Antibiotic-eluting scaffolds with responsive dual-release kinetics facilitate bone healing and eliminate S. aureus infection.

Osteomyelitis (OM) is a progressive, inflammatory infection of bone caused predominately by Staphylococcus aureus. Herein, we engineered an antibiotic-eluting collagen-hydroxyapatite scaffold capable of eliminating infection and facilitating bone healing. An iterative freeze-drying and chemical crosslinking approach was leveraged to modify antibiotic release kinetics, resulting in a layered dual-release system whereby an initial rapid release of antibiotic to clear infection was followed by a sustained controlled release to prevent reoccurrence of infection. We observed that the presence of microbial collagenase accelerated antibiotic release from the crosslinked layer of the scaffold, indicating that the material is responsive to microbial activity. As exemplar drugs, vancomycin and gentamicin-eluting scaffolds were demonstrated to be bactericidal, and supported osteogenesis in vitro. In a pilot murine model of OM, vancomycin-eluting scaffolds were observed to reduce S. aureus infection within the tibia. Finally, in a rabbit model of chronic OM, gentamicin-eluting scaffolds both facilitated radial bone defect healing and eliminated S. aureus infection. These results show that antibiotic-eluting collagen-hydroxyapatite scaffolds are a one-stage therapy for OM, which when implanted into infected bone defects simultaneously eradicate infection and facilitate bone tissue healing.

Preventive and therapeutic effects of a super-multivalent sialylated filamentous bacteriophage against the influenza virus.

The resurgence of influenza viruses as a significant global threat emphasizes the urgent need for innovative antiviral strategies beyond existing treatments. Here, we present the development and evaluation of a novel super-multivalent sialyllactosylated filamentous phage, termed t-6SLPhage, as a potent entry blocker for influenza A viruses. Structural variations in sialyllactosyl ligands, including linkage type, valency, net charge, and spacer length, were systematically explored to identify optimal binding characteristics against target hemagglutinins and influenza viruses. The selected SLPhage equipped with optimal ligands, exhibited exceptional inhibitory potency in in vitro infection inhibition assays. Furthermore, in vivo studies demonstrated its efficacy as both a preventive and therapeutic intervention, even when administered post-exposure at 2 days post-infection, under 4 lethal dose 50% conditions. Remarkably, co-administration with oseltamivir revealed a synergistic effect, suggesting potential combination therapies to enhance efficacy and mitigate resistance. Our findings highlight the efficacy and safety of sialylated filamentous bacteriophages as promising influenza inhibitors. Moreover, the versatility of M13 phages for surface modifications offers avenues for further engineering to enhance therapeutic and preventive performance.

Doxorubicin-Loaded Ultrasmall Gold Nanoparticles (1.5 nm) for Brain Tumor Therapy and Assessment of Their Biodistribution.

Ultrasmall gold nanoparticles (1.5 nm) were covalently conjugated with doxorubicin (AuDox) and AlexaFluor647 (AuAF647) to assess their biodistribution and their efficiency toward brain tumors (glioblastoma). A thorough characterization by transmission electron microscopy, small-angle X-ray scattering, and differential centrifugal sedimentation confirmed their uniform ultrasmall nature which makes them very mobile in the body. Each nanoparticle carried either 13 doxorubicin molecules (AuDox) or 2.7 AlexaFluor-647 molecules (AuAF647). The firm attachment of the ligands to the nanoparticles was demonstrated by their resilience to extensive washing, followed by centrifugation. The particles easily entered mammalian cells (HeLa, T98-G, brain endothelial cells, and human astrocytes) due to their small size. The intravenously delivered fluorescing AuAF647 nanoparticles crossed the blood-brain barrier with ∼23% accumulation in the brain tumor in an orthotopic U87 brain tumor model in nude mice. This was confirmed by elemental analysis (gold; inductively coupled plasma optical emission spectroscopy) in various organs. The doxorubicin-loaded AuDox nanoparticles inhibited brain tumor growth and prolonged animal survival without adverse side effects. Most of the nanoparticles (84%) had been excreted from the animal after 24 h, indicating a high mobility in the body.

Tomographic, Biomechanical, and In Vivo Confocal Microscopic Changes in Cornea in Chronic Gout Disease.

PURPOSE: This study aimed to evaluate the tomographic, biomechanical, and in vivo confocal microscopic (IVCM) effects of chronic gout disease on human cornea. METHODS: This prospective study included 60 eyes of 30 participants with chronic gout disease and 60 eyes of 30 healthy controls. Corneal thickness, keratometry (K) readings, and corneal aberrations were measured with Sirius 3 D corneal tomography system (Sirius, CSO, Italy). Corneal biomechanical properties (corneal hysteresis [CH], corneal resistance factor [CRF], and intraocular pressure [IOP] parameters) were assessed with an ocular response analyzer (ORA, Reichert Ophthalmic Instruments). The number and morphology of corneal endothelial cells and the number of basal epithelial cells were evaluated with IVCM (Confoscan 4.0). Tear breakup time (TBUT) was also evaluated. RESULTS: The mean diagnosis time of the patients with gout was 91.2 ± 69.6 months (12-300 month). Among corneal tomography measurements, K readings were similar between the two groups, while total and higher-order aberrations(coma, trefoil,s pherical, and quadrafoil) were significantly higher in the gout group. In the evaluation of biomechanical measurements, the CH value was significantly lower and the corneal-compensated IOP value was significantly higher in the gout group (p = 0.02, p = 0.01, respectively). The two groups did not significantly differ regarding the CRF or Goldmann IOP (p = 0.61, p = 0.15, respectively). Among the IVCM parameters, the number of corneal basal epithelial cells and the percentage of corneal endothelial hexagonality were significantly lower in the gout group, but no significant difference was detected in terms of the number of endothelial cells or polymegathism (p = 0.02, p < 0.001, p = 0.18, p = 0.59, respectively). While TBUT was significantly lower in the gout group (p < 0.001). CONCLUSION: This study showed that chronic gout disease increases the corneal aberrations and decreases the basal epithelial cell count, hexagonality ratio of endothelial cell and corneal biomechanics.

Neurofilament light chain as a biomarker of chemotherapy-induced peripheral neuropathy.

Chemotherapy-induced peripheral neuropathy (CIPN) is a common side effect of chemotherapy. The frequency of CIPN ranges from one in three to almost all patients depending on type of chemotherapy and dose. It causes symptoms that can range from sensitivity to touch and numbness to neuropathic pain in hands and feet. CIPN is notoriously difficult to grade objectively and has mostly relied on a clinician- or patient-based rating that is subjective and poorly reproducible. Thus, considerable effort has been aimed at identifying objective biomarkers of CIPN. Recent in vitro, animal, and clinical studies suggest that neurofilament light chain (NFL), a structural neuronal protein, may be an objective biomarker of CIPN. NFL released from cells to cell culture media reflects in vitro neurotoxicity, while NFL in serum reflects neuronal damage caused by chemotherapy in rodent models. Finally, NFL in serum may be a diagnostic biomarker of CIPN, but its prognostic ability to predict CIPN requires prospective evaluation. We discuss current limitations and future perspectives on the use of NFL as a preclinical and clinical biomarker of CIPN.

Traditional Chinese Manual Therapy (Tuina) Improves Knee Osteoarthritis by Regulating Chondrocyte Autophagy and Apoptosis via the PI3K/AKT/mTOR Pathway: An in vivo Rat Experiment and Machine Learning Study.

Background: Knee osteoarthritis (KOA) is on the rise due to lifestyle changes, obesity, and aging, yet effective treatments are lacking. Traditional Chinese manual therapy (Tuina) is promising for KOA. However, its mechanism remains unclear. Objective: This study aims to determine the effects of Tuina on a rat KOA model, focusing on the role of chondrocyte apoptosis and autophagy mechanisms. Methods: KOA was induced in rats by intra-articular injection of L-cysteine-activated papain into the right knee. Thirty-six male Sprague Dawley (SD) rats were randomly divided into blank, model control, Tuina, and positive drug groups. Paw withdrawal threshold tests, knee joint swelling, and passive range of motion assessed knee behavior. Cartilage tissue cytology, cytokine contents, and the mRNA and protein expression of PI3K/AKT/mTOR signaling pathway components were analyzed using HE and TUNEL staining, ELISA, RT-qPCR, and Western blotting, respectively. In addition, we used machine learning methods to conduct a secondary analysis of the dataset from the in vivo experiments in rats to verify the findings. Results: Tuina significantly relieved pain and joint swelling, and improved range of motion. Staining showed reduced articular cartilage destruction and apoptosis. Tuina reduced the serum levels of IL-1ß, IL-17, MMP-3, and MMP-13. Tuina downregulated Bax, ULK1, Beclin-1, LC3-II/I and upregulated PI3K, AKT, mTOR, and BCL-2 in cartilage tissue. The machine learning results indicated an 83.33% accuracy for the prediction model, remaining stable through both uni- and multivariate analyses. Tuina yielded the best comprehensive efficacy on KOA as well as better rat behavior and PI3K/AKT/mTOR signaling pathway improvement effect than positive drugs, while its cytokine-reducing ability was comparable to that of positive drugs. Conclusion: Tuina can alleviate cartilage tissue injury in KOA, relieve inflammation, and reduce chondrocyte apoptosis and autophagy, the underlying mechanisms of which may be associated with activation of the PI3K/AKT/mTOR signaling pathway.

The use of thermostable fluorescent proteins for live imaging in Sulfolobus acidocaldarius.

Introduction: Among hyperthermophilic organisms, in vivo protein localization is challenging due to the high growth temperatures that can disrupt proper folding and function of mostly mesophilic-derived fluorescent proteins. While protein localization in the thermophilic model archaeon S. acidocaldarius has been achieved using antibodies with fluorescent probes in fixed cells, the use of thermostable fluorescent proteins for live imaging in thermophilic archaea has so far been unsuccessful. Given the significance of live protein localization in the field of archaeal cell biology, we aimed to identify fluorescent proteins for use in S. acidocaldarius. Methods: We expressed various previously published and optimized thermostable fluorescent proteins along with fusion proteins of interest and analyzed the cells using flow cytometry and (thermo-) fluorescent microscopy. Results: Of the tested proteins, thermal green protein (TGP) exhibited the brightest fluorescence when expressed in Sulfolobus cells. By optimizing the linker between TGP and a protein of interest, we could additionally successfully fuse proteins with minimal loss of fluorescence. TGP-CdvB and TGP-PCNA1 fusions displayed localization patterns consistent with previous immunolocalization experiments. Discussion: These initial results in live protein localization in S. acidocaldarius at high temperatures, combined with recent advancements in thermomicroscopy, open new avenues in the field of archaeal cell biology. This progress finally enables localization experiments in thermophilic archaea, which have so far been limited to mesophilic organisms.

Inhibition of influenza virus infection in mice by pulmonary administration of a spray dried antiviral.

Increasing resistance to antiviral drugs approved for the treatment of influenza urges the development of novel compounds. Ideally, this should be complemented by a careful consideration of the administration route. 6'siallyllactosamine-functionalized ß-cyclodextrin (CD-6'SLN) is a novel entry inhibitor that acts as a mimic of the primary attachment receptor of influenza, sialic acid. In this study, we aimed to develop a dry powder formulation of CD-6'SLN to assess its in vivo antiviral activity after administration via the pulmonary route. By means of spray drying the compound together with trileucine, a dispersion enhancer, we created a powder that retained the antiviral effect of the drug, remained stable under elevated temperature conditions and performed well in a dry powder inhaler. To test the efficacy of the dry powder drug against influenza infection in vivo, infected mice were treated with CD-6'SLN using an aerosol generator that allowed for the controlled administration of powder formulations to the lungs of mice. CD-6'SLN was effective in mitigating the course of the disease compared to the control groups, reflected by lower disease activity scores and by the prevention of virus-induced IL-6 production. Our data show that CD-6'SLN can be formulated as a stable dry powder that is suitable for use in a dry powder inhaler and is effective when administered via the pulmonary route to influenza-infected mice.

In Vivo Models of Steroid-Induced Intraocular Hypertension.

Corticosteroids are widely utilized for their anti-inflammatory and immunosuppressive properties but often lead to ocular complications, including ocular hypertension. If untreated, ocular hypertension can progress to optic nerve atrophy and eventually result in steroid-induced glaucoma, which poses a risk of irreversible visual damage. Approximately 40% of individuals experience increased intraocular pressure after steroid use, and around 6% develop glaucoma. Although steroid-induced glaucoma is usually temporary and reversible if the treatment duration is under a year, prolonged exposure can cause permanent vision impairment. The pathogenesis of steroid-induced glaucoma is suggested to arise from increased outflow resistance of aqueous humor, primarily due to decreased expression of matrix metalloproteinases. This deficiency promotes the deposition of extracellular matrix and the dysfunction of trabecular meshwork cells. Additionally, modifications in the actin cytoskeleton increase the stiffness and alter the morphology of trabecular meshwork, further impeding aqueous humor outflow. Molecular changes, such as elevated expression of the MYOC gene, have also been implicated in restricting aqueous outflow. Various animal models, including rats, mice, primates, rabbits, cattle, sheep, cats, and dogs, have been developed to study steroid-induced glaucoma. These models exhibit pathological, pathophysiological, and molecular similarities to human disease, making them valuable for research. This review aims to summarize common animal models of steroid-induced ocular hypertension, discussing their advantages and limitations. The goal is to help researchers select appropriate models for future studies, thereby advancing the understanding of disease mechanisms and developing preventive strategies.

Stage-by-stage exploration of normal embryonic development in the Arabian killifish, Aphanius dispar.

BACKGROUND: Arabian killifish, Aphanius dispar, lives in marine coastal areas of the Middle East, as well as in streams that experience a wide range of salinities and temperatures. It has been used as a mosquito control agent and for studying the toxicities of environmental pollutants. A. dispar's eggshell (chorion) and embryos are highly transparent and are suitable for high resolution microscopic observations, offering excellent visibility of live tissues. RESULTS: In this study, the staging of normal embryonic development of A. dispar was described and investigated at different temperatures. Embryonic development was then examined under different thermal environments from 26 to 34°C. Our data suggest that temperature has a significant effect on embryonic development, with accelerated development at higher temperatures. CONCLUSION: A. dispar exhibits broad thermal tolerance and extended independent feeding capabilities, making it a promising model organism for toxicology and pathogenesis studies conducted over an extended period of time (12 days post-fertilization).

Therapeutic efficacy and in vivo distribution of human umbilical cord-derived mesenchymal stem cell spheroids transplanted via B-Ultrasound-guided percutaneous portal vein puncture in rhesus monkey models of liver fibrosis.

BACKGROUND: Liver fibrosis can progress to end-stage cirrhosis and liver cancer. Mesenchymal stem cells (MSCs) were considered the most promising therapeutic strategy, but most of the MSCs injected intravenously traditionally are trapped in the lungs, rapidly reducing their survival ability. MSC spheroids cultured in 3D have shown higher tolerance to fluid shear stress and better survival than dissociated MSCs. Simulating the route of orthotopic liver transplantation, transplanting MSC spheroids into the liver via hepatic portal vein may impact superior therapeutic effects. METHODS: In the present study, human umbilical cord-derived MSC spheroids (hUC-MSCsp) were transplanted into rhesus monkey models of liver fibrosis via B-ultrasound-guided percutaneous portal vein puncture with minimized body invasion. The therapeutic effect is evaluated through hematology, ultrasound, and pathology. To study the effect of hUC-MSCsp on gene expression in rhesus monkeys with liver injury, transcriptome sequencing analysis was performed on the livers of rhesus monkeys. The distribution of transplanted hUC-MSCsp was traced with RNA scope technology. RESULTS: We found that hUC-MSCsp significantly restored liver function, including ALT, AST, ALB, GLOB and bilirubin. hUC-MSCsp also significantly reduced liver collagen deposition and inflammatory infiltration, and promote dismission of liver ascites. Subsequently, the therapeutic effects were further validated in TGF-ß1/Smad pathway by global transcription profile. The distribution of transplanted hUC-MSCsp were also tracked, and we found that hUC-MSCsp distributed in the liver in a sphere status at 1 h after transplantation. After 16 days, the hUC-MSCsp were dispersed into dissociated cells that were predominantly distributed in the spleen, and a significant number of dissociated cells were still present in the liver. CONCLUSIONS: This study reveals the distributions of transplanted hUC-MSCsp after liver portal vein transplantation, and provides a novel approach and new insights into the molecular events of potential molecular events underlying the treatment of liver fibrosis with hUC-MSCsp.

Experimental validation of a FLUKA Monte Carlo simulation for carbon-ion radiotherapy monitoring via secondary ion tracking.

BACKGROUND: In-vivo monitoring methods of carbon ion radiotherapy (CIRT) includes explorations of nuclear reaction products generated by carbon-ion beams interacting with patient tissues. Our research group focuses on in-vivo monitoring of CIRT using silicon pixel detectors. Currently, we are conducting a prospective clinical trial as part of the In-Vivo Monitoring project (InViMo) at the Heidelberg Ion Beam Therapy Center (HIT) in Germany. We are using an innovative, in-house developed, non-contact fragment tracking system with seven mini-trackers based on the Timepix3 technology developed at CERN. PURPOSE: This article focuses on the implementation of the mini-tracker in Monte Carlo (MC) based on FLUKA simulations to monitor secondary charged nuclear fragments in CIRT. The main objective is to systematically evaluate the simulation accuracy for the InViMo project. METHODS: The implementation involved integrating the mini-tracker geometry and the scoring mechanism into the FLUKA MC simulation, utilizing the finely tuned HIT beam line. The systematic investigation included varying mini-tracker angles (from 15 ∘ $15^\circ$ to 45 ∘ $45^\circ$ in 5 ∘ $5^\circ$ steps) during the irradiation of a head-sized phantom with therapeutic carbon-ion pencil beams. To evaluate our implemented FLUKA framework, a comparison was made between the experimental data and data obtained from MC simulations. To ensure the fidelity of our comparison, experiments were performed at the HIT using the parameters and setup established in the simulations. RESULTS: Our research demonstrates high accuracy in reproducing characteristic behaviors and dependencies of the monitoring method in terms of fragment distributions in the mini-tracker, track angles, emission profiles, and fragment numbers. Discrepancies in the number of detected fragments between the experimental data and the data obtained from MC simulations are less than 4% for the angles of interest in the InViMo detection system. CONCLUSIONS: Our study confirms the potential of our simulation framework to investigate the performance of monitoring inter-fractional anatomical changes in patients undergoing CIRT using secondary nuclear charged fragments escaping from the irradiated patient.

MeCP2 Deficiency Alters the Response Selectivity of Prefrontal Cortical Neurons to Different Social Stimuli.

Rett syndrome (RTT), a severe neurodevelopmental disorder caused by mutations in the MeCP2 gene, is characterized by cognitive and social deficits. Previous studies have noted hypoactivity in the medial prefrontal cortex (mPFC) pyramidal neurons of MeCP2-deficient mice (RTT mice) in response to both social and nonsocial stimuli. To further understand the neural mechanisms behind the social deficits of RTT mice, we monitored excitatory pyramidal neurons in the prelimbic region of the mPFC during social interactions in mice. These neurons' activity was closely linked to social preference, especially in wild-type mice. However, RTT mice showed reduced social interest and corresponding hypoactivity in these neurons, indicating that impaired mPFC activity contributes to their social deficits. We identified six mPFC neural ensembles selectively tuned to various stimuli, with RTT mice recruiting fewer neurons to ensembles responsive to social interactions and consistently showing lower stimulus-ON ensemble transient rates. Despite these lower rates, RTT mice exhibited an increase in the percentage of social-ON neurons in later sessions, suggesting a compensatory mechanism for the decreased firing rate. This highlights the limited plasticity in the mPFC caused by MeCP2 deficiency and offers insights into the neural dynamics of social encoding. The presence of multifunctional neurons and those specifically responsive to social or object stimuli in the mPFC emphasizes its crucial role in complex behaviors and cognitive functions, with selective neuron engagement suggesting efficiency in neural activation that optimizes responses to environmental stimuli.

In vivo CRISPR screens identify Mga as an immunotherapy target in triple-negative breast cancer.

Immune evasion is not only critical for tumor initiation and progression, but also determines the efficacy of immunotherapies. Through iterative in vivo CRISPR screens with seven syngeneic tumor models, we identified core and context-dependent immune evasion pathways across cancer types. This valuable high-confidence dataset is available for the further understanding of tumor intrinsic immunomodulators, which may lead to the discovery of effective anticancer therapeutic targets. With a focus on triple-negative breast cancer (TNBC), we found that Mga knock-out significantly enhances antitumor immunity and inhibits tumor growth. Transcriptomics and single-cell RNA sequencing analyses revealed that Mga influences various immune-related pathways in the tumor microenvironment. Our findings suggest that Mga may play a role in modulating the tumor immune landscape, though the precise mechanisms require further investigation. Interestingly, we observed that low MGA expression in breast cancer patients correlates with a favorable prognosis, particularly in those with active interferon-γ signaling. These observations provide insights into tumor immune escape mechanisms and suggest that further exploration of MGA's function could potentially lead to effective therapeutic strategies in TNBC.

Intravital imaging of cardiac tissue utilizing tissue-stabilized heart window chamber in live animal model.

Aims: To develop and validate an optimized intravital heart microimaging protocol using a suction-based tissue motion-stabilizing cardiac imaging window to facilitate real-time observation of dynamic cellular behaviours within cardiac tissue in live mouse models. Methods and results: Intravital heart imaging was conducted using dual-mode confocal and two-photon microscopy. Mice were anesthetized, intubated, and maintained at a stable body temperature during the procedure. LysM-eGFP transgenic mice were utilized to visualize immune cell dynamics with vascular labelling by intravenous injection of anti-CD31 antibody and DiD-labelled red blood cells (RBCs). A heart imaging window chamber with a vacuum-based tissue motion stabilizer with 890-920 mbar was applied following a chest incision to expose the cardiac tissue. The suction-based heart imaging window chamber system and artificial intelligence-based motion compensation function significantly reduced motion artefacts and facilitated real-time in vivo cell analysis of immune cell and RBC trafficking, revealing a mean neutrophil movement velocity of 1.66 mm/s, which was slower compared to the RBC flow velocity of 9.22 mm/s. Intravital two-photon microscopic heart imaging enabled label-free second harmonic generation imaging of cardiac muscle structures with 820-840 nm excitation wavelength, revealing detailed biodistributions and structural variations in sarcomeres and fibrillar organization in the heart. Conclusion: The optimized intravital heart imaging protocol successfully demonstrates its capability to provide high-resolution, real-time visualization of dynamic cellular activities within live cardiac tissue.

In vivo targeted-imaging of mitochondrial acidification in an aristolochic acid I-induced nephrotoxicity mouse model by a fluorescent/photoacoustic bimodal probe.

Aristolochic acid I (AAI), a natural compound in aristolochia type Chinese medicinal herb, is generally acknowledged to have nephrotoxicity, which may be associated with mitophagy. Mitophagy is a cellular process with important functions that drive AAI-induced renal injury. Mitochondrial pH is currently measured by fluorescent probes in cell culture, but existing probes do not allow for in situ imaging of AAI-induced mitophagy in vivo. We developed a ratiometric fluorescent/PA dual-modal probe with a silicon rhodamine fluorophore and a pH-sensitive hemicyanine dye covalently linked via a short chain to obtain a FRET type probe. The probe was used to measure AAI-mediated mitochondrial acidification in live cells and in vivo. The Förster resonance energy transfer (FRET)-mediated ratiometric and bimodal method can efficiently eliminate signal variability associated with the commonly used one-emission and single detection mode by ratiometric two channels of the donor and acceptor. The probe has good water-solubility and low molecular weight with two positively charged, facilitating its precise targeting into renal mitochondria, where the fluorescent/PA changes in response to mitochondrial acidification, enabling dynamic and semi-quantitative mapping of subtle changes in mitochondrial pH in AAI-induced nephrotoxicity mouse model for the first time. Also, the joint use of L-carnitine could mitigate the mitophagy in AAI-induced nephrotoxicity.

In vivo dosimetry with an inorganic scintillation detector during multi-channel vaginal cylinder pulsed dose-rate brachytherapy: Dosimetry for pulsed dose-rate brachytherapy.

Background and purpose: In vivo dosimetry is not standard in brachytherapy and some errors go undetected. The aim of this study was to evaluate the accuracy of multi-channel vaginal cylinder pulsed dose-rate brachytherapy using in vivo dosimetry. Materials and methods: In vivo dosimetry data was collected during the years 2019-2022 for 22 patients (32 fractions) receiving multi-channel cylinder pulsed dose-rate brachytherapy. An inorganic scintillation detector was inserted in a cylinder channel. Each fraction was analysed as independent data sets. In vivo dosimetry-based source-tracking was used to determine the relative source-to-detector position. Measured dose was compared to planned and re-calculated source-tracking based doses. Assuming no change in organ and applicator geometry throughout treatment, the planned and source-tracking based dose distributions were compared in select volumes via γ-index analysis and dose-volume-histograms. Results: The mean ± SD planned vs. measured dose deviations in the first pulse were 0.8 ± 5.9 %. In 31/32 fractions the deviation was within the combined in vivo dosimetry uncertainty (averaging 9.7 %, k = 2) and planning dose calculation uncertainty (1.6 %, k = 2). The dwell-position offsets were < 2 mm for 88 % of channels, with the largest being 5.1 mm (4.0 mm uncertainty, k = 2). 3 %/2 mm γ pass-rates averaged 97.0 % (clinical target volume (CTV)), 100.0 % (rectum), 99.9 % (bladder). The mean ± SD deviation was -1. 1  ± 2.9 % for CTV D98, and -0.2 ± 0.9 % and -1.2 ± 2.5 %, for bladder and rectum D2cm3 respectively, indicating good agreement between intended and delivered dose. Conclusions: In vivo dosimetry verified accurate and stable dose delivery in multi-channel vaginal cylinder based pulsed dose-rate brachytherapy.

Phase-change ultrasound contrast agents for proton range verification: towards anin vivoapplication.

Objective.In proton therapy, range uncertainties prevent to optimally benefit from the superior depth-dose characteristics of proton beams over conventional photon-based radiotherapy. To reduce these uncertainties we recently proposed the use of phase-change ultrasound contrast agents as an affordable and effective range verification tool. In particular, superheated nanodroplets can convert into echogenic microbubbles upon proton irradiation, whereby the resulting ultrasound contrast relates to the proton range with high reproducibility. Here, we provide a firstin vivoproof-of-concept of this technology. Approach.First, thein vitrobiocompatibility of radiation-sensitive poly(vinyl) alcohol perfluorobutane nanodroplets was investigated using several colorimetric assays. Then,in vivoultrasound contrast was characterized using acoustic droplet vaporization and later using proton beam irradiations at varying energies (49.7 MeV and 62 MeV) in healthy Sprague Dawley rats. A preliminary evaluation of thein vivobiocompatibility was performed using acoustic droplet vaporization and a combination of physiology monitoring and histology. Main results.Nanodroplets were non-toxic over a wide concentration range (< 1 mM). In healthy rats, intravenously injected nanodroplets primarily accumulated in the organs of the reticuloendothelial system, where the lifetime of the generated ultrasound contrast (< 30 min) was compatible with a typical radiotherapy fraction (< 5 min). Spontaneous droplet vaporization did not result in significant background signals. Online ultrasound imaging of the liver of droplet-injected rats demonstrated an energy-dependent proton response, which can be tuned by varying the nanodroplet concentration. However, caution is warranted when deciding on the exact nanodroplet dose regimen as a mild physiological response (drop in cardiac rate, granuloma formation) was observed after acoustic droplet vaporization. Significance.These findings underline the potential of phase-change ultrasound contrast agents forin vivoproton range verification and provide the next step towards eventual clinical applications. .

Administration of Lacticaseibacillus casei CSL3 in Swiss Mice with Immunosuppression Induced by Cyclophosphamide: Effects on Immunological, Biochemical, Oxidative Stress, and Histological Parameters.

The study aimed to evaluate the effects of supplementation with Lacticaseibacillus casei CSL3 in Swiss mice immunosuppressed with cyclophosphamide on immunological, biochemical, oxidative stress, and histological parameters. The animals were distributed into four groups (control, CSL3, cyclophosphamide, and CSL3 + cyclophosphamide), where two groups were treated with L. casei CSL3 (10 log CFU mL-1) for 30 days, and two groups received chemotherapy (days 27 and 30-total dose of 250 mg kg-1). Counts of lactic acid bacteria (LAB) and bile-resistant LAB in stool samples; blood count (erythrogram, leukogram, and platelets); serum total cholesterol levels; catalase enzyme activity; and thiobarbituric acid reactive substances (TBARS) levels in liver, kidney, and brain; IL-4 expression; IL-23, TNF-α, NF-κß in the spleen; and histological changes in the liver, kidneys, and intestine were evaluated. The CSL3 + cyclophosphamide group showed a significant increase in bile-resistant LAB counts in feces (p = 0.0001), leukocyte counts, and expression of IL-23, TNF-α, and NF-κß (p < 0.05) significantly reduced total cholesterol levels (p = 0.001) and protected liver damage of supplemented animals. For oxidative stress damage, the bacterium did not influence the results. It is concluded that the bacterium is safe at a concentration of 10 log CFU mL-1 and has probiotic potential due to its positive influence on the immune response and lipid metabolism.