Quantitative lateral flow immunoassay for rapid detection and monitoring of cerebrospinal fluid leakage following incidental durotomy.

Cerebrospinal fluid (CSF) leakage due to incidental durotomy is an inherent complication of spine surgery. With appropriate treatment, complications of CSF leakage, such as headache and even meningitis, can be reduced. CSF leakage could be detected on the basis of correlated clinical symptoms; diagnosis should be based on these symptoms and appropriate imaging studies. However, the diagnosis of CSF leakage remains a challenge, especially if incidental durotomy is unrecognized during surgery; even if incidental durotomy is detected and repaired intraoperatively, the severity of the leakage and quality of the primary dural repair are difficult to evaluate postoperatively. Rapid, inexpensive, and safe methods of detecting CSF-containing samples are currently lacking; hence, the development of a point-of-care test (POCT) method to improve diagnostic efficiency is necessary. We developed a high-sensitivity lateral flow immunoassay with a stacking pad (sLFIA) for quantitative detection of ß-trace protein (BTP), a specific CSF marker. The BTP concentration in 39 clinical samples was calculated using a calibration equation for test-line intensity and evaluated by a standard laboratory method. To avoid the hook effect, we diluted each sample prior to testing. The correlation coefficient between the enzyme-linked immunosorbent assay and our BTP sLFIA method was 0.991 A 75-fold sample dilution was applied owing to the hook effect point, identified as 175 ng mL-1. We established an optimal sample-specific cutoff point at a value of 4.0 µg mL-1 for CSF leakage in subfascial drainage samples following spinal posterior decompression. The sensitivity and specificity of the BTP sLFIA method were 90% and 97%, respectively, according to a receiver operating characteristic curve analysis. In addition, clinical samples from patients who underwent primary dural repair intraoperatively were tested, and CSF leakage was successfully diagnosed using our method. Finally, the quantitation of BTP in samples collected daily provided an accurate assessment of the severity of the residual leakage. Our results demonstrate that the BTP sLFIA method possesses the potential to serve as a POCT method for screening and monitoring postoperative CSF leakage.

Assessment of Pericardial Disease with Cardiovascular MRI.

Disorders of the pericardium are common and can result in significant morbidity and mortality. Advances in multimodality imaging have enhanced our ability to diagnose and stage pericardial disease and improve our understanding of the pathophysiology of the disease. Cardiovascular MRI (CMR) can be used to define pericardial anatomy, identify the presence and extent of active pericardial inflammation, and assess the hemodynamic consequences of pericardial disease. In this way, CMR can guide the judicial use of antiinflammatory and immune modulatory medications and help with timing of pericardiectomy. CMR can also be used to diagnose congenital disorders of the pericardium. Furthermore, CMR can be used to define pericardial masses and understand their malignant potential.

Development of a multiplex and sensitive lateral flow immunoassay for the diagnosis of periprosthetic joint infection.

The diagnosis of periprosthetic joint infection (PJI) remains a challenge. However, recent studies showed that synovial fluid biomarkers have demonstrated greater diagnostic accuracy than the currently used PJI diagnostic tests. In many diagnostic tests, combining several biomarkers into panels is critical for improving diagnostic efficiency, enhancing the diagnostic precision for specific diseases, and reducing cost. In this study, we prove that combining alpha-defensin and C-reactive protein (CRP) as biomarkers possesses the potential to provide accurate PJI diagnosis. To further verify the result, we developed a multi-target lateral flow immunoassay strip (msLFIA) with staking pad design to obtain on-site rapid response for clinical diagnosis of PJI. A total of 10 synovial fluid samples were tested using the msLFIA, and the results showed that the combined measurements of synovial fluid alpha-defensin and CRP levels were consistent with those obtained from a commercial enzyme-linked immunosorbent assay kit. In addition, we developed a multi-target lateral flow immunoassay strip (msLFIA) with staking pad design to obtain on-site rapid response for clinical diagnosis of PJI, which the multi-target design is used to increase specificity and the stacking pad design is to enhance detection sensitivity. As a result, the turnaround time of the highly sensitive test can be limited from several hours to 20 min. We expect that the developed msLFIA possesses the potential for routine monitoring of PJI as a convenient, low-cost, rapid and easy to use detection device for PJI.

Postural Tachycardia Syndrome and Vasovagal Syncope: A Hidden Case of Obstructive Cardiomyopathy without Severe Septal Hypertrophy.

A 36-year-old female with symptoms of orthostatic intolerance and syncope was diagnosed with vasovagal syncope on a tilt table test and with postural tachycardia syndrome (POTS) after a repeat tilt table test. However, an echocardiogram at our institution revealed obstructive cardiomyopathy without severe septal hypertrophy, with a striking increase in left ventricular outflow tract gradient from 7 mmHg at rest to 75 mmHg during Valsalva, with a septal thickness of only 1.3 cm. Cardiac MRI showed an apically displaced multiheaded posteromedial papillary muscle with suggestion of aberrant chordal attachments to the anterior mitral leaflet contributing to systolic anterior motion of the mitral valve. She underwent surgery with reorientation of the posterior medial papillary muscle head, resection of the tethering secondary chordae to the A1 segment of the mitral valve, chordal shortening and tacking of the chordae to the A1 and A2 segments of the mitral valve, and gentle septal myectomy. After surgery, she had significant improvement in her prior symptoms. To our knowledge, this is the first reported case of obstructive cardiomyopathy without severe septal hypertrophy with abnormalities in papillary muscle and chordal attachment, in a patient diagnosed with vasovagal syncope and POTS.

The unresolved problem of beta-2 microglobulin amyloid deposits in the intervertebral discs of long-term dialysis patients.

BACKGROUND: Dialysis-related destructive spondyloarthropathy caused by beta-2 microglobulin (ß2M) amyloid deposits in intervertebral discs is a major burden for patients undergoing long-term dialysis. This study aimed to quantify the presence of ß2M amyloid deposits in the intervertebral disc tissue of such patients and analyze whether there was a significant correlation between ß2M accumulation and the duration of dialysis. METHODS: Two groups of patients who had undergone surgery for degenerative spinal pathologies were selected: the dialysis group (n = 29) with long-term dialysis and the control group (n = 10) with no renal impairment. Tissue sections were prepared from specimens of intervertebral disc tissue obtained during spinal surgery and analyzed via histological staining, including immunohistochemistry (IHC) and Congo red. RESULTS: There was a statistically significant multifold increase of ß2M expression in the disc tissue of long-term dialysis patients when compared to non-dialysis patients, as shown by both IHC (0.019 ± 0.023 µm2 vs. 0.00020 ± 0.00033 µm2, respectively; p = 0.012) and Congo red staining (0.027 ± 0.041 µm2 vs. 9.240 × 10-5 ± 5.261 × 10-5 µm2, respectively; p = 0.047). We also note a moderate strength positive correlation between the duration of dialysis and positive IHC (r = 0.39; p = 0.015) and Congo-red staining (r = 0.42; p = 0.007). CONCLUSIONS: The problem of ß2M amyloidosis in long-term dialysis patients remains unresolved even with predominant use of high-flux dialysis membranes. This highlights the insufficiency of current dialysis modalities to effectively filter ß2M.

Pericardial Effusions: Causes, Diagnosis, and Management.

The presentation of a patient with a pericardial effusion can range from an incidental finding to a life-threatening emergency. Accordingly, the causes of pericardial effusions are numerous and can generally be divided into inflammatory and non-inflammatory etiologies. For all patients with a suspected pericardial effusion, echocardiography is essential to define the location and size of an effusion. In pericardial tamponade, the hemodynamics relate to decreased pericardial compliance, ventricular interdependence, and an inspiratory decrease in the pressure gradient for left ventricular filling. Echocardiography provides insight into the pathophysiologic alterations, primarily through an assessment of chamber collapse, inferior vena cava plethora, and marked respiratory variation in mitral and tricuspid inflow. Once diagnosed, pericardiocentesis is performed in patients with tamponade, preferably with echocardiographic guidance. With a large effusion but no tamponade, pericardiocentesis is rarely needed for diagnostic purposes, though is performed if there is concern for a bacterial infection. In patients with malignancy, pericardial window is preferred given the risk for recurrence. Finally, large effusions can progress to tamponade, but can generally be followed closely until the extent of the effusion facilitates safe pericardiocentesis.

Effects of Strontium Ranelate on Spinal Interbody Fusion Surgery in an Osteoporotic Rat Model.

Osteoporosis is a bone disease that afflicts millions of people around the world, and a variety of spinal integrity issues, such as degenerative spinal stenosis and spondylolisthesis, are frequently concomitant with osteoporosis and are sometimes treated with spinal interbody fusion surgery. Previous studies have demonstrated the efficacy of strontium ranelate (SrR) treatment of osteoporosis in improving bone strength, promoting bone remodeling, and reducing the risk of fractures, but its effects on interbody fusion surgery have not been adequately investigated. SrR-treated rats subjected to interbody fusion surgery exhibited significantly higher lumbar vertebral bone mineral density after 12 weeks of treatment than rats subjected to the same surgery but not treated with SrR. Furthermore, histological and radiographic assessments showed that a greater amount of newly formed bone tissue was present and that better fusion union occurred in the SrR-treated rats than in the untreated rats. Taken together, these results show significant differences in bone mineral density, PINP level, histological score, SrR content and mechanical testing, which demonstrate a relatively moderate effect of SrR treatment on bone strength and remodeling in the specific context of recovery after an interbody fusion surgery, and suggest the potential of SrR treatment as an effective adjunct to spinal interbody fusion surgery for human patients.

Symptomatic epidural hematoma after lumbar decompression surgery.

BACKGROUND CONTEXT: Postoperative symptomatic epidural hematoma (SEH) is a serious complication of lumbar spine surgery. Despite its rarity, this uncommon complication may result in devastating neurological sequelae, including lower limb weakness. PURPOSE: A retrospective study was made to identify possible risk factors of postoperative spinal epidural hematoma by reviewing the clinical cases of this rare complication and analyzing the postoperative evaluations of patients. METHODS: From 2002 to 2010, out of 15,562 who underwent lumbar decompression procedure with/without instrumentation, 25 patients required reoperation for epidural hematoma after the initial spinal surgery. For the control group, another 75 patients were randomly selected from the pool of patients who received lumbar decompression surgery during the same period of time. The medical records of preoperative, intraoperative and postoperative factors were collected to determine possible risk factors by comparing between the cases and controls, and the postoperative evaluations of muscle power, intractable pain, saddle anesthesia, time to detection and time to evacuation were analyzed to find if there is any significant relation within the case group. Mann-Whitney U test, two-sample t test, χ (2) test and Fisher's exact test were used for statistical analysis. RESULTS: The incidence of postoperative symptomatic epidural hematoma is 0.16%. After the initial procedure, 20 (80%) patients developed progressive decrease in muscle power (MP ≤ 3), 14 (56%) patients had intractable pain (VAS ≥ 7), and 19 (76%) patients had saddle anesthesia. Preoperative diastolic blood pressure, intraoperative use of gelfoam for dura coverage and postoperative drain output were statistically significant risk factors (p < 0.01). Within the SEH case group, postoperative symptom of decreased muscle power had significant relation with blood loss, laminectomy level and fusion level (p = 0.016, 0.021, 0.010). If the symptom of decreased muscle power or perianal anesthesia was not improved after hematoma evacuation, there was a tendency for permanent leg weakness after 1-year follow-up (p = 0.001, 0.003). CONCLUSIONS: The findings suggest that preoperative diastolic blood pressure, intraoperative use of gelfoam for dura coverage and postoperative drain output are risk factors for symptomatic epidural hematoma after lumbar decompression surgery. Major blood loss and multilevel surgical procedure could result in poor recovery of muscle power. After spine decompression surgery, early detection and evacuation of hematoma are the key to avoid neurologic deterioration and have better clinical outcomes.

Increased sulfatase 1 gene expression in degenerative intervertebral disc cells.

Sulfatase 1 (SULF1) plays a key role in cell signaling involving in cell growth, differentiation, proliferation, and migration. Abnormal SULF1 expression has been implicated in the development of various cancers and diseases of the skeletal and nervous systems. The present study aims to examine the difference in SULF1 expression between degenerative and non-degenerative intervertebral discs (IVDs) to provide an enhanced understanding of disc degeneration. Degenerative and non-degenerative disc tissues were surgically harvested from patients and experimental rats. Disc degeneration-specific genes were identified by microarray analysis. The gene expression of SULF1 was measured by sulfatase assay, reverse transcription-polymerase chain reaction (RT-PCR), real-time RT-PCR, and western blotting. Also, the presence of SULF1 in human and rat discs was confirmed by immunohistochemistry. More specifically in human cells, an increase of SULF1 gene expression was observed in degenerative cells at both mRNA and protein levels, as well as in time- and dose-dependent manner in response to TNF-α treatment. Increased staining of SULF1 was detected in degenerative discs compared to non-degenerative discs for humans and rats. These findings show an upregulation of SULF1 in degenerative discs for the first time, and suggest that there is a link between SULF1 and disc degeneration.

Noncontact, low-frequency ultrasound therapy enhances neovascularization and wound healing in diabetic mice.

BACKGROUND: Chronic wounds are a major source of morbidity for patients and represent a significant health burden. Implementing noninvasive techniques that accelerate healing of these wounds would provide great benefit. Ultrasound appears to be an effective modality for the treatment of chronic wounds in humans. MIST Therapy is a noncontact, low-frequency ultrasound treatment delivered through a saline mist. A variety of mechanisms have been proposed to explain the efficacy of ultrasound therapy, but the underlying molecular and cellular pathways impacted by this technique remain unclear. The in vivo effect of noncontact, low-frequency ultrasound was therefore examined in a humanized excisional wound model. METHODS: The treatment group received noncontact, low-frequency ultrasound therapy three times per week, whereas the control group received a standard dressing change. Wounds were photographed at regular intervals to calculate healing kinetics. Wound tissue was harvested and processed for histology, quantitative polymerase chain reaction, and enzyme-linked immunosorbent assay. RESULTS: The MIST group demonstrated significantly accelerated wound healing, with 17.3 days to wound closure compared with 24 days in the controls (p < 0.05). This improvement became evident by day 9, with healing evidenced by significantly decreased mean wound area relative to original size (68 percent versus 80 percent; p < 0.01). Expression of markers of neovascularization (stromal cell-derived factor 1, vascular endothelial growth factor, and CD31) was also increased in the wound beds of noncontact, low-frequency ultrasound-treated mice compared with controls. CONCLUSION: Noncontact, low-frequency ultrasound treatment improves neovascularization and wound closure rates in excisional wounds for diabetic mice, likely because of the stimulated release of angiogenic factors.

Downregulation of protein disulfide isomerase in sepsis and its role in tumor necrosis factor-alpha release.

INTRODUCTION: Protein disulfide isomerase (PDI) is an important factor for the protein modification step in the post-translational event. PDI plays an essential role in cell survival under various stress conditions. It has been reported that PDI can serve as a negative regulator of nuclear factor-kappa-B (NF-kappaB) and that it can inhibit lipopolysaccharide (LPS)-induced proinflammatory cytokine production in macrophages. Thus, PDI may be an intracellular anti-inflammatory molecule. Although we have previously shown that Kupffer cell-derived proinflammatory cytokines cause liver injury in sepsis, the effect of sepsis on PDI expression as well as the effect of PDI inhibition on cytokine production have not been investigated. We therefore hypothesized that sepsis downregulates PDI expression and that the inhibition of PDI promotes proinflammatory cytokine production. METHOD: Adult male rats were subjected to sepsis by cecal ligation and puncture (CLP) or endotoxemia (continuous infusion of 1 microg/kg body weight LPS by an osmotic pump) for 20 hours. Hepatic tissues were collected and PDI gene expression was determined. In additional experiments, cells from a macrophage-like cell line, RAW 264.7, were treated with 100 ng/mL LPS for 4 hours and protein expressions were measured. RAW 264.7 cells were also treated with bacitracin, a specific PDI inhibitor, for 24 hours, and tumor necrosis factor-alpha (TNF-alpha) gene and protein expression as well as its release in the cell supernatant were determined. To further confirm the beneficial effect of PDI in sepsis, RAW 264.7 cells were transfected with PDI short interfering RNA (siRNA) and PDI gene expression and TNF-alpha release were measured by quantitative polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. RESULTS: PDI gene expression was significantly decreased by 28% and 69% at 20 hours after CLP or LPS infusion, respectively. LPS also decreased PDI protein expression by 33% in RAW 264.7 cells. Incubation of RAW 264.7 cells with bacitracin significantly increased TNF-alpha gene expression and TNF-alpha release as well as its cellular levels in a dose-dependent manner. Transfection of RAW 264.7 cells with PDI siRNA produced an average 36.8% inhibition of the PDI gene expression. This downregulation was correlated with a 3.19-fold increase in TNF-alpha release into the cell supernatant. CONCLUSION: Taken together, these results suggest that downregulation of PDI by sepsis significantly increases proinflammatory cytokine production. Thus, prevention of PDI downregulation in sepsis may be a novel approach to attenuate hyperinflammation and to reduce tissue injury under such conditions.