Periacetabular osteotomy restores the typically excessive range of motion in dysplastic hips with a spherical head

BACKGROUND Residual acetabular dysplasia is seen in combination with femoral pathomorphologies including an aspherical femoral head and valgus neck-shaft angle with high antetorsion. It is unclear how these femoral pathomorphologies affect range of motion (ROM) and impingement zones after periacetabular osteotomy. QUESTIONS/PURPOSES (1) Does periacetabular osteotomy (PAO) restore the typically excessive ROM in dysplastic hips compared with normal hips; (2) how do impingement locations differ in dysplastic hips before and after PAO compared with normal hips; (3) does a concomitant cam-type morphology adversely affect internal rotation; and (4) does a concomitant varus-derotation intertrochanteric osteotomy (IO) affect external rotation? METHODS Between January 1999 and March 2002, we performed 200 PAOs for dysplasia; of those, 27 hips (14%) met prespecified study inclusion criteria, including availability of a pre- and postoperative CT scan that included the hip and the distal femur. In general, we obtained those scans to evaluate the pre- and postoperative acetabular and femoral morphology, the degree of acetabular reorientation, and healing of the osteotomies. Three-dimensional surface models based on CT scans of 27 hips before and after PAO and 19 normal hips were created. Normal hips were obtained from a population of CT-based computer-assisted THAs using the contralateral hip after exclusion of symptomatic hips or hips with abnormal radiographic anatomy. Using validated and computerized methods, we then determined ROM (flexion/extension, internal- [IR]/external rotation [ER], adduction/abduction) and two motion patterns including the anterior (IR in flexion) and posterior (ER in extension) impingement tests. The computed impingement locations were assigned to anatomical locations of the pelvis and the femur. ROM was calculated separately for hips with (n = 13) and without (n = 14) a cam-type morphology and PAOs with (n = 9) and without (n = 18) a concomitant IO. A post hoc power analysis based on the primary research question with an alpha of 0.05 and a beta error of 0.20 revealed a minimal detectable difference of 4.6° of flexion. RESULTS After PAO, flexion, IR, and adduction/abduction did not differ from the nondysplastic control hips with the numbers available (p ranging from 0.061 to 0.867). Extension was decreased (19° ± 15°; range, -18° to 30° versus 28° ± 3°; range, 19°-30°; p = 0.017) and ER in 0° flexion was increased (25° ± 18°; range, -10° to 41° versus 38° ± 7°; range, 17°-41°; p = 0.002). Dysplastic hips had a higher prevalence of extraarticular impingement at the anteroinferior iliac spine compared with normal hips (48% [13 of 27 hips] versus 5% [one of 19 hips], p = 0.002). A PAO increased the prevalence of impingement for the femoral head from 30% (eight of 27 hips) preoperatively to 59% (16 of 27 hips) postoperatively (p = 0.027). IR in flexion was decreased in hips with a cam-type deformity compared with those with a spherical femoral head (p values from 0.002 to 0.047 for 95°-120° of flexion). A concomitant IO led to a normalization of ER in extension (eg, 37° ± 7° [range, 21°-41°] of ER in 0° of flexion in hips with concomitant IO compared with 38° ± 7° [range, 17°-41°] in nondysplastic control hips; p = 0.777). CONCLUSIONS Using computer simulation of hip ROM, we could show that the PAO has the potential to restore the typically excessive ROM in dysplastic hips. However, a PAO can increase the prevalence of secondary intraarticular impingement of the aspherical femoral head and extraarticular impingement of the anteroinferior iliac spines in flexion and internal rotation. A cam-type morphology can result in anterior impingement with restriction of IR. Additionally, a valgus hip with high antetorsion can result in posterior impingement with decreased ER in extension, which can be normalized with a varus derotation IO of the femur. However, indication of an additional IO needs to be weighed against its inherent morbidity and possible complications. The results are based on a limited number of hips with a pre- and postoperative CT scan after PAO. Future prospective studies are needed to verify the current results based on computer simulation and to test their clinical importance.

Osteogenic differentiation of bone marrow stromal cells is hindered by the presence of intervertebral disc cells.

BACKGROUND Clinical observations indicate that the presence of nucleus pulposus (NP) tissue during spinal fusion hinders the rate of disc ossification. While the underlying mechanism remains unknown, this observation could be due to incomplete removal of NP cells (NPCs) that secrete factors preventing disc calcification, such as bone morphogenetic protein (BMP) antagonists including noggin and members of the DAN (differential screening selected gene aberrative in neuroblastoma) family. METHODS Monolayer human bone marrow-derived mesenchymal stem cells (MSCs) were cocultured withNPCs and annulus fibrosus cells (AFCs) embedded in alginate for 21 days. At the end of coculture, MSCs were stained for mineral deposition by alizarin red, and relative expression of bone-related genes [Runt-related transcription factor 2, (RUNX2), Osteopontin (OPN), and Alkaline phosphatase (ALP)] and ALP activity were analyzed. Relative expression of three BMP antagonists, chordin (CHRD), gremlin (GREM1), and noggin (NOG), was determined in primary human NPCs and AFCs. These cells were also stained for Gremlin and Noggin by immunocytochemistry. RESULTS Alizarin red staining showed that MSC osteogenesis in monolayer cultures was inhibited by coculture with NPCs or AFCs. ALP activity and RT-PCR analyses confirmed these results and demonstrated inhibition of osteogenesis of MSC in the presence of disc cells. NOG was significantly up-regulated in MSCs after coculture. Relative gene expression of intervertebral disc (IVD) cells showed higher expression of GREM1 in NPCs than in AFCs. CONCLUSIONS We show that primary IVD cells inhibit osteogenesis of MSCs. BMP inhibitors NOG, GREM1 and CHRD were expressed in IVD cells. GREM1 appears to be differentially expressed in NPCs and AFCs. Our results have implications for the design and development of treatments for non-union in spinal fusion.

Repair of Annulus fibrosus with Genipin-enhanced Fibrin Hydrogel and silk membrane- fleece

Introduction Low back pain is often caused by a trauma causing disc herniation and /or disc degeneration. Although there are some promising approaches for nucleus pulposus repair, the inner tissue of the intervertebral disc (IVD) so far no treatment or repair is available for annulus fibrosus (AF) injuries. Here we aimed to develop a new method to seal and repair AF injuries by using a silk fleece composite and a genipin enhanced hydrogel. Methods Bovine (b) IVDs were harvested under aseptic conditions and kept in free swelling conditions for 24h in high-glucose DMEM containing 5% bovine serum for equilibration (1). A circular 2mm biopsy punch (Polymed Medical Center, Switzerland) was used to form a reproducible defect in the AF. For filling the defect and keeping the silk composite in place a human-derived fibrin gel (Baxter Tisseel, Switzerland) enhanced with 4.2mg/ml of the cross linker genipin (Wako Chemicals GmbH, Germany) was used. The silk composite consists of a mesh- and a membrane side (Spintec Engineering GmbH, Germany); the membrane is facing outwards to form a seal. bIVDs were cultured in vitro for 14 days either under dynamic load in a custom-built bioreactor under physiological conditions (0.2MPa load and ±2° torsion at 0.2Hz for 8h/day) or static diurnal load of 0.2MPa (2). At the end of culture discs were checked for seal failure, disc height, metabolic activity, cell death by necrosis (LDH assay), DNA content and glycosaminoglycan content. Results Silk composite maintained its position throughout the 14 days of culture under loaded conditions. Although repaired discs performed slightly lower in cell activity, DNA and GAG content were in the range of the control. Also LDH resulted in similar values compared to control discs (Fig 1). Height loss in repaired discs was in the same range as for static diurnal loaded control samples. For dynamically loaded samples the decrease was comparable to the injured, unrepaired discs. Fig 1 LDH of repaired discs compared to control disc after 24h in free swelling conditions for equilibration and first three loading cycles. Conclusions Silk-genipin-fibrin reinforced hydrogel is a promising approach to close AF defects as tested by two degree of freedom loading. In further experiments cytocompatibility of genipin has to be investigated. References 1. Chan SC, Gantenbein-Ritter B. Preparation of intact bovine tail intervertebral discs for organ culture. J Vis Exp 2012, Feb 2;60(60):e3490. 2. Walser J, Ferguson SJ, Gantenbein-Ritter B. Design of a mechanical loading device to culture intact bovine caudal motional segments of the spine under twisting motion. In: Davies J, editors. Replacing animal models: a practical guide to creating and using biomimetic alternatives. Chichester, UK: John Wiley & Sons, Ltd.; 2012. p. 89-105. Acknowledgements This project is funded by the Gerbert Rüf Stiftung project # GRS-028/13 and the Swiss National Science Project SNF #310030_153411.

NonViral Gene Delivery of Growth and Differentiation Factor 6 (GDF6) to whole bovine Intervertebral Disc

Question: Low back pain is an increasing global health problem, which is associated with intervertebral disc (IVD) damage and de- generation. Major changes occur in the nucleus pulposus (NP), with the degradation of the extracellular matrix (ECM) [1]. Further studies showed that growth factors from the transforming growth factor (TGF) and bone morphogenic proteins (BMP) family may induce chondrogenic differentiation of mesenchymal stem cells (MSC) [2]. Focusing on non-viral gene therapies and their possible translation into the clinics, we investigated if GDF6 (syn. BMP13 or CDMP2) can induce regeneration of degraded NP. We hypothesized that IVD transfected with plasmid over-expressing GDF6 also up-regulates other NP- and chondrogenic cell markers and enhances ECM deposition. Methods: Bovine IVD cells were isolated by pronase/collagenase II overnight digestion. After monolayer expansion up to passage 3, cells were transfected with the plasmid pGDF6 (RG211366, Origene, SF) or with green fluorescence protein (GFP) control using the NeonÒ transfection system (Invitrogen, Basel), both equipped with a Cy- tomegalovirus (CMV) promotor to induce over-expression. We tested a range of yet unpublished parameters for each of the primary disc cells to optimize efficiency. To test a non-viral gene therapy applied directly to 3D whole organ culture, bovine IVDs were harvested from fresh tails obtained from the abattoir within 5 h post-mortem [3]. Discs were then pre-incubated for 24 h in high glucose Dulbecco’s Modified Eagle Medium and 5 % fetal calf serum. Each disc was transfected by injection of 5 lg of plasmid GDF6 (Origene, RG211366) into the center by 25G needle and using Hamilton sy- ringe. Electroporation was performed using 2-needle array electrode or tweezertrodes; 8 pulses at 200mv/cm with an interval of 10 ms were applied using ECM830 Square Wave Electroporation System (Harvard Apparatus, MA) (Fig. 1). After transfection discs were cultured for 72 h to allow expression of GFP or GDF6. Discs were then fixed, cryosectioned and analysed by immunofluorescence against GDF6. Results: We successfully transfected bovine NP and AF cells in monolayer culture with the two plasmids using a 1,400 V, 20 ms and 2 pulses with a *25 % efficiency using 0.15 M cells and 3 lg DNA (Fig. 1). Organ IVD culture transfection revealed GFP6 positive staining in the centre of the disc using 2-needle array electrode. Results from tweezertrodes did not show any GFP posi- tive cells. Conclusions: We identified novel parameters to successfully transfect primary bovine IVD cells. For transfection of whole IVD explants electroporation parameters need to be further optimized. Acknowledgments: This study was supported by the Lindenhof Foundation ‘‘Forschung und Lehre’’ (Project no. 13-02-F). References 1. Roughly PJ (2004) Spine (Phila) 29:2691–2699 2. 3. Clarke LE, McConell JC, Sherratt MJ, Derby B, Richardson SM, Hoyland JA (2014) Arthritis Res Ther 16:R67 Chan SC, Gantenbein-Ritter B (2012) J Vis Exp 60(60):e3490

Self-motion direction discrimination in the visually impaired

Despite the close interrelation between vestibular and visual processing (e.g., vestibulo-ocular reflex), surprisingly little is known about vestibular function in visually impaired people. In this study, we investigated thresholds of passive whole-body motion discrimination (leftward vs. rightward) in nine visually impaired participants and nine age-matched sighted controls. Participants were rotated in yaw, tilted in roll, and translated along the interaural axis at two different frequencies (0.33 and 2 Hz) by means of a motion platform. Superior performance of visually impaired participants was found in the 0.33 Hz roll tilt condition. No differences were observed in the other motion conditions. Roll tilts stimulate the semicircular canals and otoliths simultaneously. The results could thus reflect a specific improvement in canal–otolith integration in the visually impaired and are consistent with the compensatory hypothesis, which implies that the visually impaired are able to compensate the absence of visual input.

Early reherniation of disk material in eleven dogs with surgically treated thoracolumbar intervertebral disk extrusion.

OBJECTIVE To report findings and outcomes of dogs with reherniation of nuclear material within 7 days of hemilaminectomy for acute thoracolumbar (TL) intervertebral disk extrusion. STUDY DESIGN Retrospective case series. ANIMALS Chondrodystrophic dogs (n = 11). METHODS Dogs with acute neurologic decline within 1 week of surgical decompression for TL disk extrusion were identified. Advanced imaging was used to document extradural spinal cord compression at the previous surgery site. Ten dogs had a 2nd decompressive surgery to remove extruded nuclear material. RESULTS All dogs had acute neurologic deterioration (average, 2 neurologic grades) 2-7 days after initial hemilaminectomy. Computed tomography (CT; n = 10) or myelography (n = 1) documented extradural spinal cord compression compatible with extruded disk material at the previous hemilaminectomy site. Dogs that had a 2nd surgical decompression improved neurologically within 24 hours and were paraparetic at discharge. The single dog that did not have decompressive surgery did not regain deep nociception during 185-day follow-up. CONCLUSIONS Early reherniation at the site of previous hemilaminectomy can produce acute deterioration of neurologic function and should be investigated with diagnostic imaging. Repeat decompressive surgery can lead to functional recovery.

Evaluation of the success of medical management for presumptive cervical intervertebral disk herniation in dogs.

OBJECTIVE To determine the success of medical management of presumptive cervical disk herniation in dogs and variables associated with treatment outcome. DESIGN Retrospective case series. ANIMALS Dogs (n=88) with presumptive cervical disk herniation. METHODS Dogs with presumptive cervical and thoracolumbar disk herniation were identified from medical records at 2 clinics and clients were mailed a questionnaire related to the success of therapy, clinical recurrence of signs, and quality of life (QOL) as interpreted by the owner. Signalment, duration and degree of neurologic dysfunction, and medication administration were determined from medical records. RESULTS Ninety-seven percent of dogs (84/87) with complete information were described as ambulatory at initial evaluation. Successful treatment was reported for 48.9% of dogs with 33% having recurrence of clinical signs and 18.1% having therapeutic failure. Bivariable logistic regression showed that non-steroidal anti-inflammatory drug (NSAID) administration was associated with success (P=.035; odds ratio [OR]=2.52). Duration of cage rest and glucocorticoid administration were not significantly associated with success or QOL. Dogs with less-severe neurologic dysfunction were more likely to have a successful outcome (OR=2.56), but this association was not significant (P=.051). CONCLUSIONS Medical management can lead to an acceptable outcome in many dogs with presumptive cervical disk herniation. Based on these data, NSAIDs should be considered as part of the therapeutic regimen. Cage rest duration and glucocorticoid administration do not appear to benefit these dogs, but this should be interpreted cautiously because of the retrospective data collection and use of client self-administered questionnaire follow-up. CLINICAL RELEVANCE These results provide insight into the success of medical management for presumptive cervical disk herniation in dogs and may allow for refinement of treatment protocols.

Evaluation of the success of medical management for presumptive thoracolumbar intervertebral disk herniation in dogs.

OBJECTIVE To determine the success of medical management of presumptive thoracolumbar disk herniation in dogs and the variables associated with treatment outcome. STUDY DESIGN Retrospective case series. ANIMALS Dogs (n=223) with presumptive thoracolumbar disk herniation. METHODS Medical records from 2 clinics were used to identify affected dogs, and owners were mailed a questionnaire about success of therapy, recurrence of clinical signs, and quality of life (QOL) as interpreted by the owner. Signalment, duration and degree of neurologic dysfunction, and medication administration were determined from medical records. RESULTS Eighty-three percent of dogs (185/223) were ambulatory at initial evaluation. Successful treatment was reported for 54.7% of dogs, with 30.9% having recurrence of clinical signs and 14.4% classified as therapeutic failures. From bivariable logistic regression, glucocorticoid administration was negatively associated with success (P=.008; odds ratio [OR]=.48) and QOL scores (P=.004; OR=.48). The duration of cage rest was not significantly associated with success or QOL. Nonambulatory dogs were more likely to have lower QOL scores (P=.01; OR=2.34). CONCLUSIONS Medical management can lead to an acceptable outcome in many dogs with presumptive thoracolumbar disk herniation. Cage rest duration does not seem to affect outcome and glucocorticoids may negatively impact success and QOL. The conclusions in this report should be interpreted cautiously because of the retrospective data collection and the use of client self-administered questionnaire follow-up. CLINICAL RELEVANCE These results provide an insight into the success of medical management for presumptive thoracolumbar disk herniation in dogs and may allow for refinement of treatment protocols.

Association between various physical factors and acute thoracolumbar intervertebral disk extrusion or protrusion in Dachshunds.

OBJECTIVE To determine whether body weight, body condition score, or various body dimensions were associated with acute thoracolumbar intervertebral disk extrusion or protrusion and whether any of these factors were associated with severity of clinical signs in Dachshunds. DESIGN Cross-sectional clinical study. ANIMALS 75 Dachshunds with (n = 39) or without (36) acute thoracolumbar intervertebral disk extrusion or protrusion. PROCEDURES Signalment, various body measurements, body weight, body condition score, and spinal cord injury grade were recorded at the time of initial examination. RESULTS Mean T1-S1 distance and median tuber calcaneus-to-patellar tendon (TC-PT) distance were significantly shorter in affected than in unaffected dogs. A 1-cm decrease in T1-S1 distance was associated with a 2.1-times greater odds of being affected, and a 1-cm decrease in TC-PT distance was associated with an 11.1-times greater odds of being affected. Results of multivariable logistic regression also indicated that affected dogs were taller at the withers and had a larger pelvic circumference than unaffected dogs, after adjusting for other body measurements. Results of ordinal logistic regression indicated that longer T1-S1 distance, taller height at the withers, and smaller pelvic circumference were associated with more severe spinal cord injury. CONCLUSIONS AND CLINICAL RELEVANCE Results suggest that certain body dimensions may be associated with acute thoracolumbar intervertebral disk extrusion or protrusion in Dachshunds and, in affected dogs, with severity of neurologic dysfunction.

Impairments of peripheral motion change detection during smooth pursuit eye movements

Introduction: In team sports the ability to use peripheral vision is essential to track a number of players and the ball. By using eye-tracking devices it was found that players either use fixations and saccades to process information on the pitch or use smooth pursuit eye movements (SPEM) to keep track of single objects (Schütz, Braun, & Gegenfurtner, 2011). However, it is assumed that peripheral vision can be used best when the gaze is stable while it is unknown whether motion changes can be equally well detected when SPEM are used especially because contrast sensitivity is reduced during SPEM (Schütz, Delipetkose, Braun, Kerzel, & Gegenfurtner, 2007). Therefore, peripheral motion change detection will be examined by contrasting a fixation condition with a SPEM condition. Methods: 13 participants (7 male, 6 female) were presented with a visual display consisting of 15 white and 1 red square. Participants were instructed to follow the red square with their eyes and press a button as soon as a white square begins to move. White square movements occurred either when the red square was still (fixation condition) or moving in a circular manner with 6 °/s (pursuit condition). The to-be-detected white square movements varied in eccentricity (4 °, 8 °, 16 °) and speed (1 °/s, 2 °/s, 4 °/s) while movement time of white squares was constant at 500 ms. 180 events should be detected in total. A Vicon-integrated eye-tracking system and a button press (1000 Hz) was used to control for eye-movements and measure detection rates and response times. Response times (ms) and missed detections (%) were measured as dependent variables and analysed with a 2 (manipulation) x 3 (eccentricity) x 3 (speed) ANOVA with repeated measures on all factors. Results: Significant response time effects were found for manipulation, F(1,12) = 224.31, p < .01, ηp2 = .95, eccentricity, F(2,24) = 56.43; p < .01, ηp2 = .83, and the interaction between the two factors, F(2,24) = 64.43; p < .01, ηp2 = .84. Response times increased as a function of eccentricity for SPEM only and were overall higher than in the fixation condition. Results further showed missed events effects for manipulation, F(1,12) = 37.14; p < .01, ηp2 = .76, eccentricity, F(2,24) = 44.90; p < .01, ηp2 = .79, the interaction between the two factors, F(2,24) = 39.52; p < .01, ηp2 = .77 and the three-way interaction manipulation x eccentricity x speed, F(2,24) = 3.01; p = .03, ηp2 = .20. While less than 2% of events were missed on average in the fixation condition as well as at 4° and 8° eccentricity in the SPEM condition, missed events increased for SPEM at 16 ° eccentricity with significantly more missed events in the 4 °/s speed condition (1 °/s: M = 34.69, SD = 20.52; 2 °/s: M = 33.34, SD = 19.40; 4 °/s: M = 39.67, SD = 19.40). Discussion: It could be shown that using SPEM impairs the ability to detect peripheral motion changes at the far periphery and that fixations not only help to detect these motion changes but also to respond faster. Due to high temporal constraints especially in team sports like soccer or basketball, fast reaction are necessary for successful anticipation and decision making. Thus, it is advised to anchor gaze at a specific location if peripheral changes (e.g. movements of other players) that require a motor response have to be detected. In contrast, SPEM should only be used if a single object, like the ball in cricket or baseball, is necessary for a successful motor response. References: Schütz, A. C., Braun, D. I., & Gegenfurtner, K. R. (2011). Eye movements and perception: A selective review. Journal of Vision, 11, 1-30. Schütz, A. C., Delipetkose, E., Braun, D. I., Kerzel, D., & Gegenfurtner, K. R. (2007). Temporal contrast sensitivity during smooth pursuit eye movements. Journal of Vision, 7, 1-15.

RASSF7 expression and its regulatory roles on apoptosis in human intervertebral disc degeneration

Apoptosis plays an important role in intervertebral disc degeneration (IDD). Overwhelming evidence indicates that RASSF7 is essential for cell growth and apoptosis. Recently, it has been noted that the JNK signaling can be negatively regulated by suppressing phosphorylated-MKK7 activation during pro-apoptosis. We aimed to investigate the RASSF7 expression level in human degenerative nucleus pulposus (NP) cells and non-degenerative NP cells and the link between RASSF7-JNK with NP cells apoptosis. We harvested NP tissues from 20 IDD patients as disease group and 8 cadaveric donors as normal controls. We detected RASSF7 expression by Real-time-PCR and western blotting. Consequently, we found that the expression of RASSF7 was higher in non-degenerative group than in degenerative group (P<0.05). Overexpression of RASSF7 in degenerative NP cells led to decreased apoptosis rate than that in scramble group (P<0.05). Collectively, our findings suggest that RASSF7 plays an important role in human IDD and RASSF7 might be potentially developed as a curative agent.

Motion-insensitive determination of B1+ amplitudes based on the bloch-siegert shift in single voxels of moving organs including the human heart.

PURPOSE To reliably determine the amplitude of the transmit radiofrequency ( B1+) field in moving organs like the liver and heart, where most current techniques are usually not feasible. METHODS B1+ field measurement based on the Bloch-Siegert shift induced by a pair of Fermi pulses in a double-triggered modified Point RESolved Spectroscopy (PRESS) sequence with motion-compensated crusher gradients has been developed. Performance of the sequence was tested in moving phantoms and in muscle, liver, and heart of six healthy volunteers each, using different arrangements of transmit/receive coils. RESULTS B1+ determination in a moving phantom was almost independent of type and amplitude of the motion and agreed well with theory. In vivo, repeated measurements led to very small coefficients of variance (CV) if the amplitude of the Fermi pulse was chosen above an appropriate level (CV in muscle 0.6%, liver 1.6%, heart 2.3% with moderate amplitude of the Fermi pulses and 1.2% with stronger Fermi pulses). CONCLUSION The proposed sequence shows a very robust determination of B1+ in a single voxel even under challenging conditions (transmission with a surface coil or measurements in the heart without breath-hold). Magn Reson Med, 2015. © 2015 Wiley Periodicals, Inc.