Identification of a selective manganese ionophore that enables nonlethal quantification of cellular manganese.

Available assays for measuring cellular manganese (Mn) levels require cell lysis, restricting longitudinal experiments and multiplexed outcome measures. Conducting a screen of small molecules known to alter cellular Mn levels, we report here that one of these chemicals induces rapid Mn efflux. We describe this activity and the development and implementation of an assay centered on this small molecule, named manganese-extracting small molecule (MESM). Using inductively-coupled plasma-MS, we validated that this assay, termed here "manganese-extracting small molecule estimation route" (MESMER), can accurately assess Mn in mammalian cells. Furthermore, we found evidence that MESM acts as a Mn-selective ionophore, and we observed that it has increased rates of Mn membrane transport, reduced cytotoxicity, and increased selectivity for Mn over calcium compared with two established Mn ionophores, calcimycin (A23187) and ionomycin. Finally, we applied MESMER to test whether prior Mn exposures subsequently affect cellular Mn levels. We found that cells receiving continuous, elevated extracellular Mn accumulate less Mn than cells receiving equally-elevated Mn for the first time for 24 h, indicating a compensatory cellular homeostatic response. Use of the MESMER assay versus a comparable detergent lysis-based assay, cellular Fura-2 Mn extraction assay, reduced the number of cells and materials required for performing a similar but cell lethality-based experiment to 25% of the normally required sample size. We conclude that MESMER can accurately quantify cellular Mn levels in two independent cells lines through an ionophore-based mechanism, maintaining cell viability and enabling longitudinal assessment within the same cultures.

Patient and Technical Factors Associated with Difficult Arterial Access and Ultrasound Use in the Operating Room.

Arterial catheterization enables continuous hemodynamic monitoring but has been shown to cause severe complications, especially when multiple attempts are required. The aim of this study was to explore what factors were associated with multiple attempts and ultrasound use in the operating room. We performed a retrospective analysis of patients who had arterial catheters inserted at a tertiary care children's hospital from January 2018 to March 2022, identifying clinical factors that were associated with both outcomes. A total of 3946 successful arterial catheter insertions were included. Multivariable analysis showed multiple attempts were associated with noncardiac surgery: pediatric (OR: 1.79, 95% CI: 1.30-2.51), neurologic (OR: 2.63, 95% CI: 1.89-3.57), orthopedic (OR: 3.23, 95% CI: 2.27-4.55), and non-radial artery placement (OR: 5.00, 95% CI: 3.33-7.14) (all p < 0.001). Multivariable analysis showed ultrasound use was associated with neonates (OR: 9.6, 95% CI: 4.1-22.5), infants (OR: 6.98, 95% CI: 4.67-10.42), toddlers (OR: 6.10, 95% CI: 3.8-9.8), and children (OR: 2.0, 95% CI: 1.7-2.5) compared to teenagers, with cardiac surgery being relative to other specialties-pediatric (OR: 0.48, 95% CI: 0.3-0.7), neurologic (OR: 0.27, 95% CI: 0.18-0.40), and orthopedic (OR: 0.38, 95% CI: 0.25-0.58) (all p < 0.001). In our exploratory analysis, increased odds of first-attempt arterial catheter insertion success were associated with cardiac surgery, palpation technique, and radial artery placement. Younger patient age category, ASA III and IV status, cardiac surgery, and anesthesiologist placement were associated with increased odds of ultrasound use.

Whole body potassium as a biomarker for potassium uptake using a mouse model.

Potassium is known for its effect on modifiable chronic diseases like hypertension, cardiac disease, diabetes (type-2), and bone health. In this study, a new method, neutron generator based neutron activation analysis (NAA), was utilized to measure potassium (K) in mouse carcasses. A DD110 neutron generator based NAA assembly was used for irradiation.Thirty-two postmortem mice (n= 16 males and 16 females, average weight [Formula: see text] and [Formula: see text] g) were employed for this study. Soft-tissue equivalent mouse phantoms were prepared for the calibration. All mice were irradiated for 10 minutes, and the gamma spectrum with 42K was collected using a high efficiency, high purity germanium (HPGe) detector. A lead shielding assembly was designed and developed around the HPGe detector to obtain an improved detection limit. Each mouse sample was irradiated and measured twice to reduce uncertainty. The average potassium concentration was found to be significantly higher in males [Formula: see text] compared to females [Formula: see text]. We also observed a significant correlation between potassium concentration and the weight of the mice. The detection limit for potassium quantification with the NAA system was 46 ppm. The radiation dose to the mouse was approximately 56 [Formula: see text] mSv for 10-min irradiation. In conclusion, this method is suitable for estimating individual potassium concentration in small animals. The direct evaluation of total body potassium in small animals provides a new way to estimate potassium uptake in animal models. This method can be adapted later to quantify potassium in the human hand and small animals in vivo. When used in vivo, it is also expected to be a valuable tool for longitudinal assessment, kinetics, and health outcomes.

Rodent hair is a Poor biomarker for internal manganese exposure.

Hair is used as a biomarker of manganese (Mn) exposure, yet there is limited evidence to support its utility to quantify internal vs external Mn exposure. C57BL/6 J mice and Sprague-Dawley rats were exposed in two blocks of 3 subcutaneous injections every 3 days starting on day 0 or 20. The control group received two blocks of saline (vehicle); Treatment A received the first block as Mn (50 mg/kg MnCl2 tetrahydrate), with the second block as either methylmercury (MeHg at 2.6 or 1.3 mg/kg) for mice or vehicle for rats; and Treatment B received Mn for both blocks. Hair was collected on days 0 and 60 from all treatment groups and Mn quantified by inductively coupled plasma-mass spectrometry (ICP-MS) and total Hg by Direct Mercury Analyzer (DMA). No correlation between internal Mn dose and hair Mn was observed, whereas hair Hg was significantly elevated in MeHg exposed vs non-exposed mice. Whole body Mn content at day 60 was quantified postmortem by neutron activation analysis, which detected significantly elevated Mn for Treatment B in mice and rats. Overall, we find no evidence to support the use of hair as a valid biomarker for internal exposure to Mn at a neurotoxic level.