Characterization of the metabolic heterogeneity in cell populations requires the analysis of single cells. Most current methods in single-cell analysis rely on cell manipulation, potentially altering the abundance of metabolites in individual cells. A small sample volume and the chemical diversity of metabolites are additional challenges in single-cell metabolomics. Here, we describe the combination of fiber-based laser ablation electrospray ionization (f-LAESI) with 21 T Fourier transform ion cyclotron resonance mass spectrometry (21TFTICR-MS) for in situ single-cell metabolic profiling in plant tissue. Single plant cells infected by bacteria were selected and sampled directly from the tissue without cell manipulation through mid-infrared ablation with a fine optical fiber tip for ionization by f-LAESI. Ultrahigh performance 21T-FTICR-MS enabled the simultaneous capture of isotopic fine structures (IFSs) for 47 known and 11 unknown compounds, thus elucidating their elemental compositions from single cells and providing information on metabolic heterogeneity in the cell population.

Mass spectrometry imaging (MSI) enables simultaneous spatial mapping for diverse molecules in biological tissues.M atrix-assisted laser desorption ionization (MALDI) mass spectrometry (MS) has been am ainstream MSI method for aw ide range of biomolecules.H owever,M ALDI-MSI of biological homopolymers used for energy storage and molecular feedstocki sl imited by,e .g.,p referential ionization for certain molecular classes.Matrix-free nanophotonic ionization from silicon nanopost arrays (NAPAs) is an emerging laser desorption ionization (LDI) platform with ultra-trace sensitivity and molecular imaging capabilities.H ere,w es how complementary analysis and MSI of polyhydroxybutyric acid (PHB), polyglutamic acid (PGA), and polysaccharide oligomers in soybean root nodule sections by NAPA-LDI and MALDI. ForP HB,n umber and weight average molar mass, polydispersity,and oligomer sizedistributions across the tissue section and in regions of interest were characterized by NAPA-LDI-MSI.

In plants, long-distance transport of chemicals from source to sink takes place through the transfer of sap inside complex trafficking systems. Access to this information provides insight into the physiological responses that result from the interactions between the organism and its environment. In vivo analysis offers minimal perturbation to the physiology of the organism, thus providing information that represents the native physiological state more accurately. Here we describe capillary microsampling with electrospray ionization mass spectrometry (ESI-MS) for the in vivo analysis of xylem sap directly from plants. Initially, fast MS profiling was performed by ESI from the whole sap exuding from wounds of living plants in their native environment. This sap, however, originated from the xylem and phloem and included the cytosol of damaged cells. Combining capillary microsampling with ESI-MS enabled targeted sampling of the xylem sap and single parenchymal cells in the pith, thereby differentiating their chemical compositions. With this method we analyzed soybean plants infected by nitrogen-fixing bacteria and uninfected plants to investigate the effects of symbiosis on chemical transport through the sap. Infected plants exhibited higher abundances for certain nitrogen-containing metabolites in their sap, namely allantoin, allantoic acid, hydroxymethylglutamate, and methylene glutamate, compared to uninfected plants. Using capillary microsampling, we localized these compounds to the xylem, which indicated their transport from the roots to the upper parts of the plant. Differences between metabolite levels in sap from the infected and uninfected plants indicated that the transport of nitrogen-containing and other metabolites is regulated depending on the source of nitrogen supply.

Background: Expandable endoprostheses can be used to equalize limb length for pediatric patients requiring reconstruction following large bony oncologic resections.Outcomes of the Compress ® Compliant Pre-Stress (CPS) spindle paired with an Orthopedic Salvage System expandable distal femur endoprosthesis have not been reported. Methods:We conducted a multi-institutional retrospective study of pediatric patients with distal femoral bone sarcomas reconstructed with the above endoprostheses. Statistical analysis utilized Kaplan-Meier survival technique and competing risk analysis.Results: Thirty-six patients were included from five institutions. Spindle survivorship was 86.3% (95% confidence interval [CI], 67.7-93.5) at 10 years. Two patients had a failure of osseointegration (5.7%), both within 12 months. Twenty-two (59%) patients had 70 lengthening procedures, with mean expansions of 3.2 cm (range:1-9) over 3.4 surgeries. The expandable mechanism failed in eight patients with a cumulative incidence of 16.1% (95% CI, 5.6-31.5) at 5 years. Twenty-nine patients sustained International Society of Limb Salvage failures requiring 63 unplanned surgeries. Periprosthetic joint infection occurred in six patients (16.7%). Limb preservation rate was 91% at 10 years.Conclusions: There is a high rate of osseointegration of the Compress ® spindle among pediatric patients when coupled with an expandable implant. However, there is a high rate of expansion mechanism failure and prosthetic joint infections requiring revision surgery.

Background: Although the treatment of lower-extremity bone tumors is similar between adult and pediatric patients, differences in outcomes are unknown. Outcomes for lower-extremity oncologic reconstruction have been challenging to study because of the low incidence and heterogeneity in disease and patient characteristics. The PARITY (Prophylactic Antibiotic Regimens in Tumor Surgery) trial is the largest prospective data set assembled to date for patients with lower-extremity bone tumors and presents an opportunity to investigate differences in outcomes between these groups. Methods: Patient details were acquired from the prospectively collected PARITY trial database. The 1993 Musculoskeletal Tumor Society (MSTS-93) and Toronto Extremity Salvage Score (TESS) questionnaires were administered preoperatively and at 3, 6, and 12 months postoperatively. Continuous outcomes were compared between groups with use of the Student t test, and dichotomous outcomes were compared with use of the Pearson chi-square test. Results: A total of 150 pediatric and 447 adult patients were included. Pediatric patients were more likely than adult patients to have a primary bone tumor (146 of 150 compared with 287 of 447, respectively; p < 0.001) and to have received adjuvant chemotherapy (140 of 149 compared with 195 of 441, respectively; p < 0.001). Reoperation rates were not significantly different between age groups (45 of 105 pediatric patients compared with 106 of 341 adult patients; p ≤ 0.13). Pediatric patients had higher mean MSTS-93 scores (64.7 compared with 53.8 among adult patients; p < 0.001) and TESS (73.4 compared with 60.4 among adult patients; p < 0.001) at baseline, which continued to 1 year postoperatively (mean MSTS-93 score, 82.0 compared with 76.8 among adult patients; p = 0.02; mean TESS, 87.7 compared with 78.6 among adult patients; p < 0.001). Despite the differences in outcomes between cohorts, pediatric and adult patients demonstrated similar improvement in MSTS-93 scores (mean difference, 17.4 and 20.0, respectively; p = 0.48) and TESS (mean difference, 14.1 and 14.7, respectively; p = 0.83) from baseline to 1 year postoperatively. Conclusions: Pediatric patients had significantly better functional outcomes than adult patients at nearly all of the included postoperative time points; however, pediatric and adult patients showed similar mean improvement in these outcomes at 1 year postoperatively. These findings may be utilized to help guide the postoperative expectations of patients undergoing oncologic reconstruction. Level of Evidence: Prognostic Level II. See Instructions for Authors for a complete description of levels of evidence.

Mass spectrometry imaging (MSI) enables simultaneous spatial mapping for diverse molecules in biological tissues.M atrix-assisted laser desorption ionization (MALDI) mass spectrometry (MS) has been am ainstream MSI method for aw ide range of biomolecules.H owever,M ALDI-MSI of biological homopolymers used for energy storage and molecular feedstocki sl imited by,e .g.,p referential ionization for certain molecular classes.Matrix-free nanophotonic ionization from silicon nanopost arrays (NAPAs) is an emerging laser desorption ionization (LDI) platform with ultra-trace sensitivity and molecular imaging capabilities.H ere,w es how complementary analysis and MSI of polyhydroxybutyric acid (PHB), polyglutamic acid (PGA), and polysaccharide oligomers in soybean root nodule sections by NAPA-LDI and MALDI. ForP HB,n umber and weight average molar mass, polydispersity,and oligomer sizedistributions across the tissue section and in regions of interest were characterized by NAPA-LDI-MSI.