Citations

Summary Cannabis use is becoming increasingly common, both for recreational and medical purposes. However, there is a paucity of data regarding cannabis use in the context of eosinophilic esophagitis (EoE). We aimed to determine the impact of cannabis use on presentation and treatment response in EoE. To this end, we conducted a retrospective cohort study at a large academic medical center of newly diagnosed EoE patients age ≥ 12 years. Self-reported cannabis use status, baseline characteristics, and treatment response to topical corticosteroids and dietary therapy data were extracted. Bivariate and multivariable analyses were used to compare cannabis users and non-users at time of EoE diagnosis and to assess treatment response. Of 983 EoE patients, 80 reported using cannabis, with the majority reporting daily use and administration by inhalation. Baseline symptoms and peak eosinophil count were similar between cannabis users and non-users; cannabis users were less likely to have baseline endoscopic findings of exudates, edema, and stricture, and lower total Endoscopic Reference Score. On multivariable analysis, younger age, male sex, non-White race, and psychiatric diagnosis were independently associated with history of cannabis use at EoE presentation and stricture was independently associated with cannabis non-use. Post-treatment symptom and histologic responses were similar between cannabis users and non-users though there was a higher odds of post-treatment endoscopic inflammatory features with cannabis use. In conclusion, despite presenting with milder initial endoscopic findings, cannabis users exhibited greater inflammatory findings after treatment, highlighting a potential negative influence of cannabis use on EoE management.

Abstract Traumatic brain injury (TBI) annually impacts 69 million individuals worldwide. Mild TBI constitutes approximately 90% of all TBIs. Chronic pain post-mTBI occurs in 29% to 58% of patients. This study aims to introduce a predictive model for chronic pain development in individuals diagnosed with mild traumatic brain injury (mTBI) immediately postinjury. We included individuals who had sustained mTBI in motor vehicle accident (MVA). All patients had initial assessments within the first 72 hours (representing the subacute period) after the injury and performed follow-ups for 1 year. Machine learning model was applied to the integrated measures of clinical pain, pain-related psychological parameters, mTBI clinical signs, and sociodemographic information. This study included 203 patients experiencing acute head or neck pain attributable to mTBI post-MVA. We categorized these patients into 2 groups: patients who progressed to develop chronic head or neck pain (n = 89, 43.8%) and patients who recovered (low/mild pain) (n = 114, 56.2%). Severity of the subacute neck pain, number of painful body areas, and education years were identified as the most significant factors predicting chronic pain. The optimized predictive model demonstrated high efficacy, with an accuracy of 83%, a sensitivity of 92%, and an area under the receiver operating characteristic curve of 0.8. Our findings indicate feasibility in predicting chronic post-MVA pain within the critical 72-hour window postinjury using simple bedside metrics. This approach offers a promising avenue for the early detection of individuals at increased risk for chronic pain, enabling the implementation of targeted early interventions.

Abstract One of the main reasons we have not been able to cure cancers is that treatments select for drug-resistant cells. Pest managers face similar challenges with pesticides selecting for pesticide-resistant insects, resulting in similar mechanisms of resistance. Pest managers have developed 10 principles that could be translated to controlling cancers: (i) prevent onset, (ii) monitor continuously, (iii) identify thresholds below which there will be no intervention, (iv) change interventions in response to burden, (v) preferentially select nonchemical control methods, (vi) use target-specific drugs, (vii) use the lowest effective dose, (viii) reduce cross-resistance, (ix) evaluate success based on long-term management, and (x) forecast growth and response. These principles are general to all cancers and cancer drugs and so could be employed broadly to improve oncology. Here, we review the parallel difficulties in controlling drug resistance in pests and cancer cells. We show how the principles of resistance management in pests might be applied to cancer. Integrated pest management inspired the development of adaptive therapy in oncology to increase progression-free survival and quality of life in patients with cancers where cures are unlikely. These pest management principles have the potential to inform clinical trial design.

Key Points PIEZO1 activation reduces cystogenesis: Yoda1 activates PIEZO1, raising calcium and lowering cAMP, reducing cyst growth in autosomal dominant polycystic kidney disease models. Context-dependent role of PIEZO1: PIEZO1 knockout in mice with or without polycystic kidneys does not affect cyst formation, suggesting redundancy in mechanosensitive pathways. Therapeutic potential: Findings support PIEZO1 activation as a part of combination therapy to slow cyst growth in autosomal dominant polycystic kidney disease, needing more in vivo studies. Background The disruption of calcium signaling associated with polycystin deficiency is a key factor in abnormal epithelial growth in autosomal dominant polycystic kidney disease. Calcium homeostasis can be influenced by mechanotransduction. The mechanosensitive cation channel PIEZO1 has been implicated in sensing intrarenal pressure and regulating urinary osmoregulation, but its role in kidney cystogenesis is unclear. Methods We hypothesized that altered mechanotransduction contributes to cystogenesis in autosomal dominant polycystic kidney disease and that activation of mechanosensitive cation channels could be a therapeutic strategy. Results We demonstrate that Yoda1, a PIEZO1 activator, increases intracellular calcium and reduces forskolin-induced cAMP levels in mouse inner medullary collecting duct (mIMCD3) cells. Notably, knockout of polycystin-2 attenuated the efficacy of Yoda1 in reducing cAMP levels in mIMCD3 cells. Yoda1 also reduced forskolin-induced mIMCD3 cyst surface area in vitro and cystic index in mouse metanephros ex vivo in a dose-dependent manner. However, collecting duct–specific PIEZO1 knockout neither induced cystogenesis in wild-type mice nor altered cystogenesis in the Pkd1 RC/RC mouse model. Conclusions These findings support the potential role of PIEZO1 agonists in mitigating cystogenesis by increasing intracellular calcium and reducing cAMP levels, but the unaltered in vivo cystic phenotype after PIEZO1 knockout in the collecting duct suggests possible redundancy in mechanotransductive pathways.

Abstract Background The urgent need for rapid fentanyl detection at the point-of-care originates from its involvement in a high number of overdose deaths, primarily due to illicit use. Given fentanyl's potency compared to traditional opioids, accurate and sensitive detection is critical. Current lateral flow assays (LFA) cannot detect single-digit concentrations and may yield false negatives due to their lower sensitivity. Herein, we evaluate the clinical performance of a novel LFA (InstaStrip Fentanyl Rapid Test (urine)) with 1 ng/mL cutoff for fentanyl and no cross-reactivity for norfentanyl, offering enhanced sensitivity for effective overdose prevention at the ED. Methods A total of 172 urine samples submitted for UDS analysis were used for comparison studies. Fentanyl concentrations were categorized in 5 groups to cover various concentration categories around the cutoff of the LFA (1 ng/mL). These categories are: drug-free (<0.1 ng/mL, n=68), 0.1-0.5 ng/mL (n=14), 0.5-0.9 ng/mL (n=15), 1-1.5 ng/mL (n=9) and > 1.5 ng/mL (n= 66). Concentrations were determined by in-house LC-MS/MS methods validated according to CLIA standards, with fentanyl concentrations quantitated in the ABSciex software. For LFA testing, 150 μL of urine was added to a reagent tube containing dried rabbit anti-fentanyl antibody conjugated gold nanoparticles. After 20 sec of brief shaking, a test strip was inserted into the test tube. After 5 min, qualitative results were read as follow: both the control and test lines were visible in negative results, whereas only the control lines were visible in positive results. Method comparison study of the LFA was conducted by three operators who were blinded to the LC-MS/MS results. Result comparisons were made between the LFA strip and LC-MS/MS results, as well as among different operators for the same samples. Results Of the 75 urine samples with LC-MS/MS fentanyl concentrations ≥ 1 ng/mL, 74 produced consistently positive results using the InstaStrip Fentanyl Rapid Test. Of the 97 urine samples with LC-MS/MS fentanyl concentrations < 1 ng/mL, 96 produced consistently negative results using the InstaStrip Fentanyl Rapid Test. Using the LC-MS/MS as the gold standard method and 1 ng/mL as a cutoff, the clinical sensitivity of InstaStrip Fentanyl Rapid Test was calculated to be 98.67% (95% confidence interval (CI) 92.79-99.97%), and the clinical specificity was 98.97% (95% CI 94.39-99.97%). Consistent results among three operators were obtained for 100% of the “drug free”, “0.1- 0.5 ng/mL” and > 1.5 ng/mL groups, whereas 93.3 % (14 out of 15) and 88.9% (7 out of 8) were obtained for the “0.5-0.9 ng/mL” and “1-1.5 ng/mL” groups, respectively. The two samples with a singlet discrepant result (1 false positive and 1 false negative) in the triplicate experiments contained fentanyl concentrations of 0.9 and 1.0 ng/mL respectively, which are very close to the cutoff value. Conclusions In this study, the InstaStrip Fentanyl Rapid Test (urine) demonstrated high clinical sensitivity (98.67%), high specificity (98.97%) and high reproducibility showing that it may be useful in screening and identifying patients with fentanyl overdose in minutes in an emergency setting. This would facilitate appropriate patient triaging for optimal clinical care.

Abstract Background Professional medical societies recommend that carrier screening for 113 different genetic disorders be offered during pregnancy or preconception to all individuals, regardless of ancestry, in order to provide more equitable opportunities for patients to ascertain their reproductive risks. In our clinical laboratory experience, however, the vast majority of healthcare providers continue to order carrier screening for only cystic fibrosis (CF), which is disproportionately common in white individuals of Northern European descent. Our highest volume CF screening test is performed on >10,500 diverse patients per year and utilizes a targeted genotyping array that detects >500 variants in CFTR and >1200 variants in >150 other pertinent genes. Here we report the inequities in diagnostic yield among our first 8,000 patients for whom only limited testing was ordered and examine the barriers to widespread adoption of more inclusive carrier screening. Methods Three carrier screening options (CF, CF+spinal muscular atrophy [SMA], CF+SMA+fragile X syndrome [FXS]+hemoglobinopathies) were ordered clinically and performed on DNA extracted from 8,000 whole blood samples from unique individuals utilizing a custom SNP array (Thermo Fisher GeneTitanTM platform) designed to detect >1700 pathogenic variants in 218 different genes. Data was retrospectively reviewed to determine overall detection rates for carriers of all recommended conditions vs. only the condition(s) included in the originally ordered service. Finally, reproductive health providers at nine different ordering sites across the U.S. were interviewed regarding their current carrier screening practices. Results Patients were referred for testing by 119 different clinical facilities. 98.1% (7847/8000) had CF-only testing ordered, 1.2% (99/8000) had CF+SMA testing ordered, and 0.68% (54/8000) had testing for CF+SMA+FXS+hemoglobinopathies ordered. 7.1% (568/8000) of patients had reportable variants in CFTR. Had CF+SMA testing been ordered on all 8,000 patients, however, the diagnostic yield would have been 8.9%. Of the patients who had CF-only or CF+SMA testing ordered, 11.9% (943/7946) were found to be carriers of hemoglobinopathies. Finally, had all 8000 patients had testing for all currently recommended targets on the panel, the diagnostic yield would have increased to 55% (4396/8000). Ordering providers at 9/50 different contacted sites agreed to interview requests. 100% (9/9) of respondents indicated that their patients experience significant disparities in access to carrier screening, and that the primary cause of these inequities was insufficient insurance reimbursement. 67% (6/9) of respondents indicated that >50% of their patient population identified as non-white and would have “very strongly” benefitted from “testing for other disorders beyond CF.” Conclusions Despite clinical practice guidelines urging for more population-neutral carrier screening, major gaps exist between what is recommended versus ultimately ordered. This significantly limits the clinical utility of testing, especially in patients from underrepresented backgrounds. Improved insurance coverage for more inclusive carrier screening is needed to help reduce such disparities.

Abstract Background Multiple sclerosis (MS) is an immune-mediated central nervous system (CNS) demyelinating disease which is diagnosed based on CNS lesions' spatial and temporal dissemination. The 2017 revised McDonald Criteria include cerebrospinal fluid (CSF) specific oligoclonal bands (OCBs) to indicate dissemination in time, but their assessment is labor-intensive and requires a paired serum sample. Kappa immunoglobulin free light chain in CSF (KCSF) has emerged as a more sensitive but less specific MS biomarker, measured from a single CSF sample. Our institution's MS testing panel uses KCSF initially to rule out MS, with reflex to OCB for positive confirmation if KCSF is borderline positive (≥0.06 mg/dL). However, manual correlation of laboratory and clinical data hinders the direct comparison of the diagnostic performance between these markers. Herein we report a retrospective assessment of KCSF and OCB diagnostic performance using automated chart review. Methods Between January 1st, 2021 and December 31st, 2022, there were 1,237 patients with simultaneously measured OCB and KCSF. CSF-specific OCBs were measured by separating CSF and serum proteins using an agarose isoelectric focusing gel with a pH gradient from 3-10 (Sebia). After transfer to nitrocellulose paper, antisera were applied to visualize IgG specific bands and quantify CSF-specific OCBs. KCSF was quantitated using nephelometry by applying free light chain antisera (Binding Site) to CSF samples using a Siemens BNII analyzer. An automated chart review process utilizing keyword matching was implemented to identify patients with MS. An initial filter phase required presence of the term “multiple sclerosis”, while a second required the presence of any of the following terms: “primary progressive,” “secondary progressive,” “relapsing remitting,” “progressive relapsing,” or “tumefactive” to identify patients with diagnosed MS. The automated method’s performance was then compared to a manual chart review of 200 randomly selected patients. Subsequently, the automated process was used to determine the MS diagnosis status of all 1,237 patients. Results Of the 200 manually reviewed patients, 44 MS diagnoses were found. The automated process correctly identified 41 of these with a resulting sensitivity and specificity of 93.18% and 95.51%, respectively. Applied to the entire cohort, the automated review identified 275 MS diagnoses. For the entire cohort, the area under the curve (AUC) for the receiver operating characteristic (ROC) curves for OCB and KCSF were 0.824 (95% confidence interval: 0.794, 0.854) and 0.808 (0.777, 0.839) respectively. Our institution’s diagnostic cutoff of ≥2 for OCB yielded a sensitivity and specificity of 74.2% (68.3%, 79.1%) and 84.9% (73.8%, 89.5%), respectively. The best combination for sensitivity and specificity for KCSF with the Youden index yielded a cutoff ≥0.0654 mg/dL (sensitivity of 79.6% [73.1%, 84.0%] and specificity of 72.4% [62.8%, 76.8%]), which is similar to previous publications. This supports the use of automated chart review and suggests the KCSF cutoff has been stable over time. Conclusions KCSF exhibits comparable MS diagnostic performance to OCB yet only requires one CSF sample and can be measured rapidly. When validated, mining unstructured data allows for test performance characterization in large patient cohorts without the need for manual chart review.

Abstract Background THES is a rare autosomal recessive syndromic enteropathy caused by pathogenic variants in two genes, the Tetratricopeptide Repeat Domain 37 (TTC37) and Ski2 Like RNA Helicase (SKIV2L), that encode components of the human SKI complex involved in RNA degradation. This disease is characterized by several clinical features with variable penetrance including intractable congenital diarrhea, wooly hair abnormalities, intrauterine growth restriction, facial dysmorphism, immune dysfunction, liver abnormalities, and skin hyperpigmentation. Hypogammaglobulinemia with low IgG and normal to low IgM levels have been described in these cases which may require intravenous immunoglobulin (IVIG) replacement therapy. Some cases have a reduced population of switched memory B cells, T cells with reduced IFN-γ secretion, and hyporesponsive NK cells. Our patient is a 4-year-old female reported to have a homozygous TTC37 mutation with clinical manifestations of TPN dependence, liver fibrosis with portal hypertension, splenomegaly, developmental delay, thrombocytopenia and anemia requiring transfusion, and a history of hypogammaglobinemia requiring IVIG replacement. Patient would go on to develop elevated immunoglobin levels leading to additional testing described below. Methods Serum total protein analysis performed using the using Siemens Atellica CH Total Protein II test (Weichselbaum method using biuret reagent). Serum immunoglobin (IgA, IgG and IgG), IgG subclass, and kappa/lambda free light chain level analysis performed by turbidimetric photometry assay using a Binding Site Optilite analyzer. Serum protein electrophoresis and immunotyping performed using CAPILLARYS 2 capillary zone electrophoresis and immunosubtraction with antisera. Serum protein immunofixation performed using Sebia Hydrasys 2 with Hydragel 9 IF Maxi kit. Confirmation and final determination of serum immunoproteins performed at our reference laboratory using MASS-FIX (MALDI-TOF MS) matrix-assisted laser desorption/ionization-time of flight mass spectrometry after patient serum applied to 5 separate immunoaffinity resins specific to immunoglobulin G, A, M, K, and L for purification. Results Initial testing showed a broadly migrating beta-gamma region IgG kappa monotypic restriction measuring 1.7 g/dL, with predominantly kappa subtraction and trivial lambda subtraction by immunosubtraction; serum free kappa/lambda light chain ratio was increased at 22.94. Serum immunoglobins and free light chains measured (mg/dL): IgA 738 (15 - 160), IgM 48 (50 - 240), IgG 2,805 (405 - 1,160), free kappa 20.42 (0.33 - 1.94), and free lambda 0.89 (0.57 - 2.63). IgG subclasses measured (mg/dL): IgG1 533 (320 - 950), IgG2 174 (50 - 340), IgG3 16.9 (10.0 - 120.0), IgG4 1,221.1 (2.0 - 110.0). Additional serum protein values included (g/dL): total protein 8.10 (6.00 - 8.30), albumin 2.97 (3.43 - 4.84), alpha-1 0.32 (0.21 - 0.44), alpha-2 0.52 (0.54 - 0.97), beta globulin 3.42 (0.65 - 1.03), and gamma globulin 0.87 (0.70 - 2.30). The polyclonal nature of the IgG4 spike was later demonstrated by the MASS-FIX mass spectrograph. Conclusions MASS-FIX is a useful addition to the toolset used in the diagnosis of possible monoclonal gammopathies and protein disorders, especially where broad monotypic restrictions are observed or in patients with abnormal presentations for monoclonal gammopathies. Using current standard techniques, the above case could have been incorrectly classified as a monoclonal process resulting in unnecessary cost and incorrect treatment.

Abstract Background Automated plating instruments have been shown to improve quality in Microbiology laboratories in a number of ways including consistent streaking patterns that provide better isolation of bacterial colonies compared to manual methods. The time per culture set up and automated camera imaging is dependent upon the amount of media set, such that more individual plates slow down turn around time. Conversely, smaller media surface area and the use of non-selective media may result in mixed colonies that require sub-culture for isolation before identification and susceptibilities can be performed. Based upon these findings, we attempted to reduce our media usage per culture with the implementation of automation, with the goal of using as few plates as possible while at the same time not negatively impacting the turn around time of the culture via the need to subculture poorly isolated colonies. We conducted a pre-and post-implementation study evaluating the costs per culture and turn around time delays related to subculturing in a subset. Methods In the pre-implementation period, from April-January 2023, three individual media plates (sheep blood agar, chocolate agar and CNA/MacConkey bi-plate) were routinely set for general aerobic bacterial culture sources including swabs and body fluids. In the post implementation period, from April-January 2024, a single sheep blood/chocolate agar bi-plate was set unless the gram stain showed evidence of mixed organisms (defined as more than one morphology) at which time a secondary selective media (CNA/MacConkey) was manually set. For urine cultures, in the pre-implementation period 2 plates were set (sheep blood and CNA/MacConkey bi-plate) and in the post-implementation period a single chromogenic agar/sheep blood bi-plate was set. Total numbers of cultures in both periods were collected as well as the number of cultures for which additional media were set based on gram stain results. Orders of subculture media in the laboratory information system were compared per positive cultures. Statistical analysis was performed in GraphPad Prism. Results In the pre- versus post-implementation periods, a total of 16,454 vs 17,782 general aerobic cultures and 50,680 vs 53,916 urine cultures were set, respectively. Only 3.6% of general cultures required additional media set based on gram stain. The difference in cost between pre- and post-implementation was approximately a two-thirds reduction in cost for general cultures and a 25% reduction for urine cultures, with a total savings of approximately $34,238 in media avoidance in the post-implementation period. The frequency of positive cultures requiring subculture was not significantly different between the pre- (average 36% of positive growth) and post-implementation (average 22% of positive growth) periods for both culture types and may actually have been reduced due to automation. Conclusions The implementation of reduced media concomitant with streaking via automation resulted in significant cost savings while not negatively impacting isolation, which in turn impacts culture turn around times. The automation may have reduced need for subculture, perhaps due to better isolation, thus subsequent analyses will explore this additional finding.

Abstract Background Bicarbonate stability in serum depends on time, exposure to air, temperature, tube size and sample volume. With implementation of Cobas Connection Module (CCM) automation, the workflow for bicarbonate measurement needed to be re-evaluated. The aim of this study was to determine bicarbonate stability in filled 3.5 mL and 5 mL serum-separator tubes (SSTs) and capped and uncapped aliquot tubes on the CCM automation. Methods Bicarbonate was measured on a Cobas c701 (Roche Diagnostics) at various time points. SSTs (n=8) were centrifuged 6 min at 1649g ∼30 min post-collection. Serum was aliquoted (1 mL) into capped or uncapped tubes and kept ambient (15-25°C) on the CCM track for 0, 60, 90, 150, 180, 240 and 300 minutes (n=2 per time point). Primary SSTs (5mL n=4, and 3.5mL n=5) were uncapped on the CCM track and bicarbonate measured at baseline and 30 minute intervals through 6.5 hours. An absolute difference of +/- 4 mmol/L from baseline was considered acceptable. Results Bicarbonate concentrations decrease with time at rates dependent on whether the aliquot tube is capped or uncapped or the SST is 3.5 mL or 5 mL (Figure 1).Figure 1.Average decrease in bicarbonate concentration with time for capped aliquot tubes (black square), uncapped aliquot tubes (red circle), 3.5 mL serum separator tube (blue up triangle), 5 mL serum separator tube (green down triangle). Bars represent +/- 1SD. Conclusions Bicarbonate in serum is stable for testing for 2.5 hours in uncapped 1 mL aliquot tubes, 3.5 hours in capped 1 mL aliquot tubes, 3.5 hours in uncapped full 3.5 mL SST tubes and 6 hours in uncapped full 5 mL SST tubes at ambient temperatures (15-25°C). Bicarbonate testing can be accommodated by total laboratory automation but processes such as add-ons and retesting must be done with caution.