Neutrophil-to-lymphocyte ratio associates with nutritional parameters, intratumoral immune profiles, and clinical outcomes of pembrolizumab in head and neck squamous cell carcinoma.

BACKGROUND: The relationship between the tumor-immune microenvironment and systemic inflammatory markers, including neutrophil-to-lymphocyte ratio (NLR), is unclear. METHODS: We examined the characteristics of systemic inflammatory markers and tumor immune microenvironments in relation to treatment outcomes in 29 consecutive patients with recurrent/metastatic head and neck squamous cell carcinoma (R/M HNSCC) who received pembrolizumab, using 14-marker multiplex immunohistochemistry and image cytometry. RESULTS: NLR ≥4.5 (high NLR) at pretreatment status significantly correlated with short overall survival (OS) and progression-free survival-2 (PFS2) and malnutrition status. High NLR in peripheral blood was significantly correlated with low lymphoid cell and high tumor-associated macrophage counts in tissues, especially myeloid-to-lymphoid cell ratios, suggesting an association between circulating and intratumoral immune complexity profiles. CONCLUSIONS: This study suggests a link between NLR in circulating blood, systemic nutritional status, and immune composition within the tumor.

The clinical impact of macrophage polarity after Kasai portoenterostomy in biliary atresia.

Introduction: Biliary atresia (BA) is a cholestatic hepatopathy caused by fibrosing destruction of intrahepatic and extrahepatic bile ducts, and its etiology has not been clearly revealed. In BA, liver fibrosis progression is often observed even after Kasai portoenterostomy (KPE), and more than half of cases require liver transplantation in their lifetime in Japan. Macrophages play an important role in liver fibrosis progression and are classically divided into proinflammatory (M1) and fibrotic macrophages (M2), whose phenotypic transformation is called "macrophage polarity." The polarity has been reported to reflect the tissue microenvironment. In this study, we examined the relationship between macrophage polarity and the post-KPE clinical course. Materials and methods: Thirty BA patients who underwent KPE in our institution from 2000 to 2020 were recruited. Multiple immunostainings for CD68, CD163, CK19, and α-SMA were carried out on liver biopsy specimens obtained at KPE. ROC curves were calculated based on each clinical event, and the correlation with the clinical data was analyzed. Results and discussion: The M2 ratio, defined as the proportion of M2 macrophages (CD163-positive cells), was correlated inversely with the occurrence of postoperative cholangitis (AUC: 0.7602). The patients were classified into M2 high (n = 19) and non-high (n = 11) groups based on an M2 ratio value obtained from the Youden index ( = 0.918). As a result, pathological evaluations (Metavir score, αSMA area fraction, and CK19 area fraction) were not significantly different between these groups. In mild liver fibrosis cases (Metavir score = 0-2), the M2 non-high group had a significantly lower native liver survival rate than the high group (p = 0.02). Moreover, 4 out of 8 cases in the M2 non-high group underwent early liver transplantation within 2 years after KPE. Conclusions: Non-M2 macrophages, including M1 macrophages, may be correlated with postoperative cholangitis, and the M2 non-high group in mild liver fibrosis cases had a significantly lower native liver survival rate than the high group, requiring early liver transplantation in this study. Preventing advanced liver fibrosis is a key factor in improving native liver survival for BA patients, and liver macrophages may play important roles in liver homeostasis and the promotion of inflammation and fibrosis.

Characteristic differences in the abundance of tumor-infiltrating lymphocytes and intratumoral developing T cells in thymoma, with special reference to PD-1 expression.

PURPOSE: Though programmed cell death-1 (PD-1) inhibitors mainly target tumor-infiltrating lymphocytes (TILs) expressing PD-1, developing T cells in thymus also express PD-1 in their process of maturation. To predict the therapeutic effect of PD-1 inhibitors for thymoma, it is necessary to clarify the proportions of TILs and intratumoral developing T cells. METHODS: The expressions of CD4, CD8, and PD-1 on T cells were analyzed by flow cytometry in 31 thymomas. The amount of T cell receptor excision circles (TRECs), which can be detected in newly formed naïve T cells in the thymus, was evaluated using sorted lymphocytes from thymomas by quantitative PCR. The expressions of granzyme B (GZMB) and lymphocyte activation gene-3 (LAG-3) in PD-1 + CD8 T cells were analyzed by image cytometry using multiplex immunohistochemistry. RESULTS: The PD-1 + rate in both CD4 and CD8 T cells was significantly higher in type AB/B1/B2 than in type A/B3 thymomas. The amounts of TRECs in CD4 and CD8 T cells were significantly higher in type AB/B1/B2 than in type A/B3 thymomas and comparable to normal thymus. PD-1 expression at each stage of T cell development of type AB/B1/B2 thymomas was comparable to that of normal thymus. Both the percentages and cell densities of PD-1 + CD8 T cells expressing GZMB or LAG-3, which are known to contain tumor-reactive T cells, were significantly lower in type AB/B1/B2 thymomas. CONCLUSION: Most PD-1 + T cells in type AB/B1/B2 thymomas are intratumoral developing T cells and are not TILs.

Genomic and immune microenvironment profiling in a case of metastatic intrathyroid thymic carcinoma.

Metastatic intrathyroid thymic carcinoma (ITTC) is a rare cancer with no effective drugs for controlling. This case report has shown genomic and immune microenvironment profiles in metastatic ITTC and emphasized an immunosuppression via a PD-1/PD-L1 pathway, possibly strengthening the rationale for immune checkpoint blockade as a novel treatment.

Loss of abnormal spindle-like, microcephaly-associated (Aspm) disrupts female folliculogenesis in mice during maturation and aging.

The abnormal spindle-like, microcephaly-associated (ASPM) gene is a causative gene of autosomal recessive primary microcephaly (MCPH) 5 in humans, which is characterized by a reduction in brain volume. It was previously reported that truncated Aspm proteins in transgenic mice caused major defects in the germline, a severe reduction in ovary weight and the number of follicles accompanied by reduced fertility. However; it remains unknown whether a loss of Aspm induces abnormal ovarian function, resulting in female infertility. In order to assess the ovary function, we examined vaginal smear cytology from the age of 7 weeks to 100 weeks in CAG-mediated Cre-loxP conditional Aspm-/- knockout mice and control female mice. In addition, we evaluated the ovarian size, fibrosis ratio and the number of follicles (primordial, primary, secondary, antral and atretic follicles) in mice from 15 weeks to 100 weeks old by image analyses. Mann-Whitney U-test was used for statistical analysis. The size of the ovary was significantly reduced in Aspm knockout mice at 15-20 weeks, 40-50 weeks and 70-80 weeks old compared with the control mice. Furthermore, at all stages, we found a severe decrease in the number of developing follicles at 10-15 weeks, 40-50 weeks and 70-80 weeks old, accompanied by disrupted cyclic changes of vaginal cytology and an aberrant upregulation of Foxo3, Kitl, and Lhcgr in Aspm knockout female. These results suggested that Aspm might play an important role in the folliculogenesis and estrous cyclicity of the postnatal ovary during maturation and aging.

Spatially resolved immune microenvironmental profiling for follicular thyroid carcinoma with minimal capsular invasion.

Spatial profiles of the tumor-immune microenvironment are associated with disease progression and clinicopathological factors in various cancers. Follicular thyroid carcinoma (FTC) is the second most common thyroid cancer, where the presence of capsular invasion or angioinvasion determines the pathological diagnosis; however, little is known about the immune microenvironment profiles associated with the acquisition of invasive potential of FTC. In this study, we focused on FTC with minimal capsular invasion, and the spatially resolved immune microenvironment of FTC was studied in the discovery (n = 13) and validation cohorts (n = 40). CD8+ T cells, helper T cells, regulatory T cells, B cells, natural killer cells, tumor-associated macrophages, CD66+ granulocytes, mature dendritic cells, and mast cells were quantitatively evaluated in single tissue sections, via a 12-marker multiplex immunohistochemistry and image cytometry. Cell densities and compositions of immune cells were spatially stratified by six tissue regions including tumor center, subcapsular region, capsular invasion, adjacent stroma of capsular invasion, peritumoral stroma, and adjacent normal. Lymphoid cell lineages in the tumor center and subcapsular regions were significantly lower than those in adjacent normal and peritumoral stroma, potentially related to the lymphoid lineage exclusion from the intratumoral regions of FTC. Interestingly, immune cell composition profiles in the capsular invasive front were distinct from those of intratumoral region. The ratios of T cells to CD66b+ granulocytes with capsular invasion were significantly higher than those without capsular invasion, suggesting the presence of a unique immune microenvironment at the invasive front between tumor foci and stroma. In addition, tumor cells at the capsular invasive front showed significantly higher expression of tumor programmed cell death ligand 1 (PD-L1) than those at the tumor center. This study revealed spatial immune profiles associated with capsular invasion of FTC, providing new insights into the mechanisms underlying its development and initial invasion.

Spatial Profiles of Intratumoral PD-1+ Helper T Cells Predict Prognosis in Head and Neck Squamous Cell Carcinoma.

Background: Functional interactions between immune cells and neoplastic cells in the tumor immune microenvironment have been actively pursued for both biomarker discovery for patient stratification, as well as therapeutic anti-cancer targets to improve clinical outcomes. Although accumulating evidence indicates that intratumoral infiltration of immune cells has prognostic significance, limited information is available on the spatial infiltration patterns of immune cells within intratumoral regions. This study aimed to understand the intratumoral heterogeneity and spatial distribution of immune cell infiltrates associated with cell phenotypes and prognosis in head and neck squamous cell carcinoma (HNSCC). Methods: A total of 88 specimens of oropharyngeal squamous cell carcinoma, categorized into discovery (n = 38) and validation cohorts (n = 51), were analyzed for immune contexture by multiplexed immunohistochemistry (IHC) and image cytometry-based quantification. Tissue segmentation was performed according to a mathematical morphological approach using neoplastic cell IHC images to dissect intratumoral regions into tumor cell nests versus intratumoral stroma. Results: Tissue segmentation revealed heterogeneity in intratumoral T cells, varying from tumor cell nest-polarized to intratumoral stroma-polarized distributions. Leukocyte composition analysis revealed higher ratios of TH1/TH2 in tumor cell nests with higher percentages of helper T cells, B cells, and CD66b+ granulocytes within intratumoral stroma. A discovery and validation approach revealed a high density of programmed death receptor-1 (PD-1)+ helper T cells in tumor cell nests as a negative prognostic factor for short overall survival. CD163+ tumor-associated macrophages (TAM) provided the strongest correlation with PD-1+ helper T cells, and cases with a high density of PD-1+ helper T cells and CD163+ TAM had a significantly shorter overall survival than other cases. Conclusion: This study reveals the significance of analyzing intratumoral cell nests and reports that an immune microenvironment with a high density of PD-1+ helper T cells in tumoral cell nests is a poor prognostic factor for HNSCC.

A 14-Marker Multiplexed Imaging Panel for Prognostic Biomarkers and Tumor Heterogeneity in Head and Neck Squamous Cell Carcinoma.

Recent advances made in treatment for head and neck squamous cell carcinoma (HNSCC) highlight the need for new prediction tools to guide therapeutic strategies. In this study, we aimed to develop a HNSCC-targeting multiplex immunohistochemical (IHC) panel that can evaluate prognostic factors and the intratumor heterogeneity of HNSCC. To identify IHC-based tissue biomarkers that constitute new multiplex IHC panel, a systematic review and meta-analysis were performed to analyze reported IHC biomarkers in laryngeal and pharyngeal SCC in the period of 2008-2018. The Cancer Genome Atlas (TCGA) and Reactome pathway databases were used to validate the prognostic and functional significance of the identified biomarkers. A 14-marker chromogenic multiplex IHC panel including identified biomarkers was used to analyze untreated HNSCC tissue. Forty-five high-quality studies and thirty-one candidate tissue biomarkers were identified (N = 7062). Prognostic validation in TCGA laryngeal and pharyngeal SCC cohort (N = 205) showed that ß-catenin, DKK1, PINCH1, ADAM10, and TIMP1 were significantly associated with poor prognosis, which were related to functional categories such as immune system, cellular response, cell cycle, and developmental systems. Selected biomarkers were assembled to build a 14-marker panel, evaluating heterogeneity and polarized expression of tumor biomarkers in the tissue structures, which was particularly related to activation of Wnt/ß-catenin pathway. Integrated IHC analysis based on a systemic review and meta-analysis provides an in situ proteomics tool to assess the aggressiveness and intratumor heterogeneity of HNSCC.

Expression profiles of E/P receptors and fibrosis in GnRHa-treated and -untreated women with different uterine leiomyomas.

Differential expressions of estrogen/progesterone receptors (ER/PR) and individual component of extracellular matrices derived from fibroid are reported. Information on the pattern of change in ER/PR expression and amount of tissue fibrosis after hormonal treatment is unclear. We investigated pattern of change in ER/PR expression and percentage of tissue fibrosis in different uterine leiomyomas after gonadotropin-releasing hormone agonist (GnRHa) treatment. Biopsy specimens from fibroids and adjacent myometria were collected after surgery from women with submucosal myoma (SMM, n = 18), intramural myoma (IMM, n = 16) and subserosal myoma (SSM, n = 17). A proportion of women in each group of fibroid underwent treatment with GnRHa for a variable period of 3-6 months. Tissue expression of ER and PR was analyzed by immunohistochemistry. In vitro cell proliferation effect of GnRHa on human umbilical vein endothelial cells (HUVECs) was examined. Distribution of tissue fibrosis was examined by Masson's trichrome staining with computer-captured image analysis of fibrosis derived from different types of fibroid. PR content was significantly higher than ER in tissues derived from GnRHa-untreated women with SMM and SSM (p = 0.04 for both). Comparing to untreated group, GnRHa-treatment significantly decreased either ER or PR expression in different fibroids. Exogenous treatment with GnRHa dose-dependently decreased proliferation of HUVECs. No significant difference was observed in the percentage of fibrosis in tissues collected from GnRHa-treated and -untreated women with fibroids. The distribution of fibrosis in myoma/myometria and occurrence of fibrosis in perivascular area showed an increasing trend with higher age of the women and with larger size of fibroids. Our findings suggest that despite estrogen dependency, higher PR content in GnRHa-untreated group may indicate a potential role of progesterone in leiomyoma growth. Although GnRHa therapy may shrink fibroids and reduce risk of bleeding during surgery, the occurrence of diffuse tissue fibrosis may impair effective reduction of fibroid size after hormonal treatment.

Prognostic significance of spatial immune profiles in human solid cancers.

Immune-based tumor characteristics in the context of tumor heterogeneity are associated with suppression as well as promotion of cancer progression in various tumor types. As immunity typically functions based on intercellular contacts and short-distance cytokine communications, the location and spatial relationships of the tumor immune microenvironment can provide a framework to understand the biology and potential predictive biomarkers related to disease outcomes. Immune spatial analysis is a newly emerging form of cancer research based on recent methodological advances in in situ single-cell analysis, where cell-cell interaction and the tissue architecture can be analyzed in relation to phenotyping the tumor immune heterogeneity. Spatial characteristics of tumors can be stratified into the tissue architecture level and the single-cell level. At the tissue architecture level, the prognostic significance of the density of immune cell lineages, particularly T cells, is leveraged by understanding longitudinal changes in cell distribution in the tissue architecture such as intra-tumoral and peri-tumoral regions, and invasive margins. At the single-cell level, the proximity of the tumor to the immune cells correlates with disease aggressiveness and therapeutic resistance, providing evidence to understand biological interactions and characteristics of the tumor immune microenvironment. In this review, we summarize recent findings regarding spatial information of the tumor immune microenvironment and review advances and challenges in spatial single-cell analysis toward developing tissue-based biomarkers rooted in the immune spatial landscape.

Maldevelopment of intrapulmonary bronchial cartilage in congenital diaphragmatic hernia.

BACKGROUND: Pulmonary hypoplasia is an important cause of morbidity and mortality in infants with congenital diaphragmatic hernia (CDH). This study aimed to verify our hypothesis that the abnormal development of bronchial cartilage as well as alveolar immaturity, might play a central role in hypoplasia of the lung in human CDH. METHOD: We retrospectively analyzed autopsied lungs from 10 CDH cases and compared with nine age-matched controls to assess the bronchial cartilage and alveolar maturity using morphological techniques. RESULT: Ki-67 and thyroid transcription factor-1 (TTF-1) expression in the alveoli significantly increased in bilateral lungs with CDH. The shortest distance from the bronchial cartilage to the pleura was significantly shorter in ipsilateral (left) lungs with CDH, showing a positive correlation with the radial alveolar count (RAC). Regarding the small bronchial cartilages less than 20 000 µm2 , the average cartilage area significantly decreased in left lungs with CDH, and tended to decrease in right lungs with CDH. In addition, cartilage around the bronchi less than 200 µm in diameter tended to be smaller in left lungs with CDH. In contrast, regarding the cartilage around the bronchi 200 to 400 µm in diameter, the ratio of the total cartilage area relative to the bronchial diameter tended to be higher in left lungs with CDH, although there was a large variation. CONCLUSIONS: These opposite directional cartilage abnormalities around the distal and more proximal bronchi support our hypothesis that abnormal development of bronchial cartilage might play an important role in the hypoplastic lung in CDH.

Programmed death-ligand 1 expression profiling in thymic epithelial cell tumors: Clinicopathological features and quantitative digital image analyses.

OBJECTIVES: Whether the extent of expression of programmed death-ligand 1 (PD-L1) is clinically significant remains uncertain, although immuno-oncological features have been studied in thymic epithelial cell tumors (TETs). We investigated the histological features of PD-L1 expression in TETs, and assessed PD-L1 expression using digital image analysis. MATERIALS AND METHODS: Participants comprised 66 patients with TET who underwent surgical resection between 2001 and 2016. We calculated tumor cell-positive ratio as total proportion score (TPS) with immunohistochemistry using SP263 anti-PD-L1 monoclonal antibody. PD-L1 expression was also quantified using digital image analysis of whole-slide images. We evaluated the relationship between conventional visual TPS using optical microscopy (TPS-V) and TPS from digital image analysis (TPS-IA). We further classified all TETs into high or low PD-L1 expression groups and assessed the clinical significance of PD-L1 expression level using TPS-V and TPS-IA. RESULTS: WHO histological types were Type A (n = 8), AB (n = 18), B1 (n = 5), B2 (n = 16), B3 (n = 6), metaplastic thymoma (n = 2), and thymic carcinoma (TC) (n = 11). Median TPS-Vs were 2%, 2%, 10 %, 65 %, 90 %, 1%, and 20 %, respectively. TPS-IAs correlated with TPS-Vs in TETs overall and in thymomas, but not in TCs. PD-L1 expression levels in TETs differed significantly among histological types. Whether TPS-V or TPS-IA were used, the PD-L1high group included more cases of the more aggressive histological types. Recurrence-free survival (RFS) was shorter in the PD-L1high group than in the PD-L1low group in thymoma using TPS-IA, whereas RFS of the PD-L1high group was shorter in all TETs using TPS-V. CONCLUSION: PD-L1 expression levels depended on the histological type of TET. Extensive PD-L1 expression in TETs was associated with poor prognosis. Digital image analysis is feasible for evaluating PD-L1 expression in TETs and might offer clinically relevant features of thymomas.

Biological differences between intrinsic and extrinsic adenomyosis with coexisting deep infiltrating endometriosis.

RESEARCH QUESTION: Is there a biological difference between intrinsic and extrinsic adenomyosis with coexisting deep infiltrating endometriosis (DIE)? DESIGN: In this prospective controlled study, biopsy specimens were collected after surgery from 23 women with intrinsic adenomyosis and 10 women with extrinsic adenomyosis with coexisting DIE lesions. Histological evaluation was carried out by immunoreaction to Ber-EP4 (epithelial cell marker) and CD10 (stromal cell marker). Tissue expression of oestrogen and progesterone receptors was analysed by immunohistochemistry. Tissue fibrosis was examined by Masson's trichrome staining with computer-based image analysis of fibrosis in respective samples. RESULTS: The detection rate of coexistent DIE was significantly higher in women with extrinsic adenomyosis (9/10 [90.0%]) than in women with intrinsic adenomyosis (3/23 [13.0%]; P < 0.001). The pattern of Ber-EP4-stained glands and CD10-stained stromal cells of extrinsic adenomyosis was similar to that of coexistent DIE lesions. In contrast, the pattern of gland and stromal cells was similar to the endometrium in the cases with intrinsic adenomyosis. Unlike extrinsic adenomyosis, progesterone receptor expression was significantly decreased in both gland cells (P < 0.05) and stromal cells (P < 0.05) of intrinsic adenomyosis. Although relatively more fibrosis was seen in biopsy samples of extrinsic adenomyosis and coexistent DIE than in intrinsic adenomyosis and their coexistent DIE, no significant difference was found. CONCLUSIONS: Extrinsic adenomyosis may be considered as adenomyosis externa based on a close histological and biological relationship between extrinsic adenomyosis and coexistent DIE. Our findings may contribute to the understanding of a possible biological origin of two newly classified intrinsic and extrinsic adenomyosis.

Biological differences between focal and diffuse adenomyosis and response to hormonal treatment.

RESEARCH QUESTION: Is there any difference in ovarian steroid receptor expression and pattern of fibrosis in focal and diffuse adenomyosis and response to hormonal treatment? DESIGN: Prospective controlled study where biopsy samples were prospectively collected after surgery from 30 women with focal adenomyosis, 21 women with diffuse adenomyosis and 20 women with uterine myoma. Some of these women underwent 3-6 months of treatment with gonadotrophin-releasing hormone agonist (GnRHa) before surgery. Tissue expression of oestrogen receptor (ER) and progesterone receptor (PR) was analysed by immunohistochemistry. Distribution of tissue fibrosis was examined by Masson's trichrome staining with computer-based image analysis of fibrosis in tissues derived from women with and without adenomyosis. RESULTS: There was no difference in ER/PR expression in gland cells/stromal cells of adenomyotic lesions on the ipsilateral side of focal adenomyosis and the anterior/posterior walls of diffuse adenomyosis. Compared to myoma tissues, a relatively decreased expression of ovarian steroid receptors was observed in both focal and diffuse adenomyosis. Image analysis of tissue fibrosis indicated more fibrosis in both focal and diffuse adenomyosis compared to fibrosis in the myometrium derived from women with uterine myoma. The pattern of fibrosis was no different in tissues derived from GnRHa-treated and -untreated women with focal and diffuse adenomyosis. CONCLUSIONS: No difference was found in the expression of ER/PR and entity of fibrosis between women with focal and diffuse adenomyosis regardless of GnRHa treatment. A lower expression of ER/PR compared to myoma tissue potentially clarifies the biological rationale of non-response to hormonal therapies for adenomyosis.

Label-free classification of neurons and glia in neural stem cell cultures using a hyperspectral imaging microscopy combined with machine learning.

Due to a growing demand for a viable label-free observation method in the biomedical field, many techniques, such as quantitative phase imaging and Raman spectroscopy, have been studied, and a complementary approach, hyperspectral imaging, has also been introduced. We developed a high-speed hyperspectral imaging microscopy imaging method with commercially available apparatus, employing a liquid crystal tunable bandpass filter combined with a pixel-wise machine learning classification. Next, we evaluated the feasibility of the application of this method for stem cell research utilizing neural stem cells. Employing this microscopy method, with a 562 × 562 µm2 field of view, 2048 × 2048 pixel resolution images containing 63 wavelength pixel-wise spectra could be obtained in 30 seconds. The neural stem cells were differentiated into neurons and astroglia (glia), and a four-class cell classification evaluation (including neuronal cell body, glial cell body, process and extracellular region) was conducted under co-cultured conditions. As a result, an average of 88% of the objects of interest were correctly classified, with an average precision of 94%, and more than 99% of the extracellular pixels were correctly segregated. These results indicated that the proposed hyperspectral imaging microscopy is feasible as a label-free observation method for stem cell research.

Increased Urinary Liver-Type Fatty Acid-Binding Protein Level Predicts Worsening Renal Function in Patients With Acute Heart Failure.

BACKGROUND: Urinary liver-type fatty acid-binding protein (L-FABP) is a potential biomarker for acute kidney injury, and it in turn increases cardiovascular mortality. We tested whether the urinary L-FABP level predicted short- and mid-term outcomes in patients with acute heart failure. METHODS AND RESULTS: We enrolled consecutive patients with acute heart failure, and measured their urinary L-FABP levels before acute treatment. Worsening renal function (WRF), defined as both an absolute increase in the serum creatinine level of ≥0.3mg/dL and a ≥25% relative increase in its level from baseline, occurred in 37 (26.8%) of 138 patients. Patients with a urinary L-FABP level above the upper normal limit (8.4 µg/g creatinine) (n = 49; 35.5%) were more likely than those with a urinary L-FABP level within normal limits (n = 89; 64.5%) to develop WRF (n = 26 [53.1%] vs n = 11 [12.4%]; P < .001). A urinary L-FABP level above the upper limit was independently associated with WRF (hazard ratio 1.8; P = .01). During 1 year of follow-up, 12 patients (8.7%) died, and urinary L-FABP level had no association with all-cause mortality. There was, however, a tendency toward a higher readmission rate in patients with a urinary L-FABP level above the upper normal limit who survived the index hospitalization (n = 46) than in those without an abnormal L-FABP level (n = 88; n = 13 [28.3%] vs n = 13 [14.8%]; log-rank P = .06). CONCLUSIONS: Increased urinary L-FABP level before treatment may predict WRF in patients with acute heart failure. Further investigation is warranted for its predictive ability of adverse outcomes.

Usefulness of Sweep Imaging With Fourier Transform for Evaluation of Cortical Bone in Diabetic Rats.

BACKGROUND: Diabetes decreases bone strength, possibly because of cortical bone changes. Sweep imaging with Fourier transform (SWIFT) has been reported to be useful for cortical bone evaluation. PURPOSE: To evaluate cortical bone changes in diabetic rats using SWIFT, assess the usefulness of this technique through comparisons with microcomputed tomography (µCT) and conventional MRI, and clarify the mechanism underlying cortical bone changes using histomorphometry STUDY TYPE: Animal cohort. ANIMAL MODEL: 8-week-old male Wistar/ST rats (N = 36) were divided into diabetes (induced by streptozotocin injection) and control groups. FIELD STRENGTH/SEQUENCE: 7.04T MRI, SWIFT. ASSESSMENT: Six animals from each group were sacrificed at 2, 4, and 8 weeks after injection. Tibial bones were extracted and evaluated using µCT and MRI. The cortical bone mineral density (BMD) was measured using µCT. Proton density-weighted imaging (PDWI) and SWIFT were also performed. The signal-to-noise ratio (SNR) was calculated for each acquisition. The bone formation rate was evaluated using histomorphometry. STATISTICAL TESTS: Findings at each timepoint were compared using Mann-Whitney U-tests. RESULTS: Cortical BMD was significantly lower in the diabetes group than in the control group only at 8 weeks (P < 0.05). At all timepoints, PDWI-SNR showed no significant differences between groups (P = 0.59, 0.70, and 0.82 at 2, 4, and 8 weeks, respectively). SWIFT-SNR was significantly lower in the diabetes group than in the control group (P < 0.05 at 2 and 4 weeks and P < 0.01 at 8 weeks), and the bone formation rate was significantly lower in the diabetes group than in the control group (P < 0.01 for all). DATA CONCLUSION: SWIFT can detect cortical bone changes even before a decline in the cortical BMD in a diabetic model. LEVEL OF EVIDENCE: 1 Technical Efficacy: Stage 3 J. MAGN. RESON. IMAGING 2018;48:389-397.

Longitudinal Diffusion Tensor Imaging Revealed Nerve Fiber Alterations in Aspm Mutated Microcephaly Model Mice.

Autosomal recessive primary microcephaly-5 (MCPH5) is characterized by congenital microcephaly and is caused by the mutation in the abnormal spindle-like, microcephaly-associated (ASPM) gene. This study aimed to demonstrate a correlation between radiological and pathological analyses in evaluating postnatal brain development using MCPH5-model mice, ASPM ortholog (Aspm) knockout (KO) mice. In vivo MRI was performed at two time points (postnatal 3 weeks; P3W and P10W) and complementary histopathological analyses of brains were done at P5W and P13W. In the MRI analysis, Aspm KO mice showed significantly decreased brain sizes (average 8.6% difference) with larger ventricles (average 136.4% difference) at both time points. Voxel-based statistics showed that the fractional anisotropy (FA) values were significantly lower in Aspm KO mice in both the cortex and white matter at both time points. Developmental changes in the FA values were less remarkable in the Aspm KO mice, compared with the controls. Histometric analyses revealed that the ratios of the horizontal to the vertical neurites were significantly higher in cortical layers IV, V and VI, with a remarkable increase according to maturation at P13W in the control mice (average 12.7% difference between control and KO), whereas the ratio in layer VI decreased at P13W in the KO mice. The myelin basic protein positive ratio in the white matter significantly decreased in Aspm KO mice at P5W. These results suggest that temporal FA changes are closely correlated with pathological findings such as abnormal neurite outgrowth and differentiation, which may be applicable for analyzing diseased human brain development.

Prenatal and lactational bisphenol A exposure does not alter serotonergic neurons morphologically in the murine dorsal raphe nucleus.

OBJECTIVE: There is concern that bisphenol A (BPA), an endocrine-disrupting chemical, affects brain development when exposed to a fetus and/or infant. We previously reported that increased serotonin (5-HT) and its metabolite (5-HIAA) in the dorsal raphe nucleus (DRN) in murine adult brains when they were prenatally exposed to low doses of BPA. This study investigates the morphological alteration of the dorsal raphe nucleus (DRN) in order to explain the disrupted serotonergic system after prenatal and lactational exposure to bisphenol A (BPA). METHODS: The murine dams were orally administrated with 500µg/kg/day of BPA from embryonic day 0 to postnatal 3weeks. The DRN, the main region of serotonin production, was morphometrically analyzed at 14weeks, using immunohistochemistry and image analysis combined with 3-dimensional reconstruction. RESULTS: No significant differences were revealed in the number of tryptophan hydroxylase 2-immunoreactive neurons in any of the DRN sub-regions or the morphometric parameters, including the whole volume, ventrodorsal, longitudinal, and wing lengths of the DRN among the BPA treatment and sex groups. CONCLUSIONS: The murine DRN was not morphologically affected by prenatal and lactational exposure to low doses of BPA. Further studies are necessary regarding the function of serotonergic neurons and the activity of different kinds of related receptors in the brain.

Alterations of neurotransmitter norepinephrine and gamma-aminobutyric acid correlate with murine behavioral perturbations related to bisphenol A exposure.

AIM: Humans are commonly exposed to endocrine-disrupting chemical bisphenol A (BPA), giving rise to concern over the psychobehavioral effects of BPA. The aim of this study was to investigate the effects of prenatal and lactational BPA exposure on neurotransmitters, including norepinephrine (NE), gamma-aminobutyric acid (GABA) and glutamate (Glu), and to assess the association with behavioral phenotypes. METHODS: C57BL/6J mice were orally administered with BPA (500 µg/bwkg/day) or vehicle daily from embryonic day 0 to postnatal week 3 (P3W), through their dams. The IntelliCage behavioral experiments were conducted from P11W to P15W. At around P14-16W, NE, GABA and Glu levels in nine brain regions were measured by high performance liquid chromatography. Furthermore, the associations between the neurotransmitter levels and the behavioral indices were statistically analyzed. RESULTS: In females exposed to BPA, the GABA and Glu levels in almost all regions, and the NE levels in the cortex, hypothalamus and thalamus were higher than those in the controls. In males exposed to BPA, the GABA levels in the amygdala and hippocampus showed lower values, while Glu levels were higher in some regions, compared with the controls. In regard to the associations, the number of "diurnal corner visits without drinking" was correlated with the NE levels in the cortex and thalamus in females. The "nocturnal corner visit duration without drinking" was correlated with the GABA level in the hippocampus in males. CONCLUSION: These results suggest that prenatal and lactational exposure to low doses of BPA might modulate the NE, GABA and Glu systems, resulting in behavioral alterations.