The standardized method and clinical experience may improve the reliability of visually assessed capillary refill time.

OBJECTIVE: Reliability of capillary refill time (CRT) has been questionable. The purpose of this study was to examine that a standardized method and clinical experience would improve the reliability of CRT. METHODS: This was a cross-sectional study in the emergency department (ED). Health care providers (HCPs) performed CRT without instruments (method 1) to classify patients as having normal or abnormal (≤2/>2 s) CRT. An ED attending physician quantitatively measured CRT using a chronograph (standardized visual CRT, method 2). A video camera was mounted on top of the hand tool to obtain a digital recording. The videos were used to calculate CRT via image software (image CRT, method 3) as a criterion standard of methods. Additionally, 9 HCPs reviewed the videos in a separate setting in order to visually assess CRT (video CRT, method 4). RESULTS: We enrolled 30 patients in this study. Standardized visual CRT (method 2) identified 10 abnormal patients, while two patients were identified by CRT without instruments (method 1). There was no correlation (κ value, 0.00) between CRT without instruments (method 1) and image CRT (method 3), however the correlation between standardized visual CRT (method 2) and image CRT (method 3) was strong (r = 0.64, p < 0.01). Both intra-observer reliability and correlation coefficient with image CRT (method 3) was higher in video CRT (method 4) by more experienced clinicians. CONCLUSIONS: Visual assessment is variable but a standardized method such as using a chronograph and/or clinical experience may aid clinicians to improve the reliability of visually assessed CRT.

Evaluation of accuracy of capillary refill index with pneumatic fingertip compression.

Capillary refill time (CRT) is a method of measuring a patient's peripheral perfusion status through a visual assessment performed by a clinician. We developed a new method of measuring CRT using standard pulse oximetry sensor, which was designated capillary refill index (CRI). We evaluated the accuracy of CRI in comparison to CRT image analysis. Thirty healthy adult volunteers were recruited for a derivation study and 30 patients in the emergency department (ED) were for validation. Our high fidelity mechanical device compresses and releases the fingertip to measure changes in blood volume using infrared-light (940 nm). CRT was calculated by image analysis software using recorded fingertip videos. CRI and CRT were measured at: room temperature (ROOM TEMP), 15 °C cold water (COLD), and 38 °C warm water (REWARM). Intra-rater reliability, Bland-Altman plots, and correlation coefficients were used to evaluate the accuracy of the novel CRI method. CRI (4.9 [95% CI 4.5-5.3] s) and CRT (4.0 [3.6-4.3]) in the COLD group were higher than the ROOM TEMP and REWARM groups. High intra-rater reliability was observed in both measurements (0.97 [0.95-0.98] and 0.98 [0.97-0.99], respectively). The Bland-Altman plots suggested a systematic bias: CRI was consistently higher than CRT (difference: + 1.01 s). There was a strong correlation between CRI and CRT (r = 0.89, p < 0.001). ED patients had higher CRI (3.91 [5.05-2.75]) and CRT (2.21 [3.19-1.23]) than those of healthy volunteers at room temperature. The same difference and correlation patterns were verified in the ED setting. CRI was as reliable as CRT by image analysis. The values of CRI was approximately 1 s higher than CRT.

Comparison of point-of-care peripheral perfusion assessment using pulse oximetry sensor with manual capillary refill time: clinical pilot study in the emergency department.

BACKGROUND: Traditional capillary refill time (CRT) is a manual measurement that is commonly used by clinicians to identify deterioration in peripheral perfusion status. Our study compared a novel method of measuring peripheral perfusion using an investigational device with standardized visual CRT and tested the clinical usefulness of this investigational device, using an existing pulse oximetry sensor, in an emergency department (ED) setting. MATERIAL AND METHODS: An ED attending physician quantitatively measured CRT using a chronometer (standardized visual CRT). The pulse oximetry sensor was attached to the same hand. Values obtained using the device are referred to as blood refill time (BRT). These techniques were compared in its numbers with the Bland-Altman plot and the predictability of patients' admissions. RESULTS: Thirty ED patients were recruited. Mean CRT of ED patients was 1.9 ± 0.8 s, and there was a strong correlation with BRT (r = 0.723, p < 0.001). The Bland-Altman plot showed a proportional bias pattern. The ED physician identified 3 patients with abnormal CRT (> 3 s). Area under the receiver operator characteristic curve (AUC) of BRT to predict whether or not CRT was greater than 3 s was 0.82 (95% CI, 0.58-1.00). Intra-rater reliability of BRT was 0.88 (95% CI, 0.79-0.94) and that of CRT was 0.92 (0.85-0.96). Twelve patients were admitted to the hospital. AUC to predict patients' admissions was 0.67 (95% CI, 0.46-0.87) by BRT and 0.76 (0.58-0.94) by CRT. CONCLUSIONS: BRT by a pulse oximetry sensor was an objective measurement as useful as the standardized CRT measured by the trained examiner with a chronometer at the bedside.

Can medical record reviewers reliably identify errors and adverse events in the ED?

BACKGROUND: Chart review has been the mainstay of medical quality assurance practices since its introduction more than a century ago. The validity of chart review, however, has been vitiated by a lack of methodological rigor. OBJECTIVES: By measuring the degree of interrater agreement among a 13-member review board of emergency physicians, we sought to validate the reliability of a chart review-based quality assurance process using computerized screening based on explicit case parameters. METHODS: All patients presenting to an urban, tertiary care academic medical center emergency department (annual volume of 57,000 patients) between November 2012 and November 2013 were screened electronically. Cases were programmatically flagged for review according to explicit criteria: return within 72hours, procedural evaluation, floor-to-ICU transfer within 24hours of admission, death within 24hours of admission, physician complaints, and patient complaints. Each case was reviewed independently by a 13-member emergency department quality assurance committee all of whom were board certified in emergency medicine and trained in the use of the tool. None of the reviewers were involved in the care of the specific patients reviewed by them. Reviewers used a previously validated 8-point Likert scale to rate the (1) coordination of patient care, (2) presence and severity of adverse events, (3) degree of medical error, and (4) quality of medical judgment. Agreement among reviewers was assessed with the intraclass correlation coefficient (ICC) for each parameter. RESULTS: Agreement and the degree of significance for each parameter were as follows: coordination of patient care (ICC=0.67; P<.001), presence and severity of adverse events (ICC=0.52; P=.001), degree of medical error (ICC=0.72; P<.001), and quality of medical judgment (ICC=0.67; P<.001). CONCLUSION: Agreement in the chart review process can be achieved among physician-reviewers. The degree of agreement attainable is comparable to or superior to that of similar studies reported to date. These results highlight the potential for the use of computerized screening, explicit criteria, and training of expert reviewers to improve the reliability and validity of chart review-based quality assurance.

Distinct cathepsins control necrotic cell death mediated by pyroptosis inducers and lysosome-destabilizing agents.

Necrotic cell death triggers a range of biological responses including a strong adaptive immune response, yet we know little about the cellular pathways that control necrotic cell death. Inhibitor studies suggest that proteases, and in particular cathepsins, drive necrotic cell death. The cathepsin B-selective inhibitor CA-074-Me blocks all forms of programmed necrosis by an unknown mechanism. We found that cathepsin B deficiency does not prevent induction of pyroptosis and lysosome-mediated necrosis suggesting that CA-074-Me blocks necrotic cell death by targeting cathepsins other than cathepsin B. A single cathepsin, cathepsin C, drives necrotic cell death mediated by the lysosome-destabilizing agent Leu-Leu-OMe (LLOMe). Here we present evidence that cathepsin C-deficiency and CA-074-Me block LLOMe killing in a distinct and cell type-specific fashion. Cathepsin C-deficiency and CA-074-Me block LLOMe killing of all myeloid cells, except for neutrophils. Cathepsin C-deficiency, but not CA-074-Me, blocks LLOMe killing of neutrophils suggesting that CA-074-Me does not target cathepsin C directly, consistent with inhibitor studies using recombinant cathepsin C. Unlike other cathepsins, cathepsin C lacks endoproteolytic activity, and requires activation by other lysosomal proteases, such as cathepsin D. Consistent with this theory, we found that lysosomotropic agents and cathepsin D downregulation by siRNA block LLOMe-mediated necrosis. Our findings indicate that a proteolytic cascade, involving cathepsins C and D, controls LLOMe-mediated necrosis. In contrast, cathepsins C and D were not required for pyroptotic cell death suggesting that distinct cathepsins control pyroptosis and lysosome-mediated necrosis.

A proteolytic cascade controls lysosome rupture and necrotic cell death mediated by lysosome-destabilizing adjuvants.

Recent studies have linked necrotic cell death and proteolysis of inflammatory proteins to the adaptive immune response mediated by the lysosome-destabilizing adjuvants, alum and Leu-Leu-OMe (LLOMe). However, the mechanism by which lysosome-destabilizing agents trigger necrosis and proteolysis of inflammatory proteins is poorly understood. The proteasome is a cellular complex that has been shown to regulate both necrotic cell death and proteolysis of inflammatory proteins. We found that the peptide aldehyde proteasome inhibitors, MG115 and MG132, block lysosome rupture, degradation of inflammatory proteins and necrotic cell death mediated by the lysosome-destabilizing peptide LLOMe. However, non-aldehyde proteasome inhibitors failed to prevent LLOMe-induced cell death suggesting that aldehyde proteasome inhibitors triggered a pleotropic effect. We have previously shown that cathepsin C controls lysosome rupture, necrotic cell death and the adaptive immune response mediated by LLOMe. Using recombinant cathepsin C, we found that aldehyde proteasome inhibitors directly block cathepsin C, which presumably prevents LLOMe toxicity. The cathepsin B inhibitor CA-074-Me also blocks lysosome rupture and necrotic cell death mediated by a wide range of necrosis inducers, including LLOMe. Using cathepsin-deficient cells and recombinant cathepsins, we demonstrate that the cathepsins B and C are not required for the CA-074-Me block of necrotic cell death. Taken together, our findings demonstrate that lysosome-destabilizing adjuvants trigger an early proteolytic cascade, involving cathepsin C and a CA-074-Me-dependent protease. Identification of these early events leading to lysosome rupture will be crucial in our understanding of processes controlling necrotic cell death and immune responses mediated by lysosome-destabilizing adjuvants.

Role of lysosome rupture in controlling Nlrp3 signaling and necrotic cell death.

The Nod-like receptor, Nlrp3, has been linked to inflammatory diseases and adjuvant-mediated immune responses. A wide array of structurally diverse agents does not interact directly with Nlrp3, but is thought to activate the Nlrp3 inflammasome by inducing a common upstream signal, such as lysosome rupture. To test the connection between lysosome integrity and Nlrp3 signaling, we analyzed inflammasome activation following stimulation of murine macrophages with lysosome-destabilizing agents and pyroptosis inducers. Here we provide evidence that lysosomal rupture and the corresponding release of lysosomal hydrolases is an early event in macrophages exposed to the lysosome-destabilizing adjuvants LLOMe and alum. Lysosome rupture preceded cell death induction mediated by these agents and was associated with the degradation of low-molecular weight proteins, including the inflammasome component caspase-1. Proteolysis of caspase-1 was controlled by specific cathepsins, but was independent of autocatalytic processes and Nlrp3 signaling. Consistent with these findings, lysosome-disrupting agents triggered only minimal caspase-1 activation and failed to cause caspase-1-dependent cell death (pyroptosis), generally associated with Nlrp3 signaling. In contrast, lysosome rupture was a late event in macrophages exposed to prototypical pyroptosis inducers. These agents triggered extensive Nlrp3 signaling prior to lysosome rupture with only minimal impact on the cellular proteome. Taken together, our findings suggest that lysosome impairment triggers a cascade of events culminating in cell death but is not crucial for Nlrp3 signaling. The significant differences observed between lysosome-disrupting agents and pyroptosis inducers might explain the distinct immunologic responses associated with these compounds.

Cathepsin-mediated necrosis controls the adaptive immune response by Th2 (T helper type 2)-associated adjuvants.

Immunologic adjuvants are critical components of vaccines, but it remains unclear how prototypical adjuvants enhance the adaptive immune response. Recent studies have shown that necrotic cells could trigger an immune response. Although most adjuvants have been shown to be cytotoxic, this activity has traditionally been considered a side effect. We set out to test the role of adjuvant-mediated cell death in immunity and found that alum, the most commonly used adjuvant worldwide, triggers a novel form of cell death in myeloid leukocytes characterized by cathepsin-dependent lysosome-disruption. We demonstrated that direct lysosome-permeabilization with a soluble peptide, Leu-Leu-OMe, mimics the alum-like form of necrotic cell death in terms of cathepsin dependence and cell-type specificity. Using a combination of a haploid genetic screen and cathepsin-deficient cells, we identified specific cathepsins that control lysosome-mediated necrosis. We identified cathepsin C as critical for Leu-Leu-OMe-induced cell death, whereas cathepsins B and S were required for alum-mediated necrosis. Consistent with a role of necrotic cell death in adjuvant effects, Leu-Leu-OMe replicated an alum-like immune response in vivo, characterized by dendritic cell activation, granulocyte recruitment, and production of Th2-associated antibodies. Strikingly, cathepsin C deficiency not only blocked Leu-Leu-OMe-mediated necrosis but also impaired Leu-Leu-OMe-enhanced immunity. Together our findings suggest that necrotic cell death is a powerful mediator of a Th2-associated immune response.

Proteasome inhibitors prevent caspase-1-mediated disease in rodents challenged with anthrax lethal toxin.

NOD-like receptors (NLRs) and caspase-1 are critical components of innate immunity, yet their over-activation has been linked to a long list of microbial and inflammatory diseases, including anthrax. The Bacillus anthracis lethal toxin (LT) has been shown to activate the NLR Nalp1b and caspase-1 and to induce many symptoms of the anthrax disease in susceptible murine strains. In this study we tested whether it is possible to prevent LT-mediated disease by pharmacological inhibition of caspase-1. We found that caspase-1 and proteasome inhibitors blocked LT-mediated caspase-1 activation and cytolysis of LT-sensitive (Fischer and Brown-Norway) rat macrophages. The proteasome inhibitor NPI-0052 also prevented disease progression and death in susceptible Fischer rats and increased survival in BALB/c mice after LT challenge. In addition, NPI-0052 blocked rapid disease progression and death in susceptible Fischer rats and BALB/c mice challenged with LT. In contrast, Lewis rats, which harbor LT-resistant macrophages, showed no signs of caspase-1 activation after LT injection and did not exhibit rapid disease progression. Taken together, our findings indicate that caspase-1 activation is critical for rapid disease progression in rodents challenged with LT. Our studies indicate that pharmacological inhibition of NLR signaling and caspase-1 can be used to treat inflammatory diseases.