Hydroxytyrosol has various pharmacological properties, including anti-oxidative stress and anti-inflammatory activities, preventing hyperglycemia, insulin resistance, and the metabolic syndrome. The present study is focused on the anti-adipogenic and lipolytic activity of hydroxytyrosol on primary human visceral adipocytes. Pre-adipocytes were analyzed after 10 (P10) and 20 (P20) days of treatment during differentiation and after 7 (A7) days of treatment when they reached mature shape. The treatment with hydroxytyrosol extract significantly (P < 0.001) increased apoptosis in P10 and P20 cells in comparison to control and A7 cells; significantly (P < 0.001) reduced triglyceride accumulation in P20 cells compared to P10 and control cells; and significantly (P < 0.001) increased lipolysis in P20 cells in comparison to control cells and A7 mature adipocytes. Hydroxytyrosol-treated P20 cells significantly (P < 0.05) increased expression of genes involved in inhibition of adipogenesis, such as GATA2, GATA3, WNT3A, SFRP5, HES1, and SIRT1. In contrast, genes involved in promoting adipogenesis such as LEP, FGF1, CCND1, and SREBF1 were significantly down-regulated by hydroxytyrosol treatment. These data suggest that hydroxytyrosol promotes lipolysis and apoptotic activity in primary human visceral pre-adipocytes during differentiation and does not affect already mature adipocytes.

Mesonephric-like adenocarcinomas are rare neoplasms occurring in the uterine corpus and ovary which bear a close morphologic resemblance to cervical mesonephric adenocarcinomas. They also have a similar immunophenotype and harbor similar molecular abnormalities to mesonephric adenocarcinomas and it is debated whether they are truly of mesonephric origin or represent Mullerian neoplasms closely mimicking mesonephric adenocarcinomas. We report an unusual case with bilateral ovarian serous borderline tumors and extraovarian low-grade serous carcinoma (invasive implants). In one ovary, there was a component of mesonephric-like adenocarcinoma. The immunophenotypes of the serous and the mesonephric-like components were distinct and as expected for the individual tumor types (serous component diffusely positive with WT1 and estrogen receptor and negative with GATA3, TTF1 and CD10; mesonephric-like component WT1 and estrogen receptor negative and GATA3, TTF1, and CD10 positive; both components diffusely positive with PAX8 and exhibiting "wild-type" p53 immunoreactivity). In all components (bilateral serous borderline tumors, low-grade serous carcinoma and mesonephric-like adenocarcinoma), an identical KRAS mutation was detected (NM_004985.4): c.35G>A, p.(G12D) proving a clonal association between the serous and mesonephric-like components and excluding a collision neoplasm. This represents the second reported case of a combined ovarian low-grade serous tumor and mesonephric-like adenocarcinoma; in the previously reported case, an identical NRAS mutation was present in both components. These 2 cases provide evidence that ovarian mesonephric-like adenocarcinomas have, at least in some cases, a Mullerian origin and differentiate along mesonephric lines. We present additional evidence for this by reviewing associated findings in published and unpublished ovarian mesonephric-like adenocarcinomas; 8 of 11 of these neoplasms contained other Mullerian lesions in the same ovary, mainly endometriosis and adenomas/adenofibromas.

Background: Mesonephric-like adenocarcinoma (M-LAC) is a rare, recently described tumor occurring in the uterine corpus and ovary, which shares the same morphological and immunohistochemical features with the more common mesonephric adenocarcinoma (MAC), which mostly arises the uterine cervix. Despite the similarities between these tumors, the histogenesis of M-LAC is still disputable.

Case presentation: Sixty-one-year-old woman presented with an advanced tumor of the left ovary with intraabdominal spread and liver metastases. After receiving 5 cycles of neoadjuvant chemotherapy, she underwent a hysterectomy with bilateral salpingo-oophorectomy, and resection of the liver metastasis, omentum, and appendix. Histologically, the ovarian tumor consisted of two components, whose morphology and immunohistochemical results were typical of either a serous borderline tumor (immunohistochemical positivity for PAX8, WT1, ER and PR) or a mesonephric-like carcinoma (immunohistochemical positivity for PAX8, TTF1 and GATA3). Only the component of the mesonephric-like adenocarcinoma metastasized to the omentum and liver. A molecular analysis with a panel of 271 genes (size 1020 kbp) was performed separately on samples from the borderline tumor, primary ovarian mesonephric-like adenocarcinoma, and liver metastasis. The results showed the clonal origin of all samples, which shared the same KRAS (NM_004985.3:c.34G > T, p.(G12C)) and PIK3CA (NM_006218.2:c.1633G > A, p.(E545K)) somatic mutations. Moreover, in the sample from the primary mesonephric-like carcinoma and its liver metastasis a likely pathogenic somatic MYCN mutation (NM_005378.4:c.131C > T, p.(P44L) was found. In all samples, the deletion of exons 9-10 in the CHEK2 gene was present, which is in concordance with the previously performed genetic testing of the blood specimen which revealed the hereditary CHEK2 mutation in this patient.

Conclusions: Our result support the theory that at least some mesonephric-like ovarian adenocarcinomas are of Müllerian origin. The serous borderline tumor seems to be a precursor of mesonephric-like adenocarcinoma, which has been proven in our case by both tumors sharing the same mutations, and the presence of cumulative molecular aberrations in the mesonephric-like adenocarcinoma.

Keywords: Case report; Mesonephric-like adenocarcinoma; Ovary; Serous borderline tumor.

Renal cell carcinoma (RCC) is the most common type of kidney cancer in adults and accounts for ~80% of all kidney cancer cases. However, the pathogenesis of RCC has not yet been fully elucidated. To interpret the pathogenesis of RCC at the molecular level, gene expression data and bio-informatics methods were used to identify RCC associated genes. Gene expression data was downloaded from Gene Expression Omnibus (GEO) database and identified differentially coexpressed genes (DCGs) and dysfunctional pathways in RCC patients compared with controls. In addition, a regulatory network was constructed using the known regulatory data between transcription factors (TFs) and target genes in the University of California Santa Cruz (UCSC) Genome Browser (http://genome.ucsc.edu) and the regulatory impact factor of each TF was calculated. A total of 258,0427 pairs of DCGs were identified. The regulatory network contained 1,525 pairs of regulatory associations between 126 TFs and 1,259 target genes and these genes were mainly enriched in cancer pathways, ErbB and MAPK. In the regulatory network, the 10 most strongly associated TFs were FOXC1, GATA3, ESR1, FOXL1, PATZ1, MYB, STAT5A, EGR2, EGR3 and PELP1. GATA3, ERG and MYB serve important roles in RCC while FOXC1, ESR1, FOXL1, PATZ1, STAT5A and PELP1 may be potential genes associated with RCC. In conclusion, the present study constructed a regulatory network and screened out several TFs that may be used as molecular biomarkers of RCC. However, future studies are needed to confirm the findings of the present study.

The intimate link between location of fat accumulation and metabolic disease risk and depot-specific differences is well established, but how these differences between depots are regulated at the molecular level remains largely unclear. Here we show that TRIP-Br2 mediates endoplasmic reticulum (ER) stress-induced inflammatory responses in visceral fat. Using in vitro, ex vivo and in vivo approaches, we demonstrate that obesity-induced circulating factors upregulate TRIP-Br2 specifically in visceral fat via the ER stress pathway. We find that ablation of TRIP-Br2 ameliorates both chemical and physiological ER stress-induced inflammatory and acute phase response in adipocytes, leading to lower circulating levels of inflammatory cytokines. Using promoter assays, as well as molecular and pharmacological experiments, we show that the transcription factor GATA3 is responsible for the ER stress-induced TRIP-Br2 expression in visceral fat. Taken together, our study identifies molecular regulators of inflammatory response in visceral fat that-given that these pathways are conserved in humans-might serve as potential therapeutic targets in obesity.

The functions of POU class 5 transcription factor 1 (Oct-4) and caudal-type homeobox 2 (Cdx2) in the differentiation of the murine inner cell mass (ICM) and trophectoderm (TE) have been described in detail. However, little is known about the roles of OCT-4 and CDX2 in preimplantation bovine embryos. To elucidate their functions during early development in bovine embryos, we performed OCT-4 and CDX2 downregulation using RNA interference. We injected OCT-4- or CDX2-specific short interfering RNAs (siRNAs) into bovine zygotes. The rate of blastocyst development of OCT-4-downregulated embryos was lower compared with uninjected or control siRNA-injected embryos. Gene expression analysis revealed decreased CDX2 and fibroblast growth factor 4 expression in OCT-4-downregulated embryos. CDX2-downregulated embryos developed to the blastocyst stage; however, in most cases, blastocoel formation was delayed. Gene expression analysis revealed decreased GATA3 expression and elevated NANOG expression in CDX2-downregulated embryos. In conclusion, OCT-4 and CDX2 are essential for early development and gene expression involved in differentiation of ICM and TE lineages in bovine embryos.

GATA transcription factors are implicated in establishing cell fate during mammalian development. In early mammalian embryos, GATA3 is selectively expressed in the extraembryonic trophoblast lineage and regulates gene expression to promote trophoblast fate. However, trophoblast-specific GATA3 function is dispensable for early mammalian development. Here, using dual conditional knockout mice, we show that genetic redundancy of Gata3 with paralog Gata2 in trophoblast progenitors ensures the successful progression of both pre- and postimplantation mammalian development. Stage-specific gene deletion in trophoblasts reveals that loss of both GATA genes, but not either alone, leads to embryonic lethality prior to the onset of their expression within the embryo proper. Using ChIP-seq and RNA-seq analyses, we define the global targets of GATA2/GATA3 and show that they directly regulate a large number of common genes to orchestrate stem versus differentiated trophoblast fate. In trophoblast progenitors, GATA factors directly regulate BMP4, Nodal and Wnt signaling components that promote embryonic-extraembryonic signaling cross-talk, which is essential for the development of the embryo proper. Our study provides genetic evidence that impairment of trophoblast-specific GATA2/GATA3 function could lead to early pregnancy failure.

The NuRD chromatin remodeling and deacetylation complex, which includes MTA1, MBD3, CHD4 and HDAC1 among other components, is of importance for development and cancer progression. The oncogenic MUC1-C protein activates EZH2 and BMI1 in the epigenetic reprogramming of triple-negative breast cancer (TNBC). However, there is no known link between MUC1-C and chromatin remodeling complexes. Here we showed that MUC1-C binds directly to the MYC HLH-LZ domain and identified a previously unrecognized MUC1-C->MYC pathway that regulates the NuRD complex. MUC1-C/MYC complexes selectively activated the MTA1 and MBD3 genes and post-transcriptionally induced CHD4 expression in basal- but not luminal-type BC cells. In turn, MUC1-C formed complexes with these NuRD components on the ESR1 promoter. Downregulating MUC1-C decreased MTA1/MBD3/CHD4/HDAC1 occupancy and increased H3K27 acetylation on the ESR1 promoter with induction of ESR1 expression and downstream estrogen response pathways. Targeting MUC1-C and these NuRD components also induced expression of FOXA1, GATA3 and other markers associated with the luminal phenotype. These findings support a model in which MUC1-C activates the NuRD complex to drive dedifferentiation and reprogramming of TNBC cells.

LSD1 (KDM1A) is a histone demethylase that plays both oncogenic and tumor suppressor roles in breast cancer. However, the exact contexts under which it plays these opposite functions remain largely elusive. By characterizing its role in luminal breast epithelial cells, here we show that inhibition of LSD1 by both genetic and pharmacological approaches increases their invasion and migration, whereas its inhibition by genetic approach, but not by pharmacological approach, impairs their proliferation/survival. Induced loss of LSD1 in luminal cells in a mouse model of luminal breast cancer, MMTV-PyMT, leads to a profound increase in lung metastasis. Mechanistically, LSD1 interacts with GATA3, a key luminal-specific transcription factor (TF), and their common target genes are highly related to breast cancer. LSD1 positively regulates GATA3 expression. It also represses expression of TRIM37, a breast epithelial oncogene encoding a histone H2A ubiquitin ligase, and ELF5, a key TF gene for luminal progenitors and alveolar luminal cells. LSD1-loss also leads to reduced expression of several cell-cell adhesion genes (e.g., CDH1, VCL, CTNNA1), possibly via TRIM37-upregulation and subsequently TRIM37-mediated repression. Collectively, our data suggest LSD1 largely plays a tumor suppressor role in luminal breast cancer and the oncogenic program associated with LSD1-inhibition may be suppressed via TRIM37-inhibition.

Dysfunction of noradrenergic (NA) neurons is associated with a number of neuronal disorders. Diverse neuronal subtypes can be generated by direct reprogramming. However, it is still unknown how to convert non-neuronal cells into NA neurons. Here, we show that seven transcription factors (TFs) (Ascl1, Phox2b, AP-2α, Gata3, Hand2, Nurr1, and Phox2a) are able to convert astrocytes and fibroblasts into induced NA (iNA) neurons. These iNA neurons express the genes required for the biosynthesis, release, and re-uptake of noradrenaline. Moreover, iNA neurons fire action potentials, receive synaptic inputs, and control the beating rate of co-cultured ventricular myocytes. Furthermore, iNA neurons survive and integrate into neural circuits after transplantation. Last, human fibroblasts can be converted into functional iNA neurons as well. Together, iNA neurons are generated by direct reprogramming, and they could be potentially useful for disease modeling and cell-based therapies.

At the center of fibrosing diseases is the aberrant activation of tissue fibroblasts. The cellular and molecular mechanisms of how the immune system augments fibroblast activation have been described; however, little is known about how the immune system controls fibroblast function in tissues. Here, we identify regulatory T cells (T_regs) as important regulators of fibroblast activation in skin. Bulk cell and single-cell analysis of T_regs in murine skin and lungs revealed that T_regs in skin are transcriptionally distinct and skewed toward T helper 2 (T_Hregs in lung, skin T_regs preferentially expressed high levels of GATA3, the master T_HGATA3 were highly enriched in skin "T_Hreg" subsets. In functional experiments, T_reg depletion resulted in a preferential increase in T_Hregs resulted in spontaneous skin fibroblast activation, profibrotic gene expression, and dermal fibrosis, all of which were exacerbated in a bleomycin-induced murine model of skin sclerosis. Lineage-specific deletion of Gata3 in T_regs resulted in an exacerbation of T_Hregs play a critical role in regulating fibroblast activation in skin and do so by expressing a unique tissue-restricted transcriptional program that is mediated, at least in part, by GATA3.

Although atrial natriuretic peptide (ANP) has been well recognized for its role in the regulation of volume-pressure homeostasis in cardiovascular system, its impact on respiratory system, particularly on the pathogenesis of acute allergic asthma, is yet to be elucidated. In the present report, we induced mice with OVA for onset of acute allergic asthma along with the administration of recombinant ANP or A71915 (an antagonist for ANP/natriuretic peptide receptor A, NPRA). It was noted that treatment of mice with ANP significantly promoted inflammatory infiltration in the airway and the production of inflammatory cytokines in the bronchoalveolar lavage fluid (BALF) and lung homogenates, and the number of inflammatory cells in the BALF was significantly higher as compared with that of PBS treated asthmatic mice. Moreover, blockade of ANP/NPRA signaling by A71915 almost completely attenuated the effect of ANP administration. Mechanistic studies revealed that ANP repressed the expression of Th1 transcription factor T-bet, but enhanced Th2 transcription GATA3 expression. Together, our data provided feasible evidence suggesting that ANP/NPRA signaling predominantly induces a Th2-type response in favor of pathological processes during the course of acute allergic asthma.

Triple negative breast cancers (TNBCs) are known to express low PGR, ESR1, and ERBB2, and high KRT5, KRT14, and KRT17. However, the reasons behind the increased expressions of KRT5, KRT14, KRT17 and decreased expressions of PGR, ESR1, and ERBB2 in TNBCs are not fully understood. Here we show that, expression of chromosome 19 miRNA cluster (C19MC) specifically marks human TNBCs. Low REST and high CEBPB correlate with expression of C19MC, KRT5, KRT14, and KRT17 and enhancers of these genes/cluster are regulated by CEBPB and REST binding sites. The C19MC miRNAs in turn can potentially target REST to offer a positive feedback loop, and might target PGR, ESR1, ERBB2, GATA3, SCUBE2, TFF3 mRNAs to contribute towards TNBC phenotype. Thus our study demonstrates that C19MC miRNA expression marks TNBCs and that C19MC miRNAs and CEBPB might together determine the TNBC marker expression pattern.

OBJECTIVE

New immunohistochemical (IHC) markers of urothelial carcinoma (UCa) and prostatic adenocarcinoma (PCa) have emerged in recent years, yet comparative studies to establish markers remain lacking. We aimed to identify an effective but parsimonious approach for poorly differentiated bladder neck lesions, to establish a best practice panel approach in a setting simulating prospective use.

METHODS

We tested the performance of a panel of IHC markers on whole sections of a consecutive cohort of transurethral resection specimens of poorly differentiated, challenging bladder neck resections (n=36).

RESULTS

In the setting of poorly differentiated bladder neck carcinomas, biomarker sensitivities for UCa were as follows: GATA3, 100%; S100P, 88%; p63, 75%; and cytokeratin (CK) 5/6, 56%; specificities of each were 100%. CK7 and CK20 showed sensitivities of 75% and 63%, though these were only 85% and 80% specific. For PCa markers, NKX3.1, p501S, prostate-specific membrane antigen, and androgen receptor (AR) each showed 100% sensitivity, outperforming ERG (35%) and prostate-specific antigen (PSA; 25%). All the prostate histogenesis markers were 100% specific, except for AR, which was positive in 13% of the UCa cases.

CONCLUSIONS

Novel IHC markers show improved diagnostic performance that enables positive and negative support for identifying histogenesis with the use of as few as two markers for this critical therapeutic distinction. PSA underperforms newer markers.

GATA3 has been recognized as a promising marker for primary urothelial carcinoma (UC), consistently showing higher expression levels than urothelial markers thrombomodulin and uroplakin III. However, expression of GATA3 in comparison with UC-associated markers CK7 and p63 has not been systematically studied. Moreover, no studies have been conducted to establish GATA3 sensitivity in regional metastases. In this study, high-density tissue microarrays were constructed from 69 matched paired primary and metastatic bladder tumors including pure urothelial UCs with papillary (n=48) or flat phenotype (n=9), mixed tumors with micropapillary, glandular, small cell, squamous, giant cell, and plasmacytoid features (n=9), and 3 adenocarcinomas. GATA3 was expressed in 62/69 (90%) primary UC and 64/69 (93%) metastases, with significantly higher staining intensity in nodal metastases (P=0.03). In primary tumors, GATA3 was positive in 44/48 (92%) papillary UCs, 9/9 (100%) flat UCs, 8/9 (89%) mixed UCs, and 1/3 (33%) adenocarcinomas, whereas in metastases these numbers were 45/48 (94%), 9/9 (100%), 8/9 (89%), and 2/3 (67%), respectively. The majority of positive cases showed strong diffuse nuclear reactivity: 75% of primary UCs and 79% of metastases. GATA3 sensitivity in primary and metastatic UCs was comparable to that of CK7 and superior to that of p63 (P<0.05). GATA3 specificity was computed in comparison with its morphologic mimics expressing CK7 and p63, including 208 primary and 24 metastatic tumors from the lung, cervix, and head and neck regions. Strong GATA3 expression was present in 2/51 (4%) cervical carcinomas, whereas weak GATA3 expression was present in 7/51 (14%) cervical, 6/74 (8%) head and neck cancers, and 2/83 (3%) lung carcinomas. Remaining 191 primary and 24 metastatic tumors were GATA3 negative. Therefore, specificity of GATA3 calculated on the basis of morphologic and immunophenotypic UC mimics from lung, cervix, head and neck was 92%. Our findings demonstrate high sensitivity and specificity of the GATA3 diagnostic marker, with not only maintained but increased expression in regional metastases.

GATA binding protein 3 (GATA3) is a transcription factor, which belongs to a distinct family of tumor suppressor genes. It is involved in human cancer cell growth and differentiation, and plays an important role in cell proliferation and apoptosis. Although, its expression has been reported in various cancers, there are limited data in genitourinary malignancies. Recent studies found GATA3 to be a sensitive marker for urothelial carcinoma (UC) and associated with prognostic pathologic features. Its level of expression was found to be an independent factor predicting cancer recurrence.

In this article, immunohistochemical evaluation of GATA3 expression in genitourinary malignancies (invasive UC, renal cell carcinoma, and prostatic adenocarcinomas) was performed.

GATA3 was positive in 56/79 (70.8%) of invasive UC, and was negative in all renal cell carcinoma and prostatic adenocarcinomas. The pattern of GATA3 staining, when positive, was intensely nuclear within the clusters of malignant cells. No cytoplasmic staining was noted. Negative controls were all negative. High GATA3 expression was associated with larger tumor size in invasive UC (3.19cm vs. 1.65cm, P = 0.01). GATA3 expression did not correlate with other clinicopathologic parameters in UC.

This data suggest that GATA3 is a sensitive marker in confirming invasive UC, and may be helpful in differentiating it from metastatic tumors of renal and prostatic origin. Furthermore, strong GATA3 expression was noted to have an effect on tumor size in patients with UC.

Helminths and their products can shape immune responses by modulating immune cells, which are dysfunctional in inflammatory diseases such as asthma. We previously identified SJMHE1, a small molecule peptide from the HSP60 protein of Schistosoma japonicum. SJMHE1 can inhibit delayed-type hypersensitivity and collagen-induced arthritis in mice. In the present study, we evaluated this peptide's potential intervention effect and mechanism on ovalbumin-induced asthma in mice. SJMHE1 treatment suppressed airway inflammation in allergic mice, decreased the infiltrating inflammatory cells in the lungs and bronchoalveolar lavage fluid, modulated the production of pro-inflammatory and anti-inflammatory cytokines in the splenocytes and lungs of allergic mice, reduced the percentage of Th2 cells and increased the proportion of Th1 and regulatory T cells (Tregs). At the same time, Foxp3 and T-bet expression increased, and GATA3 and RORγt decreased in the lungs of allergic mice. We proved that SJMHE1 can interrupt the development of asthma by diminishing airway inflammation in mice. The down-regulation of Th2 response and the up-regulation of Th1 and Tregs response may contribute to the protection induced by SJMHE1 in allergic mice. SJMHE1 can serve as a novel therapy for asthma and other allergic or inflammatory diseases.

Trophectoderm (TE) lineage development is pivotal for proper implantation, placentation, and healthy pregnancy. However, only a few TE-specific transcription factors (TFs) have been systematically characterized, hindering our understanding of the process. To elucidate regulatory mechanisms underlying TE development, here we map super-enhancers (SEs) in trophoblast stem cells (TSCs) as a model. We find both prominent TE-specific master TFs (Cdx2, Gata3, and Tead4), and >150 TFs that had not been previously implicated in TE lineage, that are SE-associated. Mapping targets of 27 SE-predicted TFs reveals a highly intertwined transcriptional regulatory circuitry. Intriguingly, SE-predicted TFs show 4 distinct expression patterns with dynamic alterations of their targets during TSC differentiation. Furthermore, depletion of a subset of TFs results in dysregulation of the markers for specialized cell types in placenta, suggesting a role during TE differentiation. Collectively, we characterize an expanded TE-specific regulatory network, providing a framework for understanding TE lineage development and placentation.

Neuroendocrine breast carcinomas represent a rare subtype of breast cancer. Their definition, prevalence, and prognosis remain controversial in the literature. The 2012 WHO classification of breast cancer categorizes neuroendocrine carcinomas into three morphologically distinct subtypes: well-differentiated neuroendocrine tumors, poorly differentiated neuroendocrine carcinomas, and invasive breast carcinomas with neuroendocrine differentiation. We aimed to gain insight into the clinical, morphologic, phenotypic, and molecular features of 47 neuroendocrine breast carcinomas. Targeted next-generation sequencing by an AmpliSeq 22 cancer gene hotspot panel and the Prosigna assay were performed on 42/47 and 35/47 cases, respectively. Average age at diagnosis was 69 years. All tumors were estrogen receptor-positive and the large majority expressed progesterone receptor (89%), GATA3 (98%), FOXA1 (96%), and CK8/18 (98%). There was an almost equal distribution of luminal A (52%) and B (48%) carcinomas. Almost half of the cohort (49%) displayed a high risk of recurrence score with the Prosigna test. Patients with a neuroendocrine carcinoma had a shorter disease-free survival compared with those affected by carcinomas of no special type matched for age, size, grade, and estrogen receptor status. No significant differences were observed in terms of overall survival. Stratification of neuroendocrine carcinomas using the 2012 WHO criteria did not reveal statistically significant differences among the distinct categories (well-differentiated neuroendocrine tumors, poorly differentiated neuroendocrine carcinomas, and invasive breast carcinomas with neuroendocrine differentiation), in terms of either progression-free or overall survival. Our targeted sequencing analysis found three cases (7%) harboring a PIK3CA mutation, and in three other cases (7%) TP53 mutations were detected. This study showed that neuroendocrine breast carcinoma is a distinct subtype of luminal carcinoma with a low rate of PIK3CA mutations and with an aggressive clinical behavior. An accurate identification of neuroendocrine differentiation may be useful to better tailor patient adjuvant therapy within luminal carcinomas.

Allergic asthma is a chronic inflammatory disease induced by the inhalation of allergens, which trigger the activation of T helper type 2 (Th2) cells that release Th2 cytokines. Recently, herbal medicines are being considered a major source of novel agents to treat various diseases. In the present study, we evaluated the anti-asthmatic effects of a Codonopsis lanceolata extract (CLE) and the mechanisms involved in its anti-inflammatory effects. Treatment with CLE reduced infiltration of inflammatory cells, especially eosinophils, and the production of mucus in lung tissues. Levels of Th2 cytokines, such as IL-4, IL-5, and IL-13, and chemokines were also decreased following treatment with CLE. Moreover, Th2 cell proportion in vivo and differentiation in vitro were reduced as evidenced by the decreased expression of GATA3⁺. Furthermore, the expression of superoxide dismutase (SOD)2, a mitochondrial ROS (mROS) scavenger, was increased, which was related to Th2 cell regulation. Interestingly, treatment with CLE increased the number of macrophages in the lungs and enhanced the immune-suppressive property of macrophages. Our findings indicate that CLE has potential as a novel therapeutic agent to inhibit Th2 cell differentiation by regulating mROS scavenging.

Estrogen receptor (ER)-positive progesterone receptor (PR)-negative breast cancers are infrequent but clinically challenging. Despite the volume of genomic data available on these tumors, their biology remains poorly understood. Here, we aimed to identify clinically relevant subclasses of ER+/PR- breast cancers based on their mutational landscape. The Cancer Genomics Data Server was interrogated for mutational and clinical data of all ER+ breast cancers with information on PR status from The Cancer Genome Atlas (TCGA), Memorial Sloan Kettering (MSK), and Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) projects. Clustering analysis was performed using gplots, ggplot2, and ComplexHeatmap packages. Comparisons between groups were performed using the Student's t-test and the test of Equal or Given Proportions. Survival curves were built according to the Kaplan⁻Meier method; differences in survival were assessed with the log-rank test. A total of 3570 ER+ breast cancers (PR- n = 959, 27%; PR+ n = 2611, 73%) were analyzed. Mutations in well-known cancer genes such as TP53, GATA3, CDH1, HER2, CDH1, and BRAF were private to or enriched for in PR- tumors. Mutual exclusivity analysis revealed the presence of four molecular clusters with significantly different prognosis on the basis of PIK3CA and TP53 status. ER+/PR- breast cancers are genetically heterogeneous and encompass a variety of distinct entities in terms of prognostic and predictive information.

Much research effort has been focused on investigating new compounds derived from low-cost sources, such as natural products, for treating leishmaniasis. Oleuropein derived from numerous plants, particularly from the olive tree, Olea europaea L. (Oleaceae), is a biophenol with many biological activities. Our previous findings showed that oleuropein exhibits leishmanicidal effects against three Leishmania spp. in vitro, and minimizes the parasite burden in L. donovani-infected BALB/c mice. The aim of the present study is to investigate the possible mechanism(s) that mediate this leishmanicidal activity.

We determined the efficacy of oleuropein in elevating ROS and NO production in L. donovani-infected J774A.1 macrophages and in explanted splenocytes and hepatocytes obtained from L. donovani-infected BALB/c mice. We also assessed the expression of genes that are related to inflammation, T-cell polarization and antioxidant defense, in splenocytes. Finally, we determined the ratios of specific IgG2a/IgG1 antibodies and DTH reactions in L. donovani-infected BALB/c mice treated with oleuropein.

Oleuropein was able to elevate ROS production in both in vitro and in vivo models of visceral leishmaniasis and raised NO production in ex vivo cultures of splenocytes and hepatocytes. The extensive oxidative stress found in oleuropein-treated mice was obviated by the upregulation of the host's antioxidant enzyme (mGCLC) and the simultaneous downregulation of the corresponding enzyme of the parasite (LdGCLC). Moreover, oleuropein was able to mount a significant Th1 polarization characterized by the expression of immune genes (IL-12β, IL-10, TGF-β1, IFN-γ) and transcription factors (Tbx21 and GATA3). Moreover, this immunomodulatory effect was also correlated with an inhibitory effect on IL-1β gene expression, rather than with the expression of IL-1α, IL-1rn and TNF-α. Furthermore, oleuropein-treated BALB/c mice mounted a delayed-type hypersensitivity (DTH) response and an elevated Leishmania-specific IgG2a/IgG1 ratio that clearly demonstrated an in vivo protective mechanism.

The ability of Oleuropein to promote a Th1 type immune response in L. donovani-infected BALB/c mice points towards the candidacy of this bioactive compound as an immunomodulatory agent that may complement therapeutic approaches to leishmaniasis.