Characterization of the three-dimensional kinematics and failure of human spinal segments

Spine disorders are one of the most prevalent and costly problems facing modern medicine, with an estimated annual cost of over $95 billion. Improved understanding of the biomechanical properties of the normal and pathological spine is essential for prevention of such disorders and treatment of traumatic injury, disc degeneration, or other ailments. However, the complex structure of the spine makes experimental testing that is relevant to physiological behavior difficult. Hence, new testing methods are needed to provide more accurate descriptions of in vivo behavior. Therefore, three principal aims are pursued in this work. The purpose of the first aim was to investigate the biomechanical performance of the human cadaver spine by applying a pure bending moment in conjunction with a range of compressive axial loads. This study showed that a minimum of 500 N compressive axial preload along the spinal curvature produces more comparable results to in-vivo studies, providing an important guideline for experiments investigating range of motion and spinal stability. The purpose of the second aim was to determine the motion response of the spinal joint to pure bending in any plane around its circumference. Towards that goal, the three-dimensional motion envelope of two-level spinal segments was analyzed. Since the posterior elements were intact, restriction in motion caused by zygapophyseal joints resulted a smaller displacement for extension than in flexion. Characterization of the motion envelope can provide a better understanding of the complex joint kinematics and assist in the design of new implants with the goal of restoring joint function. The purpose of the third aim was to investigate the strength of a single vertebral body during compression fracture by following the path of least resistance. In vivo, the spinal column continuously maintains equilibrium and minimizes stress via a complex system of muscles, tendons and ligaments such that each vertebra experiences a predominantly axial load. In a typical experiment, compressive loads are also applied axially but unwanted shear forces and moments are generated as well. Using a novel testing approach, identification of the weakest region of a vertral body, and following this path of least resistance, a true measure of vertebral strength, we were able to demonstrate the effect of current experimental limitations in overpredicting bone strength

Innate immune dysfunctions in aged mice facilitate the systemic dissemination of methicillin-resistant S. aureus.

Elderly humans show increased susceptibility to invasive staphylococcal disease after skin and soft tissue infection. However, it is not understood how host immunity changes with aging, and how that predisposes to invasive disease. In a model of severe skin infection, we showed that aged mice (16- to 20-month-old) exhibit dramatic bacterial dissemination compared with young adult mice (2-month-old). Bacterial dissemination was associated with significant reductions of CXCL1 (KC), polymorphonuclear cells (PMNs), and extracellular DNA traps (NETs) at the infection site. PMNs and primary skin fibroblasts isolated from aged mice showed decreased secretion of CXCL2 (MIP-2) and KC in response to MRSA, and in vitro analyses of mitochondrial functions revealed that the mitochondrial electron transport chain complex I plays a significant role in induction of chemokines in the cells isolated from young but not old mice. Additionally, PMNs isolated from aged mice have reduced ability to form NETs and to kill MRSA. Expression of nuclease by S. aureus led to increased bacterial systemic dissemination in young but not old mice, suggesting that defective NETs formation in elderly mice permitted nuclease and non-nuclease expressing S. aureus to disseminate equally well. Overall, these findings suggest that gross impairment of both skin barrier function and innate immunity contributes to the propensity for MRSA to disseminate in aged mice. Furthermore, the study indicates that contribution of bacterial factors to pathogenicity may vary with host age

A Translational Porcine Model for Human Cell-Based Therapies in the Treatment of Posttraumatic Osteoarthritis After Anterior Cruciate Ligament Injury.

BACKGROUND: There is a high incidence of posttraumatic osteoarthritis (PTOA) after anterior cruciate ligament (ACL) injury, and these injuries represent an enormous health care economic burden. In an effort to address this unmet clinical need, there has been increasing interest in cell-based therapies. PURPOSE: To establish a translational large animal model of PTOA and demonstrate the feasibility of intra-articular human cell-based interventions. STUDY DESIGN: Descriptive laboratory study. METHODS: Nine Yucatan mini-pigs underwent unilateral ACL transection and were monitored for up to 12 weeks after injury. Interleukin 1 beta (IL-1β) levels and collagen breakdown were evaluated longitudinally using enzyme-linked immunosorbent assays of synovial fluid, serum, and urine. Animals were euthanized at 4 weeks (n = 3) or 12 weeks (n = 3) after injury, and injured and uninjured limbs underwent magnetic resonance imaging (MRI) and histologic analysis. At 2 days after ACL injury, an additional 3 animals received an intra-articular injection of 107 human bone marrow-derived mesenchymal stem cells (hBM-MSCs) combined with a fibrin carrier. These cells were labeled with the luciferase reporter gene (hBM-MSCs-Luc) as well as fluorescent markers and intracellular iron nanoparticles. These animals were euthanized on day 0 (n = 1) or day 14 (n = 2) after injection. hBM-MSC-Luc viability and localization were assessed using ex vivo bioluminescence imaging, fluorescence imaging, and MRI. RESULTS: PTOA was detected as early as 4 weeks after injury. At 12 weeks after injury, osteoarthritis could be detected grossly as well as on histologic analysis. Synovial fluid analysis showed elevation of IL-1β shortly after ACL injury, with subsequent resolution by 2 weeks after injury. Collagen type II protein fragments were elevated in the synovial fluid and serum after injury. hBM-MSCs-Luc were detected immediately after injection and at 2 weeks after injection using fluorescence imaging, MRI, and bioluminescence imaging. CONCLUSION: This study demonstrates the feasibility of reproducing the chondral changes, intra-articular cytokine alterations, and body fluid biomarker findings consistent with PTOA after ACL injury in a large animal model. Furthermore, we have demonstrated the ability of hBM-MSCs to survive and express transgene within the knee joint of porcine hosts without immunosuppression for at least 2 weeks. CLINICAL RELEVANCE: This model holds great potential to significantly contribute to investigations focused on the development of cell-based therapies for human ACL injury-associated PTOA in the future (see Appendix Figure A1, available online)

Detection of low back pain using pH level‐dependent imaging of the intervertebral disc using the ratio of R1ρ dispersion and −OH chemical exchange saturation transfer (RROC)

Purpose: Low pH is associated with intervertebral disc (IVD)-generated low back pain (LBP). The purpose of this work was to develop an in vivo pH level-dependent magnetic resonance imaging (MRI) method for detecting discogenic LBP, without using exogenous contrast agents. Methods: The ratio of R1ρ dispersion and chemical exchange saturation transfer (CEST) (RROC) was used for pH-level dependent imaging of the IVD while eliminating the effect of labile proton concentration. The technique was validated by numerical simulations and studies on phantoms and ex vivo porcine spines. Four male (ages 42.8 ± 18.3) and two female patients (ages 55.5 ± 2.1) with LBP and scheduled for discography were examined with the method on a 3.0 Tesla MR scanner. RROC measurements were compared with discography outcomes using paired t-test. Results: Simulation and phantom results indicated RROC is a concentration independent and pH level-dependent technique. Porcine spine study results found higher RROC value was related to lower pH level. Painful discs based on discography had significant higher RROC values than those with negative diagnosis (P < 0.05). Conclusion: RROC imaging is a promising pH level dependent MRI technique that has the potential to be a noninvasive imaging tool to detect painful IVDs in vivo. © 2014 Wiley Periodicals, Inc

MRSA infection induces lower KC and PMNs at the infection site in aged mice.

<p>Age- and gender- matched mice were infected subcutaneously (s.c.) with 10⁹ CFUs of CST9. On day 3 post-infection, the mice were sacrificed, and (A) skin lesion size, (B) skin CFUs, and (C) kidney CFUs were determined. (D) MIP-2, (E) KC, and (F) PMNs from the lesions were quantitated at 3 hours post-infection. At least 3 mice were infected for each time point for IVIS analysis. At least 3 mice in each age group were infected for the chemokine, cytokine, and PMN analyses. *<i>p</i><0.05, **<i>p</i><0.01 for comparisons of young and aged mice.</p

Mitochondrial electron transporter chain complex I plays an important role in the induction of MIP-2 in primary cells isolated from young but not old mice.

<p>(A) PMN KC at 18 h post-infection; (B) PMN MIP-2 at 18 h post-infection. (C) PMN membrane potential at 3 hours post-infection. (D) PMN membrane potential at 18 hours post-infection. The fluorescence of uninfected PMNs isolated from young mice is arbitrary set as 100%. (E–G) Primary cells were infected for 18 h with MRSA in the presence or absence of rotenone, and culture supernatants were collected for chemokine analyses. (E) PMN MIP-2, (F) PMN KC, and (G) skin fibroblast MIP-2. *<i>p</i><0.05, **<i>p</i><0.01, ***<i>p</i><0.005. #<i>p</i><0.05 for membrane potentials of young versus aged PMNs after infection with MRSA across a range of MOIs.</p

Aged mice show defective NETs formation in response to MRSA.

<p>(A) PMNs show reduced bactericidal activities against MRSA at 1 hours post infection, <i>in vitro</i> (n = 3). (B–C) PMNs from young and aged mice were incubated with or without 20 nM PMA for 1 h, and then with either MRSA at MOI of 10 or PBS for another 2 h. (B) Quantitative analysis of percentage of cells that is NETs positive (n = 3). (C) Representative image of <i>in vitro</i> NETs formation. Cells were stained for histone (Red) and DNA (Blue). (D–E) PMNs from young and aged mice were incubated with 20 nM PMA or MRSA at MOI of 10 with or without 100 ng of DNase I for 2 h. (D) Representative images of <i>in vitro</i> NETs formation with or without DNase treatment. Cells were stained for histone (Red) and DNA (Blue). (E) Quantitative analysis of percentage of cells that are NETs positive (n = 3). (F) NETs formation within skin lesions on day 3 post-infection. Infected tissues were stained for staphylococcal protein A (Spa) (green) and DNA (blue) (400×). (G) Immunochemical stain of elastase within skin lesions on day 3 post-infection. Infected tissues were stained for elastase and counter stained with hematoxylin (1000×). *<i>p</i><0.05, **<i>p</i><0.01, ***<i>p</i><0.005.</p