Innate Immune Training Initiates Efferocytosis to Protect against Lung Injury.

Innate immune training involves myelopoiesis, dynamic gene modulation, and functional reprogramming of myeloid cells in response to secondary heterologous challenges. The present study evaluates whether systemic innate immune training can protect tissues from local injury. Systemic pretreatment of mice with ß-glucan, a trained immunity agonist, reduces the mortality rate of mice with bleomycin-induced lung injury and fibrosis, as well as decreasing collagen deposition in the lungs. ß-Glucan pretreatment induces neutrophil accumulation in the lungs and enhances efferocytosis. Training of mice with ß-glucan results in histone modification in both alveolar macrophages (AMs) and neighboring lung epithelial cells. Training also increases the production of RvD1 and soluble mediators by AMs and efferocytes. Efferocytosis increases trained immunity in AMs by stimulating RvD1 release, thus inducing SIRT1 expression in neighboring lung epithelial cells. Elevated epithelial SIRT1 expression is associated with decreased epithelial cell apoptosis after lung injury, attenuating tissue damage. Further, neutrophil depletion dampens the effects of ß-glucan on macrophage accumulation, epigenetic modification in lung macrophages, epithelial SIRT1 expression, and injury-mediated fibrosis in the lung. These findings provide mechanistic insights into innate immune training and clues to the potential ability of centrally trained immunity to protect peripheral organs against injury-mediated disorders.

Panoramic dental tomosynthesis imaging by use of CBCT projection data.

Dental CBCT and panoramic images are important imaging modalities used in dental diagnosis and treatment planning. In order to acquire a panoramic image without an additional panoramic scan, in this study, we proposed a method of reconstructing a panoramic image by extracting panoramic projection data from dental CBCT projection data. After specifying the patient's dental arch from the patient's CBCT image, panoramic projection data are extracted from the CBCT projection data along the appropriate panoramic scan trajectory that fits the dental arch. A total of 40 clinical human datasets and one head phantom dataset were used to test the proposed method. The clinical human dataset used in this study includes cases in which it is difficult to reconstruct panoramic images from CBCT images, such as data with severe metal artifacts or data without teeth. As a result of applying the panoramic image reconstruction method proposed in this study, we were able to successfully acquire panoramic images from the CBCT projection data of various patients. The proposed method acquires a universally applicable panoramic image that is less affected by CBCT image quality and metal artifacts by extracting panoramic projection data from dental CBCT data and reconstructing a panoramic image.

Special Issue: Resistance to Targeted Therapies in Human Cancer.

Cancer is the second leading cause of death worldwide, accounting for approximately 10 million deaths in 2020 [...].

MicroRNA-34a-5p as a promising early circulating preclinical biomarker of doxorubicin-induced chronic cardiotoxicity.

Cardiotoxicity is a serious adverse effect of an anticancer drug, doxorubicin (DOX), which can occur within a year or decades after completion of therapy. The present study was designed to address a knowledge gap concerning a lack of circulating biomarkers capable of predicting the risk of cardiotoxicity induced by DOX. Profiling of 2083 microRNAs (miRNAs) in mouse plasma revealed 81 differentially expressed miRNAs 1 week after 6, 9, 12, 18, or 24 mg/kg total cumulative DOX doses (early-onset model) or saline (SAL). Among these, the expression of seven miRNAs was altered prior to the onset of myocardial injury at 12 mg/kg and higher cumulative doses. The expression of only miR-34a-5p was significantly (false discovery rate [FDR] < 0.1) elevated at all total cumulative doses compared with concurrent SAL-treated controls and showed a statistically significant dose-related response. The trend in plasma miR-34a-5p expression levels during DOX exposures also correlated with a significant dose-related increase in cardiac expression of miR-34a-5p in these mice. Administration of a cardioprotective drug, dexrazoxane, to mice before DOX treatment, significantly mitigated miR-34a-5p expression in both plasma and heart in conjunction with attenuation of cardiac pathology. This association between plasma and heart may suggest miR-34a-5p as a potential early circulating marker of early-onset DOX cardiotoxicity. In addition, higher expression of miR-34a-5p (FDR < 0.1) in plasma and heart compared with SAL-treated controls 24 weeks after 24 mg/kg total cumulative DOX dose, when cardiac function was altered in our recently established delayed-onset cardiotoxicity model, indicated its potential as an early biomarker of delayed-onset cardiotoxicity.

Doxorubicin-induced delayed-onset subclinical cardiotoxicity in mice.

Subclinical cardiotoxicity at low total cumulative doxorubicin (DOX) doses can manifest into cardiomyopathy in long-term cancer survivors. However, the underlying mechanisms are poorly understood. In male B6C3F1 mice, assessment of cardiac function by echocardiography was performed at 1, 4, 10, 17, and 24 weeks after exposure to 6, 9, 12, and 24 mg/kg total cumulative DOX doses or saline (SAL) to monitor development of delayed-onset cardiotoxicity. The 6- or 9-mg/kg total cumulative doses resulted in a significant time-dependent decline in systolic function (left ventricular ejection fraction (LVEF) and fractional shortening (FS)) during the 24-week recovery although there was not a significant alteration in % LVEF or % FS at any specific time point during the recovery. A significant decline in systolic function was elicited by the cardiotoxic cumulative DOX dose (24 mg/kg) during the 4- to 24-week period after treatment compared to SAL-treated counterparts. At 24 weeks after DOX treatment, a significant dose-related decrease in the expression of genes and proteins involved in sarcoplasmic reticulum (SR) calcium homeostasis (Ryr2 and Serca2) was associated with a dose-related increase in the transcript level of Casp12 (SR-specific apoptosis) in hearts. These mice also showed enhanced apoptotic activity in hearts indicated by a significant dose-related elevation in the number of apoptotic cardiomyocytes compared to SAL-treated counterparts. These findings collectively suggest that a steady decline in SR calcium handling and apoptosis might be involved in the development of subclinical cardiotoxicity that can evolve into irreversible cardiomyopathy later in life.

Production of Membrane Vesicles in Listeria monocytogenes Cultured with or without Sub-Inhibitory Concentrations of Antibiotics and Their Innate Immune Responses In Vitro.

Listeriosis is a food-borne illness caused by Listeria monocytogenes. Ampicillin (AMP) alone or in combination with gentamicin (GEN) is the first-line treatment option. Membrane vesicle (MV) production in L. monocytogenes under antibiotic stress conditions and pathologic roles of these MVs in hosts have not been reported yet. Thus, the aim of this study was to investigate the production of MVs in L. monocytogenes cultured with sub-minimum inhibitory concentrations (MICs) of AMP, GEN, or trimethoprim/sulfamethoxazole (SXT) and determine pathologic effects of these MVs in colon epithelial Caco-2 cells. L. monocytogenes cultured in tryptic soy broth with 1/2 MIC of AMP, GEN, or SXT produced 6.0, 2.9, or 1.5 times more MV particles, respectively, than bacteria cultured without antibiotics. MVs from L. monocytogenes cultured with AMP (MVAMP), GEN (MVGEN), or SXT (MVSXT) were more cytotoxic to Caco-2 cell than MVs obtained from cultivation without antibiotics (MVTSB). MVAMP induced more expression of tumor necrosis factor (TNF)-α gene than MVTSB, MVGEN and MVSXT, whereas MVTSB induced more expression of interleukin (IL)-1ß and IL-8 genes than other MVs. Expression of pro-inflammatory cytokine genes by L. monocytogenes MVs was significantly inhibited by proteinase K treatment of MVs. In conclusion, antibiotic stress can trigger the biogenesis of MVs in L. monocytogenes and MVs produced by L. monocytogenes exposed to sub-MIC of AMP can induce strong pro-inflammatory responses by expressing TNF-α gene in host cells, which may contribute to the pathology of listeriosis.

A synthesizing method for signal-enhanced and artifact-reduced mammogram from digital breast tomosynthesis.

In this paper, we propose a method for compositing a synthetic mammogram (SM) from digital breast tomosynthesis (DBT) slice images. The method consists of four parts. The first part is image reconstruction of DBT from the acquired projection data by use of backprojection-filtration (BPF) algorithm with a low-frequency boosting scheme and a high-density object reduction technique embedded. Also, a few expectation-maximization (EM) iterations have been additively implemented on top of the BPF algorithm to prepare a separate volume image. The second is generating three kinds of intermediate SMs. A forward projection image and a linear structure weighted forward projection image were computed. A maximum intensity projection of the BPF reconstructed volume image was also generated. The third part is integrating three intermediate SMs. The last is the post-processing of the SM. We scanned two physical phantoms in a prototype DBT scanner, and we have evaluated the performance of the proposed method. We also performed a clinical data study by use of 30 patient data who went through both DBT and digital mammography (DM) scans. Three experienced radiologists have read the SMs generated by several component techniques and also read the DM of each patient, and evaluated the generated SMs. The experimental phantom study and the clinical reader study consistently demonstrated the usefulness of the proposed method.

Wnt-PLC-IP3-Connexin-Ca2+ axis maintains ependymal motile cilia in zebrafish spinal cord.

Ependymal cells (ECs) are multiciliated neuroepithelial cells that line the ventricles of the brain and the central canal of the spinal cord (SC). How ependymal motile cilia are maintained remains largely unexplored. Here we show that zebrafish embryos deficient in Wnt signaling have defective motile cilia, yet harbor intact basal bodies. With respect to maintenance of ependymal motile cilia, plcδ3a is a target gene of Wnt signaling. Lack of Connexin43 (Cx43), especially its channel function, decreases motile cilia and intercellular Ca2+ wave (ICW) propagation. Genetic ablation of cx43 in zebrafish and mice diminished motile cilia. Finally, Cx43 is also expressed in ECs of the human SC. Taken together, our findings indicate that gap junction mediated ICWs play an important role in the maintenance of ependymal motile cilia, and suggest that the enhancement of functional gap junctions by pharmacological or genetic manipulations may be adopted to ameliorate motile ciliopathy.

Del-1, an Endogenous Inhibitor of TGF-ß Activation, Attenuates Fibrosis.

Uncontrolled activation of transforming growth factor (TGF)-ß results in a wide range of pathologic conditions. Therapeutic interventions to regulate TGF-ß signaling during fibrosis have been developed but the effectiveness is still limited. Here, we show that developmental endothelial locus-1 (Del-1) ameliorates fibrosis in mice by inhibiting αv integrin-mediated activation of TGF-ß. Del-1 bound to αvß6 integrin, an important activator of TGF-ß, and inhibited the binding of αvß6 integrin to the latency-associated peptide (LAP), thereby suppressing αv integrin-mediated activation of TGF-ß. Lack of Del-1 increased colocalization of αv integrin and LAP in the lungs, which was reversed by Del-1 supplementation. The crucial role of Del-1 in regulating TGF-ß activity was recapitulated in a mouse model of fibrosis using an adenovirus expressing inactive TGF-ß1. Del-1 supplementation improved the pathological characteristics of the mice and reduced mortality. Thus, we propose that Del-1 is a negative regulator of TGF-ß activation and a potential anti-fibrotic factor.

The effect of the type of anesthesia on the quality of postoperative recovery after orthopedic forearm surgery.

BACKGROUND: Although the quality of postoperative recovery may be affected by factors, there are few investigations whether the type of anesthesia also affects it. In this single-blinded, prospective, observational study, we compared the quality of postoperative recovery in patients undergoing orthopedic forearm surgery under general or regional anesthesia (brachial plexus block). METHODS: Ninety-seven subjects, aged 18-65 years and American Society of Anesthesiologists physical status I or II, undergoing orthopedic forearm surgery, were allocated to general or regional anesthesia group. The quality of postoperative recovery was assessed using a validated Korean version of Quality of Recovery-40 (QoR-40K) questionnaire. Patients were surveyed three times, the day before surgery (baseline) and 1st and 7th day after the surgery, and the scores of both groups were compared. RESULTS: We analyzed 47 and 50 patients in general and regional anesthesia, respectively. The global QoR-40K score and those of each of its five dimensions were not significantly different between the two groups at baseline, 1st and 7th day postoperatively. In two-way RM ANOVA, the global QoR-40K score at postoperative 1st day was significantly lower than that of baseline (P < 0.001) and postoperative 7th day (P < 0.001), respectively, in both general and regional anesthesia groups. However, there was no significant difference at each timepoint between the two groups. CONCLUSIONS: The present study suggests that brachial plexus block with intravenous dexmedetomidine infusion does not improve the quality of postoperative recovery compared to sevoflurane inhalation anesthesia with remifentanil infusion in patients undergoing orthopedic forearm surgery.

Combination therapy of low-dose cyclosporine and steroid in adults with IgA nephropathy
.

BACKGROUND: A few clinical trials in IgA nephropathy (IgAN) have shown that cyclosporine A (CyA) had therapeutic efficacy in reducing proteinuria. MATERIALS AND METHODS: This is a retrospective study, and all cases were selected based on kidney biopsy-proven IgAN. We reviewed the data of IgAN patients in the glomerulonephritis registry at Kyung Hee University Medical center and collected data on 86 patients with urinary protein/Cr ratio (PCR; g/g) > 0.5 and estimated GFR (eGFR) of > 50 mL/min/1.73m2 who were treated with combination therapy of low-dose CyA plus low-dose steroid (C+P; n = 37) and high-dose steroid single therapy (P; n = 49). RESULTS: In the C+P group, the mean duration of therapy was 14.5 ± 13.1 months, and the mean duration of follow-up 66.2 ± 36.3 months. In the C+P group, the urine PCR levels significantly declined after treatment (< 0.05). After 6 months of treatment, 12 (32%) patients were in complete remission and 7 (19%) in partial remission in the C+P group, compared with 21 (42%) and 11 (22%) in the P group, respectively. Urine PCR levels were also significantly reduced in 12 patients in the C+P group who had initial urine PCR between 0.5 and 1.0. The degree of hematuria was significantly reduced after treatment in the C+P group. These effects of C+P therapy on proteinuria and hematuria were very comparable to high-dose P therapy. After 2 years, a decline in renal function, > 25% decrease in eGFR from baseline levels, developed in 3 (8.1%) in the C+P group, compared with 4 (8.2%) in the P group. The rate of decline in renal function during follow-up was -0.14 ± 0.40 mL/min/1.73m2/month in the C+P group compared with -0.12 ± 0.22 mL/min/1.73m2/month in the P group. There were no changes of mean eGFR during the first 24 months, but the eGFR significantly decreased at last follow-up in both groups. When patients in the C+P group were divided into progressive (n = 9) and nonprogressive (n = 28) groups, a significant reduction in the amount of proteinuria after treatment was observed in the nonprogressive group, in contrast to the progressive group. In the C+P group, there were no severe adverse effects, especially no acute renal impairment, requiring discontinuation of CyA in this study. The incidence of infection was much lower in the C+P group than that in the P group. The limitation is that CyA acts to nonspecifically reduce proteinuria, so it requires long-term follow-up off CyA therapy for more than 2 years to determine. CONCLUSION: Our retrospective uncontrolled study provides only weak evidence that combination therapy of low-dose C+P could be an alternative to high-dose P therapy and be safe in adult IgAN patients with relatively normal renal function and proteinuria of > 0.5 g/g. Development of safe and effective therapy is still a major challenge requiring well-controlled prospective studies with this or other combination therapies.

Different epithelial cell response to membrane vesicles produced by Listeria monocytogenes cultured with or without salt stress.

We have previously shown that Listeria monocytogenes, a causative agent of listeriosis, can produce membrane vesicles (MVs) during in vitro culture. The aim of this study was to investigate the ability of MVs from L. monocytogenes cultured with or without salt stress to induce cytotoxicity and pro-inflammatory responses in colon epithelial Caco-2 cells. MVs were purified from wild-type L. monocytogenes 10403S strain and an isogenic ΔsigB mutant strain. MVs from both wild-type and ΔsigB mutant strains increased viability of Caco-2 cells regardless of salt stress. Both MVs from wild-type and ΔsigB mutant strains stimulated expression of pro-inflammatory cytokine and chemokine genes in Caco-2 cells. Expression levels of pro-inflammatory cytokine genes in cells treated with MVs from bacteria cultured without salt stress were significantly higher than those in cells treated with MVs from bacteria cultured with salt stress. However, expression levels of chemokine genes in cells treated with MVs from bacteria cultured with salt stress were significantly higher than those in cells treated with MVs from bacteria cultured without salt stress. In addition, expression levels of interleukin (IL)-1ß and IL-8 genes were partially inhibited by either lysozyme-treated MVs or ethylenediaminetetraacetic acid-treated MVs compared to those after treatment with intact MVs. Our results suggest that salt stress can affect the production of L. monocytogenes MVs, thus causing different pro-inflammatory responses in host cells.

Candidate early predictive plasma protein markers of doxorubicin-induced chronic cardiotoxicity in B6C3F1 mice.

Cardiotoxicity is a serious adverse effect of doxorubicin (DOX) treatment in cancer patients. Currently, there is a lack of sensitive biomarkers to predict the risk of DOX-induced cardiotoxicity. Using SOMAmer-based proteomic technology, 1129 proteins were profiled to identify potential early biomarkers of cardiotoxicity in plasma from male B6C3F1 mice given a weekly intravenous dose of 3 mg/kg DOX or saline (SAL) for 2, 3, 4, 6, or 8 weeks (6, 9, 12, 18, or 24 mg/kg cumulative DOX doses, respectively). Also, a group of mice received the cardio-protectant, dexrazoxane (DXZ; 60 mg/kg; intraperitoneal) 30 min before a weekly DOX or SAL dose. Proteomic analysis in plasma collected a week after the last dose showed a significant ≥1.2-fold change in level of 18 proteins in DOX-treated mice compared to SAL-treated counterparts during 8-week exposure. Of these, neurogenic locus notch homolog protein 1 (NOTCH1), von Willebrand factor (vWF), mitochondrial glutamate carrier 2, Wnt inhibitory factor 1, legumain, and mannan-binding lectin serine protease 1 were increased in plasma at 6 mg/kg cumulative DOX dose, prior to the release of myocardial injury marker, cardiac troponin I at 12 mg/kg and higher cumulative doses. These six proteins also remained significantly elevated following myocardial injury or pathology at 24 mg/kg. Pretreatment of mice with DXZ significantly attenuated DOX-induced elevated levels of only NOTCH1 and vWF with mitigation of cardiotoxicity. This suggests NOTCH1 and vWF as candidate early biomarkers of DOX cardiotoxicity, which may help in addressing a clinically important question of identifying cancer patients at risk for cardiotoxicity.

The Relationship between Epidemiologic Factors and Usage Pattern of Hair Care Products in Korea.

BACKGROUND: For hair care products that are used almost everyday, it is important to estimate the cumulative dosage of long-term exposure and to assess the effects on the human body. Little data are available to evaluate actual daily usage in Asian populations. OBJECTIVE: Reliable exposure data for hair care products is essential to conduct safety assessments. METHODS: We evaluated the actual usage pattern and amounts by checking the daily log over a 2-week period, to obtain all the data regarding the participants' hair care preferences. And, statistical analyses were conducted to analyze the daily use amount (g/d) and daily usage per hair length (g/cm/d), and other variables by sex, age group, and hair oiliness. RESULTS: Throughout this study, we found that female users consumed significantly larger daily amounts of shampoo and rinse. Male groups used more hair gel and spray than female groups. Interestingly, all the hair care products studied scored higher levels of usage among men when calibrated per unit length. Koreans tend to use lesser amount of rinse although their hairs are usually thicker than the Western hairs. CONCLUSION: This study provides exposure information for commonly used hair care products, which will be useful for risk assessment purposes.

Socioeconomic factors influencing cosmetic usage patterns.

Despite daily exposure to chemicals in cosmetic products, there are few studies on the exposure levels to cosmetics particularly in Asians. We sought to investigate the usage pattern of cosmetics, including eye makeup products, fragrances, color makeups, and hair and nail care products, in Koreans. An online survey of 1,800 participants (908 males and 892 females, aged 15-59 years) from 5 Metropolitan cities (Seoul, Incheon, Daejeon, Daegu, and Busan) in Korea was conducted. For overall types of cosmetics, the proportion of users was 7.1% (range: 0.0-46.3%) in males and 30.7% (range: 1.0-82.8%) in females. The most prevalently used product was perfume (46.3%) in males and lipstick (82.8%) in females. In addition, the mean number of application for overall types of cosmetics was 22.7 (range: 4.6-49.4) times per month in male users and 24.8 (range: 4.2-62.0) in female users. The usage pattern was significantly altered according to demographic and socioeconomic factors, including age group, occupation, and income. This work estimated the prevalences and frequencies of use of a wide collection of cosmetics from a large number of Koreans and found that the usage pattern was significantly associated with demographic and socioeconomic factors.

Optimal dose reduction algorithm using an attenuation-based tube current modulation method for cone-beam CT imaging.

To reduce the radiation dose given to patients, a tube current modulation (TCM) method has been widely used in diagnostic CT systems. However, the TCM method has not yet been applied to a kV-CBCT system on a LINAC machine. The purpose of this study is to investigate if a TCM method would be desirable in a kV-CBCT system for image-guided radiation therapy (IGRT) or not. We have developed an attenuation-based TCM method using prior knowledge from planning CT images of patients. The TCM method can provide optimized dose reductions without degrading image quality for kV-CBCT imaging. Here, we investigate whether or not our suggested TCM method is desirable to use in kV-CBCT systems to confirm and revise the exact position of a patient for IGRT. Patients go through diagnostic CT scans for RT planning; therefore, using information from prior CT images can enable estimations of the total X-ray attenuation through a patient's body in a CBCT setting for radiation treatment. Having this planning CT image allows to use the proposed TCM method in RT. The proposed TCM method provides a minimal amount of current for each projection, as well as total current, required to reconstruct the current modulated CBCT image with an image quality similar to that of CBCT. After applying a calculated TCM current for each projection, projection images were acquired and the current modulated CBCT image was reconstructed using a FDK algorithm. To validate the proposed approach, we used a numerical XCAT phantom and a real ATOM phantom and evaluated the performance of the proposed method via visual and quantitative image quality metrics. The organ dose due to imaging radiation was calculated in both cases and compared using the GATE simulation toolkit. As shown in the quantitative evaluation, normalized noise and SSIM values of the TCM were similar to those of conventional CBCT images. In addition, the proposed TCM method yielded comparable image quality to that of conventional CBCT images for both simulations and experimental studies as organ doses were decreased. We have successfully demonstrated the feasibility and dosimetric merit of a prototypical TCM method for kV-CBCT via simulations and experimental study. The results indicate that the proposed TCM method and overall framework can be a viable option for CBCT imaging that utilizes an optimal dose reduction without degrading image quality. Thus, this method reduces the probability for side effects due to radiation exposure.

Salt stress affects global protein expression profiles of extracellular membrane-derived vesicles of Listeria monocytogenes.

Our previous study has suggested that Listeria monocytogenes produces extracellular membrane vesicles (MVs) and its general stress transcription factor sigma B (σB) affects the production of MVs under energy stress. The objective of this study was to evaluate the production of MVs and perform global protein profiling for MVs with or without salt stress to understand the function of MVs in the pathogenesis of L. monocytogenes. When cells of L. monocytogenes were grown under 0.5 M salt stress, protein concentrations of MVs derived from wild-type strain and its isogenic ΔsigB mutant were approximately doubled compared to those of MVs derived from cells without salt stress. Proteomic analyses showed that the number of MV proteins expressed in wild-type strain was similar to that in ΔsigB mutant under salt stress. However, global protein expression profiles were dramatically changed under salt stress compared to those without salt stress. Fifteen σB dependent proteins were expressed in MVs of wild-type strain under salt stress, including osmolyte transporter OpuCABCD. In addition, MVs produced by salt stressed wild-type and ΔsigB mutant inhibited biofilm formation abilities of both strains. Taken together, our results suggest that salt stress can promote the production of MVs involved in carnitine transporter proteins, with σB playing a pivotal role in biological event.

A Feasibility Study of Low-Dose Single-Scan Dual-Energy Cone-Beam CT in Many-View Under-Sampling Framework.

A single-scan dual-energy low-dose cone-beam CT (CBCT) imaging technique that exploits a multi-slit filter is proposed in this paper. The multi-slit filter installed between the x-ray source and the scanned object is reciprocated during a scan. The x-ray beams through the slits would generate relatively low-energy x-ray projection data, while the filtered beams would make high-energy projection data. An iterative image reconstruction algorithm that uses an adaptive-steepest-descent method to minimize image total-variation under the constraint of data fidelity was applied to reconstructing the image from the low-energy projection data. Since the high-energy projection data suffer from a substantially high noise level due to the beam filtration, we have developed a new algorithm that exploits the joint sparsity between the low- and high-energy CT images for image reconstruction of the high-energy CT image. The proposed image reconstruction algorithm uses a gradient magnitude image (GMI) of the low-energy CT image by regularizing the difference of GMIs of the low- and high-energy CT images to be minimized. The feasibility of the proposed technique has been demonstrated by the use of various phantoms in the experimental CBCT setup. Furthermore, based on the proposed dual-energy imaging, a material differentiation was performed and its potential utility has been shown. The proposed imaging technique produced promising results for its potential application to a low-dose single-scan dual-energy CBCT.

Both absolute and relative quantification of urinary mRNA are useful for non-invasive diagnosis of acute kidney allograft rejection.

Urinary mRNA analysis with three-gene set (18S rRNA, CD3ε, and IP-10) has been suggested as a non-invasive biomarker of acute rejection (AR) in kidney transplant recipients using quantitative real-time PCR (qPCR). Application of droplet digital PCR (ddPCR), which has been suggested to provide higher sensitivity, accuracy, and absolute quantification without standard curves, could be a useful method for the quantifying low concentration of urinary mRNA. We investigated the urinary expression of these three genes in Korean patients with kidney transplantation and also evaluated the usefulness of ddPCR. 90 urine samples were collected at time of allograft biopsy in kidney recipients (n = 67) and from patients with stable renal function more than 10 years (n = 23). Absolute quantification with both PCR system showed significant higher mRNA levels of CD3ε and IP-10 in AR patients compared with stable transplants (STA), but there was no difference in 18S rRNA expression across the patient groups. To evaluate discrimination between AR and STA, ROC curve analyses of CTOT-4 formula yielded area under the curve values of 0.72 (95% CI 0.60-0.83) and 0.77 (95% CI 0.66-0.88) for qPCR and ddPCR, respectively. However, 18S normalization of absolute quantification and relative quantification with 18S showed better discrimination of AR from STA than those of the absolute method. Our data indicate that ddPCR system without standard curve would be useful to determine the absolute quantification of urinary mRNA from kidney transplant recipients. However, comparative method also could be useful and convenient in both qPCR and ddPCR analysis.