Multiregional radiomic model for breast cancer diagnosis: value of ultrasound-based peritumoral and parenchymal radiomics.

Background: Breast cancer consists not only of neoplastic cells but also of significant changes in the surrounding and parenchymal stroma, which can be reflected in radiomics. This study aimed to perform breast lesion classification through an ultrasound-based multiregional (intratumoral, peritumoral, and parenchymal) radiomic model. Methods: We retrospectively reviewed ultrasound images of breast lesions from institution #1 (n=485) and institution #2 (n=106). Radiomic features were extracted from different regions (intratumoral, peritumoral, and ipsilateral breast parenchymal) and selected to train the random forest classifier with the training cohort (n=339, a subset of the institution #1 dataset). Then, the intratumoral, peritumoral, and parenchymal, intratumoral & peritumoral (In&Peri), intratumoral & parenchymal (In&P), and intratumoral & peritumoral & parenchymal (In&Peri&P) models were developed and validated on the internal (n=146, another subset of institution 1) and external (n=106, institution #2 dataset) test cohorts. Discrimination was evaluated using the area under the curve (AUC). Calibration curve and Hosmer-Lemeshow test assessed calibration. Integrated discrimination improvement (IDI) was used to assess performance improvement. Results: The performance of the In&Peri (AUC values 0.892 and 0.866), In&P (0.866 and 0.863), and In&Peri&P (0.929 and 0.911) models was significantly better than that of the intratumoral model (0.849 and 0.838) in the internal and external test cohorts (IDI test, all P<0.05). The intratumoral, In&Peri and In&Peri&P models showed good calibration (Hosmer-Lemeshow test, all P>0.05). The multiregional (In&Peri&P) model had the highest discrimination among the 6 radiomic models in the test cohorts, respectively. Conclusions: The multiregional model combining radiomic information of intratumoral, peritumoral, and ipsilateral parenchymal regions yielded better performance than the intratumoral model in distinguishing malignant breast lesions from benign lesions.

The value of contrast-enhanced ultrasound in the diagnosis of BI-RADS-US 4a lesions less than 2 cm in diameter.

BACKGROUND: Breast cancer is the most common malignant tumor in women. Early diagnosis of benign and malignant breast tumors is of great significance. OBJECTIVE: To retrospectively analyze the value of contrast-enhanced ultrasonography (CEUS) in the diagnosis of Breast Imaging-Reporting and Data System (BI-RADS) 4a breast lesions less than 2 cm in diameter. METHODS: CEUS was performed for 143 breast masses less than 2 cm in diameter that were diagnosed as BI-RADS 4a by ultrasound and reclassified. Considering pathological diagnosis as the gold standard, the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of reclassified lesions after CEUS for the diagnosis of benign and malignant masses were analyzed. RESULTS: BI-RADS 4a breast masses with a diameter less than 2 cm (n = 143) were confirmed by pathology; 103 and 40 were classified as benign and malignant, respectively. The sensitivity, specificity, PPV, and NPV of CEUS for the diagnosis were 90%, 86%, 72%, and 95%, respectively. The area under the receiver operating characteristic (ROC) curve of CEUS for the diagnosis of benign and malignant tumors after CEUS was 0.904. CONCLUSION: CEUS can help to improve the diagnostic accuracy of BI-RADS 4a masses with a diameter less than 2 cm.

MiRNA-1202 promotes the TGF-ß1-induced proliferation, differentiation and collagen production of cardiac fibroblasts by targeting nNOS.

BACKGROUND: Atrial fibrillation (AF) is a clinically common arrhythmia that affects human health. Myocardial fibrosis serves as an important contributor to AF. Recently, miRNA-1202 have been reported to be up-regulated in AF. However, the role of miRNA-1202 and its mechanism in myocardial fibrosis remain unclear. METHODS: Human cardiac fibroblasts (HCFs) were used to construct a fibrosis model by TGF-ß1 induction. The expression of miR-1202 was measured by qRT-PCR. Cell proliferation was assessed by CCK-8 assays. Protein expression levels were measured by western blot. Collagen accumulation was measured by ELISA. The relationship between miR-1202 and nNOS was investigated by luciferase reporter assays. RESULTS: MiR-1202 expression was obviously increased in HCFs and was both time- and dose-independent. MiR-1202 could increase the proliferation and collagen I, collagen III, and α-SMA levels with or without TGF-ß1. MiR-1202 could also increase TGF-ß1 and p-Smad2/3 protein levels in comparison to the control group. However, they were obviously decreased after inhibitor transfection. MiR-1202 targets nNOS for negative regulation of HCFs fibrosis by decreasing cell differentiation, collagen deposition and the activity of the TGF-ß1/Smad2/3 pathway. Co-transfection of miR-1202 inhibitor and siRNA of nNOS inhibited nNOS protein expression, thereby enhancing the HCFs proliferation. Furthermore, co-transfection of the miR-1202 inhibitor and siRNA of nNOS significantly promoted collagen I, collagen III, TGF-ß1, Smad2/3 and α-SMA protein expression and Smad2/3 protein phosphorylation. These findings suggested that miR-1202 promotes HCFs transformation to a pro-fibrotic phenotype by targeting nNOS through activating the TGF-ß1/Smad2/3 pathway.

hsa-miR-4443 inhibits myocardial fibroblast proliferation by targeting THBS1 to regulate TGF-ß1/α-SMA/collagen signaling in atrial fibrillation.

Fibrosis caused by the increase in extracellular matrix in cardiac fibroblasts plays an important role in the occurrence and development of atrial fibrillation (AF). The aim of this study was to investigate the role of hsa-miR-4443 in AF, human cardiac fibroblast (HCFB) proliferation, and extracellular matrix remodeling. TaqMan Stem-loop miRNA assay was used to measure hsa-miR-4443 expression in patients with persistent AF (n=123) and healthy controls (n=100). Patients with AF were confirmed to have atrial fibrosis by late gadolinium enhancement. At the cellular level, after hsa-miR-4443 mimic and inhibitor were transfected with HCFBs, proliferation, apoptosis, migration, and invasion were analyzed. Lastly, hsa-miR-4443-targeted gene and transforming growth factor (TGF)-ß1/α-SMA/collagen pathway were evaluated by dual-luciferase reporter assay and western blot, respectively. In patients with AF, hsa-miR-4443 decreased significantly and collagen metabolism level increased significantly. Logistic regression analysis showed that low hsa-miR-4443 level was a risk factor of AF (P<0.001). The receiver operating characteristic curve revealed that hsa-miR-4443 was useful for predicting AF (area under the curve: 0.828, sensitivity: 0.71, specificity: 0.78, P<0.001). In HCFBs, hsa-miR-4443 targeted thrombospondin-1 (THBS1) and downregulated TGF-ß1/α-SMA/collagen pathway. The inhibition of hsa-miR-4443 expression promoted HCFB proliferation, migration, invasion, myofibroblast differentiation, and collagen production. The significant reduction of hsa-miR-4443 can be used as a biomarker for AF. hsa-miR-4443 protected AF by targeting THBS1 and regulated TGF-ß1/α-SMA/collagen pathway to inhibit HCFB proliferation and collagen synthesis.

Localization, Gene Expression, and Functions of Glutamine Synthetase Isozymes in Wheat Grain (Triticum aestivum L.).

Glutamine synthetase (GS) plays a major role in plant nitrogen metabolism, but the roles of individual GS isoforms in grains are unknown. Here, the localization and expression of individual TaGS isozymes in wheat grain were probed with TaGS isoenzyme-specific antibodies, and the nitrogen metabolism of grain during the grain filling stage were investigated. Immunofluorescence revealed that TaGS1;1, TaGS1;3, and TaGS2 were expressed in different regions of the embryo. In grain transporting tissues, TaGS1;2 was localized in vascular bundle; TaGS1;2 and TaGS1;1 were in chalaza and placentochalaza; TaGS1;1 and TaGS1;3 were in endosperm transfer cells; and TaGS1;3 and TaGS2 were in aleurone layer. GS exhibited maximum activity and expression at 8 days after flowering (DAF) with peak glutamine content in grains; from then, NH 4 + increased largely from NO 3 - reduction, glutamate dehydrogenase (GDH) aminating activity increased continuously, and the activities of GS and glutamate synthase (GOGAT) decreased, while only TaGS1;3 kept a stable expression in different TaGS isozymes. Hence, GS-GOGAT cycle and GDH play different roles in NH 4 + assimilation of grain in different stages of grain development; TaGS1;3, located in aleurone layer and endosperm transfer cells, plays a key role in Gln into endosperm for gluten synthesis. At 30 DAF, grain amino acids are mainly transported from maternal phloem.

hsa-miR-4443 inhibits myocardial fibroblast proliferation by targeting THBS1 to regulate TGF-ß1/α-SMA/collagen signaling in atrial fibrillation

Fibrosis caused by the increase in extracellular matrix in cardiac fibroblasts plays an important role in the occurrence and development of atrial fibrillation (AF). The aim of this study was to investigate the role of hsa-miR-4443 in AF, human cardiac fibroblast (HCFB) proliferation, and extracellular matrix remodeling. TaqMan Stem-loop miRNA assay was used to measure hsa-miR-4443 expression in patients with persistent AF (n=123) and healthy controls (n=100). Patients with AF were confirmed to have atrial fibrosis by late gadolinium enhancement. At the cellular level, after hsa-miR-4443 mimic and inhibitor were transfected with HCFBs, proliferation, apoptosis, migration, and invasion were analyzed. Lastly, hsa-miR-4443-targeted gene and transforming growth factor (TGF)-β1/α-SMA/collagen pathway were evaluated by dual-luciferase reporter assay and western blot, respectively. In patients with AF, hsa-miR-4443 decreased significantly and collagen metabolism level increased significantly. Logistic regression analysis showed that low hsa-miR-4443 level was a risk factor of AF (P<0.001). The receiver operating characteristic curve revealed that hsa-miR-4443 was useful for predicting AF (area under the curve: 0.828, sensitivity: 0.71, specificity: 0.78, P<0.001). In HCFBs, hsa-miR-4443 targeted thrombospondin-1 (THBS1) and downregulated TGF-β1/α-SMA/collagen pathway. The inhibition of hsa-miR-4443 expression promoted HCFB proliferation, migration, invasion, myofibroblast differentiation, and collagen production. The significant reduction of hsa-miR-4443 can be used as a biomarker for AF. hsa-miR-4443 protected AF by targeting THBS1 and regulated TGF-β1/α-SMA/collagen pathway to inhibit HCFB proliferation and collagen synthesis.

Nitrogen Regulating the Expression and Localization of Four Glutamine Synthetase Isoforms in Wheat (Triticum aestivum L.).

Glutamine synthetase (GS), the key enzyme in plant nitrogen assimilation, is strictly regulated at multiple levels, but the most relevant reports focus on the mRNA level. Using specific antibodies as probes, the effects of nitrogen on the expression and localization of individual wheat GS (TaGS) isoforms were studied. In addition to TaGS2, TaGS1;1 with high affinity to substrate and TaGS1;3 with high catalytic activity were also localized in mesophyll, and may participate in cytoplasmic assimilation of ammonium (NH4+) released from photorespiration or absorbed by roots; TaGS1;2 was localized in xylem of leaves. In roots, although there were hundreds of times more TaGS1;1 than TaGS1;2 transcripts, the amount of TaGS1;1 subunit was not higher than that of TaGS1;2; NH4+ inhibited TaGS1;1 expression but stimulated TaGS1;3 expression. In root tips, nitrate stimulated TaGS1;1, TaGS1;3, and TaGS2 expression in meristem, while NH4+ promoted tissue differentiation and TaGS1;2 expression in endodermis and vascular tissue. Only TaGS1;2 was located in vascular tissue of leaves and roots, and was activated by glutamine, suggesting a role in nitrogen transport. TaGS1;3 was induced by NH4+ in root endodermis and mesophyll, suggesting a function in relieving NH4+ toxicity. Thus, TaGS isoforms play distinct roles in nitrogen assimilation for their different kinetic properties, tissue locations, and response to nitrogen regimes.

Propofol inhibits growth and invasion of pancreatic cancer cells through regulation of the miR-21/Slug signaling pathway.

AIM: Propofol, an intravenous anesthetic agent, has been found to inhibit invasion and growth of pancreatic cancer cells in vitro. However, the mechanisms underlying these tumor-promoting phenotypes are not known. The microRNA miR-21 has been reported to be overexpressed in pancreatic cancer, and overexpression of miR-21 confers a poor prognosis to patients with pancreatic cancer. Further studies have identified the E-cadherin transcription repressor Slug as a direct target of miR-21. In this study, we assessed whether propofol inhibits invasion and growth of pancreatic cancer cells by regulation of miR-21/Slug signaling. METHODS: PANC-1 pancreatic cancer cells were treated with different concentrations of propofol (1, 5 or 10 µg/mL) for 48 h, or 10 µg/mL propofol for 12, 24 or 36 h. Cell survival and apoptosis were detected by LDH release, BrdU cell proliferation and flow cytometry assays; cell invasion and migration were detected by transwell migration assays. miR-21 mimic (miR-21), Slug cDNA, PUMA siRNA and E-cadherin siRNA transfection was used to assess the signaling pathway in which propofol functions in PANC-1 cells. Protein and mRNA expression, respectively, were detected by western blotting and quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) assays. RESULTS: Propofol inhibited growth and invasion, and induced apoptosis, in a dose- and time-dependent manner in PANC-1 cells. Propofol inhibited miR-21 levels and decreased Slug expression, resulting in an increase in Slug-dependent PUMA and E-cadherin expression in PANC-1 cells. miR-21 overexpression or PUMA or E-cadherin silencing impaired propofol-induced cell apoptosis, growth and invasion. Re-expression of Slug attenuated the expression of PUMA and E-cadherin that was induced by propofol treatment, the reduction of growth and invasion, and the increase in cell apoptosis. CONCLUSIONS: Propofol can effectively inhibit invasion and induce apoptosis of PANC-1 cells by regulating miR-21/Slug signals.

Knockdown of S100A4 blocks growth and metastasis of anaplastic thyroid cancer cells in vitro and in vivo.

Anaplastic thyroid cancer (ATC) is a locally aggressive type of thyroid tumor with high rate of distant metastases. It is often incurable because it does not respond to radioiodine, radiotherapy, or chemotherapy. With conventional treatment, the median survival is about 6 months; therefore, new treatment options are needed. S100A4 is a calcium-binding protein related to the metastatic potential of carcinoma. Previous study has found S100A4 was overexpressed in human papillary thyroid carcinomas (PTC) tissues, and overexpression of S100A4 is associated with thyroid tumour invasion and metastasis. In the present study, we first examined S100A4 protein expression in 14 ATC tissues, 20 PTC tissues and 14 normal thyroid tissue by immunohistochemistry analysis. We then knocked down of S100A4 expression by RNA interference (S100A4 siRNA) and investigated its effects on growth and metastasis in two human ATC cell lines 8505C (BRAFV600E) and Cal-62 (BRAFwt) in vitro and in vivo. S100A4 and BRAFV600E protein expression was evaluated by western blot assay and immunohistochemistry analysis. Using immunohistochemistry, we found that high levels of S100A4 were detected in ATC specimens and PTC specimens. No S100A4 staining was observed in normal thyroid tissues. S100A4 siRNA significantly decreased proliferation and increased apoptosis, and inhibited the invasive potential of the two cells in vitro. In addition, S100A4 siRNA could effectively inhibit BRAFV600E expression in the 8505C cells, and treatment with 100 ng/ml human recombinant BRAF V600E in S100A4 siRNA/8505C cells could partly restore its proliferative and invasive ability. Results of implantation in vivo showed S100A4 shRNA could significantly inhibit abdominal cavity metastasis and tumor growth in vivo. Furthermore, knockdown of S100A4 has significant role on invasion, metastasis and growth inhibition in the 8505C cells than that of in the Cal-62 cells. These results support the hypothesis that S100A4 contributes significantly to growth and metastasis, and that down-regulation of S100A4 expression decreases the metastatic potential of ATC cells. Furthermore, down-regulation of S100A4 expression is more marked in BRAFV600E cells than that of in the BRAFwt cells.