Comparative histopathological analysis of age-associated intervertebral disc degeneration in CD-1 and C57BL/6 mice: Anatomical and sex-based differences.

Background: Intervertebral disc (IVD) degeneration is a major contributor to back pain and disability. The cause of IVD degeneration is multifactorial, with no disease-modifying treatments. Mouse models are commonly used to study IVD degeneration; however, the effects of anatomical location, strain, and sex on the progression of age-associated degeneration are poorly understood. Methods: A longitudinal study was conducted to characterize age-, anatomical-, and sex-specific differences in IVD degeneration in two commonly used strains of mice, C57BL/6 and CD-1. Histopathological evaluation of the cervical, thoracic, lumbar, and caudal regions of mice at 6, 12, 20, and 24 months of age was conducted by two blinded observers at each IVD for the nucleus pulposus (NP), annulus fibrosus (AF), and the NP/AF boundary compartments, enabling analysis of scores by tissue compartment, summed scores for each IVD, or averaged scores for each anatomical region. Results: C57BL/6 mice displayed mild IVD degeneration until 24 months of age; at this point, the lumbar spine demonstrated the most degeneration compared to other regions. Degeneration was detected earlier in the CD-1 mice (20 months of age) in both the thoracic and lumbar spine. In CD-1 mice, moderate to severe degeneration was noted in the cervical spine at all time points assessed. In both strains, age-associated IVD degeneration in the thoracic and lumbar spine was associated with increased histopathological scores in all IVD compartments. In both strains, minimal degeneration was detected in caudal IVDs out to 24 months of age. Both C57BL/6 and CD-1 mice displayed sex-specific differences in the presentation and progression of age-associated IVD degeneration. Conclusions: These results showed that the progression and severity of age-associated degeneration in mouse models is associated with marked differences based on anatomical region, sex, and strain. This information provides a fundamental baseline characterization for users of mouse models to enable effective and appropriate experimental design, interpretation, and comparison between studies.

Emergency radiologic approach to sinus disease.

Sinus disease is commonly encountered, especially in the acute care setting. Imaging can support a diagnosis of sinusitis, help identify an etiology, and delineate intracranial and extracranial complications. Suspicion of complicated rhinosinusitis is an indication for contrast-enhanced computed tomography or magnetic resonance imaging. It is important for radiologists to be familiar with patient risk factors that predispose to uncommon but aggressive forms of sinus disease such as invasive fungal sinusitis. Lastly, many conditions, ranging from benign to malignant, can mimic rhinosinusitis clinically and on imaging. Radiologists can help by recognizing these entities and facilitating appropriate referral and follow-up. This article reviews the breadth of sinus disease commonly encountered in the emergency setting, potential complications, and mimics.

Smooth Muscle Conditions of the Chest: A Clinical, Radiologic, and Pathologic Review.

Smooth muscle conditions of the chest have diverse clinical and imaging manifestations and may involve nearly every thoracic structure. Differentiation among these conditions requires the integration of clinical, radiologic, and histopathologic data. Histologic examination in conjunction with immunohistochemistry is essential for differentiation from other spindle cell neoplastic mimics. Familiarity with these entities will ensure the inclusion of smooth muscle conditions in the differential diagnosis of thoracic soft tissue lesions and potentially guide the clinician in appropriate management. We review the clinical, imaging, and histopathologic features of thoracic smooth muscle-related conditions organized by the anatomic structures affected.

Thoracic endometriosis presenting as a catamenial hemothorax with discordant video-assisted thoracoscopic surgery.

Thoracic endometriosis is uncommon and may be overlooked, resulting in a delay in diagnosis. We describe the case of a 47-year-old woman presenting with acute onset pleuritic pain and hemothorax secondary to this rare entity. The diagnosis of thoracic endometriosis is driven by a compatible clinical history coupled with supportive imaging and immunohistochemical findings. Imaging features lack specificity, however, computed tomography and magnetic resonance imaging play an important role in identifying pleural/diaphragmatic involvement and excluding other more common diseases. Immunohistochemical pleural fluid analysis can confirm the presence of hormone receptor-positive endometrial glands and stroma. We illustrate a few potential diagnostic pitfalls, specifically the inconsistency in diagnostic yield of video-assisted thoracoscopic surgery/thoracentesis and the variable temporal association of patients' symptoms and pathology with menstruation. Prompt identification of thoracic endometriosis is important as it enables early institution of therapy and limits future complications.

Transcriptional profiling of the murine intervertebral disc and age-associated changes in the nucleus pulposus.

Purpose/Aim: The intervertebral disc (IVD) is composed of cell types whose subtle phenotypic differences allow for the formation of distinct tissues. The role of the nucleus pulposus (NP) in the initiation and progression of IVD degeneration is well established; however, the genes and pathways associated with NP degeneration are poorly characterized.Materials and Methods: Using a genetic strategy for IVD lineage-specific fluorescent reporter expression to isolate cells, gene expression and bioinformatic analysis was conducted on the murine NP at 2.5, 6, and 21 months-of-age and the annulus fibrosus (AF) at 2.5 and 6 months-of-age. A subset of differentially regulated genes was validated by qRT-PCR.Results: Transcriptome analysis identified distinct profiles of NP and AF gene expression that were remarkably consistent at 2.5 and 6 months-of-age. Prg4, Cilp, Ibsp and Comp were increased >50-fold in the AF relative to NP. The most highly enriched NP genes included Dsc3 and Cdh6, members of the cadherin superfamily, and microRNAs mir218-1 and mir490. Changes in the NP between 2.5 and 6 months-of-age were associated with up-regulation of molecular functions linked to laminin and Bmp receptor binding (including up-regulation of Bmp5 & 7), with the most up-regulated genes being Mir703, Shh, and Sfrp5. NP degeneration was associated with molecular functions linked to alpha-actinin binding (including up-regulation of Ttn & Myot) and cytoskeletal protein binding, with the overall most up-regulated genes being Rnu3a, Snora2b and Mir669h.Conclusions: This study provided insight into the phenotypes of NP and AF cells, and identified candidate pathways that may regulate degeneration.

The role of PPARγ in childhood obesity-induced fractures.

Globally, obesity is on the rise with ~ 30% of the world's population now obese, and childhood obesity is following similar trends. Childhood obesity has been associated with numerous chronic conditions, including musculoskeletal disorders. This review highlights the effects of childhood adiposity on bone density by way of analyzing clinical studies and further describing two severe skeletal conditions, slipped capital femoral epiphysis and Blount's disease. The latter half of this review discusses bone remodeling and cell types that mediate bone growth and strength, including key growth factors and transcription factors that help orchestrate this complex pathology. In particular, the transcriptional factor peroxisome proliferator-activated receptor gamma (PPARγ) is examined as it is a master regulator of adipocyte differentiation in mesenchymal stem cells (MSCs) that can also influence osteoblast populations. Obese individuals are known to have higher levels of PPARγ expression which contributes to their increased adipocyte numbers and decreased bone density. Modulating PPAR*gamma* signaling can have significant effects on adipogenesis, thereby directing MSCs down the osteoblastogenesis pathway and in turn increasing bone mineral density. Lastly, we explore the potential of PPARγ as a druggable target to decrease adiposity, increase bone density, and be a treatment for children with obesity-induced bone fractures.

Allogeneic mesenchymal stem cells for treatment of severe burn injury.

The most important determinant of survival post-burn injury is wound healing. For decades, allogeneic mesenchymal stem cells (MSCs) have been suggested as a potential treatment for severe burn injuries. This report describes a patient with a severe burn injury whose wounds did not heal with over 18 months of conventional burn care. When treated with allogeneic MSCs, wound healing accelerated with no adverse treatment complications. Wound sites showed no evidence of keloids or hypertrophic formation during a 6-year follow-up period. This therapeutic use of allogeneic MSCs for large non-healing burn wounds was deemed safe and effective and has great treatment potential.

Earlier tracheostomy and percutaneous endoscopic gastrostomy in patients with hemorrhagic stroke: associated factors and effects on hospitalization.

OBJECTIVE: Existing literature supports benefits of early tracheostomy and percutaneous endoscopic gastrostomy (PEG) in certain patient populations. The aim of this study was to review tracheostomy and PEG placement data in patients with hemorrhagic stroke in order to identify factors associated with earlier placement and to evaluate outcomes. METHODS: The authors performed a retrospective review of consecutive patients treated for hemorrhagic stroke between June 1, 2011, and June 1, 2015. Data were analyzed by logistic and multiple linear regression. RESULTS: Of 240 patients diagnosed with hemorrhagic stroke, 31.25% underwent tracheostomy and 35.83% underwent PEG tube placement. Factors significantly associated with tracheostomy and PEG included the presence of pneumonia on admission and subarachnoid hemorrhage. Earlier tracheostomy was significantly associated with shorter ICU length of stay; earlier tracheostomy and PEG placement were associated with shorter overall hospitalization. Timing of tracheostomy and PEG was not significantly associated with patient survival or the incidence of complications in this population. CONCLUSIONS: This study identified patient risk factors associated with increased likelihood of tracheostomy and PEG in patients with hemorrhagic stroke who were critically ill. Additionally, we found that the timing of tracheostomy was associated with length of ICU stay and overall hospital stay, and that the timing of PEG was associated with overall length of hospitalization. Complication rates related to tracheostomy and PEG in this population were minimal. This retrospective data set supports some benefit to earlier tracheostomy and PEG placement in this population and justifies the need for further prospective study.

Burn injury and multiple sclerosis: A retrospective case-control study.

OBJECTIVES: The purpose of this study was to determine whether having a previous diagnosis of multiple sclerosis (MS) changed acute care needs in burn-injured patients. METHODS: This was a retrospective case-control study that included adult (aged ≥18years) patients with an acute burn injury. Control patients were matched with eleven patients with a history of MS at a 4:1 ratio. Outcomes included fluid resuscitation volumes, temperature, heart rate, mean arterial pressure, in-hospital complications, and hospital length of stay (LOS). RESULTS: There were fifty-five patients included and of those, eleven had a documented history of MS. Fluid resuscitation volumes, temperature, heart rate, and mean arterial pressure were similar between groups during the resuscitation period (p>0.05). LOS was similar between both groups (12, IQR: 2-17 vs. median 16, IQR: 12-21; p=0.090). However, when normalized to % TBSA burn, patients with MS had a significantly higher median LOS/% TBSA burned (1.2, IQR: 0.7-2.0 vs. 2.1, IQR: 1.1-7.1; p=0.031). CONCLUSIONS: Patients with concurrent burn injuries and MS have a significantly longer LOS/% TBSA burn suggesting that more time is required to heal their wounds. Surprisingly, there were no other significant differences in the after the burn acute phase between these two cohorts.

Contributors to the length-of-stay trajectory in burn-injured patients.

OBJECTIVES: Burn patients have a highly variable length-of-stay (LOS) due to the complexity of the injury itself. The LOS for burn patients is estimated as one day per percent total body surface area (TBSA) burn. To focus care expectation and prognosis we aimed to identify key factors that contribute to prolonged LOS. METHODS: This was a retrospective cohort-study (2006-2016) in an adult burn-centre that included patients with ≥10% TBSA burn. Patients were stratified into expected-LOS (<2 days LOS/%TBSA) and longer-than-expected-LOS (≥2 days LOS/%TBSA). We assessed demographics, comorbidities, and in-hospital complications. Logistic regression and propensity matching was utilized. RESULTS: Of the 583 total patients, 477 had an expected-LOS whereas 106 a longer-than-expected-LOS. Non-modifiable factors such as age, 3rd degree TBSA%, inhalation injuries and comorbidities were greater in the exceeded LOS patients. Subsequent matched analysis revealed factors like number of procedures performed, days ventilated and in-hospital complications (bacteremia, pneumonia, sepsis, graft loss, and respiratory failure) were significantly increased in the longer-than-expected-LOS group. CONCLUSIONS: Progress has been made to update the conventional one day/%TBSA to better aid health care providers in giving appropriate outcomes for patients and their families and to supply intensive care units with valuable data to assess quality of care and to improve patient prognosis.

Histological and reference system for the analysis of mouse intervertebral disc.

A new scoring system based on histo-morphology of mouse intervertebral disc (IVD) was established to assess changes in different mouse models of IVD degeneration and repair. IVDs from mouse strains of different ages, transgenic mice, or models of artificially induced IVD degeneration were assessed. Morphological features consistently observed in normal, and early/later stages of degeneration were categorized into a scoring system focused on nucleus pulposus (NP) and annulus fibrosus (AF) changes. "Normal NP" exhibited a highly cellularized cell mass that decreased with natural ageing and in disc degeneration. "Normal AF" consisted of distinct concentric lamellar structures, which was disrupted in severe degeneration. NP/AF clefts indicated more severe changes. Consistent scores were obtained between experienced and new users. Altogether, our scoring system effectively differentiated IVD changes in various strains of wild-type and genetically modified mice and in induced models of IVD degeneration, and is applicable from the post-natal stage to the aged mouse. This scoring tool and reference resource addresses a pressing need in the field for studying IVD changes and cross-study comparisons in mice, and facilitates a means to normalize mouse IVD assessment between different laboratories. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:233-243, 2018.

Whole-body vibration of mice induces progressive degeneration of intervertebral discs associated with increased expression of Il-1ß and multiple matrix degrading enzymes.

OBJECTIVE: Whole-body vibration (WBV) is a popular fitness trend based on claims of increased muscle mass, weight loss and reduced joint pain. Following its original implementation as a treatment to increase bone mass in patients with osteoporosis, WBV has been incorporated into clinical practice for musculoskeletal disorders, including back pain. However, our recent studies revealed damaging effects of WBV on joint health in a murine model. In this report, we examined potential mechanisms underlying disc degeneration following exposure of mice to WBV. METHODS: Ten-week-old male mice were exposed to WBV (45 Hz, 0.3 g peak acceleration, 30 min/day, 5 days/week) for 4 weeks, 8 weeks, or 4 weeks WBV followed by 4 weeks recovery. Micro-computed tomography (micro-CT), histological, and gene expression analyses were used to assess the effects of WBV on spinal tissues. RESULTS: Exposure of mice to 4 or 8 weeks of WBV did not alter total body composition or induce significant changes in vertebral bone density. On the other hand, WBV-induced intervertebral disc (IVD) degeneration, associated with decreased disc height and degenerative changes in the annulus fibrosus (AF) that did not recover within 4 weeks after cessation of WBV. Gene expression analysis showed that WBV for 8 weeks induced expression of Mmp3, Mmp13, and Adamts5 in IVD tissues, changes preceded by increased expression of Il-1ß. CONCLUSIONS: Progressive IVD degeneration induced by WBV was associated with increased expression of Il-1ß within the IVD that preceded Mmp and Adamts gene induction. Moreover, WBV-induced IVD degeneration is not reversed following cessation of vibration.

Notochord Cells in Intervertebral Disc Development and Degeneration.

The intervertebral disc is a complex structure responsible for flexibility, multi-axial motion, and load transmission throughout the spine. Importantly, degeneration of the intervertebral disc is thought to be an initiating factor for back pain. Due to a lack of understanding of the pathways that govern disc degeneration, there are currently no disease-modifying treatments to delay or prevent degenerative disc disease. This review presents an overview of our current understanding of the developmental processes that regulate intervertebral disc formation, with particular emphasis on the role of the notochord and notochord-derived cells in disc homeostasis and how their loss can result in degeneration. We then describe the role of small animal models in understanding the development of the disc and their use to interrogate disc degeneration and associated pathologies. Finally, we highlight essential development pathways that are associated with disc degeneration and/or implicated in the reparative response of the tissue that might serve as targets for future therapeutic approaches.

Repeated exposure to high-frequency low-amplitude vibration induces degeneration of murine intervertebral discs and knee joints.

OBJECTIVE: High-frequency, low-amplitude whole-body vibration (WBV) is being used to treat a range of musculoskeletal disorders; however, there is surprisingly limited knowledge regarding its effect(s) on joint tissues. This study was undertaken to examine the effects of repeated exposure to WBV on bone and joint tissues in an in vivo mouse model. METHODS: Ten-week-old male mice were exposed to vertical sinusoidal vibration under conditions that mimic those used clinically in humans (30 minutes per day, 5 days per week, at 45 Hz with peak acceleration at 0.3g). Following WBV, skeletal tissues were examined by micro-computed tomography, histologic analysis, and immunohistochemistry, and gene expression was quantified using real-time polymerase chain reaction. RESULTS: Following 4 weeks of WBV, intervertebral discs showed histologic hallmarks of degeneration in the annulus fibrosus, disruption of collagen organization, and increased cell death. Greater Mmp3 expression in the intervertebral disc, accompanied by enhanced collagen and aggrecan degradation, was found in mice exposed to WBV as compared to controls. Examination of the knee joints after 4 weeks of WBV revealed meniscal tears and focal damage to the articular cartilage, changes resembling osteoarthritis. Moreover, mice exposed to WBV also demonstrated greater Mmp13 gene expression and enhanced matrix metalloproteinase-mediated collagen and aggrecan degradation in articular cartilage as compared to controls. No changes in trabecular bone microarchitecture or density were detected in the proximal tibia. CONCLUSION: Our experiments reveal significant negative effects of WBV on joint tissues in a mouse model. These findings suggest the need for future studies of the effects of WBV on joint health in humans.

Proteomic signature of the murine intervertebral disc.

Low back pain is the most common musculoskeletal problem and the single most common cause of disability, often attributed to degeneration of the intervertebral disc. Lack of effective treatment is directly related to our limited understanding of the pathways responsible for maintaining disc health. While transcriptional analysis has permitted initial insights into the biology of the intervertebral disc, complete proteomic characterization is required. We therefore employed liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) protein/peptide separation and mass spectrometric analyses to characterize the protein content of intervertebral discs from skeletally mature wild-type mice. A total of 1360 proteins were identified and categorized using PANTHER. Identified proteins were primarily intracellular/plasma membrane (35%), organelle (30%), macromolecular complex (10%), extracellular region (9%). Molecular function categorization resulted in three distinct categories: catalytic activity (33%), binding (molecule interactions) (29%), and structural activity (13%). To validate our list, we confirmed the presence of 14 of 20 previously identified IVD-associated markers, including matrix proteins, transcriptional regulators, and secreted proteins. Immunohistochemical analysis confirmed distinct localization patterns of select protein with the intervertebral disc. Characterization of the protein composition of healthy intervertebral disc tissue is an important first step in identifying cellular processes and pathways disrupted during aging or disease progression.

Impaired intervertebral disc development and premature disc degeneration in mice with notochord-specific deletion of CCN2.

OBJECTIVE: Currently, our ability to treat intervertebral disc (IVD) degeneration is hampered by an incomplete understanding of disc development and aging. The specific function of matricellular proteins, including CCN2, during these processes remains an enigma. The aim of this study was to determine the tissue-specific localization of CCN proteins and to characterize their role in IVD tissues during embryonic development and age-related degeneration by using a mouse model of notochord-specific CCN2 deletion. METHODS: Expression of CCN proteins was assessed in IVD tissues from wild-type mice beginning on embryonic day 15.5 to 17 months of age. Given the enrichment of CCN2 in notochord-derived tissues, we generated notochord-specific CCN2-null mice to assess the impact on the IVD structure and extracellular matrix composition. Using a combination of histologic evaluation and magnetic resonance imaging (MRI), IVD health was assessed. RESULTS: Loss of the CCN2 gene in notochord-derived cells disrupted the formation of IVDs in embryonic and newborn mice, resulting in decreased levels of aggrecan and type II collagen and concomitantly increased levels of type I collagen within the nucleus pulposus. CCN2-knockout mice also had altered expression of CCN1 (Cyr61) and CCN3 (Nov). Mirroring its role during early development, notochord-specific CCN2 deletion accelerated age-associated degeneration of IVDs. CONCLUSION: Using a notochord-specific gene targeting strategy, this study demonstrates that CCN2 expression by nucleus pulposus cells is essential to the regulation of IVD development and age-associated tissue maintenance. The ability of CCN2 to regulate the composition of the intervertebral disc suggests that it may represent an intriguing clinical target for the treatment of disc degeneration.

Acute vibration induces transient expression of anabolic genes in the murine intervertebral disc.

OBJECTIVE: Low-amplitude whole-body vibration has been adopted for the treatment of back pain and spinal disorders. However, there is limited knowledge of the impact of vibration on the intervertebral disc (IVD). This study was undertaken to examine the effects of acute vibration on anabolic and catabolic pathways in the IVD and to characterize the dependence of these changes on time and frequency. METHODS: Custom-designed platforms were developed to apply acute vibration to ex vivo and in vivo mouse models. Spinal segments (ex vivo) or mice (in vivo) were subjected to vibration (for 30 minutes at 15-90 Hz with peak acceleration at 0.3g), and IVDs were examined at specific time points after vibration. Gene expression was quantified using real-time polymerase chain reaction, and protein levels were examined by quantitative mass spectrometry and immunofluorescence. RESULTS: In the ex vivo model, acute vibration at 15 Hz induced expression of anabolic genes (aggrecan, biglycan, decorin, type I collagen, and Sox9) and suppressed expression of Mmp13, with the most pronounced changes detected 6 hours following vibration. These beneficial effects were frequency dependent and were no longer evident between 45 and 90 Hz. In vivo, the effects on anabolic gene expression were even more robust and were accompanied by decreased expression of Adamts4, Adamts5, and Mmp3. Moreover, significant increases in the protein levels of aggrecan, biglycan, decorin, and type I collagen were detected in vivo. CONCLUSION: These findings demonstrate dramatic anabolic effects of acute vibration on IVD tissue, responses that are dependent on frequency. The similarity of the in vivo and ex vivo responses indicates that at least some effects of vibration are tissue autonomous.