Cross-α/ß polymorphism of PSMα3 fibrils.

The formation of ordered cross-ß amyloid protein aggregates is associated with a variety of human disorders. While conventional infrared methods serve as sensitive reporters of the presence of these amyloids, the recently discovered amyloid secondary structure of cross-α fibrils presents new questions and challenges. Herein, we report results using Fourier transform infrared spectroscopy and two-dimensional infrared spectroscopy to monitor the aggregation of one such cross-α-forming peptide, phenol soluble modulin alpha 3 (PSMα3). Phenol soluble modulins (PSMs) are involved in the formation and stabilization of Staphylococcus aureus biofilms, making sensitive methods of detecting and characterizing these fibrils a pressing need. Our experimental data coupled with spectroscopic simulations reveals the simultaneous presence of cross-α and cross-ß polymorphs within samples of PSMα3 fibrils. We also report a new spectroscopic feature indicative of cross-α fibrils.

Timing-dependent synergies between motor cortex and posterior spinal stimulation in humans.

Volitional movement requires descending input from the motor cortex and sensory feedback through the spinal cord. We previously developed a paired brain and spinal electrical stimulation approach in rats that relies on convergence of the descending motor and spinal sensory stimuli in the cervical cord. This approach strengthened sensorimotor circuits and improved volitional movement through associative plasticity. In humans, it is not known whether posterior epidural spinal cord stimulation targeted at the sensorimotor interface or anterior epidural spinal cord stimulation targeted within the motor system is effective at facilitating brain evoked responses. In 59 individuals undergoing elective cervical spine decompression surgery, the motor cortex was stimulated with scalp electrodes and the spinal cord was stimulated with epidural electrodes, with muscle responses being recorded in arm and leg muscles. Spinal electrodes were placed either posteriorly or anteriorly, and the interval between cortex and spinal cord stimulation was varied. Pairing stimulation between the motor cortex and spinal sensory (posterior) but not spinal motor (anterior) stimulation produced motor evoked potentials that were over five times larger than brain stimulation alone. This strong augmentation occurred only when descending motor and spinal afferent stimuli were timed to converge in the spinal cord. Paired stimulation also increased the selectivity of muscle responses relative to unpaired brain or spinal cord stimulation. Finally, clinical signs suggest that facilitation was observed in both injured and uninjured segments of the spinal cord. The large effect size of this paired stimulation makes it a promising candidate for therapeutic neuromodulation. KEY POINTS: Pairs of stimuli designed to alter nervous system function typically target the motor system, or one targets the sensory system and the other targets the motor system for convergence in cortex. In humans undergoing clinically indicated surgery, we tested paired brain and spinal cord stimulation that we developed in rats aiming to target sensorimotor convergence in the cervical cord. Arm and hand muscle responses to paired sensorimotor stimulation were more than five times larger than brain or spinal cord stimulation alone when applied to the posterior but not anterior spinal cord. Arm and hand muscle responses to paired stimulation were more selective for targeted muscles than the brain- or spinal-only conditions, especially at latencies that produced the strongest effects of paired stimulation. Measures of clinical evidence of compression were only weakly related to the paired stimulation effect, suggesting that it could be applied as therapy in people affected by disorders of the central nervous system.

Ultra-rapid Electrophilic Cysteine Arylation.

Rapid bond-forming reactions are crucial for efficient bioconjugation. We describe a simple and practical strategy for facilitating ultra-rapid electrophilic cysteine arylation. Using a variety of sulfone-activated pyridinium salts, this uncatalyzed reaction proceeds with exceptionally high rate constants, ranging from 9800 to 320,000 M-1·s-1, in pH 7.0 aqueous buffer at 25 °C. Such reactions allow for stoichiometric bioconjugation of micromolar cysteine within minutes or even seconds. Even though the arylation is extremely fast, the chemistry exhibits excellent selectivity, thus furnishing functionalized peptides and proteins with both high conversion and purity.

Intraoperative electrical stimulation of the human dorsal spinal cord reveals a map of arm and hand muscle responses.

Although epidural stimulation of the lumbar spinal cord has emerged as a powerful modality for recovery of movement, how it should be targeted to the cervical spinal cord to activate arm and hand muscles is not well understood, particularly in humans. We sought to map muscle responses to posterior epidural cervical spinal cord stimulation in humans. We hypothesized that lateral stimulation over the dorsal root entry zone would be most effective and responses would be strongest in the muscles innervated by the stimulated segment. Twenty-six people undergoing clinically indicated cervical spine surgery consented to mapping of motor responses. During surgery, stimulation was performed in midline and lateral positions at multiple exposed segments; six arm and three leg muscles were recorded on each side of the body. Across all segments and muscles tested, lateral stimulation produced stronger muscle responses than midline despite similar latency and shape of responses. Muscles innervated at a cervical segment had the largest responses from stimulation at that segment, but responses were also observed in muscles innervated at other cervical segments and in leg muscles. The cervical responses were clustered in rostral (C4-C6) and caudal (C7-T1) cervical segments. Strong responses to lateral stimulation are likely due to the proximity of stimulation to afferent axons. Small changes in response sizes to stimulation of adjacent cervical segments argue for local circuit integration, and distant muscle responses suggest activation of long propriospinal connections. This map can help guide cervical stimulation to improve arm and hand function.NEW & NOTEWORTHY A map of muscle responses to cervical epidural stimulation during clinically indicated surgery revealed strongest activation when stimulating laterally compared to midline and revealed differences to be weaker than expected across different segments. In contrast, waveform shapes and latencies were most similar when stimulating midline and laterally, indicating activation of overlapping circuitry. Thus, a map of the cervical spinal cord reveals organization and may help guide stimulation to activate arm and hand muscles strongly and selectively.

Decompression alone versus decompression with fusion in patients with lumbar spinal stenosis with degenerative spondylolisthesis: a systematic review and meta-analysis.

INTRODUCTION: Surgical decompression is standard care in the treatment of degenerative spondylolisthesis in patients with symptomatic lumbar spinal stenosis, but there remains controversy over the benefits of adding fusion. The persistent lack of consensus on this matter and the availability of new data warrants a contemporary systematic review and meta-analysis of the literature. METHODS: Multiple online databases were systematically searched up to October 2022 for randomized controlled trials (RCTs) and prospective studies comparing outcomes of decompression alone versus decompression with fusion for lumbar spinal stenosis in patients with degenerative spondylolisthesis. Primary outcome was the Oswestry Disability Index. Secondary outcomes included leg and back pain, surgical outcomes, and radiological outcomes. Pooled effect estimates were calculated and presented as mean differences (MD) with their 95% confidence intervals (CI) at two-year follow-up. RESULTS: Of the identified 2403 studies, eventually five RCTs and two prospective studies were included. Overall, most studies had a low or unclear risk of selection bias and most studies were focused on low grade degenerative spondylolisthesis. All patient-reported outcomes showed low statistical heterogeneity. Overall, there was high-quality evidence suggesting no difference in functionality at two years of follow-up (MD - 0.31, 95% CI - 3.81 to 3.19). Furthermore, there was high-quality evidence of no difference in leg pain (MD - 1.79, 95% CI - 5.08 to 1.50) or back pain (MD - 2.54, 95% CI - 6.76 to 1.67) between patients undergoing decompression vs. decompression with fusion. Pooled surgical outcomes showed less blood loss after decompression only, shorter length of hospital stay, and a similar reoperation rate compared to decompression with fusion. CONCLUSION: Based on the current literature, there is high-quality evidence of no difference in functionality after decompression alone compared to decompression with fusion in patients with degenerative lumbar spondylolisthesis at 2 years of follow-up. Further studies should focus on long-term comparative outcomes, health economic evaluations, and identifying those patients that may benefit more from decompression with fusion instead of decompression alone. This review was registered at Prospero (CRD42021291603).

Telemedical Support Using Smartphones for Spine Surgery in Low- and Middle-Income Countries.

Objective: Low- and middle-income countries (LMICs) face many challenges compared to industrialized nations, most notably in regard to the health care system. Patients often have to travel long distances to receive medical care with few reliable transportation mechanisms. In time-critical emergencies, this is a significant disadvantage. One specialty that is particularly affected by this is spine surgery. Within this field, traumatic injuries and acutely compressive pathologies are often time-critical. Increasing global networking capabilities through internet access offers the possibility for telemedical support in remote regions. Recently, high-performance cameras and processors became available in commercially available smartphones. Due to their wide availability and ease of use, this could provide a unique opportunity to offer telemedical support in LMICs. Methods: We conducted a feasibility study with a neurosurgical institution in east Africa. To ensure telemedical support, a commercially available smartphone was selected as the experimental hardware. Preoperatively, resolution, contrast, brightness, and color reproduction were assessed under theoretical conditions using a test chart. Intraoperatively, the image quality was assessed under different conditions. In the first step, the instrumentation table was displayed, and the mentor surgeon marked an instrument that the mentee surgeon should recognize correctly. In the next evaluation step, the surgical field was shown on film and the mentor surgeon marked an anatomical structure, and in the last evaluation step, the screen of the X-ray machine was captured, and the mentor surgeon again marked an anatomical structure. Subjective image quality was rated by two independent reviewers using the similar modified Likert scale as before on a scale of 1-5, with 1 indicating inadequate quality and 5 indicating excellent quality. Results: The image quality during the video calls was rated as sufficient overall. When evaluating the test charts, a quality of 97% ± 5 on average was found for the chart with the white background and a quality of 84% ± 5 on average for the chart with the black background. The color reproduction, the contrast, and the reproduction of brightness were rated excellent. Intraoperatively, the visualization of the instrument table was also rated excellent. Visualization of the operative site was rated 1.5 ± 0.5 on average and it was not possible to recognize relevant anatomical structures with the required confidence for surgical procedures. Image quality of the X-ray screen was rated 1.5 ± 0.9 on average. Conclusion: Current generation smartphones have high imaging performance, high computing power, and excellent connectivity. However, relevant anatomical structures during spine surgery procedures and on the X-ray screen in the operating room could not be identified with reliability to provide adequate surgical support. Nevertheless, our study showed the potential in smartphones supporting surgical procedures in LMICs, which could be helpful in other surgical fields.

Rapid Electrophilic Cysteine Arylation with Pyridinium Salts.

Here, we present a series of fluorinated cationic reagents that enable rapid arylation of cysteine under mild conditions compatible with proteins and peptides. The highly polarized C-F bond and attractive nucleophile-electrophile Coulombic interactions substantially accelerate cysteine arylation, leading to unusually high rate constants on the order of 100 M-1·s-1 and allowing for equimolar labeling of substrates at micromolar concentrations. The synthetic modularity of this approach promotes the direct coupling of structurally diverse phenol-containing functional motifs to cysteine residues of biomacromolecules with high efficiency. This user-friendly chemistry enables fast bond formation between commonly used bioconjugation partners, thus greatly streamlining the synthetic chemistry workflow, and can be easily developed as convenient kits for chemical biology and medicinal chemistry applications.

Feasibility of smart glasses in supporting spinal surgical procedures in low- and middle-income countries: experiences from East Africa.

OBJECTIVE: Telemedicine technology has been developed to allow surgeons in countries with limited resources to access expert technical guidance during surgical procedures. The authors report their initial experience using state-of-the-art wearable smart glasses with wireless capability to transmit intraoperative video content during spine surgery from sub-Saharan Africa to experts in the US. METHODS: A novel smart glasses system with integrated camera and microphone was worn by a spine surgeon in Dar es Salaam, Tanzania, during 3 scoliosis correction surgeries. The images were transmitted wirelessly through a compatible software system to a computer viewed by a group of fellowship-trained spine surgeons in New York City. Visual clarity was determined using a modified Snellen chart, and a percentage score was determined on the smallest line that could be read from the 8-line chart on white and black backgrounds. A 1- to 5-point scale (from 1 = unrecognizable to 5 = optimal clarity) was used to score other visual metrics assessed using a color test card including hue, contrast, and brightness. The same scoring system was used by the group to reach a consensus on visual quality of 3 intraoperative points including instruments, radiographs (ability to see pedicle screws relative to bony anatomy), and intraoperative surgical field (ability to identify bony landmarks such as transverse processes, pedicle screw starting point, laminar edge). RESULTS: All surgeries accomplished the defined goals safely with no intraoperative complications. The average download and upload connection speeds achieved in Dar es Salaam were 45.21 and 58.89 Mbps, respectively. Visual clarity with the modified white and black Snellen chart was 70.8% and 62.5%, respectively. The average scores for hue, contrast, and brightness were 2.67, 3.33, and 2.67, respectively. Visualization quality of instruments, radiographs, and intraoperative surgical field were 3.67, 1, and 1, respectively. CONCLUSIONS: Application of smart glasses for telemedicine offers a promising tool for surgical education and remote training, especially in low- and middle-income countries. However, this study highlights some limitations of this technology, including optical resolution, intraoperative lighting, and internet connection challenges. With continued collaboration between clinicians and industry, future iterations of smart glasses technology will need to address these issues to stimulate robust clinical utilization.

Mechanisms Underlying Long-Term Synaptic Zinc Plasticity at Mouse Dorsal Cochlear Nucleus Glutamatergic Synapses.

In many brain areas, such as the neocortex, limbic structures, and auditory brainstem, synaptic zinc is released from presynaptic terminals to modulate neurotransmission. As such, synaptic zinc signaling modulates sensory processing and enhances acuity for discrimination of different sensory stimuli. Whereas sensory experience causes long-term changes in synaptic zinc signaling, the mechanisms underlying this long-term synaptic zinc plasticity remain unknown. To study these mechanisms in male and female mice, we used in vitro and in vivo models of zinc plasticity observed at the zinc-rich glutamatergic dorsal cochlear nucleus (DCN) parallel fiber synapses onto cartwheel cells. High-frequency stimulation of DCN parallel fiber synapses induced LTD of synaptic zinc signaling (Z-LTD), evidenced by reduced zinc-mediated inhibition of EPSCs. Low-frequency stimulation induced LTP of synaptic zinc signaling (Z-LTP), evidenced by enhanced zinc-mediated inhibition of EPSCs. Pharmacological manipulations of Group 1 metabotropic glutamate receptors (G1 mGluRs) demonstrated that G1 mGluR activation is necessary and sufficient for inducing Z-LTD and Z-LTP. Pharmacological manipulations of Ca2+ dynamics indicated that rises in postsynaptic Ca2+ are necessary and sufficient for Z-LTD induction. Electrophysiological measurements assessing postsynaptic expression mechanisms, and imaging studies with a ratiometric extracellular zinc sensor probing zinc release, supported that Z-LTD is expressed, at least in part, via reductions in presynaptic zinc release. Finally, exposure of mice to loud sound caused G1 mGluR-dependent Z-LTD at DCN parallel fiber synapses, thus validating our in vitro results. Together, our results reveal a novel mechanism underlying activity- and experience-dependent plasticity of synaptic zinc signaling.SIGNIFICANCE STATEMENT In the neocortex, limbic structures, and auditory brainstem, glutamatergic nerve terminals corelease zinc to modulate excitatory neurotransmission and sensory responses. Moreover, sensory experience causes bidirectional, long-term changes in synaptic zinc signaling. However, the mechanisms of this long-term synaptic zinc plasticity remain unknown. Here, we identified a novel Group 1 mGluR-dependent mechanism that causes bidirectional, long-term changes in synaptic zinc signaling. Our results highlight new mechanisms of brain adaptation during sensory processing, and potentially point to mechanisms of disorders associated with pathologic adaptation, such as tinnitus.

Impact of the COVID-19 pandemic on neuro-oncology outcomes.

INTRODUCTION: The Coronavirus disease 2019 (COVID-19) pandemic has uprooted healthcare systems worldwide, disrupting care and increasing dependence on alternative forms of health care delivery. It is yet to be determined how the pandemic affected neuro-oncology patient outcomes, given that the majority of even "elective" neurosurgical oncology procedures are time-sensitive. This study quantifies changes in neuro-oncological care during the height of the pandemic and investigates patient outcomes in 2020 compared to a historical control. METHODS: We performed a retrospective review of patients with malignant brain tumor diagnoses who were seen at our institution between March 13 and May 1 of 2020 and 2019. Alterations in care, including shift from in-person to telehealth, delays in evaluation and intervention, and treatment modifications were evaluated. These variables were analyzed with respect to brain tumor control and mortality. RESULTS: 112 patients from 2020 to 166 patients from 2019 were included. There was no significant difference in outcomes between the cohorts, despite significantly more treatment delays (p = 0.0160) and use of telehealth (p < 0.0001) in 2020. Patients in 2020 who utilized telehealth visits had significantly more stable tumor control than those who had office visits (p = 0.0124), consistent with appropriate use of in-person visits for patients with progression. CONCLUSIONS: Our study showed that use of telehealth and selective alterations in neuro-oncological care during the COVID-19 pandemic did not lead to adverse patient outcomes. This suggests that adaptive physician-led changes were successful and may inform management during the ongoing pandemic, especially with the emergence of the Delta variant.

The effect of surgery on radiation necrosis in irradiated brain metastases: extent of resection and long-term clinical and radiographic outcomes.

OBJECTIVE: Radiation therapy is a cornerstone of brain metastasis (BrM) management but carries the risk of radiation necrosis (RN), which can require resection for palliation or diagnosis. We sought to determine the relationship between extent of resection (EOR) of pathologically-confirmed RN and postoperative radiographic and symptomatic outcomes. METHODS: A single-center retrospective review was performed at an NCI-designated Comprehensive Cancer Center to identify all surgically-resected, previously-irradiated necrotic BrM without admixed recurrent malignancy from 2003 to 2018. Clinical, pathologic and radiographic parameters were collected. Volumetric analysis determined EOR and longitudinally evaluated perilesional T2-FLAIR signal preoperatively, postoperatively, and at 3-, 6-, 12-, and 24-months postoperatively when available. Rates of time to 50% T2-FLAIR reduction was calculated using cumulative incidence in the competing risks setting with last follow-up and death as competing events. The Spearman method was used to calculate correlation coefficients, and continuous variables for T2-FLAIR signal change, including EOR, were compared across groups. RESULTS: Forty-six patients were included. Most underwent prior stereotactic radiosurgery with or without whole-brain irradiation (N = 42, 91%). Twenty-seven operations resulted in gross-total resection (59%; GTR). For the full cohort, T2-FLAIR edema decreased by a mean of 78% by 6 months postoperatively that was durable to last follow-up (p < 0.05). EOR correlated with edema reduction at last follow-up, with significantly greater T2-FLAIR reduction with GTR versus subtotal resection (p < 0.05). Among surviving patients, a significant proportion were able to decrease their steroid use: steroid-dependency decreased from 54% preoperatively to 15% at 12 months postoperatively (p = 0.001). CONCLUSIONS: RN resection conferred both durable T2-FLAIR reduction, which correlated with EOR; and reduced steroid dependency.

Multilevel anterior cervical osteotomies with uncinatectomies to correct a fixed kyphotic deformity associated with ankylosing spondylitis: technical note and operative video.

Ankylosing spondylitis (AS) is an inflammatory disorder leading to ossification of joints and ligaments, resulting in autofusion throughout the spinal column. In patients with fixed, kyphotic cervical deformities, which cause an impaired horizontal gaze and severe neck pain, surgical intervention is warranted. Although several articles have described the anterior and/or posterior surgical treatments used to address the fixed kyphosis, few sources present the key operative steps and technical nuances. The purpose of this technical report was to provide detailed surgical steps, representative photographs, and an operative video demonstrating multilevel anterior cervical osteotomies, uncinatectomies, and a posterior osteotomy for the correction of a fixed cervical deformity secondary to AS.

Native Zinc Catalyzes Selective and Traceless Release of Small Molecules in ß-Cells.

The loss of insulin-producing ß-cells is the central pathological event in type 1 and 2 diabetes, which has led to efforts to identify molecules to promote ß-cell proliferation, protection, and imaging. However, the lack of ß-cell specificity of these molecules jeopardizes their therapeutic potential. A general platform for selective release of small-molecule cargoes in ß-cells over other islet cells ex vivo or other cell-types in an organismal context will be immensely valuable in advancing diabetes research and therapeutic development. Here, we leverage the unusually high Zn(II) concentration in ß-cells to develop a Zn(II)-based prodrug system to selectively and tracelessly deliver bioactive small molecules and fluorophores to ß-cells. The Zn(II)-targeting mechanism enriches the inactive cargo in ß-cells as compared to other pancreatic cells; importantly, Zn(II)-mediated hydrolysis triggers cargo activation. This prodrug system, with modular components that allow for fine-tuning selectivity, should enable the safer and more effective targeting of ß-cells.

Photoactivatable Sensors for Detecting Mobile Zinc.

Fluorescent sensors for mobile zinc are valuable for studying complex biological systems. Because these sensors typically bind zinc rapidly and tightly, there has been little temporal control over the activity of the probe after its application to a sample. The ability to control the activity of a zinc sensor in vivo during imaging experiments would greatly improve the time resolution of the measurement. Here, we describe photoactivatable zinc sensors that can be triggered with short pulses of UV light. These probes are prepared by functionalizing a zinc sensor with protecting groups that render the probe insensitive to metal ions. Photoinduced removal of the protecting groups restores the binding site, allowing for zinc-responsive changes in fluorescence that can be observed in live cells and tissues.

Challenges and Opportunities in Brain Bioinorganic Chemistry.

Metal ions play critical roles in neurotransmission, memory formation, and sensory perception. Understanding the molecular details of these processes is the Holy Grail of metalloneurochemistry. Here we describe five challenges for collaborative teams of chemists, biologists, and neuroscientists to help make this dream a reality.

AMPA receptor inhibition by synaptically released zinc.

The vast amount of fast excitatory neurotransmission in the mammalian central nervous system is mediated by AMPA-subtype glutamate receptors (AMPARs). As a result, AMPAR-mediated synaptic transmission is implicated in nearly all aspects of brain development, function, and plasticity. Despite the central role of AMPARs in neurobiology, the fine-tuning of synaptic AMPA responses by endogenous modulators remains poorly understood. Here we provide evidence that endogenous zinc, released by single presynaptic action potentials, inhibits synaptic AMPA currents in the dorsal cochlear nucleus (DCN) and hippocampus. Exposure to loud sound reduces presynaptic zinc levels in the DCN and abolishes zinc inhibition, implicating zinc in experience-dependent AMPAR synaptic plasticity. Our results establish zinc as an activity-dependent, endogenous modulator of AMPARs that tunes fast excitatory neurotransmission and plasticity in glutamatergic synapses.

CF2H, a Hydrogen Bond Donor.

The CF2H group, a potential surrogate for the OH group, can act as an unusual hydrogen bond donor, as confirmed by crystallographic, spectroscopic, and computational methods. Here, we demonstrate the bioisosterism of the OH and CF2H groups and the important roles of CF2-H···O hydrogen bonds in influencing intermolecular interactions and conformational preferences. Experimental evidence, corroborated by theory, reveals the distinctive nature of CF2H hydrogen bonding interactions relative to their normal OH hydrogen bonding counterparts.

Metalloneurochemistry and the Pierian Spring: 'Shallow Draughts Intoxicate the Brain'.

Metal ions perform critical and diverse functions in nervous system physiology and pathology. The field of metalloneurochemistry aims to understand the mechanistic bases for these varied roles at the molecular level. Here, we review several areas of research that illustrate progress toward achieving this ambitious goal and identify key challenges for the future. We examine the use of lithium as a mood stabilizer, the roles of mobile zinc and copper in the synapse, the interplay of nitric oxide and metals in retrograde signaling, and the regulation of iron homeostasis in the brain. These topics were chosen to demonstrate not only the breadth of the field, but also to highlight opportunities for discovery by studying such complex systems in greater detail. We are beginning to uncover the principles by which receptors and transmitters utilize metal ions to modulate neurotransmission. These advances have revealed exciting new insights into the intricate mechanisms that give rise to learning, memory, and sensory perception, while opening many new avenues for further exploration.

Comparison of pain outcomes between two anti-GD2 antibodies in patients with neuroblastoma.

BACKGROUND: Addition of anti-GD2 antibody ch14.18 to the treatment of neuroblastoma has improved outcomes. The most common side effect of ch14.18 is neuropathic pain, which may in part be complement-mediated. Hu14.18K322A is a humanized anti-GD2 antibody designed to diminish complement activation and induce less pain. We compare the pain outcomes in patients treated with ch14.18 and those treated with hu14.18K322A, and explore dose-dependent relationships between pain scores, opioid requirements, and complement levels in patients treated with hu14.18K322A. PROCEDURE: Opioid (morphine equivalent mg/kg) and anxiolytic requirements during course 1 (4 days) in patients treated with hu14.18K322A and ch14.18 were reviewed. Correlations between antibody dose and pain scores, opioid requirements, and complement levels were examined for patients receiving hu14.18K322A. RESULTS: Patients treated with hu14.18K322A (n = 19) had lower opioid requirements than those who received ch14.18 (n = 9). The differences in median opioid requirements (mg/kg) were statistically significant for the overall course (1.57 vs. 2.41, P = 0.019) as well as for Days 3 (0.34 vs. 0.65, P = 0.005), and 4 (0.32 vs. 0.64, P = 0.010). No difference in anxiolytic use was observed between the two groups. In the group treated with hu14.18K322A, we found a positive correlation between antibody dose administered and pain scores, but no correlation between antibody dose and opioid requirements or changes in complement levels. CONCLUSIONS: In this retrospective analysis, hu14.18K322A induced less pain than ch14.18 based on opioid requirements. Pediatr Blood Cancer 2015;62:224-228. © 2014 Wiley Periodicals, Inc.

Thioamide-based fluorescent protease sensors.

Thioamide quenchers can be paired with compact fluorophores to design "turn-on" fluorescent protease substrates. We have used this method to study a variety of serine-, cysteine-, carboxyl-, and metallo-proteases, including trypsin, chymotrypsin, pepsin, thermolysin, papain, and calpain. Since thioamides quench some fluorophores red-shifted from those naturally occurring in proteins, this technique can be used for real time monitoring of protease activity in crude preparations of virtually any protease. We demonstrate the value of this method in three model applications: (1) characterization of papain enzyme kinetics using rapid-mixing experiments, (2) selective monitoring of cleavage at a single site in a peptide with multiple proteolytic sites, and (3) analysis of the specificity of an inhibitor of calpain in cell lysates.