Using ultrasound to diagnose long bone fractures.

Point-of-care ultrasound may be an alternative to radiographs for diagnosing long bone fractures when medical resources are limited. Safe and timesaving, ultrasound also can be used in the ED as a screening tool for suspected long bone fractures and can detect associated musculoskeletal injuries. Ultrasound can be used in radiation-sensitive patients such as children and pregnant patients.Studies have found that clinicians using ultrasound can detect long bone fractures with an average 90% sensitivity and specificity after an average of 1 to 4 hours total of didactic and practical training. More research is needed to determine standards for ultrasound training, patient morbidity outcomes, cost effectiveness, and insurance benefits.

The evolving role of transplantation in multiple myeloma: the need for a heterogeneous approach to a heterogeneous disease.

Autologous stem cell transplant (ASCT) is an established frontline standard of care for the younger, fitter patients with newly diagnosed multiple myeloma (NDMM) who are eligible for the procedure, and has contributed to improved overall survival. In the current era of novel therapies, the treatment landscape and prognosis have changed. The outstanding efficacy seen with regimens based on novel agents has led to a questioning of the frontline treatment paradigm with respect to ASCT. A key current question is whether to use transplant early or to collect stem cells early but save ASCT for salvage therapy. In this review, we evaluate the clinical data for each approach as well as the arguments in favor of early or delayed ASCT. We also consider the clinical/clonal heterogeneity of myeloma and review the evidence regarding which patient subgroups may benefit most from each approach. We summarize current treatment guidelines for transplant-eligible patients with NDMM and review the evolving role of minimal residual disease evaluation and its potential effect on the debate over early vs delayed ASCT. We conclude that frontline ASCT remains a standard of care for a substantial proportion of patients; however, delayed/salvage ASCT is increasingly being used in the context of highly active frontline regimens based on novel agents and the ongoing personalization of myeloma treatment.

Purine salvage promotes treatment resistance in H3K27M-mutant diffuse midline glioma.

BACKGROUND: Diffuse midline gliomas (DMG), including diffuse intrinsic pontine gliomas (DIPGs), are a fatal form of brain cancer. These tumors often carry a driver mutation on histone H3 converting lysine 27 to methionine (H3K27M). DMG-H3K27M are characterized by altered metabolism and resistance to standard of care radiation (RT) but how the H3K27M mediates the metabolic response to radiation and consequent treatment resistance is uncertain. METHODS: We performed metabolomics on irradiated and untreated H3K27M isogenic DMG cell lines and observed an H3K27M-specific enrichment for purine synthesis pathways. We profiled the expression of purine synthesis enzymes in publicly available patient data and our models, quantified purine synthesis using stable isotope tracing, and characterized the in vitro and in vivo response to de novo and salvage purine synthesis inhibition in combination with RT. RESULTS: DMG-H3K27M cells activate purine metabolism in an H3K27M-specific fashion. In the absence of genotoxic treatment, H3K27M-expressing cells have higher relative activity of de novo synthesis and apparent lower activity of purine salvage demonstrated via stable isotope tracing of key metabolites in purine synthesis and by lower expression of hypoxanthine-guanine phosphoribosyltransferase (HGPRT), the rate-limiting enzyme of purine salvage into IMP and GMP. Inhibition of de novo guanylate synthesis radiosensitized DMG-H3K27M cells in vitro and in vivo. Irradiated H3K27M cells upregulated HGPRT expression and hypoxanthine-derived guanylate salvage but maintained high levels of guanine-derived salvage. Exogenous guanine supplementation decreased radiosensitization in cells treated with combination RT and de novo purine synthesis inhibition. Silencing HGPRT combined with RT markedly suppressed DMG-H3K27M tumor growth in vivo. CONCLUSIONS: Our results indicate that DMG-H3K27M cells rely on highly active purine synthesis, both from the de novo and salvage synthesis pathways. However, highly active salvage of free purine bases into mature guanylates can bypass inhibition of the de novo synthetic pathway. We conclude that inhibiting purine salvage may be a promising strategy to overcome treatment resistance in DMG-H3K27M tumors.

GTP Signaling Links Metabolism, DNA Repair, and Responses to Genotoxic Stress.

How cell metabolism regulates DNA repair is incompletely understood. Here, we define a GTP-mediated signaling cascade that links metabolism to DNA repair and has significant therapeutic implications. GTP, but not other nucleotides, regulates the activity of Rac1, a guanine nucleotide-binding protein, which promotes the dephosphorylation of serine 323 on Abl-interactor 1 (Abi-1) by protein phosphatase 5 (PP5). Dephosphorylated Abi-1, a protein previously not known to activate DNA repair, promotes nonhomologous end joining. In patients and mouse models of glioblastoma, Rac1 and dephosphorylated Abi-1 mediate DNA repair and resistance to standard-of-care genotoxic treatments. The GTP-Rac1-PP5-Abi-1 signaling axis is not limited to brain cancer, as GTP supplementation promotes DNA repair and Abi-1-S323 dephosphorylation in nonmalignant cells and protects mouse tissues from genotoxic insult. This unexpected ability of GTP to regulate DNA repair independently of deoxynucleotide pools has important implications for normal physiology and cancer treatment. SIGNIFICANCE: A newly described GTP-dependent signaling axis is an unexpected link between nucleotide metabolism and DNA repair. Disrupting this pathway can overcome cancer resistance to genotoxic therapy while augmenting it can mitigate genotoxic injury of normal tissues. This article is featured in Selected Articles from This Issue, p. 5.

Rewiring of cortical glucose metabolism fuels human brain cancer growth.

The brain avidly consumes glucose to fuel neurophysiology. Cancers of the brain, such as glioblastoma (GBM), lose aspects of normal biology and gain the ability to proliferate and invade healthy tissue. How brain cancers rewire glucose utilization to fuel these processes is poorly understood. Here we perform infusions of 13 C-labeled glucose into patients and mice with brain cancer to define the metabolic fates of glucose-derived carbon in tumor and cortex. By combining these measurements with quantitative metabolic flux analysis, we find that human cortex funnels glucose-derived carbons towards physiologic processes including TCA cycle oxidation and neurotransmitter synthesis. In contrast, brain cancers downregulate these physiologic processes, scavenge alternative carbon sources from the environment, and instead use glucose-derived carbons to produce molecules needed for proliferation and invasion. Targeting this metabolic rewiring in mice through dietary modulation selectively alters GBM metabolism and slows tumor growth. Significance: This study is the first to directly measure biosynthetic flux in both glioma and cortical tissue in human brain cancer patients. Brain tumors rewire glucose carbon utilization away from oxidation and neurotransmitter production towards biosynthesis to fuel growth. Blocking these metabolic adaptations with dietary interventions slows brain cancer growth with minimal effects on cortical metabolism.

Diagnosis of ecto- and endoparasites in laboratory rats and mice.

Internal and external parasites remain a significant concern in laboratory rodent facilities, and many research facilities harbor some parasitized animals. Before embarking on an examination of animals for parasites, two things should be considered. One: what use will be made of the information collected, and two: which test is the most appropriate. Knowing that animals are parasitized may be something that the facility accepts, but there is often a need to treat animals and then to determine the efficacy of treatment. Parasites may be detected in animals through various techniques, including samples taken from live or euthanized animals. Historically, the tests with the greatest diagnostic sensitivity required euthanasia of the animal, although PCR has allowed high-sensitivity testing for several types of parasite. This article demonstrates procedures for the detection of endo- and ectoparasites in mice and rats. The same procedures are applicable to other rodents, although the species of parasites found will differ.

Diagnostic necropsy and selected tissue and sample collection in rats and mice.

There are multiple sample types that may be collected from a euthanized animal in order to help diagnose or discover infectious agents in an animal colony. Proper collection of tissues for further histological processing can impact the quality of testing results. This article describes the conduct of a basic gross examination including identification of heart, liver, lungs, kidneys, and spleen, as well as how to collect those organs. Additionally four of the more difficult tissue/sample collection techniques are demonstrated. Lung collection and perfusion can be particularly challenging as the tissue needs to be properly inflated with a fixative in order for inside of the tissue to fix properly and to enable thorough histologic evaluation. This article demonstrates the step by step technique to remove the lung and inflate it with fixative in order to achieve optimal fixation of the tissue within 24 hours. Brain collection can be similarly challenging as the tissue is soft and easily damaged. This article demonstrates the step by step technique to expose and remove the brain from the skull with minimal damage to the tissue. The mesenteric lymph node is a good sample type in which to detect many common infectious agents as enteric viruses persist longer in the lymph node than they are shed in feces. This article demonstrates the step by step procedure for locating and aseptically removing the mesenteric lymph node. Finally, identification of infectious agents of the respiratory tract may be performed by bacterial culture or PCR testing of nasal and/or bronchial fluid aspirates taken at necropsy. This procedure describes obtaining and preparing the respiratory aspirate sample for bacterial culture and PCR testing.

The meningo-orbital foramen in a Scottish population.

The meningo-orbital foramen is a small opening in the orbit lateral to the lateral end of the superior orbital fissure. It is widely reported to contain an orbital branch of the middle meningeal artery. The foramen may be single or multiple and may occur in the posterosuperior part of the lateral orbital wall or in the posterolateral part of the orbital roof. There is a lack of clarity in the literature as to whether foramina occurring in the orbital roof are the same entity as those occurring in the lateral wall. The disposition of the lesser wing of the sphenoid at the anterior limit of the middle cranial fossa makes it difficult to see how a foramen communicating with the anterior cranial fossa could transmit a branch of the middle meningeal artery. Our study contained 16 meningo-orbital foramina in the orbital roof that would transmit a fine probe. Fourteen of these passed into canals that tracked posteriorly in the bone to open into the middle cranial fossa close to the lateral extremity of the superior orbital fissure. The other two of these foramina communicated with the anterior cranial fossa and both were associated with a more posterior foramen that communicated with the middle cranial fossa. We hope this study clarifies an issue with relevance to surgery in the anterior cranial fossa.