Deep learning model to predict Ki-67 expression of breast cancer using digital breast tomosynthesis.

BACKGROUND: Developing a deep learning (DL) model for digital breast tomosynthesis (DBT) images to predict Ki-67 expression. METHODS: The institutional review board approved this retrospective study and waived the requirement for informed consent from the patients. Initially, 499 patients (mean age: 50.5 years, range: 29-90 years) referred to our hospital for breast cancer were participated, 126 patients with pathologically confirmed breast cancer were selected and their Ki-67 expression measured. The Xception architecture was used in the DL model to predict Ki-67 expression levels. The high Ki-67 vs low Ki-67 expression diagnostic performance of our DL model was assessed by accuracy, sensitivity, specificity, areas under the receiver operating characteristic curve (AUC), and by using sub-datasets divided by the radiological characteristics of breast cancer. RESULTS: The average accuracy, sensitivity, specificity, and AUC were 0.912, 0.629, 0.985, and 0.883, respectively. The AUC of the four subgroups separated by radiological findings for the mass, calcification, distortion, and focal asymmetric density sub-datasets were 0.890, 0.750, 0.870, and 0.660, respectively. CONCLUSIONS: Our results suggest the potential application of our DL model to predict the expression of Ki-67 using DBT, which may be useful for preoperatively determining the treatment strategy for breast cancer.

[Evaluation of the Effectiveness of a Neck Fixation Device for Improving Image Quality in Diffusion-weighted Whole-body Imaging with Background Suppression].

The effectiveness of a neck fixation device to improve the image quality of DWIBS was investigated. Healthy volunteers were examined while chewing with and without a neck fixation device using a 3-T MRI system. Distance of mandibular movement was measured using true-fast imaging of steady-state precession (true FISP). Signal-to-noise ratio (SNR) and apparent diffusion coefficient (ADC) of DWIBS were measured. Image quality of DWIBS was scored by visual evaluation. These values were compared with and without a neck fixation device. Regarding results, the mandibular displacement and ADC were decreased, and the SNR and visual score were increased by the use of the fixation device. There is a significant difference between with and without a neck fixation device in each measurement. The technique using a neck fixation device helps improve image quality of DWIBS in the head and neck region.

Automated volume measurement of abdominal adipose tissue from entire abdominal cavity in Dixon MR images using deep learning.

The purpose of this study was to realize an automated volume measurement of abdominal adipose tissue from the entire abdominal cavity in Dixon magnetic resonance (MR) images using deep learning. Our algorithm involves a combination of extraction of the abdominal cavity and body trunk regions using deep learning and extraction of a fat region based on automatic thresholding. To evaluate the proposed method, we calculated the Dice coefficient (DC) between the extracted regions using deep learning and labeled images. We also compared the visceral adipose tissue (VAT) and subcutaneous adipose tissue volumes calculated by employing the proposed method with those calculated from computed tomography (CT) images scanned on the same day using the automatic calculation method previously developed by our group. We implemented our method as a plug-in in a web-based medical image processing platform. The DCs of the abdominal cavity and body trunk regions were 0.952 ± 0.014 and 0.995 ± 0.002, respectively. The VAT volume measured from MR images using the proposed method was almost equivalent to that measured from CT images. The time required for our plug-in to process the test set was 118.9 ± 28.0 s. Using our proposed method, the VAT volume measured from MR images can be an alternative to that measured from CT images.

Fast B1 Mapping Based on Double-Angle Method with T1 Correction Using Standard Pulse Sequence.

Radiofrequency (RF) field (B1) mapping by combining the double-angle method (DAM) and T1 correction was investigated. The signal intensities S1 and S2 acquired by flip angle (FA) α and double FA 2α at short repetition time (TR) were converted to a signal intensity at TR=∞ by T1 correction. Then, these were used for DAM calculation. The T1 values are measured from two different images acquired with different TRs based on the saturation recovery (SR) method preliminarily. The effects of imaging parameters for T1 estimation and measured FA were investigated using CuSO4-doped water phantoms. A two-dimensional gradient echo type echo planar imaging pulse sequence was used. T1 values obtained by the 2-SR method were underestimated compared to the multipoint inversion recovery method. FA error was less than 5% when the appropriate imaging parameters were used. The acquisition time could be shortened to under 25 s by the use of T1-corrected DAM.

A potential bat adenovirus-based oncolytic virus targeting canine cancers.

Although a canine adenovirus (CAdV)-based oncolytic virus (OV) candidate targeting canine tumors has been reported, its oncolytic effect could be attenuated by CAdV vaccine-induced neutralizing antibodies in dog patients. To circumvent this issue, we focused on the bat adenovirus (BtAdV) strain, which was previously isolated from healthy microbats. We previously showed that this virus replicated efficiently in canine cell lines and did not serologically cross-react with CAdVs, suggesting that it may offer the possibility of an OV candidate for canine tumors. Here, we tested the growth properties and cytotoxicity of the BtAdV Mm32 strain in a panel of canine tumor cells and found that its characteristics were equivalent to those of CAdVs. To produce an Mm32 construct with enhanced tumor specificity, we established a novel reverse genetics system for BtAdV based on bacterial artificial chromosomes, and generated a recombinant virus, Mm32-E1Ap + cTERTp, by inserting a tumor-specific canine telomerase reverse transcriptase promoter into its E1A regulatory region. The growth and cytotoxicity of this recombinant were superior to those of wild-type Mm32 in canine tumor cells, unlike in normal canine cells. These data suggest that Mm32-E1Ap + cTERTp could be a promising OV for alternative canine cancer therapies.

[Imaging Parameter Optimization of Non-contrast Three-dimensional Time-of-flight Magnetic Resonance Angiography for Patients with Intracranial Stents Using a 1.5 T Magnetic Resonance Imaging System].

The imaging parameters of non-contrast three-dimensional time-of-flight magnetic resonance angiography (3D TOF-MRA) were optimized to improve the image quality for patients treated using stent-assisted coiling. A simulated blood flow phantom with three types of stents (Enterprise 2, Neuroform Atlas, and LVIS) was imaged by changing echo time (TE), band width (BW), flip angle (FA), and matrix (phase, frequency). The difference between the signal intensity in the simulated vessel and the background was measured at each imaging condition. The ratio of this difference with and without the stent was evaluated as the relative in-stent signal (RIS). In addition, the error ratio of the stent lumen diameter was assessed by comparing the full width at half maximum (FWHM) to that measured by 3D X-ray angiography. The RIS was higher in order of LVIS, Neuroform Atlas, and Enterprise 2 in all conditions. The RIS was higher in imaging conditions with short TE, narrow BW, high FA, and large phase matrix. The highest RIS was seen with a frequency matrix of 320 in the Enterprise 2 and 256 in the others. FWHM error ratio was smaller in the same order as the RIS. FWHM error ratio was smaller in imaging conditions with short TE, large frequency matrix (>384), large phase matrix (>224), and high FA (>20°). Imaging conditions of 3D TOF-MRA that were effective to improve the image quality for stent lumen evaluation were short TE and high spatial resolution.

Characterization of a novel species of adenovirus from Japanese microbat and role of CXADR as its entry factor.

Recently, bat adenoviruses (BtAdVs) of genus Mastadenovirus have been isolated from various bat species, some of them displaying a wide host range in cell culture. In this study, we isolated two BtAdVs from Japanese wild microbats. While one isolate was classified as Bat mastadenovirus A, the other was phylogenetically independent of other BtAdVs. It was rather related to, but serologically different from, canine adenoviruses. We propose that the latter, isolated from Asian parti-colored bat, should be assigned to a novel species of Bat mastadenovirus. Both isolates replicated in various mammalian cell lines, implying their wide cell tropism. To gain insight into cell tropism of these BtAdVs, we investigated the coxsackievirus and adenovirus receptor (CXADR) for virus entry to the cells. We prepared CXADR-knockout canine kidney cells and found that replication of BtAdVs was significantly hampered in these cells. For confirmation, their replication in canine CXADR-addback cells was rescued to the levels with the original cells. We also found that viral replication was corrected in human or bat CXADR-transduced cells to similar levels as in canine CXADR-addback cells. These results suggest that BtAdVs were able to use several mammalian-derived CXADRs as entry factors.

A PB1-K577E Mutation in H9N2 Influenza Virus Increases Polymerase Activity and Pathogenicity in Mice.

H9N2 avian influenza viruses are present in poultry worldwide. These viruses are considered to have pandemic potential, because recent isolates can recognize human-type receptor and several sporadic human infections have been reported. In this study, we aimed to identify mutations related to mammalian adaptation of H9N2 influenza virus. We found that mouse-adapted viruses had several mutations in hemagglutinin (HA), PB2, PA, and PB1. Among the detected mutations, PB1-K577E was a novel mutation that had not been previously reported to involve mammalian adaptation. A recombinant H9N2 virus bearing only the PB1-K577E mutation showed enhanced pathogenicity in mice, with increased virus titers in nasal turbinates compared to that in mice infected with the wild-type virus. In addition, the PB1-K577E mutation increased virus polymerase activity in human cell culture at a lower temperature. These data suggest that the PB1-K577E mutation is a novel pathogenicity determinant of H9N2 virus in mice and could be a signature for mammalian adaptation.

Antigenic heterogeneity among phylogenetic clusters of influenza D viruses.

Influenza (flu) D virus, a possible causative agent of bovine respiratory disease, is genetically classified into three clusters: D/OK-, D/660-, and D/Japan-lineages. To evaluate antigenic heterogeneity among these clusters, we compared antibody titers to each lineage virus using bovine sera collected over time following virus infection. Antibody titers to D/Japan-lineage virus rose rapidly in the acute phase of infection, and were 4 times higher than those to the other clustered viruses. In the later phase of infection, titers to D/Japan-lineage virus were equivalent to those to D/OK-lineage virus, and still higher than those to D/660-lineage virus. These results suggest the existence of common and lineage-specific antigenic epitopes in the hemagglutinin-esterase-fusion protein of flu D viruses.

Detection of bat hepatitis E virus RNA in microbats in Japan.

Several recent studies have reported that various bat species harbor bat hepatitis E viruses (BatHEV) belonging to the family Hepeviridae, which also contains human hepatitis E virus (HEV). The distribution and ecology of BatHEV are not well known. Here, we collected and screened 81 bat fecal samples from nine bat species in Japan to detect BatHEV RNA by RT-PCR using HEV-specific primers, and detected three positive samples. Sequence and phylogenetic analyses indicated that these three viruses were BatHEVs belonging to genus Orthohepevirus D like other BatHEV strains reported earlier in various countries. These data support the first detection of BatHEVs in Japanese microbats, indicating their wide geographical distribution among multiple bat species.

Heparan Sulfate Proteoglycan Is an Important Attachment Factor for Cell Entry of Akabane and Schmallenberg Viruses.

Akabane virus (AKAV) and Schmallenberg virus (SBV) are members of the genus Orthobunyavirus, which are transmitted by arthropod vectors with a broad cellular tropism in vitro as well as in vivo Both AKAV and SBV cause arthrogryposis-hydranencephaly syndrome in ruminants. The main cellular receptor and attachment factor for entry of these orthobunyaviruses are unknown. Here, we found that AKAV and SBV infections were inhibited by the addition of heparin or enzymatic removal of cell surface heparan sulfates. To confirm this finding, we prepared heparan sulfate proteoglycan (HSPG)-knockout (KO) cells by using a clustered regularly interspaced short palindromic repeat (CRISPR)-Cas9 system and measured the quantities of binding of these viruses to cell surfaces. We observed a substantial reduction in AKAV and SBV binding to cells, limiting the infections by these viruses. These data demonstrate that HSPGs are important cellular attachment factors for AKAV and SBV, at least in vitro, to promote virus replication in susceptible cells.IMPORTANCE AKAV and SBV are the etiological agents of arthrogryposis-hydranencephaly syndrome in ruminants, which causes considerable economic losses in the livestock industry. Here, we identified heparan sulfate proteoglycan as a major cellular attachment factor for the entry of AKAV and SBV. Moreover, we found that heparin is a strong inhibitor of AKAV and SBV infections. Revealing the molecular mechanisms of virus-host interactions is critical in order to understand virus biology and develop novel live attenuated vaccines.

Nationwide Distribution of Bovine Influenza D Virus Infection in Japan.

Cattle are major reservoirs of the provisionally named influenza D virus, which is potentially involved in the bovine respiratory disease complex. Here, we conducted a serological survey for the influenza D virus in Japan, using archived bovine serum samples collected during 2010-2016 from several herds of apparently healthy cattle in various regions of the country. We found sero-positive cattle across all years and in all the prefectural regions tested, with a total positivity rate of 30.5%, although the positivity rates varied among regions (13.5-50.0%). There was no significant difference in positivity rates for Holstein and Japanese Black cattle. Positivity rates tended to increase with cattle age. The herds were clearly divided into two groups: those with a high positive rate and those with a low (or no) positive rate, indicating that horizontal transmission of the virus occurs readily within a herd. These data demonstrate that bovine influenza D viruses have been in circulation for at least 5 years countrywide, emphasizing its ubiquitous distribution in the cattle population of Japan.

Hepatic Relaxation Times from Postmortem MR Imaging of Adult Humans.

PURPOSE: To measure T1 and T2 values of hepatic postmortem magnetic resonance (PMMR) imaging. MATERIALS AND METHODS: We performed hepatic PMMR imaging of 22 deceased adults (16 men, 6 women; mean age, 56.3 years) whose deaths were for reasons other than liver injury or disease at a mean of 27.7 hours after death. Before imaging, the bodies were kept in cold storage at 4°C (mean rectal temperature, 17.6°C). We measured T1 and T2 values in the liver at two sites (the anterior segment of the right lobe and the lateral segment of the left lobe). We also investigated the influence of the body temperature and postmortem interval on T1 and T2 values. RESULTS: In the anterior segment of the right lobe and the lateral segment of the left lobe, T1 values of PMMR imaging were 524 ± 112 ms and 472 ± 104 ms (mean ± standard deviation), respectively; while T2 values were 42 ± 6 ms and 43 ± 8 ms, respectively. T1 and T2 values did not differ significantly between the two sites (P ≧ 0.05). Regarding temperature, the T2 values of hepatic PMMR imaging were linearly correlated with the body temperature, but the T1 values were not. The T1 and T2 values of the two sites in the liver did not correlate with the postmortem interval. CONCLUSION: Reduction in body temperature after death is considered to induce T1 and T2 value changes in the liver on PMMR imaging.

Characterization of two endoglucanases for the classification of the earthworm, Eisenia fetida Waki.

Eisenia fetida and Eisenia andrei are vermicomposting species that are used as model animals for testing chemical material toxicology. Eisenia spp. are grown commercially in various fields in Japan. However, these two species have not been classified because it is difficult to distinguish them morphologically; thus, all bred earthworms are called E. fetida. However, it has been proposed that these two species have different expression regulation mechanisms. Here, we classified a sample of earthworms purchased from several farms, confirming that both E. fetida and E. andrei are present in Japanese earthworm breeding programs. We also characterized two highly active endoglucanases (EfEG1 and EfEG2) from the E. fetida Waki strain, which contained strong fibrinolytic enzymes for improving human health. We confirmed that EfEG1 is 1371 bp long and belongs to GHF9. Thus, E. fetida Waki may have commercial application for biomass utilization and as a dietary health supplement.

Possibility of visualization of gastrothorax based on unenhanced postmortem computed tomography/PMCT.

We present a fatal case of a gastrothorax due to an acute gastric volvulus resulting from a Bochdalek hernia. A 5-year-old boy without previous medical history was brought to our institution in a state of cardiopulmonary arrest and was subsequently pronounced dead. Postmortem computed tomography (PMCT) of the torso showed abdominal organs involving the lower section of the esophagus up to the entire stomach, the left side of the transverse colon, the entire spleen, and the tail of the pancreas herniated into the left thoracic cavity. The stomach was markedly expanded and a mesentero-axial (rotation along the short axis) volvulus was observed, displacing mediastinal structures to the right side and depressing the diaphragmatic contour. A PMCT of the thorax at the lung window setting revealed displacement of bilateral lungs. The bilateral lungs were severely atelectatic and congested. The PMCT findings mentioned above were consistent with the autopsy findings. PMCT can provide useful information for the diagnosis in cases we initially cannot predict any significant changes, for example, organ displacement.

Optimization of Inversion Time for Postmortem Fluid-attenuated Inversion Recovery (FLAIR) MR Imaging at 1.5T: Temperature-based Suppression of Cerebrospinal Fluid.

PURPOSE: Signal intensity (SI) and image contrast on postmortem magnetic resonance (MR) imaging are different from those of imaging of living bodies. We sought to suppress the SI of cerebrospinal fluid (CSF) sufficiently for fluid-attenuated inversion recovery (FLAIR) sequence in postmortem MR (PMMR) imaging by optimizing inversion time (TI). MATERIALS AND METHODS: We subject 28 deceased patients to PMMR imaging 3 to 113 hours after confirmation of death (mean, 27.4 hrs.). PMMR imaging was performed at 1.5 tesla, and T1 values of CSF were measured with maps of relaxation time. Rectal temperatures (RT) measured immediately after PMMR imaging ranged from 6 to 32°C (mean, 15.4°C). We analyzed the relationship between T1 and RT statistically using Pearson's correlation coefficient. We obtained FLAIR images from one cadaver using both a TI routinely used for living bodies and an optimized TI calculated from the RT. RESULTS: T1 values of CSF ranged from 2159 to 4063 ms (mean 2962.4), and there was a significantly positive correlation between T1 and RT (r = 0.96, P < 0.0001). The regression expression for the relationship was T1 = 74.4 * RT + 1813 for a magnetic field strength of 1.5T. The SI of CSF was effectively suppressed with the optimized TI (0.693 * T1), namely, TI = 0.693 * (77.4 * RT + 1813). CONCLUSION: Use of the TI calculated from the linear regression of the T1 and RT optimizes the FLAIR sequence of PMMR imaging.

Cerebral relaxation times from postmortem MR imaging of adults.

PURPOSE: We measured T1 and T2 values of cerebral postmortem magnetic resonance (PMMR) imaging and compared the data of cadavers with that of living human subjects. MATERIALS AND METHODS: We performed PMMR imaging of the brains of 30 adults (22 men, 8 women; mean age, 58.2 years) whose deaths were for reasons other than brain injury or disease at a mean of 29.4 hours after death. Before imaging, the bodies were kept in cold storage at 4°C (mean rectal temperature, 15.6°C). We measured T1 and T2 values in the brain bilaterally at 5 sites (bilateral caudate nucleus, putamen, thalamus and gray matter and white matter of the frontal lobe) and compared the data of PMMR imaging with that from MR imaging of the corresponding sites in 24 healthy volunteers (9 men, 15 women; mean age, 51.8 years). We also investigated the influence of body temperature on T1 and T2 values. RESULTS: Compared with MR imaging findings in the living subjects, PMMR imaging showed significantly shorter T1 values in the caudate nucleus, putamen, thalamus and gray matter and white matter of the frontal lobe and significantly longer T2 values in the gray matter and white matter of the frontal lobe; T2 values in the caudate nucleus, putamen, and thalamus showed no such differences. T1 values correlated significantly with body temperature in all 5 brain sites measured, but T2 values did not. CONCLUSION: Compared with findings of cerebral MR imaging in living adult subjects, those of PMMR imaging tended to demonstrate shorter T1 values and longer T2 values. We attribute this to increased water content of tissue, reduced pH, and reduced body temperature after death.

Group B betacoronavirus in rhinolophid bats, Japan.

We report group B Betacoronavirus infection in little Japanese horseshoe bats in Iwate prefecture. We then used reverse-transcription PCR to look for the coronavirus RNA-dependent RNA polymerase gene in fecal samples collected from 27 little Japanese horseshoe bats and found eight were provisionally positive. We had a success in the nucleotide sequencing of six of the eight positive samples and compared them with those of authentic coronaviruses. We found that these six samples were positive in coronavirus infection, and they belonged to the group B Betacornavirus by phylogenetic analysis. Virus isolation using the Vero cell culture was unsuccessful. Pathogenic trait of these bat coronaviruses remained unexplored.

High-resolution 3D-MRI of postmortem brain specimens fixed by formalin and gadoteridol.

PURPOSE: The purpose of this investigation was to compare magnetization-prepared rapid gradient echo (MP-RAGE) images with T1-weighted images (T1WI) and T2-weighted images (T1W2) of postmortem brain tissue fixed by admixtures of formalin and gadoteridol. We additionally sought to explore the feasibility of using fixed brain magnetic resonance imaging (MRIs) in forensic practices. METHODS: Specimens included in the study were eight whole brains that had been removed during forensic autopsy. Brain specimens were randomly divided into three groups and MRIs were performed either (A) the day of autopsy (n=2) on unfixed tissue, (B) after immersion fixation in 20% formalin (n=3), or (C) after immersion fixation in 20% formalin mixed with 4 mL/L ProHance® (gadoteridol) (n=3). T1WI, T2WI, and MP-RAGE images of all group samples were acquired with a 3T clinical MR scanner. Gray and white matter contrasts of the cortex and basal nucleus in every fixation group and image sequence were then visually compared. RESULTS: Gray/white matter contrasts of the cortex were good in all images obtained by MP-RAGE, and T1WIs of specimens fixed by formalin and gadoteridol-mixed formalin. Additionally, gray/white matter contrast in the basal nucleus was sufficient in the MP-RAGE sequence of specimens fixed by gadoteridol-mixed formalin. CONCLUSIONS: MRI of brains immersion-fixed in formalin and gadolinium could serve as a promising tool for neuropathological assessment in forensic practices.