the plate thickness. This means that phosphors in commercial LEDs operate well within an intriguing albedo range around 0.7. We discuss how salient parameters that can be derived from first principles control the optical properties of a white LED.

The role of cryopreservation in the biomechanical properties of the intervertebral disc.

Implantation of intervertebral disc (IVD) allograft or tissue engineered disc constructs in the spine has emerged as an alternative to artificial disc replacement for the treatment of severe degenerative disc disease (DDD). Establishment of a bank of cryopreserved IVD allografts enables size matching and facilitates logistics for effective clinical management. However, the biomechanical properties of cryopreserved IVDs have not been previously reported. This study aimed to assess if cryopreservation with different concentrations of cryopreservant agents (CPA) would affect the dynamic viscoelastic properties of the IVD. Whole porcine lumbar IVDs (n = 40) were harvested and processed using various concentrations of CPA, 0 % CPA, 10 % CPA and 20 % CPA. The discs were cryopreserved using a stepwise freezing protocol and stored in liquid nitrogen. After four weeks of storage, the cryopreserved IVDs were quickly thawed at 37 °C for dynamic viscoelastic testing. The apparent modulus, elastic modulus (G'), viscous modulus (G") and loss modulus (G"/G') were calculated and compared to a fresh control group. Cryopreserved IVD without cryopreservants was significantly stiffer than the control. In the dynamic viscoelastic testing, cryopreservation with the use of CPA was able to preserve both G' and G" of an IVD. No significant differences were found between fresh IVD and IVD cryopreserved with 10 % CPA or 20 % CPA. This study demonstrated that CPAs at an optimal concentration could preserve the mechanical properties of the IVD allograft and can provide further credence for the application of long-term storage of IVD allografts for disc transplantation or tissue engineered construct applications.

Mesenchymal stem cell-based repair of articular cartilage with polyglycolic acid-hydroxyapatite biphasic scaffold.

This study investigates the capacity of a composite scaffold composed of polyglycolic acid-hydroxyapatite (PGA-HA) and autologous mesenchymal stem cells (MSCs) to promote repair of osteochondral defects. MSCs from culture-expanded rabbits were seeded onto a PGA and HA scaffold. After a 72-hour co-culture period, the cell-adhered PGA and HA were joined together, forming an MSCs-PGA-HA composite. Full-thickness cartilage defects in the intercondylar fossa of the femur were then implanted with the MSC-PGA-HA composite, the PGA-HA scaffold only, or they were left empty (n=20). Animals were sacrificed 16 or 32 weeks after surgery and the gross appearance of the defects was evaluated. The specimens were examined histologically for morphologic features, and stained immunohistochemically for type 2 collagen. Specimens of the MSCs-PGA-HA composite implantation group demonstrated hyaline cartilage and a complete subchondral bone formation. At 16 weeks post-implantation, significant integration of the newly formed tissue with surrounding normal cartilage and subchondral bone was observed when compared to the two control groups. At 32 weeks, no sign of progressive degeneration of the newly formed tissue was found. A significant difference in histological grading score was found compared with the control groups. The novel MSCs-seeded, PGA-HA biphasic graft facilitated both articular cartilage and subchondral bone regeneration in an animal model and might serve as a new approach for clinical applications.

Age-related degeneration of lumbar intervertebral discs in rabbits revealed by deuterium oxide-assisted MRI.

OBJECTIVES: Intervertebral disc (IVD) degeneration is associated with a loss of disc water content and change in biochemical composition of the disc. Rabbit is a frequently used model to evaluate the efficacy of therapeutics for disc degeneration. This study addresses whether rabbits undergo age-related disc degeneration, assessed using deuterium oxide-assisted magnetic resonance imaging (MRI) of the lumbar IVDs. MATERIALS AND METHODS: The lumbar spines of adolescent, adult, and aged rabbits (6-36 months) were subjected to T2-weighted/short-tau inversion recovery (STIR) MRI scan along with water-deuterium oxide (H(2)O:D(2)O) dilutions. The total and maximum H(2)O:D(2)O index (HDi) of the lumbar IVDs were determined and compared between disc levels at different ages. RESULTS: Adolescent rabbit lumbar discs had similar total HDi, suggesting the hydration and biochemical composition was similar among the lumbar levels. With the use of H(2)O:D(2)O reference, the discs were shown to undergo continual decrease in signal with aging which non-calibrated measurement method could not reveal. The HDi decrease rate was higher at the caudal than cranial levels. CONCLUSION: This study provided in vivo evidence of age-related progressive disc degenerative change in rabbit lumbar discs, suggesting aged rabbits can be considered as a natural disc degeneration model in disc regeneration studies. However, it is important to select proper disc levels as intra-subject controls due to different rates of degenerative changes between caudal and cranial levels.

Ultrasound bladder measurements in children with severe primary nocturnal enuresis: pretreatment and posttreatment evaluation and its correlation with treatment outcome.

PURPOSE: Results from our previous study demonstrated a high predictive value using ultrasound bladder measurements to identify abnormal bladder function in children with enuresis or urinary tract infection. We prospectively evaluated the role of ultrasound measured bladder parameters for the assessment of bladder dysfunction and posttreatment bladder functional changes (if any), and their correlation with treatment outcome in children with primary nocturnal enuresis. MATERIALS AND METHODS: Patients presenting with severe primary nocturnal enuresis (more than 3 wet nights weekly) were prospectively recruited. At study entry each patient underwent ultrasound, and natural and conventional filling cystometric studies. Bladder volume and wall thickness index was calculated based on ultrasound studies and classified as thick (less than 70), normal (70 to 130) or thin (more than 130). The criteria for diagnosing urodynamic patterns included normal, overactive and underactive detrusor activity. Correlation between the ultrasound measured parameters and urodynamic findings was then evaluated. Patients were treated based on our standardized treatment protocol. Bladder measurements were repeated in those children who had completed treatment. The McNemar test was used for comparing posttreatment changes in bladder measurements corresponding to treatment outcome in different groups, and p values less than 0.05 were regarded as statistically significant. RESULTS: A total of 35 children (23 males, 12 females; mean age 9.03 years) were prospectively recruited. At study entry bladder volume and wall thickness index was normal in 8 patients, less than 70 in 24 and more than 130 in 3. When bladder volume and wall thickness index was correlated with ultrasound 87.5% of the patients with a normal index exhibited a normal bladder pattern on imaging. In addition, 96% of the patients with an index of less than 70 exhibited bladder overactivity on ultrasound. All of the children with a normal index had either a complete or good response to treatment, whereas 62.5% of those with an index of less than 70 did not respond to treatment. On followup bladder dysfunction had resolved in 37.5% of the children with an initial index of less than 70, all of whom had a good response to the treatment. Bladder dysfunction persisted in 62.5% of the patients, all of whom had partial or no response to treatment (p <0.001). CONCLUSIONS: Ultrasound measured bladder parameters correlated well with ultrasound findings, changes in bladder function and treatment outcome in children with primary nocturnal enuresis. This study further confirms that this specially designed ultrasound protocol can provide useful predictive clues that may be helpful in differentiating between various treatment subtypes, guiding clinical management and minimizing the need for invasive urodynamic studies.

Deep venous thrombosis caused by femoral venous catheters in critically ill adult patients.

STUDY OBJECTIVES: To determine the frequency of and potential risk factors for catheter-related deep venous thrombosis (DVT) in critically ill adult patients. DESIGN: Prospective, controlled, observational cohort study. SETTING: A mixed medical and surgical ICU in a university hospital. PATIENTS: All adult patients undergoing femoral vein catheterization. INTERVENTIONS: None. MEASUREMENTS: ICU diagnosis, underlying disease, demographic data, type of catheter, complications during cannulation, use of anticoagulants, coagulation status, medications infused, and duration of catheterization were recorded. Compression and duplex Doppler ultrasound studies of both femoral veins were performed prior to insertion, at 12 h after insertion, and daily until catheter removal. Follow-up investigation was performed at 24 h and 1 week after removal. RESULTS: Of 140 cases entered into the study, 124 were evaluated. Fourteen patients developed iliofemoral vein DVTs. Two were clinically obvious. Twelve (9.6%) were line related (uncannulated leg normal) and two (1.6%) occurred only in the uncannulated leg (p = 0.011; relative risk, 6.0; confidence interval, 1.5 to 23.5). Line-related DVT can occur any time from the day after insertion to 1 week after removal. The incidence of catheter-related DVT was unrelated to number of insertion attempts, arterial puncture or hematoma, duration of catheterization, coagulation status, or type of infused medications. No other predisposing or protective factors were identified. Three of the 12 patients with catheter-related DVT died. In no patient was clinical pulmonary embolus suspected. CONCLUSION: Although the femoral route is convenient and has potential advantages, the use of femoral lines increases the risk of iliofemoral DVT. Catheter-related DVT may occur as soon as 1 day after cannulation and is usually asymptomatic. This increased risk should be carefully considered when the femoral route of cannulation is chosen.