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Objective: To investigate the effect of microgravity-mediated Notch1 signaling on macrophage polarization on bone homeostasis. Methods: The animal model was constructed by tail-limb suspension （HLS） to simulate the microgravity environment. The animals were grouped into Control group, HLS group, HLS+NC group, HLS+si group, HLS+rhNF-κB group. ELISA was used to detect the content of TNF-α and IL-1β in serum of rats. TUNEL staining was used to detect the apoptosis of bone tissue. Immunofluorescence was used to detect the polarization of macrophages in bone tissue. The rat osteoblast CP-R091 microgravity model was constructed by simulating the microgravity environment with a rotating wall bioreactor. The cell experiments were divided into Control group, HLS group, HLS+NC group, HLS+si group, HLS+rhNF-κB group. CCK-8 test was used to detect the proliferation activity of cells in each group, and AO test was used to test the apoptosis rate of cells in each group. PCR was used to detect the expression of osteogenesis- related genes in bone tissues and cells. Western blot was used to detect the expression of Notch1, hair division-related enhancer-1 （HES-1）, and Notch pathway ligand 1 （Jagged1） in bone tissues and cells of each group. Results: Compared with control group, the contents of TNF-α and IL-1β in the serum of the rats in the HLS group, the apoptosis rate, and the proportion of M1 macrophages were significantly increased. Compared with HLS group, the HLS+si group could obviously partially reverse the change trend of the above parameters, while HLS+rhNF-κB group significantly changed the above parameter values. Compared with control group, the proliferation activity of the cells in the HLS group was significantly reduced, and the apoptosis rate was significantly increased. Compared with HLS group, the HLS+si group could obviously partially reverse the change trend of the above parameters, while the HLS+ rhNF-κB group made the above parameter values worse. The expressions of the osteogenesis-related genes collagen type Ⅰ（COL1）, osteocalcin （OCN） and Runt-related transcription factor 2 （RUNX2） in bone tissues and cells in the microgravity environment were significantly decreased, while the expressions of Notch-1, Hes-1 and Jagged1 were significantly increased, and the differences were statistically significant （all P<0.05）. Conclusion: Microgravity-mediated Notch1 signaling regulates M1/M2 polarization of macrophages, participates in cell proliferation and apoptosis in bone tissue, and affects the progress of bone homeostasis. [ABSTRACT FROM AUTHOR]

A group of 27 patients diagnosed with metastatic triple-negative breast cancer (mTNBC) was randomly distributed into two groups and underwent different lines of metronomic treatment (mCHT). The former group (N 14) received first-line mCHT and showed a higher overall survival rate than the second group (N 13), which underwent second-line mCHT. Analysis of one patient still alive from the first group, diagnosed with mTNBC in 2019, showed a complete metabolic response (CMR) after a composite approach implicating first-line mCHT followed by second-line epirubicin and third-line nab-paclitaxel, and was chosen for subsequent molecular characterization. We found altered expression in the cancer stemness-associated gene NOTCH-1 and its corresponding protein. Additionally, we found changes in the expression of oncogenes, such as MYC and AKT, along with their respective proteins. Overall, our data suggest that a first-line treatment with mCHT followed by MTD might be effective by negatively regulating stemness traits usually associated with the emergence of drug resistance. [ABSTRACT FROM AUTHOR]

Notch signal plays an important role in regulating cell–cell interactions with the adjacent cells. However, it remains unknown whether Jagged1 (JAG‐1) mediated Notch signaling regulates bone cancer pain (BCP) via the spinal cell interactions mechanism. Here, we showed that intramedullary injection of Walker 256 breast cancer cells increased the expression of JAG‐1 in spinal astrocytes and knockdown of JAG‐1 reduced BCP. The supplementation of exogenous JAG‐1 to the spinal cord induced BCP‐like behavior and promoted expression of c‐Fos and hairy and enhancer of split homolog‐1 (Hes‐1) in the spinal cord of the naïve rats. These effects were reversed when the rats were administered intrathecal injections of N‐[N‐(3,5‐difluorophenacetyl)‐l‐alanyl]‐S‐phenylglycine t‐butyl ester (DAPT). The intrathecal injection of DAPT reduced BCP and inhibited Hes‐1 and c‐Fos expression in the spinal cord. Furthermore, our results showed that JAG‐1 up‐regulated Hes‐1 expression by inducing the recruitment of Notch intracellular domain (NICD) to the RBP‐J/CSL‐binding site located within the Hes‐1 promoter sequence. Finally, the intrathecal injection of c‐Fos‐antisense oligonucleotides (c‐Fos‐ASO) and administration of sh‐Hes‐1 to the spinal dorsal horn also alleviated BCP. The study indicates that inhibition of the JAG‐1/Notch signaling axis may be a potential strategy for the treatment of BCP. [ABSTRACT FROM AUTHOR]

A disintegrin and metalloproteinase 10 (ADAM10) plays an essential role in the regulation of survival, proliferation, migration, and differentiation of various neural cells. Nevertheless, the role of ADAM10 in oligodendrocyte precursors (OPCs) and myelination in the central nervous system (CNS) of developing and adult mouse brains is still unknown. We generated ADAM10 conditional knockout (ADAM10 cKO) mice lacking the ADAM10 gene primarily in OPCs by crossing NG2-Cre mice with ADAM10 loxp/loxp mice. We found that OPCs expressed ADAM10 in the mouse corpus callosum and the hippocampus. ADAM10 cKO mice showed significant loss of back hair and reduction in weight and length on postnatal (30 ± 2.1) day, died at (65 ± 5) days after birth, and exhibited the "anxiety and depression-like" performances. Conditional knockout of ADAM10 in OPCs resulted in a prominent increase in myelination and a decrease in the number of OPCs in the corpus callosum at P30 owing to premyelination and lack of proliferation of OPCs. Moreover, the number of proliferating OPCs and mature oligodendrocytes (OLs) also decreased with age in the corpus callosum of ADAM10 cKO mice from P30 to P60. Western blot and RT-PCR results showed that the activation of Notch-1 and its four target genes, Hes1, Hes5, Hey1, and Hey2, was inhibited in the corpus callosum tissue of ADAM10 knockout mice. In our study, we provided experimental evidence to demonstrate that ADAM10 is essential for modulating CNS myelination and OPC development by activating Notch-1 signaling in the developing and adult mouse brain. [ABSTRACT FROM AUTHOR]

A group of 27 patients diagnosed with metastatic triple-negative breast cancer (mTNBC) was randomly distributed into two groups and underwent different lines of metronomic treatment (mCHT). The former group (N 14) received first-line mCHT and showed a higher overall survival rate than the second group (N 13), which underwent second-line mCHT. Analysis of one patient still alive from the first group, diagnosed with mTNBC in 2019, showed a complete metabolic response (CMR) after a composite approach implicating first-line mCHT followed by second-line epirubicin and third-line nab-paclitaxel, and was chosen for subsequent molecular characterization. We found altered expression in the cancer stemness-associated gene NOTCH-1 and its corresponding protein. Additionally, we found changes in the expression of oncogenes, such as MYC and AKT, along with their respective proteins. Overall, our data suggest that a first-line treatment with mCHT followed by MTD might be effective by negatively regulating stemness traits usually associated with the emergence of drug resistance.

Microglia-mediated neuroinflammation is known to play a pivotal role in the pathogenesis of different neurological diseases. Gastrodin, a phenolic glucoside, has been reported to exert anti-inflammatory effects in activated microglia challenged with lipopolysaccharide (LPS); however, the underlying mechanism has remained obscure. The present study aimed to ascertain if Gastrodin would regulate the Notch signaling pathway involved in microglia activation. We show here that LPS increased the expression of various members of the Notch-1 pathway, including intracellular Notch receptor domain (NICD), recombining binding protein suppressor of hairless (RBP-Jκ) and transcription factor hairy and enhancer of split-1 (Hes-1) in microglia in postnatal rat brain and in BV-2 microglia. Remarkably, Gastrodin was found to markedly attenuate the expression of the above various biomarkers both in vivo and in vitro. Moreover, increased phosphorylation level of ERK, JNK and P38 induced by LPS was attenuated with pretreatment of Notch-1 signaling inhibitor, N-[N-(3,5-difluorophenacetyl)-1-alany1-Sphenyglycinet-butylester (DAPT) as well as Gastrodin. Gastrodin mimicked the effects of DAPT by inhibiting the LPS-induced expression of IL-1β, IL-6, IL-23, TNF-α and NO. Moreover, lentivirus transfection mediated NICD overexpression inhibited the anti-inflammatory effects of Gastrodin. Furthermore, the activation of Notch-1 signaling promoted microglia migration and Gastrodin could inhibit the migration of activated BV-2 microglia by regulating the Notch-1 signaling pathway. In light of the above, our results indicate that Notch-1 signaling pathway is involved in the anti-inflammatory effects of Gastrodin against LPS-induced microglia activation. These findings provide a new biological target of Gastrodin for the treatment of neuroinflammatory disorders. [ABSTRACT FROM AUTHOR]

Aims: The aim of our study is to investigate the effect induced by alternated mechanical loading on Notch-1 in mandibular condylar cartilage (MCC) of growing rabbits. Methods: A total of 64 ten-day-old rabbits were randomly divided into two groups according to dietary hardness: normal diet group (pellet) and soft diet group (powder). In each group, the rabbits were further divided into four subgroups by feeding time: two weeks, four weeks, six weeks, and eight weeks. Animals would be injected 5-bromo-2′-deoxyuridine (BrdU) every day for one week before sacrificing. Histomorphometric analysis of MCC thickness was performed through haematoxylin and eosin (HE) staining. Immunochemical analysis was done to test BrdU and Notch-1. The quantitative real-time polymerase chain reaction (qRT-PCR) and western blot were used to measure expression of Notch-1, Jagged-1, and Delta-like 1 (Dll-1). Results: The thickness of MCC in the soft diet group was thinner than the one in normal diet group. Notch-1 was restricted in fibrous layer, proliferative layer, and hypertrophic layer. The expression of Notch-1 increased from two weeks to six weeks and then fell down. Notch-1 in normal diet group was higher than that in soft diet group in anterior part of MCC. The statistical differences of Notch-1 were shown at two, four, and six weeks (p < 0.05). The result of western blot and quantitative real-time PCR (qRT-PCR) showed the expression of Dll-1 and Jagged-1 rose from two to four weeks and started to decrease at four weeks. BrdU distributed in all layers of cartilage and subchondral bone. The number of BrdU-positive cells, which were less in soft diet group, was decreasing along with the experiment period. The significant difference was found at four, six, and eight weeks in anterior and posterior parts (p < 0.05). Conclusion: The structure and proliferation of MCC in rabbits were sensitive to dietary loading changes. The proper mechanical loading was essential for transduction of Notch signalling pathway and development of mandibular condylar cartilage.

Sunitinib (SUN) is a chemotherapeutic agent clinically approved for treatment of metastatic renal carcinoma. Despite its remarkable benefits, various renal toxicities have been reported that limit its clinical uses. Oleuropein (OLE) is the main polyphenolic constituent of olive tree and mediates the majority of its valuable pharmacological activities. The current study examined the probable renoprotective effects of OLE against SUN-induced nephrotoxicity. Adult male albino rats were co-treated by SUN (25 mg/kg, 3 times/week, PO) with either a drug vehicle or OLE (60 mg/kg/day, daily, PO) for four weeks. A control group comprising of age-matched rats was used. Four weeks later, blood specimens were collected to assess kidney functions. Kidneys were harvested for biochemical and histopathological analyses. Administration of SUN induced kidney dysfunction, along with marked rises in endothelin-1 (ET-1) and monocyte chemotactic protein-1 (MCP-1) levels in renal tissues. Histological abnormalities were also detected in kidneys of SUN-treated rats including glomerular and tubular interstitial congestion along with interstitial fibrosis. On molecular levels, there was a decline in renal SIRT6 expression along with significant up-regulation of Notch-1, NLRP-3, interleukin -1β (IL-1β) and cleaved caspsase-3. All these changes were almost alleviated by OLE co-treatment. These findings suggest the implication of SIRT6/Notch-1/NLRP3/IL-1β axis in the pathogenesis of SUN-induced nephrotoxicity and highlight OLE as a prospective renoprotective agent during SUN chemotherapy to halt its renal toxicity likely through promotion of SIRT6 and suppression of Notch-1/NLRP3/IL-1β signaling pathway. [Display omitted] • Oleuropein alleviates the nephrotoxicity of sunitinib chemotherapy. • Oleuropein up-regulates SIRT6 expression in kidneys of sunitinib-treated rats. • Oleuropein offsets Notch-1 expression in kidneys of sunitinib-treated rats. • Oleuropein suppresses NLRP-3 inflammasome in kidneys of sunitinib-treated rats. • Oleuropein ameliorates apoptosis & interstitial fibrosis in sunitinib-treated rats. [ABSTRACT FROM AUTHOR]

• Notch-1 pathway plays an important role in lung carcinoma. • Lung parenchyma from Smokers exhibited higher expression of Notch-1. • CSE increases Notch-1 activation and Hes-1 target gene in lung cancer cells. • Notch-1 modulation affects ki-67 and survivin expression in lung cancer cells. Notch-1 pathway plays an important role in lung carcinoma, stem cell regulation, cellular communication, growth and differentiation. Cigarette smoke is involved in the regulation of Notch signaling. However, current data regarding the impact of cigarette smoke on the Notch pathway in lung cancer progression are limited. The present study aimed to explore whether cigarette smoke exposure altered Notch-1 pathway in ex-vivo (surgical samples of lung parenchyma from non-smoker and smoker patients with lung adenocarcinoma) and in vitro (adenocarcinoma A549 cell line) approaches. The expression of Notch-1, Jagged-1 and CD133 in surgical samples was evaluated by immunohistochemistry. A549 were exposed to cigarette smoke extracts (2.5 % and 5 % CSE for 6, 24 and 48 h) and the expression of Notch-1, Jagged-1 and Hes-1 was evaluated by Real-Time PCR and Western Blot (nuclear fractions). Expression and localization of Notch-1, Hes-1, CD133 and ABCG2 were assessed by immunofluorescence. The expression of survivin and Ki-67 was assessed by flow cytometry following CSE exposure and inhibition of Notch-1 signaling. Smokers lung parenchyma exhibited higher expression of Notch-1. CSE exposure increased Notch-1 and Hes-1 gene and nuclear protein expression in A549. Immunofluorescence confirmed higher expression of nuclear Hes-1 in CSE-stimulated A549 cells. CSE increased both survivin and Ki-67 expression and this effect was reverted by inhibition of the Notch-1 pathway. In conclusion, these data show that cigarette smoke may promote adenocarcinoma progression by activating the Notch-1 pathway thus supporting its role as hallmark of lung cancer progression and as a new target for lung cancer treatment. [ABSTRACT FROM AUTHOR]

Abstract Background The involvement of lncRNA FEZF1-AS1 has been analyzed in many types of cancers, while its roles in non-small cell lung cancer (NSCLC) remains unclear. We then explored the role of FEZF1-AS1 in NSCLC. Methods qPCR and western blot were performed to measure gene expression. FEZF1-AS1, miR-34a, and NOTCH-1 were overexpressed to analyze the relationship between them. Transwell assays were performed to analyze the effects of transfections on cell invasion and migration. Results FEZF1-AS1 was up-regulated in NSCLC patients. Increased expression levels of FEZF1-AS1 were observed with the increase in clinical stages. Bioinformatics analysis showed that miR-34a can bind with FEZF1-AS1. In NSCLC tissues, NOTCH-1 and FEZF1-AS1 were positively correlated. In NSCLC cells, over-expression of FEZF1-AS1 resulted in up-regulated expressions of NOTCH-1, while miR-34a over-expression mediated down-regulated expressions of NOTCH-1. In addition, FEZF1-AS1 and miR-34a did not alter each other, while bioinformatics analysis showed that miR-34a can bind FEZF1-AS1. Analysis of cell migration and invasion showed increased cell invasion and migration rates after FEZF1-AS1 and NOTCH-1 over-expression. MiR-34a played the opposite role and reduced the effects of FEZF1-AS1 over-expression. Conclusions FEZF1-AS1 promoted NSCLC cell migration and invasion through the up-regulation of NOTCH1 by serving as a sponge of miR-34a.

Lun Luo,1 Yuan Zhang,2 Haiyong He,1 Chuan Chen,1 Baoyu Zhang,1 Meiqin Cai1 1Department of Neurosurgery, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong Province 510630, People’s Republic of China; 2Department of Obstetrics, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong Province 510630, People’s Republic of ChinaCorrespondence: Lun Luo; Meiqin CaiDepartment of Neurosurgery, The Third Affiliated Hospital of Sun Yat-sen University, No. 600 Tianhe Road, Tianhe District, Guangzhou, Guangdong Province 510630, People’s Republic of ChinaTel +86-20-85253333Email yv6440@163.com; wlbexvwotgje41@163.comIntroduction: LncRNA FEZF1-AS1 has been reported to be an oncogene in many types of cancer, while its role in glioblastoma (GBM) is unknown. This study aimed to investigate the potential involvement of FEZF1-AS1 in GBM.Methods: FEZF1-AS1 expression in paired GBM and non-tumor tissues from GBM patients was determined by RT-qPCR. A 2-year follow-up was performed to analyze the prognostic value of FEZF1-AS1 for GBM. Cell transfections were performed to analyze the interactions between FEZF1-AS1, miR-34a and Notch-1. Transwell assay was performed to analyze the role of FEZF1-AS1, miR-34a and Notch-1 in regulating GBM cell invasion and migration.Results: In this study, analysis of TCGA dataset revealed the upregulation of FEZF1-AS1 in GBM, and the overexpression of FEZF1-AS1 in GBM was further confirmed using GBM tissues from GBM patients included in this study. High levels of FEZF1-AS1 were correlated with poor survival. FEZF1-AS1 was predicted to form base pairing with miR-34a. However, overexpression of FEZF1-AS1 and miR-34a failed to affect the expression of each other. However, upregulation of Notch-1, a target of miR-34a, was observed after FEZF1-AS1 in GBM cells. Moreover, increased invasion and migration rates of GBM cells were observed after FEZF1-AS1 and Notch-1 overexpression. MiR-34a played an opposite role and reduced the effects of FEZF1-AS1 and Notch-1 overexpression.Conclusion: FEZF1-AS1 may sponge miR-34a to upregulate Notch-1 in GBM, thereby promoting cancer cell invasion and migration.Keywords: glioblastoma, FEZF1-AS1, miR-34a, Notch-1

Abstract This study aimed to evaluate the specific regulatory roles of microRNA-146a (miRNA-146a) in temporal lobe epilepsy (TLE) and explore the related regulatory mechanisms. A rat model of TLE was established by intraperitoneal injection of lithium chloride-pilocarpine. These model rats were injected intracerebroventricularly with an miRNA-146a inhibitor and Notch-1 siRNA. Then, neuronal damage and cell apoptosis in the cornu ammonis (CA) 1 and 3 regions of the hippocampus were assessed. SOD and MDA levels in the hippocampus were detected by chromatometry, and IL-1β, IL-6, and IL-18 levels were detected by ELISA. Then, we evaluated the expression levels of caspase-9, GFAP, Notch-1, and Hes-1 in the hippocampus. The interaction between Notch-1 and miRNA-146a was assessed by a dual luciferase reporter gene assay. A rat model of TLE was successfully established, which exhibited significantly increased miRNA-146a expression in the hippocampus. Silencing of miRNA-146a significantly alleviated the neuronal damage and cell apoptosis in the CA1 and CA3 regions of the hippocampus in TLE rats and decreased MDA, IL-1β, IL-6, and IL-18 levels and increased SOD levels in the hippocampus of TLE rats. In addition, silencing of miRNA-146a significantly decreased the expression levels of caspase-9, GFAP, Notch-1, and Hes-1 in the hippocampus of TLE rats. Notch-1 was identified as a target of miRNA-146a and silencing of Notch-1 aggravated the neuronal damage in the CA1 and CA3 regions. Silencing of miRNA-146a alleviated the neuronal damage in the hippocampus of TLE rats by down-regulating Notch-1.

Despite the therapeutic advances in treating malignancies, the efficient radiotherapeutic approaches with deprived adverse reactions still represent a potential clinical inquiry. The current study aims to elucidate the role of gallic acid (GA) in modifying the hazardous renal cytotoxicity induced by acute exposure to radiation. The MTT test was used to evaluate the viability of Vero cells exposed to 2 Gy gamma radiation with or without incubation of GA. In an in vivo model, male Wistar rats were divided into four experimental groups (n = 6): Control, Irradiated (IRR, 5 Gy), GA (100 mg/kg, i.p.) + IRR, and Glycogen synthase kinase inhibitor (GSKI, 3 mg/kg, i.p.) + IRR. Based on the MTT toxicity assay, from 0 and up to 5 μM dosages of GA did not demonstrate any cytotoxicity to Vero cells. The optimal GA dose that could protect the cells from radiation was 5 μM. Furthermore, GA exerted a protective effect from gamma radiation on renal tissue as indicated by corrected renal functions, decreased LDH level in serum, and balanced oxidative status, which is indicated by decreased tissue contents of NOx and TBARS with a significant increase of reduced GSH. These outcomes were inferred by the upregulation of nuclear factor erythroid 2-related factor 2 (Nrf2) expression. The overall molecular impact of radiation in damaging the renal tissue may be explained by modifying the upstream AKT activity and its downstream targets GSK-3β/Notch-1. Here, we concluded that the anticipated adverse reaction in the course of radiation exposure could be protected by daily administration of GA., (© 2024 Wiley Periodicals LLC.)

The habit of drinking tea is highly prevalent and could be utilized to introduce more health benefits through fortification with medicinal plants. The purpose of this analysis was to assess the nutritional quality and health benefits of fortified Ziziphus tea (ZT) with green tea (GT) against obesity and non‐alcoholic fatty liver disease (NAFLD). Proximate analysis and sensory evaluation were carried out on the fortified tea. In the in vivo study, 15 SD rats were used for each group. Flow cytometry was utilized for caspase 3 analysis. ELISA was used for the detection of tumor necrosis factor‐alpha and adiponectin levels. Real‐time PCR was used to detect Notch‐1 and Hes‐1 gene expression. The composition of fortified (GT+ZT) showed a significant improvement in the nutritional value represented by the increase in overall protein, crude fat, crude fiber, ash, carbohydrate, mineral contents, and antioxidant capacity. Treatment with GT+ZT restored the disturbance in body weight, lipid profile, liver function, glucose, insulin sensitivity index, and oxidative status. It reversed the changes in TNF‐α and adiponectin levels. Their protective effects against NAFLD were indicated by the inhibition of hepatic caspase‐3 activity, suppression of Notch‐1, and Hes‐1 gene expression and amelioration of high‐fat diet (HFD)‐induced histological alterations. Collectively, our findings, elucidate the precise mechanism of fortified ZT+GT for the attenuation of obesity‐induced metabolic disorders and NAFLD via regulating lipolysis, TNF‐α, adiponectin, apoptosis, and Notch‐1 signaling pathways, and provide a foundation for an easily implemented healthy habit of drinking. Practical applications: The incorporation of bioactive compounds into functional foods is a growing market. Consumer attention in well‐being has increased rapidly toward a fortified diet that provides additional health effects. The fortified (GT+ZT) tea may potentially serve as an easily implemented healthy drinking habit to prevent and manage obesity and NAFLD and reduce the risk of other diseases. Fortification with ZT improved the health‐promoting functionality of GT through the enhancement of total protein, carbohydrates, antioxidant, and mineral contents. This was reflected by their synergetic therapeutic activity in ameliorating the disturbance in obesity‐related disorders and NAFLD via regulating lipolysis, inflammation, oxidative stress, apoptosis, and Notch‐1 signaling pathways. Therefore, (GT+ZT) could be considered functional foods which attribute to functional improvement and reduction in disease risk. [ABSTRACT FROM AUTHOR]

Surgical decompression is the primary treatment for cervical spondylotic myelopathy (CSM) patients with compressive spinal cord injury (CSCI). However, the prognosis of patients with CSCI varies, and the pathophysiological changes following decompression remain poor. This study aimed to investigate the pathophysiological changes and the role of Notch-1 activation after decompression in a rat CSCI model. Surgical decompression was conducted at 1 week post-injury (wpi). DAPT was intraperitoneally injected to down-regulate Notch-1 expression. Basso, Beattie, and Bresnahan scores and an inclined plane test were used to evaluate the motor function recovery. Hematoxylin and eosin staining was performed to assess pathophysiological changes, while hypoxia-inducible factor 1 alpha, vascular endothelial growth factor (VEGF), von Willebrand factor (vWF), matrix metalloproteinase (MMP)-9, MMP-2, Notch-1, and Hes-1 expression in the spinal cord were examined by immunohistochemical analysis or quantitative PCR. The results show that early decompression can partially promote motor function recovery. Improvements in structural and cellular damage and hypoxic levels were also observed in the decompressed spinal cord. Moreover, decompression resulted in increased VEGF and vWF expression, but decreased MMP-9 and MMP-2 expression at 3 wpi. Expression levels of Notch-1 and its downstream gene Hes-1 were increased after decompression, and the inhibition of Notch-1 significantly reduced the decompression-induced motor function recovery. This exploratory study revealed preliminary pathophysiological changes in the compressed and decompressed rat spinal cord. Furthermore, we confirmed that early surgical decompression partially promotes motor function recovery may via activation of the Notch-1 signaling pathway after CSCI. These results could provide new insights for the development of drug therapy to enhance recovery following surgery.

The cross-talk between axon and glial cells during development and in adulthood is mediated by several molecules. Among them are neurotransmitters and their receptors, which are involved in the control of myelinating and non-myelinating glial cell development and physiology. Our previous studies largely demonstrate the functional expression of cholinergic muscarinic receptors in Schwann cells. In particular, the M2 muscarinic receptor subtype, the most abundant cholinergic receptor expressed in Schwann cells, inhibits cell proliferation downregulating proteins expressed in the immature phenotype and triggers promyelinating differentiation genes. In this study, we analysed the in vitro modulation of the Neuregulin-1 (NRG1)/erbB pathway, mediated by the M2 receptor activation, through the selective agonist arecaidine propargyl ester (APE). M2 agonist treatment significantly downregulates NRG1 and erbB receptors expression, both at transcriptional and protein level, and causes the internalization and intracellular accumulation of the erbB2 receptor. Additionally, starting from our previous results concerning the negative modulation of Notch-active fragment NICD by M2 receptor activation, in this work, we clearly demonstrate that the M2 receptor subtype inhibits erbB2 receptors by Notch-1/NICD downregulation. Our data, together with our previous results, demonstrate the existence of a cross-interaction between the M2 receptor and NRG1/erbB pathway-Notch1 mediated, and that it is responsible for the modulation of Schwann cell proliferation/differentiation.

Surgical decompression is the primary treatment for cervical spondylotic myelopathy (CSM) patients with compressive spinal cord injury (CSCI). However, the prognosis of patients with CSCI varies, and the pathophysiological changes following decompression remain poor. This study aimed to investigate the pathophysiological changes and the role of Notch-1 activation after decompression in a rat CSCI model. Surgical decompression was conducted at 1 week post-injury (wpi). DAPT was intraperitoneally injected to down-regulate Notch-1 expression. Basso, Beattie, and Bresnahan scores and an inclined plane test were used to evaluate the motor function recovery. Hematoxylin and eosin staining was performed to assess pathophysiological changes, while hypoxia-inducible factor 1 alpha, vascular endothelial growth factor (VEGF), von Willebrand factor (vWF), matrix metalloproteinase (MMP)-9, MMP-2, Notch-1, and Hes-1 expression in the spinal cord were examined by immunohistochemical analysis or quantitative PCR. The results show that early decompression can partially promote motor function recovery. Improvements in structural and cellular damage and hypoxic levels were also observed in the decompressed spinal cord. Moreover, decompression resulted in increased VEGF and vWF expression, but decreased MMP-9 and MMP-2 expression at 3 wpi. Expression levels of Notch-1 and its downstream gene Hes-1 were increased after decompression, and the inhibition of Notch-1 significantly reduced the decompression-induced motor function recovery. This exploratory study revealed preliminary pathophysiological changes in the compressed and decompressed rat spinal cord. Furthermore, we confirmed that early surgical decompression partially promotes motor function recovery may via activation of the Notch-1 signaling pathway after CSCI. These results could provide new insights for the development of drug therapy to enhance recovery following surgery. [ABSTRACT FROM AUTHOR]

Endogenous repair after chronic compressive spinal cord injury (CCSCI) is of great clinical interest. Ischemia-hypoxia-induced angiogenesis has been proposed to play an important role during this repair process. Emerging evidence indicates that long non-coding RNAs (lncRNAs) are involved in the pathophysiological processes of various diseases. Here, we identified a lncRNA (Xist; X-inactive specific transcript) with upregulated expression in cervical spine lesions during endogenous neurological repair in CCSCI rats. Therapeutically, the introduction of Xist to rats increased neurological function in vivo as assayed using the Basso, Beattie, and Bresnahan (BBB) score and inclined plane test (IPT). We found that the introduction of Xist enhanced endogenous neurological repair by promoting angiogenesis and microvessel density after CCSCI, while depletion of Xist inhibited angiogenesis and cell sprouting and migration. Mechanistically, Xist promoted angiogenesis by sponging miR-32-5p and modulating Notch-1 expression both in vitro and in vivo. These findings suggest a role of the Xist/miR-32-5p/Notch-1 axis in endogenous repair and provide a potential molecular target for the treatment of ischemia-related central nervous system (CNS) diseases.

Abstract Background Abnormal apoptosis of lung endothelial cells has been observed in emphysematous lung tissue and has been suggested to be an important upstream event in the pathogenesis of chronic obstructive pulmonary disease (COPD). Studies have shown that microRNAs (miRNAs) contribute to the pathogenesis of pulmonary diseases by regulating cell apoptosis. The present study was designed to investigate the expression of microRNA-34a (miR-34a) in human pulmonary microvascular endothelial cells (HPMECs) exposed to cigarette smoke extract (CSE), and the potential regulatory role of miR-34a in endothelial cell apoptosis. Results Our results showed that the expression of miR-34a was significantly increased in CSE-treated HPMECs, and inhibiting miR-34a attenuated CSE-induced HPMEC apoptosis. Furthermore, expression of Notch-1, a receptor protein in the Notch signalling pathway, was decreased and was inversely correlated with miR-34a expression in HPMECs treated with CSE. Computational miRNA target prediction confirmed that Notch-1 is a target of miR-34a. Luciferase reporter assay further confirmed the direct interaction between miR-34a and the 3’-untranslated region (UTR) of Notch-1. Restoration of Notch-1 pathway was able to partially block the effect of miR-34a on HPMEC apoptosis. These results indicate that Notch-1 is a critical downstream target of miR-34a in regulating the CSE-induced HPMEC apoptosis. Conclusions Our results suggest that miR-34a plays a key role in CSE-induced endothelial cell apoptosis by directly regulating its target gene Notch-1 in endothelial cells.

• Notch-1 intervenes in the reparative processes of the airway mucosa. • cigarette smoke exposure limits Notch-1 signaling activation in bronchial epithelial cells. • cigarette smoke exposure hinders repair processes in bronchial epithelial cells. Notch-1 intervenes in the reparative processes of mucosa by controlling cell proliferation, differentiation and stem cell maintenance. Cigarette smoke alters airway epithelial homeostasis. The present study explored whether: Smokers showed altered Notch-1 expression; and whether in bronchial epithelial cells (16HBE): a) cigarette smoke extracts (CSE) altered the expression of Notch-1, of its ligand Jagged-1 (Jag-1) and the nuclear translocation of Notch-1; b) Notch-1 signaling activation as well as CSE modified Ki67, PCNA, p21, IL-33 expression, cell proliferation and repair processes. Notch-1 expression was assessed in the epithelium from large airway surgical samples from non-smoker and smoker subjects by immunohistochemistry.16HBE were cultured with/without CSE and Jag-1. A Notch-1 inhibitor (DAPT) was used as control. The expression of Notch-1, Jag-1, Ki67, PCNA, p21, IL-33 and cell proliferation (by CFSE) were all assessed by flow cytometry. Notch-1 nuclear expression was evaluated by immunofluorescence and western blot analysis. Repair processes were assessed by wound assay. Smokers had cytoplasmic but not nuclear Notch-1 expression. Although CSE increased Notch-1 expression, it counteracted Notch-1 signaling activation since it reduced Jag-1 expression and Notch-1 nuclear translocation. Notch-1 signaling activation by Jag-1 increased Ki67, PCNA and repair processes but reduced intracellular IL-33 and p21 expression without affecting cell proliferation. DAPT counteracted the effects of Notch-1 activation on PCNA and IL-33. CSE increased Ki67, PCNA, p21 and IL-33 expression but reduced cell proliferation and repair processes. In conclusion, cigarette smoke exposure, limiting Notch-1 signaling activation and hindering repair processes, amplifies injury processes in bronchial epithelial cells. [ABSTRACT FROM AUTHOR]

Background: The involvement of lncRNA FEZF1-AS1 has been analyzed in many types of cancers, while its roles in non-small cell lung cancer (NSCLC) remains unclear. We then explored the role of FEZF1-AS1 in NSCLC.Methods: qPCR and western blot were performed to measure gene expression. FEZF1-AS1, miR-34a, and NOTCH-1 were overexpressed to analyze the relationship between them. Transwell assays were performed to analyze the effects of transfections on cell invasion and migration.Results: FEZF1-AS1 was up-regulated in NSCLC patients. Increased expression levels of FEZF1-AS1 were observed with the increase in clinical stages. Bioinformatics analysis showed that miR-34a can bind with FEZF1-AS1. In NSCLC tissues, NOTCH-1 and FEZF1-AS1 were positively correlated. In NSCLC cells, over-expression of FEZF1-AS1 resulted in up-regulated expressions of NOTCH-1, while miR-34a over-expression mediated down-regulated expressions of NOTCH-1. In addition, FEZF1-AS1 and miR-34a did not alter each other, while bioinformatics analysis showed that miR-34a can bind FEZF1-AS1. Analysis of cell migration and invasion showed increased cell invasion and migration rates after FEZF1-AS1 and NOTCH-1 over-expression. MiR-34a played the opposite role and reduced the effects of FEZF1-AS1 over-expression.Conclusions: FEZF1-AS1 promoted NSCLC cell migration and invasion through the up-regulation of NOTCH1 by serving as a sponge of miR-34a. [ABSTRACT FROM AUTHOR]

Cancer stem-like cells are rare immortal cells within tumor, which are thought to play important roles in ionizing radiation (IR) therapy-resistance. Quercetin is a natural flavonoid with potential anti-cancer properties without significant cytotoxicity in normal tissues. In this study, we demonstrated that quercetin-IR combination treatment exhibited more dramatic anti-cancer effect than either quercetin or IR treatment alone via targeting colon cancer stem cells (CSCs) and inhibiting the Notch-1 signaling. These effects were further verified by in vivo studies which showed remarkable decrease of the CSCs markers and the expression of Notch-1 signaling proteins in human colon cancer xenografts in nude mice. Co-treatment with quercetin and low dose of radiation significantly reduced the expressions of all five proteins of γ-secretase complex in HT-29 and DLD-1 cells. In addition, ectopic expression of the Notch intracellular domain (NICD) partly reversed the inhibition effects by the combination therapy. In conclusion, our results indicated that the combination of quercetin (20 μM) and IR (5Gy) might be a promising therapeutic strategy for colon cancer treatment by targeting colon cancer stem-like cells and inhibiting the Notch-1 signaling. In future studies, we intend to further explore the potential therapeutic efficacy of the quercetin-radiation combination treatment in clinical trials. Image 1 • Quercetin and IR can target colon cancer stem cells. • Quercetin and IR inhibit the Notch-1. • Quercetin and IR inhibit γ-secretase complex. [ABSTRACT FROM AUTHOR]

Natural dietary molecules such as flavonoids have been recognized for their immense potential in cancer therapeutics with several health benefits. Hes-1 and Notch-1 overexpression has been associated with the progression of cervical cancer. However, the apoptosis-inducing potential of one such potent flavanol against these two key components of the Notch signaling pathway in cervical cancer has not been elucidated to date. Therefore, in this study, we performed several in vitro assays to gain detailed insight about the apoptotic inducing effect of rutin as well as its modulatory effect on Notch-1 and Hes-1 in cervical cancer cells. The results indicated that rutin led to a dose-dependent antiproliferative effects on Caski cervical cancer cells. DAPI and Mitotracker red staining revealed that rutin induced significant apoptotic effects via caspase-3/9 activation, ROS generation, and alteration in Bax/Bcl2 mRNA expression. Cell cycle analysis resulted in the arrest of cell cycle progression in G0/G1 that was associated with a reduced expression of CDK4 and Cyclin D1. The gene expression analysis further revealed that rutin treatment decreases Notch-1 and Hes-1 mRNA expression. Altogether, these results showed that rutin showed potent anticancer effects in human cervical cancer Caski cells by triggering apoptosis, G0/G1 phase arrest, and downregulating the level of Notch-1 and Hes-1 of the Notch signaling pathway.

Objective: Notch-1 signaling is significantly associated with the occurrence and development of atherosclerosis (AS). However, the molecular mechanisms underlying the specific deletion of Notch-1 in AS-associated macrophages are not fully understood. This study aimed to investigate the effects of Notch-1 in AS., Methods and Results: Tissue samples were obtained from atherosclerotic segments of human carotid arteries. Immunofluorescence staining showed that Notch-1 was significantly colocalized with macrophages (CD68+), and Notch-1 staining was increased in human vulnerable plaques. Notch-1 MAC-KO /ApoE -/- mice were generated in which Notch-1 was selectively inactivated in macrophages, and WT for littermate control mice (ApoE -/- /Notch-1 WT ). A control group was then established. All mice fed with a high-fat and Oil Red O, Movat, a-SMA, CD68, and Sirius red staining were used to evaluate the morphology. Specific deletion of Notch-1 in macrophages repressed the pathophysiology of AS. Immunofluorescent staining and Western blotting revealed that Notch-1 MAC-KO repressed M1 and M2 responses in AS. Here, GSEA revealed that Notch-1 activation and PI3K signaling were statistically significantly correlated with each other, and Notch-1 was involved in the regulation of the PI3K signaling pathway. In the in vitro experiments, the secretion of Arg-1 and exosomes was classified by peritoneal macrophages of Notch-1 MAC-KO /ApoE -/- and Notch-1 WT /ApoE -/- mice. Immunohistochemistry staining and Western blotting were used to measure the expression levels of Notch1, PI3K, p-PI3K, AKT, p-AKT, Arg-1, IL-6, CD36, SREBP-1, CD206, iNOS, cleaved-caspase-3/-9, Bax, CD9, Alix and TSG101 in the peritoneal macrophages and exosomes, respectively., Conclusions: The specific deletion of Notch-1 in macrophage represses the formation and development of AS via the PI3K/AKT signaling pathway.

Objective: To investigate the effect of Notch-1 signaling on NAFLD and its molecular mechanism., Methods: The lipid deposition in liver tissues was detected by oil red O staining. Western blotting was performed to detect the expressions of SREBP1C, SREBP2, LXR, IL-1β, IL-18, NLRP3, Notch-1, NOX2, NOX4, p-PI3K and p-SHP2 in macrophages, and the expressions of ALIX, CD9, IL-1β and SREBP1C in exosomes. Macrophages in the Notch-1 MAC-KO group and Notch-1 WT group were treated with FFA, and those in the Notch-1 WT +FFA group and Notch-1 MAC-KO +FFA group were treated with SHP2 inhibitors PHPS1 and Relaxin., Results: It was observed by oil red O staining that lipid deposition in mice with NAFLD was reduced in the Notch-1 MAC-KO group. The results of Western blotting showed that the expressions of ALIX, CD9, IL-1β and SREBP1C in macrophage exosomes were significantly lower in the Notch-1 MAC-KO group than in the Notch-1 WT group. In macrophages, the expressions of SREBP1C, SREBP2, LXR, IL-1β, IL-18, Notch-1, NOX2, NOX4 and p-PI3K significantly decreased, while the expression of p-SHP2 significantly increased in the Notch-1 MAC-KO group compared with the Notch-1 WT group. The Notch-1 MAC-KO +FFA group had significantly decreased expressions of SREBP1C, NLRP3, IL-1β, IL-18, SREBP2, NOX2, NOX4 and p-PI3K and a significantly increased expression of p-SHP2 compared with the Notch-1 WT +FFA group. However, the differences in the above proteins were all eliminated after PHPS1 and Relaxin were added., Conclusion: Specific knockout of Notch-1 attenuates NAFLD, and reduces inflammation and lipid deposition in the liver by promoting SHP2 phosphorylation.

Pancreatic cancer is well known to be the most deadly malignancy with the worst survival rate of all cancers. High temperature requirement factor A1 (HtrA1) plays an important role in cancer cell proliferation, migration, apoptosis, and differentiation. This study aimed to explore the function of HtrA1 in pancreatic cancer cell growth and its underlying mechanism. We found that the expression of HtrA1 was lower in pancreatic cancer tissue compared to the adjacent normal tissue. Consistently, HtrA1 levels were also decreased in two human pancreatic cancer cell lines, PANC-1 and BXPC-3. Moreover, enforced expression of HtrA1 inhibited cell viability and colony formation of PANC-1 and BXPC-3 cells. Overexpression of HtrA1 promoted apoptosis and suppressed migratory ability of tumor cells. On the contrary, siRNA-mediated knockdown of HtrA1 promoted the growth potential of pancreatic cancer cells. In addition, we found that up-regulation of HtrA1 reduced the expression of Notch-1 in pancreatic cancer cells. On the contrary, knockdown of HtrA1 increased the expression levels of Notch-1. Furthermore, overexpression of Notch-1 abolished the anti-proliferative effect of HtrA1 on pancreatic cancer cells. Taken together, our findings demonstrated that HtrA1 could inhibit pancreatic cancer cell growth via regulating Notch-1 expression, which implied that HtrA1 might be developed as a novel molecular target for pancreatic cancer therapy.

Mixed‐lineage kinase 3 (MLK3), the mitogen‐activated protein kinase kinase kinase (MAP3K), has been recognized as a player in tumorigenesis and oncogenic signalling, yet its detailed functions and signalling in cervical cancer have not been fully elucidated. Here, we identify that cervical cancer cells display higher mRNA and protein levels of MLK3 than normal cervical epithelial squamous cells. In HeLa and SiHa cell, MLK3 knockdown using siRNA remarkably suppressed cell survival and promoted cell apoptosis, with increased expression of the apoptosis‐related protein Bax and reduced Bcl‐2. Moreover, MLK3 knockdown promoted cell autophagy, demonstrated by increased ratio of autophagy‐related proteins LC3II/LC3I and decreased p62 expression in MLK3 depletion cells. Furthermore, MLK3 knockdown remarkably abolished Notch‐1 expression in cervical cancer cells. By co‐treating Hela cells with MLK3 specific siRNA and pcDNA3.1‐Notch‐1 overexpression plasmid or autophagy inhibitor 3‐MA, we found that MLK3 played its role in cervical cancer cells via the Notch‐1/autophagy network. Our results demonstrate the importance of MLK3 in cervical cancer progression via modulating the Notch‐1/autophagy network, and suggest that MLK3 is a promising therapeutic target for cervical cancer. [ABSTRACT FROM AUTHOR]

Neuroblastoma is a childhood tumor arising from developing a sympathetic nervous system and causes around 10% of pediatric tumors. Despite advancement in the use of sophisticated techniques in molecular biology, neuroblastoma patient's survivability rate is very less. Notch pathway is significant in upholding cell maintenance and developmental process of organs. Notch-1 proteins are a ligand-activated transmembrane receptor which decides the fate of the cell. Notch signaling leads to transcription of genes which indulged in numerous diseases including tumor progression. Ganoderic acid, a lanosterol triterpene, isolated from fungus Ganoderma lucidum with a wide range of medicinal values. In the present study, various isoforms of the ganoderic acid and natural inhibitors were docked by molecular docking using Maestro 9 in the Notch-1 signaling pathway. The receptor-based molecular docking exposed the best binding interaction of Notch-1 with ganoderic acid A with GScore (− 8.088), kcal/mol, Lipophilic EvdW (− 1.74), Electro (− 1.18), Glide emodel (− 89.944) with the active participation of Arg 189, Arg 199, Glu 232 residues. On the other hand natural inhibitor, curcumin has GScore (− 7.644), kcal/mol, Lipophilic EvdW (− 2.19), Electro (− 0.73), Glide emodel (− 70.957) with Arg 75 residues involved in docking. The ligand binding affinity of ganoderic acid A in Notch-1 is calculated using MM-GBSA (− 76.782), whereas curcumin has (− 72.815) kcal/mol. The QikProp analyzed the various drug-likeness parameters such as absorption, distribution, metabolism, excretion, and toxicity (ADME/T) and isoforms of ganoderic acid require some modification to fall under Lipinski rule. The ganoderic acid A and curcumin were the best-docked among different compounds and exhibits downregulation in Notch-1 mRNA expression and inhibits proliferation, viability, and ROS activity in IMR-32 cells. [ABSTRACT FROM AUTHOR]

Background: Alterations in the nasal epithelial barrier homeostasis and increased interleukin 33 (IL‐33) expression contribute to the pathogenesis of chronic rhinosinusitis with nasal polyps (CRSwNP). Aims: As Notch‐1 signaling is crucial in repair processes of mucosa, the current study assessed Notch‐1/Jagged‐1 signaling and IL‐33 in the epithelium of nasal polyps biopsies from allergic (A‐CRSwNP; n = 9) and not allergic (NA‐CRSwNP; n = 9) subjects by immunohistochemistry. We also assessed, in a model of nasal epithelial cells, the effects of stimulation of Notch‐1 with Jagged‐1 on the expression of IL‐33 (by flow cytometry, immunofluorescence, and immunocytochemistry), Jagged‐1 (by flow cytometry), and p‐CREB transcription factor (by western blot analysis). Results: Ex vivo (a) in normal epithelium, the expression of Notch‐1 and IL‐33 were higher in NA‐CRSwNP than in A‐CRSwNP; (b) in metaplastic epithelium, the expression of Notch‐1, Jagged‐1, and IL‐33 were higher in NA‐CRSwNP than in A‐CRSwNP; (c) in hyperplastic epithelium, the expression of Notch‐1, Jagged‐1, and IL‐33 were higher in A‐CRSwNP than in NA‐CRSwNP; and (d) in basal epithelial cells, no differences were observed in the expression of Jagged‐1, IL‐33, and Notch‐1. The expression of Notch‐1 significantly correlated with the expression of IL‐33. In vitro, stimulation of Notch‐1 with Jagged‐1 induced the expression of (a) Jagged‐1; (b) IL‐33; and (c) p‐CREB transcription factor. The inhibitor of Notch‐1, DAPT, reduced all the effects of Jagged‐1 on nasal epithelial cells. Conclusions: The data herein provided support, for the first time, a putative role of Notch‐1/Jagged‐1 signaling in the overexpression of IL‐33 in the epithelium of nasal polyps from patients with CRSwNP. The current study assessed for the first time Notch‐1, Jagged‐1, and interleukin 33 expression in the epithelium of CRSwNP from allergic and not allergic exsmoker subjects. The data demonstrated a role of Notch‐1/Jagged‐1 signaling in a positive feedback upregulating Jagged‐1 expression and in promoting proinflammatory responses inducing p‐CREB and IL‐33 expression in nasal epithelial cells. [ABSTRACT FROM AUTHOR]

Expression changes of Notch-1 and nuclear factor-κB (NF-κB) in cardiomyocytes from coronary heart disease (CHD) rats were investigated. The CHD model established by high-fat diet in 48 clean SD rats was set as the observation group, and another 48 healthy rats routinely fed were the control group. Rats were sacrificed on the 1st, 3rd, 7th, and 14th days after successful modeling. The rat myocardial cells were harvested to examine the changes of Notch-1 and NF-κB using immunoblot (western blot analysis) and TUNEL assay. Cardiomyocyte apoptotic rate, by Pearson's correlation test was used to analyze the correlation between Notch-1 and NF-κB. With the prolongation of the course of CHD in rats, the expression levels of Notch-1 and NF-κB proteins gradually increased, and the expression in the observation group was significantly higher than that in the control group (P<0.01). On the seventh day, the expression levels of Notch-1 and NF-κB protein in the rats in the observation group showed significant difference from those on day 1 and day 3 (P<0.05), and they were significantly different on the 7th and 14th days (P<0.05). There was no correlation between the expression level of the two proteins with the age and sex of the rats. Pearson's correlation analysis showed that Notch-1 was positively correlated with NF-κB protein (r=0.745, P=0.005). The results of myocardial apoptosis test showed that with the prolongation of the CHD course in the rats of the observation group, the cardiomyocyte apoptosis rate did not differ from the control group. The expression of Notch-1 and NF-κB protein is increased in cardiomyocytes of CHD rats. Notch-1 and NF-κB participate in the occurrence and development of CHD. [ABSTRACT FROM AUTHOR]

The present study aimed to investigate whether rhamnetin induced apoptosis in human breast cancer cells and the underlying molecular mechanism of this anti cancer effect. The treatment of MCF-7 cells with rhamnetin was able to significantly inhibit cell proliferation and induce caspase-3/9 activity in a dose- and time-dependent manner, compared with untreated cells. In addition, treatment with rhamnetin was able to significantly promote the expression of p53 protein and microRNA (miR-)34a compared with untreated cells. The treatment with rhamnetin also suppressed the expression of Notch1 protein in MCF-7 cells compared with untreated cells. Subsequently, miR-24a expression was promoted in rhamnetin-treated MCF-7 cells using a miR-34a plasmid. The overexpression of miR-34a was able to significantly inhibit cell viability and induce caspase-3/9 activity in MCF-7 cells following treatment with rhamnetin. Furthermore, the overexpression of miR-34a was able to significantly promote the expression of p53 protein and miR-34a, and suppress the expression of Notch1 protein in rhamnetin-treated MCF-7 cells. Therefore, the results of the present study demonstrated that rhamnetin induced apoptosis in human breast cancer cells via the miR-34a/Notch-1 signaling pathway. [ABSTRACT FROM AUTHOR]

Cancer stem cells (CSCs) constitute a subpopulation of transformed cells that possess intrinsic ability to undergo selfrenewal and differentiation, which drive tumour resistance and cancer recurrence. It has been reported that CSCs possess enhanced protection against oxidative stress induced by reactive oxygen species compared with nonstem-like cancer cells. In the present work, we investigated the role of heme oxygenase-1 (HO-1), a representative antioxidant enzyme, on the stemness and selfrenewal of human breast CSCs. We found that pharmacologic or genetic inhibition of HO-1 attenuated the sphere formation, whereas HO-1 inducers enhanced the number and the size of tumourspheres in breast CSCs. Carbon monoxide (CO) is endogenously generated as a consequence of degradation of heme by HO-1. The proportion of populations of CD44+/CD24− cells retaining CSC properties was increased in MDA-MB-231 cells treated with a CO-releasing molecule (CORM-2). Following CORM-2 treatment, the expression of Notch-1 and related genes Jagged-1 and Hes1 was increased, which was accompanied by the mammosphere formation. Taken together, these findings suggest that HO-1-derived CO production stimulates the formation of mammospheres in breast cancer cells through activation of Notch-1 signalling. [ABSTRACT FROM AUTHOR]

The aim of the present study was to investigate the role of long non-coding (lnc)RNA HOTAIR in premature ovarian failure. A total of 69 women with spontaneous premature ovarian failure and 48 healthy women were enrolled in the present study. Ovarian tissues and blood samples were harvested from each participant. The expression of HOTAIR in ovarian tissues and sera was detected using reverse transcription-quantitative polymerase chain reaction. A receiver operator characteristic curve analysis was performed to assess the diagnostic value of HOTAIR expression in ovarian tissues and sera for premature ovarian failure. Hamster ovary cell lines overexpressing HOTAIR were established and cell apoptosis were assessed using an MTT assay. The effect of HOTAIR upregulation on the expression of Notch-1 was measured using western blotting. The expression of HOTAIR in ovarian tissues and serum samples was significantly lower in patients with premature ovarian failure compared with healthy controls. These results suggest that ovarian and serum HOTAIR expression levels may be used to accurately predict the risk of premature ovarian failure. It was also demonstrated that HOTAIR overexpression upregulates Notch-1 protein expression in hamster ovary cells and reduced apoptosis, whereas the Notch inhibitor L685458 ameliorated these effects. In conclusion, the results of the present study suggest that LncRNA HOTAIR overexpression improves premature ovarian failure by upregulating the expression of Notch-1. [ABSTRACT FROM AUTHOR]

Abstract Background Nasal polyposis (NP) and sinonasal inverted papillomas (SIP) are considered benign lesions capable of recurrence or malignant transformation although not with the same prevalence. Since fluctuations of Caveolin-1 and Notch-1 proteins expression have been reported in many pathologies, the current study aimed to investigate their involvement in the epithelial transformation observed in SIPs compared to NP. Methods Immunohistochemical expression of Caveolin-1 and Notch-1 proteins was assessed in 104 patients with sinonasal lesions (45 NP, 45 SIP and 14 NP with SIP), semiquantively (percentage times intensity). Proteins expression profiles were evaluated statistically for their correlation with patients demographic and clinicopathological variables (grade of dysplasia, inflammation, recurrence) as well as with markers of proliferation (Ki67) and apoptosis (7-AAD) as determined by flow cytometry analysis. Results SIP lesions presented increased Caveolin-1 immunopositivity compared to NP (62.2%, vs 40.9%; p = 0.045). Cytoplasmic staining was observed only in epithelium's basal and suprabasal layers. Caveolin-1 positivity was not related to Ki67 expression, apoptosis, inflammation or dysplasia, eventhough 81.8% of highly immunopositive lesions were dysplastic (p = 0.03). Also, smokers presented significantly increased immunopositivy (p = 0.03). In contrast SIP lesions presented reduced Notch-1 expression compared to NP (68.9% vs 100%; p < 0.001). Dysplastic lesions presented low Notch-1 immunopositivity (p < 0.001). Enhancement of Notch-1 gene expression was also associated with inflammation. Conclusions The herein presented data suggest that the expression profiles of Caveolin-1 and Notch-1 proteins in sinonasal pathologies are distinctive and that could be explored as potential targets for the development of alternative therapeutic approaches. [ABSTRACT FROM AUTHOR]

Introduction: Pancreatic cancer is a complex disease, with an extremely poor response to chemotherapy. Emerging evidence indicates that the tumor microenvironment (TME) might play an important role in mediating chemoresistance. Areas covered: The evaluated study by Geller and collaborators describes several bacterial species within pancreatic tumor tissues and TME and investigated their roles in gemcitabine chemoresistance. Intratumor bacteria express the enzyme cytidine deaminase (CDD), whose long form (CDDL) was shown to metabolize gemcitabine into its inactive metabolite. CDDL is mostly expressed by Gammaproteobacteria and this was among the most common species in pancreatic cancer tissues. Interestingly, mouse models of colorectal cancer injected with bacterial CDDL displayed a reduced response to gemcitabine, but this resistance was neutralized by the antibiotic ciprofloxacin. Expert Commentary: The increased knowledge on the microbiome in pancreatic tissues, as well as its role in chemoresistance, will provide innovative prognostic and therapeutic strategies. [ABSTRACT FROM AUTHOR]

Ｏｂｊｅｃｔｉｖｅ　Ｔｏ ｉｎｖｅｓｔｉｇａｔｅ ｔｈｅ ｒｏｌｅｓ ａｎｄ ａｎｔｉ⁃ｃａｎｃｅｒ ｍｅｃｈａｎｉｓｍ ｏｆ ａｒｔｉｆｉｃｉａｌｌｙ ｓｙｎｔｈｅｓｉｚｅｄ ＥＧＦ⁃ｃｏｎｔａｉｎｉｎｇ ｆｉｂｕｌｉｎ⁃ｌｉｋｅ ｅｘｔｒａｃｅｌｌｕｌａｒ ｍａｔｒｉｘ ｐｒｏｔｅｉｎ （ＥＦＥＭＰ１） ｄｅｒｉｖｅｄ ｔｕｍｏｒ ｓｕｐｐｒｅｓｓｏｒ ＺＲ３０ ｐｒｏｔｅｉｎ ｉｎ ｇｌｉｏｍａ （ＧＢＭ）． Ｍｅｔｈｏｄｓ　ＺＲ３０ ｐｒｏｔｅｉｎ ｗｅｒｅ ｉｎ ｖｉｔｒｏ ｅｘｐｒｅｓｓｅｄ ｕｓｉｎｇ ａ ｗｈｅａｔ ｇｅｒｍ ｃｅｌｌ⁃ｆｒｅｅ ｓｙｓｔｅｍ． ＧＢＭ ｃｅｌｌ ｌｉｎｅｓ （Ｕ２５１， Ｕ２５１ＮＳ， ａｎｄ Ｕ８７） ｗｅｒｅ ｃｕｌｔｕｒｅｄ ｆｏｒ ２⁃３ ｄａｙｓ ｉｎ ｔｈｅ ｐｒｅｓｅｎｃｅ ｏｒ ａｂｓｅｎｃｅ ｏｆ ＺＲ３０ ｔｒｅａｔｍｅｎｔ． ＭＭＰ⁃２ ｌｅｖｅｌ ｗａｓ ｄｅｔｅｃｔｅｄ ｂｙ ｇｅｌａｔｉｎ ｚｙｍｏｇｒａｐｈｙ ａｓｓａｙ， ｍｏｒｅｏｖｅｒ， ｔｈｅ ｅｘｐｒｅｓｓｉｏｎ ｏｆ ＥＧＦＲ， Ｎｏｔｃｈ⁃１ ａｎｄ ｐ⁃Ａｋｔ／ Ａｋｔ ｌｅｖｅｌｓ ｗｅｒｅ ｄｅｔｅｒｍｉｎｅｄ ｂｙ ｗｅｓｔｅｒｎ ｂｌｏｔ． Ａｄｄｉｔｉｏｎａｌｌｙ， ＭＴＴ ａｓｓａｙ ｗａｓ ｕｓｅｄ ｔｏ ｍｅａｓｕｒｅ ＺＲ３０′ｓ ｅｆｆｅｃｔ ｏｎ ｔｈｅ ｃｅｌｌ ｐｒｏｌｉｆｅｒａｔｉｏｎ ｏｆ Ｕ２５１ ａｎｄ Ｕ２５１ＮＳ ｃｅｌｌｓ． Ｆｕｒｔｈｅｒｍｏｒｅ， ｐｒｅ⁃ｍｉｘｅｄ Ｕ２５１⁃ ＧＦＰ ａｎｄ Ｕ２５１ＮＳ⁃ＲＦＰ ｃｅｌｌｓ （１ ∶ ９） ｗｅｒｅ ｉｎｊｅｃｔｅｄ ｉｎｔｏ ｔｈｅ ｂｒａｉｎ ｏｆ ｎｕｄｅ ｍｉｃｅ， ａｎｄ ｔｈｅｎ ＺＲ３０ ｏｒ ＰＢＳ ｗａｓ ｉｎｊｅｃｔｅｄ ｉｎｔｏ ｔｈｅ ｉｎｔｒａ⁃ｔｕｍｏｒ ａｆｔｅｒ １０ ａｎｄ ２１ ｄａｙｓ， ｒｅｓｐｅｃｔｉｖｅｌｙ． Ｔｈｅｎ ＤＮＡ ｗａｓ ｅｘｔｒａｃｔｅｄ ｆｒｏｍ ｔｈｅ ｒｉｇｈｔ ｂｒａｉｎ ｏｆ ｎｕｄｅ ｍｉｃｅ ｉｎ ｅａｃｈ ｇｒｏｕｐ． Ｃｏｍｐａｒａｔｉｖｅ ｑｕａｎｔｉｔａｔｉｖｅ ｐｏｌｙｍｅｒａｓｅ ｃｈａｉｎ ｒｅａｃｔｉｏｎ （ＣＱ⁃ＰＣＲ） ｗａｓ ｕｓｅｄ ｔｏ ｅｘａｍｉｎｅ ｔｈｅ ｃｏｐｙ ｎｕｍｂｅｒｓ ｏｆ ｈｕｍａｎ ｇｅｎｅ ｈＳＰＡＧ１６， ｍｏｕｓｅ ｇｅｎｅ ｍＳｐａｇ１６， ＧＦＰ ａｎｄ ＲＦＰ． Ｔｈｅ ｓｕｒｖｉｖａｌ ｓｔａｔｕｓ ｏｆ ｅａｃｈ ｇｒｏｕｐ ｏｆ ｎｕｄｅ ｍｉｃｅ ｗａｓ ａｌｓｏ ｏｂｓｅｒｖｅｄ． Ｒｅｓｕｌｔｓ　Ｔｈｅ ｌｅｖｅｌｓ ｏｆ ａｃｔｉｖａｔｅｄ ＭＭＰ⁃２ ｉｎ Ｕ８７ ａｎｄ Ｕ２５１ ｃｅｌｌｓ ｗｅｒｅ ｌｏｗｅｒ ａｆｔｅｒ １０， ５０ ａｎｄ １００ ｎｇ／ ｍｌ ＺＲ３０ ｔｒｅａｔｍｅｎｔ ｆｏｒ ２⁃３ ｄａｙｓ． Ｗｅｓｔｅｒｎ ｂｌｏｔ ａｎａｌｙｓｉｓ ｓｈｏｗｅｄ ｔｈａｔ ＺＲ３０ ｔｒｅａｔｍｅｎｔ ｒｅｄｕｃｅｄ ｔｈｅ ｅｘｐｒｅｓｓｉｏｎ ｏｆ ＥＧＦＲ， Ｎｏｔｃｈ⁃１ ａｎｄ ｐ⁃Ａｋｔ／ Ａｋｔ ｉｎ Ｕ２５１ ｃｅｌｌｓ， ａｎｄ ｉｎｈｉｂｉｔｅｄ Ｎｏｔｃｈ⁃１ ａｎｄ ｐ⁃Ａｋｔ／ Ａｋｔ ｅｘｐｒｅｓｓｉｏｎ ｉｎ Ｕ２５１ＮＳ ｃｅｌｌｓ， ａｎｄ ｔｈｅｎ ｄｅｃｒｅａｓｅｄ ｔｈｅ ｒｅｓｐｏｎｓｅ ｏｆ Ｕ２５１ ｃｅｌｌｓ ｔｏ ＥＧＦ ｓｔｉｍｕｌａｔｉｏｎ． Ｍｏｒｅｏｖｅｒ， ＺＲ３０ ｉｎｈｉｂｉｔｅｄ ｔｈｅ ｃｅｌｌ ｐｒｏｌｉｆｅｒａｔｉｏｎ ｏｆ Ｕ２５１ ａｎｄ Ｕ２５１ＮＳ ｔｗｏ ｄａｙｓ ａｆｔｅｒ ｅｘｐｏｓｕｒｅ． Ｔｈｅ ｉｎ ｖｉｖｏ ｏｒｔｈｏｔｏｐｉｃ ＧＢＭ ｘｅｎｏｇｒａｆｔｓ ｗｅｒｅ ｓｕｃｃｅｓｓｆｕｌｌｙ ｃｏｎｓｔｒｕｃｔｅｄ． ＣＱ⁃ＰＣＲ ｒｅｓｕｌｔｓ ｉｎｄｉｃａｔｅｄ ｔｈａｔ ｔｈｅ ｈＳＰＡＧ１６／ ｍＳｐａｇ１６ ｒａｔｉｏｓ ｏｆ ｍｉｃｅ ｉｎ ＰＢＳ ｇｒｏｕｐ ａｎｄ ＺＲ３０ ｔｒｅａｔｍｅｎｔ ｇｒｏｕｐｓ ａｔ １８０， ７００， ａｎｄ １ ８００ ｎｇ ｄｏｓａｇｅｓ ｗｅｒｅ ３．６７±２．８２， １．１８±０．９７， １．７５±１．５５ ａｎｄ １．３８±１．１７， ｒｅｓｐｅｃｔｉｖｅｌｙ， ａｎｄ ＺＲ３０ ｔｒｅａｔｍｅｎｔ ｇｒｏｕｐｓ ｓｈｏｗｅｄ ｓｉｇｎｉｆｉｃａｎｔｌｙ ｌｏｗｅｒ ｒａｔｉｏｓ ｔｈａｎ ｔｈｅ ＰＢＳ ｇｒｏｕｐ （Ｐ＜０．０５ ｆｏｒ ａｌｌ）． Ｃｏｒｒｅｓｐｏｎｄｉｎｇｌｙ， ｔｈｅ ｒａｔｉｏｓ ｏｆ ＧＦＰ／ ＲＦＰ ｉｎ ｅａｃｈ ｇｒｏｕｐ ｗｅｒｅ １．９７±０．８０， １．９７±０．８５， １．４８±０．７１ ａｎｄ １．７３±０．７７， ｒｅｓｐｅｃｔｉｖｅｌｙ， ｓｈｏｗｉｎｇ ｎｏ ｓｔａｔｉｓｔｉｃａｌ ｓｉｇｎｉｆｉｃａｎｃｅ （Ｐ＞０．０５ ｆｏｒ ａｌｌ）． Ｗｈｅｎ ｔｒｅａｔｍｅｎｔ ｗａｓ ｐｅｒｆｏｒｍｅｄ １０ ｄ ａｆｔｅｒ ｃｅｌｌ ｉｍｐｌａｎｔａｔｉｏｎ， ａｎｄ ｔｈｅ ｍｅｄｉａｎ ｓｕｒｖｉｖａｌ ｔｉｍｅ ｏｆ ｍｉｃｅ ｉｎ ＰＢＳ ｇｒｏｕｐ ａｎｄ ＺＲ３０ ｇｒｏｕｐ ｗａｓ ４０．５ ｄａｙｓ ａｎｄ ５９．０ ｄａｙｓ， ｒｅｓｐｅｃｔｉｖｅｌｙ． Ｗｈｅｎ ｔｒｅａｔｍｅｎｔ ｗａｓ ｐｅｒｆｏｒｍｅｄ ２１ ｄ ａｆｔｅｒ ｃｅｌｌ ｉｍｐｌａｎｔａｔｉｏｎ， ｔｈｅ ｍｅｄｉａｎ ｓｕｒｖｉｖａｌ ｔｉｍｅ ｏｆ ｍｉｃｅ ｉｎ ＰＢＳ ｇｒｏｕｐ ａｎｄ ＺＲ３０ ｇｒｏｕｐ ｗａｓ ｅｘｔｅｎｄｅｄ ｔｏ ５７．０ ｄａｙｓ ａｎｄ ７４．５ ｄａｙｓ， ｒｅｓｐｅｃｔｉｖｅｌｙ． Ｔｈｅ ｍｅｄｉａｎ ｓｕｒｖｉｖａｌ ｔｉｍｅ ｏｆ ＺＲ３０ ｔｒｅａｔｍｅｎｔ ｇｒｏｕｐｓ ｓｉｇｎｉｆｉｃａｎｔｌｙ ｐｒｏｌｏｎｇｅｄ （Ｐ＜０．０５ ｆｏｒ ａｌｌ）． Ｃｏｎｃｌｕｓｉｏｎｓ　ＺＲ３０ ｉｎｈｉｂｉｔｓ ｉｎ ｖｉｔｒｏ ｃｅｌｌ ｇｒｏｗｔｈ， ｉｎｖａｓｉｏｎ， ａｎｇｉｏｇｅｎｅｓｉｓ ａｎｄ ｓｔｅｍｎｅｓｓ ｍａｉｎｔｅｎａｎｃｅ ｉｎ ｇｌｉｏｍａ ｖｉａ ｓｕｐｐｒｅｓｓｉｎｇ ａｃｔｉｖａｔｅｄ ＭＭＰ⁃２， ＥＧＦＲ， ｐ⁃Ａｋｔ／ Ａｋｔ ａｎｄ Ｎｏｔｃｈ⁃１ ｐｒｏｔｅｉｎｓ． Ｉｎ ｖｉｖｏ， ＺＲ３０ ｍａｒｋｅｄｌｙ ｉｎｃｒｅａｓｅｄ ｓｕｒｖｉｖａｌ ｏｆ ｍｉｃｅ ｈａｒｂｏｒｉｎｇ ｇｌｉｏｍａ ｘｅｎｏｇｒａｆｔｓ， ｅｖｅｎ ｆｏｒ ｏｎｌｙ ｏｎｅ ｉｎｔｒａ⁃ｔｕｍｏｒａｌ ｉｎｊｅｃｔｉｏｎ ａｔ ｔｈｅ ｔｉｍｅ ｏｆ ｅａｒｌｙ ｔｕｍｏｒ ｆｏｒｍａｔｉｏｎ． Ｏｖｅｒａｌｌ， ｔｈｅ ｉｎ ｖｉｖｏ ａｎｄ ｉｎ ｖｉｔｒｏ ｅｘｐｅｒｉｍｅｎｔｓ ｓｕｐｐｏｒｔｅｄ ｔｈｅ ｔｈｅｒａｐｅｕｔｉｃ ｐｏｔｅｎｔｉａｌ ｏｆ ＺＲ３０ ｆｏｒ ＧＢＭ. [ABSTRACT FROM AUTHOR]

Fengqin Xu,1,* Zhi-qiang Zhang,2,* Yong-chao Fang,2 Xiao-lei Li,2 Yu Sun,2 Chuan-zhi Xiong,2 Lian-qi Yan,2 Qiang Wang2 1Department of Orthopaedics, Hongquan Hospital, 2Department of Orthopaedics, Subei People’s Hospital, Yangzhou, Jiangsu Province, People’s Republic of China *These authors contributed equally to this work Background: Metastasis-associated lung adenocarcinoma transcript 1 (MALAT-1) is identified to be overexpressed in several cancers. However, the role of MALAT-1 in chondrosarcoma is poorly understood.Methods: The expression of MALAT-1 and Notch-1 signaling pathway was detected in chondrosarcoma tissues and chondrosarcoma cells by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot. 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay was performed to examine the cell viability of chondrosarcoma cells transfected with si-MALAT-1 or pcDNA-MALAT-1. Then the expression of Notch-1 signaling pathway was detected when MALAT-1 was upregulated or downregulated in chondrosarcoma cells. A subcutaneous chondrosarcoma cells xenograft model was used to confirm the effect of MALAT-1 on tumor growth in vivo.Results: We found the increased expression of MALAT-1 and Notch-1 signaling pathway in chondrosarcoma tissue and cells. MALAT-1 promoted the proliferation of chondrosarcoma cells. In addition, MALAT-1 activated the Notch-1 signaling pathway at posttranscriptional level in chondrosarcoma cells. Meanwhile, overexpression of Notch-1 reversed the effect of si-MALAT-1 on the proliferation of chondrosarcoma cells. Finally, we found that MALAT-1 promoted the tumor growth in a subcutaneous chondrosarcoma cells xenograft model, which confirmed the promoted effect of MALAT-1 on the tumor growth in vivo.Conclusion: Taken together, our study demonstrated that MALAT-1 promoted the proliferation of chondrosarcoma cell via activating Notch-1 signaling pathway. Keywords: MALAT-1, cell proliferation, chondrosarcoma, Notch-1, molecular pathophysiology

Ponatinib, a third-generation tyrosine kinase inhibitor (TKI), is the only approved TKI that is effective against T315I mutations in patients with chronic myeloid leukemia (CML). Specific activation of Notch signaling in CML cells by ponatinib can be considered as the “on-target effect” on the tumor and represents a therapeutic approach for CML. Nevertheless, ponatinib-induced vascular toxicity remains a serious concern, with underlying mechanisms being poorly understood. We aimed to determine the mechanisms of ponatinib-induced vascular toxicity, defining associated signaling pathways and identifying potential rescue strategies. We exposed human umbilical endothelial cells (HUVECs) to ponatinib or vehicle in the presence or absence of the neutralizing factor anti-Notch-1 antibody for exposure times of 0−72 h. Label-free proteomics and network analysis showed that protein cargo of HUVECs treated with ponatinib triggered apoptosis and inhibited vasculature development. We validated the proteomic data showing the inhibition of matrigel tube formation, an up-regulation of cleaved caspase-3 and a downregulation of phosphorylated AKT and phosphorylated eNOS. We delineated the signaling of ponatinib-induced vascular toxicity, demonstrating that ponatinib inhibits endothelial survival, reduces angiogenesis and induces endothelial senescence and apoptosis via the Notch-1 pathway. Ponatinib induced endothelial toxicity in vitro. Hyperactivation of Notch-1 in the vessels can lead to abnormal vascular development and vascular dysfunction. By hyperactivating Notch-1 in the vessels, ponatinib exerts an “on-target off tumor effect”, which leads to deleterious effects and may explain the drug’s vasculotoxicity. Selective blockade of Notch-1 prevented ponatinib-induced vascular toxicity.

ADAM10 metalloprotease is required for activation of Notch-1, a transmembrane receptor regulating cell differentiation, proliferation and apoptosis, whose intracellular proteolytic fragment NICD mediates some key cardiovascular effects of the hormone relaxin (RLX). This study demonstrates the involvement of ADAM10 and PI3K/Akt signaling in mediating RLX-induced Notch-1 activation. H9c2 cardiomyocytes and NIH3T3 fibroblasts were incubated with human RLX-2 (17 nmol/l, 24 h) in presence or absence of the PI3K or Akt inhibitors wortmannin (WT, 100 nmol/l) and triciribine (TCN, 1 μmol/l). Cyclohexanedione-inactivated RLX (iRLX) served as negative control. RLX significantly increased Akt phosphorylation, ADAM10 and NICD expression, which were abolished by WT or TCN and did not occur with iRLX. These findings highlight a new receptor-specific signal transduction pathway of RLX. [ABSTRACT FROM AUTHOR]

The antifibrotic potential of platelet-rich plasma (PRP) is controversial. This study examined the effects of PRP on in vitro transforming growth factor (TGF)-β1-induced differentiation of fibroblasts into myofibroblasts, the main drivers of fibrosis, and the involvement of vascular endothelial growth factor (VEGF)-A in mediating PRP-induced responses. The impact of PRP alone on fibroblast differentiation was also assessed. Myofibroblastic phenotype was evaluated by confocal fluorescence microscopy and western blotting analyses of α-smooth muscle actin (sma) and type-1 collagen expression, vinculin-rich focal adhesion clustering, and stress fiber assembly. Notch-1, connexin 43, and VEGF-A expression were also analyzed by RT-PCR. PRP negatively regulated fibroblast-myofibroblast transition via VEGF-A/VEGF receptor (VEGFR)-1-mediated inhibition of TGF-β1/Smad3 signaling. Indeed TGF-β1/PRP co-treated fibroblasts showed a robust attenuation of the myofibroblastic phenotype concomitant with a decrease of Smad3 expression levels. The VEGFR-1 inhibition by KRN633 or blocking antibodies, or VEGF-A neutralization in these cells prevented the PRP-promoted effects. Moreover PRP abrogated the TGF-β1-induced reduction of VEGF-A and VEGFR-1 cell expression. The role of VEGF-A signaling in counteracting myofibroblast generation was confirmed by cell treatment with soluble VEGF-A. PRP as single treatment did not induce fibroblast myodifferentiation. This study provides new insights into cellular and molecular mechanisms underpinning PRP antifibrotic action. [ABSTRACT FROM AUTHOR]

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive cancers. Recent research has demonstrated that chronic pancreatitis (CP) is associated with an increased risk of PDAC, partly due to acinar‐to‐ductal metaplasia (ADM). Baicalein has been shown to exert anti‐inflammatory and anti‐tumor effects for CP or PDAC, respectively. The aim of our study was to investigate the effect of baicalein, and the putative underlying mechanism, on inflammatory cytokines‐induced ADM of rat pancreatic acinar cell line AR42J. To investigate ADM and baicalein effects in vitro, AR42J were treated with recombinant rat Tumor Necrosis Factor alpha (rTNFα) with or without baicalein for 5 days. Results showed that rTNFα‐induced AR42J cells switched their phenotype from dominantly amylase‐positive acinar cells to dominantly cytokeratin 19‐positive ductal cells. Moreover, expression of the transcripts for TNFα or Hes‐1, a Notch target, was up‐regulated in these cells. Interestingly, baicalein reduced the population of ADM as well as cytokines gene expression but not Hes‐1. Baicalein inhibited NF‐κB activation induced by rTNFα in AR42J, but no effect on Notch 1activation. Moreover, baicalein suppressed the secretion of TNFα and Nitric Oxide (NO) in macrophages stimulated with LPS and further inhibited ADM of conditional medium‐treated AR42J cells. Baicalein also suppressed the inflammatory response of LPS‐activated macrophages, thereby inhibited ADM of AR42J by altering their microenvironment. Taken together, our study indicates that baicalein reduces rTNFα‐induced ADM of AR42J cells by inhibiting NF‐κB activation. It also sheds new light on Chinese material medica therapy of pancreatitis and thereby prevention of PDAC. [ABSTRACT FROM AUTHOR]