Contactin-associated protein-like 2 (CNTNAP2) mutations impair the essential α-secretase cleavages, leading to autism-like phenotypes.

Mutations in the Contactin-associated protein-like 2 (CNTNAP2) gene are associated with autism spectrum disorder (ASD), and ectodomain shedding of the CNTNAP2 protein plays a role in its function. However, key enzymes involved in the C-terminal cleavage of CNTNAP2 remain largely unknown, and the effect of ASD-associated mutations on this process and its role in ASD pathogenesis remain elusive. In this report we showed that CNTNAP2 undergoes sequential cleavages by furin, ADAM10/17-dependent α-secretase and presenilin-dependent γ-secretase. We identified that the cleavage sites of ADAM10 and ADAM17 in CNTNAP2 locate at its C-terminal residue I79 and L96, and the main α-cleavage product C79 by ADAM10 is required for the subsequent γ-secretase cleavage to generate CNTNAP2 intracellular domain (CICD). ASD-associated CNTNAP2 mutations impair the α-cleavage to generate C79, and the inhibition leads to ASD-like repetitive and social behavior abnormalities in the Cntnap2-I1254T knock-in mice. Finally, exogenous expression of C79 improves autism-like phenotypes in the Cntnap2-I1254T knock-in and Cntnap2-/- knockout mice. This data demonstrates that the α-secretase is essential for CNTNAP2 processing and its function. Our study indicates that inhibition of the cleavage by pathogenic mutations underlies ASD pathogenesis, and upregulation of its C-terminal fragments could have therapeutical potentials for ASD treatment.

Enhanced Gasdermin-E-mediated Pyroptosis in Alzheimer's Disease.

Amyloid ß protein (Aß) is a critical factor in the pathogenesis of Alzheimer's disease (AD). Aß induces apoptosis, and gasdermin-E (GSDME) expression can switch apoptosis to pyroptosis. In this study, we demonstrated that GSDME was highly expressed in the hippocampus of APP23/PS45 mouse models compared to that in age-matched wild-type mice. Aß treatment induced pyroptosis by active caspase-3/GSDME in SH-SY5Y cells. Furthermore, the knockdown of GSDME improved the cognitive impairments of APP23/PS45 mice by alleviating inflammatory response. Our findings reveal that GSDME, as a modulator of Aß and pyroptosis, plays a potential role in Alzheimer's disease pathogenesis and shows that GSDME is a therapeutic target for AD.

Evaluation of InSAR Tropospheric Delay Correction Methods in the Plateau Monsoon Climate Region Considering Spatial-Temporal Variability.

The tropospheric delay caused by the temporal and spatial variation of meteorological parameters is the main error source in interferometric synthetic aperture radar (InSAR) applications for geodesy. To minimize the impact of tropospheric delay errors, it is necessary to select the appropriate tropospheric delay correction method for different regions. In this study, the interferogram results of the InSAR, corrected for tropospheric delay using the Linear, Generic Atmospheric Correction Online Service for InSAR (GACOS) and ERA-5 atmospheric reanalysis dataset (ERA5) methods, are presented for the study area of the junction of the Hengduan Mountains and the Yunnan-Kweichow Plateau, which is significantly influenced by the plateau monsoon climate. Four representative regions, Eryuan, Binchuan, Dali, and Yangbi, are selected for the study and analysis. The phase standard deviation (STD), phase-height correlation, and global navigation satellite system (GNSS) data were used to evaluate the effect of tropospheric delay correction by integrating topographic, seasonal, and meteorological factors. The results show that all three methods can attenuate the tropospheric delay, but the correction effect varies with spatial and temporal characteristics.

Transcriptional activation of CSTB gene expression by transcription factor Sp3.

CSTB has been reported to be associated with the pathogenesis of many malignant tumors, especially hepatocellular carcinoma (HCC). However, how the expression of this gene is regulated is largely unknown. We initially cloned and analyzed the promoter region of the CSTB gene by luciferase assay and the Sp3 binding site (CCCCGCCCCGCG) was found in it. The results of electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) experiments verified that the transcription factor, Sp3 could bind to the " CCCCGCCCCGCG ″ site of the CSTB gene promoter. We showed that the overexpression of Sp3 significantly increased the endogenous mRNA and protein expression levels of CSTB, whereas knockdown of Sp3 decreased the mRNA and protein expression levels according to quantitative real-time PCR (qRT‒PCR) and western blotting. In conclusion, CSTB gene expression is closely regulated by transcription factor Sp3, which may be a potential mechanism for the dysregulation of CSTB expression in HCC.

GPSAdb: a comprehensive web resource for interactive exploration of genetic perturbation RNA-seq datasets.

Gene knock-out/down methods are commonly used to explore the functions of genes of interest, but a database that systematically collects perturbed data is not available currently. Manual curation of all the available human cell line perturbed RNA-seq datasets enabled us to develop a comprehensive human perturbation database (GPSAdb, https://www.gpsadb.com/). The current version of GPSAdb collected 3048 RNA-seq datasets associated with 1458 genes, which were knocked out/down by siRNA, shRNA, CRISPR/Cas9, or CRISPRi. The database provides full exploration of these datasets and generated 6096 new perturbed gene sets (up and down separately). GPSAdb integrated the gene sets and developed an online tool, genetic perturbation similarity analysis (GPSA), to identify candidate causal perturbations from differential gene expression data. In summary, GPSAdb is a powerful platform that aims to assist life science researchers to easily access and analyze public perturbed data and explore differential gene expression data in depth.

Astrocyte-Ablation of Mtnr1b Increases Anxiety-Like Behavior in Adult Male Mice.

BACKGROUND: Astrocytes are essential for synaptic transmission, and their dysfunction can result in neuropsychiatric disorders such as anxiety and depression. Many studies have shown that global knockout of Melatonin receptor 2 (Mtnr1b) is associated with the development of various mental disorders. AIM: This study aimed to investigate the effects of astrocyte ablation of Mtnr1b on cognitive function and anxiety-like behavior in mice, as well as the potential biological mechanisms. METHODS: A conditional Cre-loxP system allowing deletion of Mtnr1b from astrocytes was developed to investigate the specific role Mtnr1b. Control and Mtnr1b cKO𝐺𝑓𝑎𝑝 mice were selected for cognitive function behavioral testing (Morris water maze test, novel object recognition test) and emotion-related behavioral testing (open field, elevated plus maze). After testing, brain tissue was collected and examined by immunofluorescence for the expression of neuronal nuclei (NeuN), glutamate decarboxylase 67 (GAD67), and vesicular glutamate transporter 1 (vGluT1). RNA-seq was performed on hippocampal tissue from control and Mtnr1b cKO𝐺𝑓𝑎𝑝 mice to identify differentially expressed genes. Additional confirmation of differential gene expression was performed using real-time quantitative polymerase chain reaction (qRT-PCR). RESULTS: Mtnr1b cKO𝐺𝑓𝑎𝑝 mice were not significantly different from control mice in the Morris water maze and novel object recognition tests. Results from the open field and elevated plus maze tests showed that Mtnr1b cKO𝐺𝑓𝑎𝑝 mice exhibited significantly more anxiety-like behavior than did controls. Immunofluorescence revealed that the number of mature neurons did not differ significantly between Mtnr1b cKO𝐺𝑓𝑎𝑝 mice and controls. The expression of GAD67 in the hippocampal CA1 and CA3 areas of Mtnr1b cKO𝐺𝑓𝑎𝑝 mice was significantly lower than in the control group, but no significant difference was detected for vGluT1 expression. RNA-seq and qRT-PCR results showed that Mtnr1b knockout in astrocytes led to a decrease in the levels of gamma-aminobutyric acid sub-type A (GABAA) receptors and Kir2.2. CONCLUSIONS: The astrocyte-specific knockout in Mtnr1b cKO𝐺𝑓𝑎𝑝 mice results in anxiety-like behavior, which is caused by down-regulation of gamma-aminobutyric acid-ergic (GABAergic) synaptic function.

Amyotrophic lateral sclerosis with primary progressive aphasia: a case report and literature review.

The association between amyotrophic lateral sclerosis (ALS) and primary progressive aphasia (PPA) is rarely seen in patients. A case of ALS-PPA with a possible reticulon 2 (RTN2) mutation was reported in this study. Moreover, we systematically reviewed the previous reports of 28 ALS cases with progressive non-fluent aphasia (PNFA) and semantic dementia (SD) to identified the unique pathologic features and strong heritability of ALS-PPA. There is a different heritability among the ALS-SD, ALS-PNFA, and the ALS-unclassified PPA groups (p=0.003). Males are more prone to have ALS-PPA than females in all the three groups (p=0.028). PPA-ALS usually starts with cognitive impairment, and the onset most often involves the bulbar. In addition, chromosome 9 open reading frame 72(C9ORF72) and TANK-binding kinase 1 (TBK1) are important pathogenic genes of PPA-ALS. Overall, heritability is of high certainty in ALS-SD, ALS-PNFA, and the ALS-unclassified PPA groups. TAR (Trans-Activator Regulatory) DNA-binding Protein 43 (TDP43) is a 100% predictive pathologic protein of ALS-PPA. C9ORF72 and TBK1 are important pathogenic genes of PPA-ALS.

Sp4 Regulates PTTG1IP Gene Transcription and Expression.

Pituitary tumor-transforming gene 1 protein (PTTG)-interacting protein, also known as PTTG-binding factor (PBF), is encoded by a proto-oncogene PTTG1IP. PBF has been identified through its interaction with PTTG. Similar to PTTG, PBF has been implicated in the etiology of several tumors, including pituitary, thyroid, and breast cancer. PBF can induce the translocation of PTTG into the nucleus, and then lead to tumorigenesis. Studies have shown that PBF plays a vital and complex role in increasing tumor development. However, the transcriptional regulation of PTTG1IP gene remains undefined. In this study, we have cloned a 467-bp fragment of the 5' flanking region of the human PTTG1IP gene and identified the region (-212 to +7 bp) necessary for PTTG1IP gene promoter activity by luciferase assay. Electrophoretic mobility shift assay revealed PTTG1IP gene promoter containing Sp4 response elements. Overexpression of Sp4 increased PTTG1IP gene transcription and expression in HeLa cells. Our study demonstrates that Sp4 regulates PTTG1IP gene transcription and expression.

Regulation of the Human IL-10RB Gene Expression by Sp8 and Sp9.

BACKGROUND: Interleukin-10 (IL-10) is a classic anti-inflammatory cytokine that exerts its effects via the receptor complexes IL-10RA and IL-10RB. Loss of IL-10RB results in many diseases. Moreover, IL-10RB is closely associated with neuronal survival and synaptic formation. However, the regulation of IL-10RB gene expression remains elusive. OBJECTIVE: To investigate whether the expression of IL-10RB gene is increased in brain of Alzheimer's disease (AD) and its transcriptional regulation. METHODS: We examined the gene expression of AD patient brain from public database and detected the protein expression of AD model mouse brain by western blot. We constructed a variety of reporter gene plasmids with different lengths or mutation sites, tested the promoter activity and defined the functional region of the promoter with the luciferase reporter assay. The protein-DNA binding between transcription factors and the promoter was analyzed using chromatin immunoprecipitation (ChIP) and electrophoretic mobility shift assay (EMSA). RESULTS: We found that the IL-10RB is elevated in the brain of AD patient and AD model mice. The minimal promoter of the IL-10RB gene is located in the -90 to +51 bp region (relative to the transcriptional start site) and is sufficient for high-level expression of the IL-10RB gene. Transcription factors Sp8 and Sp9 bind to the IL-10RB promoter in vitro. The overexpression or knockdown of Sp8 and Sp9 affected the IL-10RB promoter activity and its gene expression. CONCLUSION: Our study functionally characterized the promoter of the IL-10RB gene and demonstrated that Sp8 and Sp9 regulated its expression.

Clinical Profile and Treatment Outcome in MOGAD: A Single-Center Case-Series Study in Guiyang, China.

Background: The clinical spectrum of myelin oligodendrocyte glycoprotein antibody-associated disease (MOGAD) is expanding over time. However, the long-term management and prognosis of this disorder are still controversial. Therefore, this study aimed to report the clinical profiles and treatment outcomes of MOGAD in our center. Methods: This was a single-center case-series study. Clinical and para-clinical data, along with treatment outcomes of patients with MOGAD were analyzed. Results: A total of 27 patients were identified, of which 19 (70%) patients were women, and the median age at disease onset was 40 years (range 20-67). A total of 47 episodes were observed, with optic neuritis (53%) being the most frequent presentation and 60% of them were unilateral. Other presentations included rhombencephalitis (RE) (17%), limbic encephalitis (9%), simultaneous optic neuritis and myelitis (9%), acute disseminated encephalomyelitis (ADEM)-like presentation (6%), myelitis (4%), and ADEM (2%). One patient presenting with RE also met the diagnostic criteria of area postrema syndrome (APS). Another patient with RE presented with imaging characteristics of chronic lymphocytic inflammation with pontine perivascular enhancement responsive to steroids (CLIPPERS). A total of 29 lumbar punctures were recorded, among which an elevated protein level was found in 34% of the samples, pleocytosis was found in 14% of the samples, and positive intrathecal oligoclonal bands were found in 19% of the patients. One patient was found to have anti-N-methyl-D-aspartate receptor antibodies both in his serum and cerebrospinal fluid. Intravenous methylprednisolone (IVMP) was administrated for 85% of the attacks while both IVMP and intravenous immunoglobulin were for 6% of the attacks. Moreover, nine patients received maintenance therapy. Among them, six patients were treated with mycophenolate mofetil, three patients were treated with prednisone, rituximab, and teriflunomide, respectively. The median follow-up period was 20 months (range 6-127). At follow-up, twelve (44%) patients experienced a relapsing course, and the median time to the first relapse was 9.5 months (range 2-120). The median Expanded Disability Status Scale score at nadir was 3.5 (range 2-8) and was 0 (range 0-3) at the last follow-up. Conclusion: The clinical spectrum of MOGAD is heterogenous, wherein APS and CLIPPERS-form can occur. The long-term outcome of MOGAD seems benign. Further studies are warranted to determine the risk factors of relapse and identify the optimal steroid-sparing agents.

Chronic Alcohol Exposure Alters Gene Expression and Neurodegeneration Pathways in the Brain of Adult Mice.

BACKGROUND: Chronic alcohol consumption can alter the structure of the central nervous system and disrupt cognitive function. Alcoholics are more likely to develop neurodegenerative disorders such as Alzheimer's disease (AD) and Parkinson's disease (PD). However, the role of alcohol in promoting neurotoxicity and neurodegeneration remains unclear. OBJECTIVE: In this study, we aimed at estimating the effects of chronic binge alcohol exposure on brain transcriptome and behavior changes in a chronic "Drinking in the Dark" (DID) mouse model. METHODS: The adult C57BL/6J male mice were exposed to alcohol for 4 weeks. RNA-seq was applied to assess the effects of chronic alcohol exposure on transcriptome in brain. The open field test and novel object recognition test were used to assess the changes of anxiety level, locomotive function, and short-term memory induced by alcohol. RNA-seq analysis revealed that chronic alcohol exposure caused significant change in the brain transcriptome, especially in prefrontal cortex. RESULTS: The gene dysregulation caused by chronic alcohol exposure includes pathways related to mitochondrial energy metabolism (such as oxidative phosphorylation) and multiple neurodegenerative diseases (such as AD and PD). Furthermore, the pathway and network analyses suggest that the genes involved in mitochondrial energy metabolism, ubiquitin-proteasome system, Wnt signaling pathway, and microtubules may attribute to the neurotoxicity and neurodegeneration caused by chronic alcohol consumption. Additionally, locomotive function was also significantly impaired. CONCLUSION: This work provides gene transcriptional profile data for future research on alcohol-induced neurodegenerative diseases, especially AD and PD.

JAM2 predicts a good prognosis and inhibits invasion and migration by suppressing EMT pathway in breast cancer.

OBJECTIVES: Large-scale epidemiological surveys have shown that patients with Down syndrome, which is caused by a chromosomal abnormality (an extra chromosome 21), are significantly less likely to develop solid tumors, including breast cancer, than those without. This feature has prompted the search for oncogenes located on chromosome 21. Junctional adhesion molecule 2 (JAM2), which is located on chromosome 21, is expressed at low levels in breast cancer and is associated with a good prognosis. These findings strongly suggest that JAM2 may be a potential oncogene suppressor in breast cancer. However, the role and function of JAM2 in breast cancer are not yet clear. Therefore, this study aimed to explore the biological functions and mechanisms of JAM2 in breast cancer. METHODS: Several databases were used to explore JAM2 expression in breast cancer and to analyze its diagnostic and prognostic value in breast cancer. Changes in relevant markers were examined at the gene and protein levels using RT-qPCR and Western blot techniques, in addition, cell migration and invasion abilities were identified by scratch assays and transwell assays. Untargeted metabolomics, transcriptome sequencing and Luminex liquid suspension chip detection were performed in combination to study the mechanisms. RESULTS: JAM2 is expressed at low levels in breast cancer, and patients with high JAM2 expression have a good prognosis, indicating that JAM2 has good clinical diagnostic and prognostic value. Overexpression of JAM2 can block the invasion and migration of breast cancer cells, and the mechanism may be that JAM2 inhibits the EMT pathway. Finally, combined multiomics analysis revealed that JAM2 may affect the immune microenvironment of breast cancer by influencing the secretion of CXCL9/10 from tumor cells.

Coexisting amyotrophic lateral sclerosis and chorea: A case report and literature review.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) coexisting with chorea is very rare. CASE REPORT: We present the case of a 48-year-old man with ALS and chorea; the diagnostic certainty was high based on clinical examination results. Combining the data from literature, we analyzed the characteristics of patients with ALS and chorea. We found that ALS coexisting with chorea is very rare, but is often hereditary with a genetic mutation. Most patients with ALS and chorea are caused by abnormal amplification of a CAG sequence in the HTT gene, and these patients have a mild course of disease. The FUS, VCP, and SETX genes also have low mutation frequencies in patients with ALS and chorea. CONCLUSION: The abnormal amplification of a CAG sequence in the HTT gene in ALS with chorea has an obvious familial genetic tendency, and most patients have a mild disease course.

A Chinese CADASIL family with p.R578C mutation at exon 11 of the NOTCH3 gene.

OBJECTIVE: To analyze one clinical case of cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy(CADASIL), and to perform analysis of the related gene mutation for the proband and her family. METHODS: Analysis of clinical data from the patient diagnosed with CADASIL, including clinical manifestations, blood test results and brain imaging results, followed by high-throughput sequencing of blood samples. Pathogenicity assessment of the gene mutation, and first generation verification were performed on some family members according to genetic variation interpretation standards and guidelines of the American College of Medical Genetics and Genomics (ACMG). RESULTS: Onset of the proband occurred younger than 50-years-old with recurrent migraine attacks and positive family history of migraine and stroke, but without risk factors for cerebrovascular diseases. The craniocerebral magnetic resonance imaging (MRI) results showed diffusive white matter lesions and thus clinically met criteria for CADASIL diagnosis. NOTCH3 gene analysis showed a p.R578C mutation (1732 C > T) at the11th exon on chromosome 19 of the proband and some family members. CONCLUSIONS: NOTCH3 mutation is related to CADASIL. In this study, we observed a rather rare familial NOTCH3 mutation in China. This report further support the mutation site is pathogenic.

[Corrigendum] HORMAD1 promotes docetaxel resistance in triple negative breast cancer by enhancing DNA damage tolerance.

Following the publication of this article, the authors realized that the published version of Fig. 4A contained an erroneous label; essentially, the information purported to relate to experiments having been performed with docetaxel should not have been included in this figure. The correctly labelled version of Fig. 4 is shown with the remainder of Fig. 4 on the next page. This change does not affect the data shown in the paper, and the text in the published article did accurately describe the information shown in this figure. The authors sincerely apologize for the error that was introduced during the preparation of this figure, and thank the Editor for allowing them the opportunity to publish a Corrigendum. Furthermore, they regret any inconvenience caused. [the original article was published in Oncology Reports 46: Article no. 138, 2021; DOI: 10.3892/or.2021.8089].

HORMAD1 promotes docetaxel resistance in triple negative breast cancer by enhancing DNA damage tolerance.

HORMA domain­containing protein 1 (HORMAD1), is normally expressed only in the germline, but is frequently re­activated in human triple­negative breast cancer (TNBC); however, its function in TNBC is largely unknown. In the present study, the expression and biological significance of HORMAD1 in human TNBC was evaluated. Bioinformatics analysis and reverse transcription­quantitative PCR were used to evaluate HORMAD1 expression in datasets and cell lines. HORMAD1 protein expression was detected in TNBC samples using immunohistochemical assays, and the effect of HORMAD1 on cell proliferation was determined using Cell Counting Kit­8, plate colony formation and standard growth curve assays. Cell cycle, reactive oxygen species (ROS) and apoptosis analyses were conducted using flow cytometry. The activity of caspases was measured using caspase activity assay kit. The levels of key apoptosis regulators and autophagy markers were detected by western blot analysis. TNBC cell survival and apoptosis were not influenced by small interfering RNA targeting HORMAD1 alone; however, HORMAD1 knockdown enhanced autophagy and docetaxel (Doc)­induced apoptosis, compared with the control group. Furthermore, higher ROS levels and caspase­3, ­8 and ­9 activity were detected in MDA­MB­436 TNBC cells with HORMAD1 knockdown upon exposure to Doc. The levels of the induced DNA damage marker γH2AX were also higher, while those of the DNA repair protein RAD51 were lower in TNBC cells with HORMAD1 knockdown compared with the controls. Furthermore, the expression of the autophagy marker P62 was enhanced in MDA­MB­231 cells in response to HORMAD1 overexpression. Notably, Doc­induced apoptosis was similarly increased by both HORMAD1 overexpression and treatment with the autophagy inhibitor, 3­methyladenine (3MA); however, the Doc­induced increase in autophagy was not inhibited by 3MA. The present data indicated that HORMAD1 was involved in autophagy and that the inhibition of autophagy can partially enhance the induction of apoptosis by Doc. The role of HORMAD1 in the DNA damage tolerance of tumor cells may be the main reason for Doc resistance; hence, HORMAD1 could be an important therapeutic target in TNBC.

Forest structure dependency analysis of L-band SAR backscatter.

Forest structure plays an important role in forest biomass inversion using synthetic aperture radar (SAR) backscatter. Synthetic aperture radar (SAR) sensors with long-wavelength have the potentiality to provide reliable and timely forest biomass inversion for their ability of deep penetration into the forest. L-band SAR backscatter shows useful for forest above-ground biomass (AGB) estimation. However, the way that forest structure mediating the biomass-backscatter affects the improvement of the related biomass estimation accuracy. In this paper, we have investigated L-band SAR backscatter sensitivity to forests with different mean canopy density, mean tree height and mean DBH (diameter at breast height) at the sub-compartment level. The forest species effects on their relationship were also considered in this study. The linear correlation coefficient R, non-linear correlation parameter, Maximal Information Coefficient (MIC), and the determination coefficient R2 from linear function, Logarithmic function and Quadratic function were used in this study to analyze forest structural properties effects on L-band SAR backscatter. The HV channel, which is more sensitive than HH to forest structure parameters, was chosen as the representative of SAR backscatter. 6037 sub-compartment were involved in the analysis. Canopy density showed a great influence on L-band backscatter than mean forest height and DBH. All of the R between canopy density and L-band backscatter were greater than 0.7 during the forest growth cycle. The sensitivity of L-band backscatter to mean forest height depends on forest canopy density. When canopy density was lower than 0.4, R values between mean forest height are smaller than 0.5. In contrast, the values of R were greater than 0.8 if canopy density was higher than 0.4. The sensitivity SAR backscatter to DBH fluctuated with canopy density, but it only showed obvious sensitivity when canopy density equals to 0.6, where both the linear and non-liner correlation values are higher than others. However, their effects on L-bang HV backscatter are affected by forest species, the effects on three forest structural parameters depend on tree species.

AGR3 promotes estrogen receptor-positive breast cancer cell proliferation in an estrogen-dependent manner.

Breast cancer is one of the most common malignancies and the leading cause of cancer-associated death among women. Anterior gradient 3 (AGR3) is a cancer-associated gene and is similar to its homologous oncogene AGR2. However, whether AGR3 participates in breast cancer progression remains unclear. The present study aimed to investigate the function of AGR3 in ER-positive breast cancer. In the present study, reverse transcription-quantitative PCR was used to detect AGR3 mRNA expression in breast cancer tissues and cell lines; linear correlation analysis was used to investigate the correlation between AGR3 and estrogen receptor 1 (ESR1) expression in breast cancer via GEO dataset analysis; western blotting was used to assess the levels of AGR3, ER and GAPDH; small interfering (si)RNA transfection was used to knock down AGR3 and ESR1 expression; and finally the Cell Counting Kit-8 assay was used to evaluate cell viability. In the present study, AGR3 expression was markedly increased in estrogen receptor (ER)-positive breast cancer tissues and cell lines compared with that in ER-negative breast cancer. AGR3 expression was upregulated in estrogen-treated T47D cells, whereas 4-hydroxytamoxifen, an inhibitor of estrogen-ER activity in breast cancer cells, downregulated AGR3 expression in T47D cells. Functional assays demonstrated that knockdown of AGR3 using siRNAs inhibited T47D cell proliferation compared with that of the negative control group. Additionally, AGR3 expression was decreased after knocking down ESR1. The present results suggested that AGR3 may serve an important role in estrogen-mediated cell proliferation in breast cancer and that AGR3 knockdown may be a potential therapeutic strategy for ER-positive breast cancer.

Although c­MYC contributes to tamoxifen resistance, it improves cisplatin sensitivity in ER­positive breast cancer.

Tamoxifen (TAM) resistance is a major challenge in the treatment of estrogen receptor­positive (ER+) breast cancer. To date, to the best of our knowledge, there are only a few studies available examining the response of patients with TAM­resistant breast cancer to chemotherapy, and the guidelines do not specify recommended drugs for these patients. In the present study, TAM­resistant cells were shown to exhibit increased proliferation and invasion compared with the parent cells, and the increased expression of c­MYC was demonstrated to play an important role in TAM resistance. Furthermore, the TAM­resistant cells were significantly more sensitive to cisplatin compared with the parent cells, and the silencing of c­MYC expression desensitized the cells to cisplatin through the inhibition of the cell cycle. An increased c­MYC expression was observed in 28 pairs of primary and metastatic tumors from patients treated with TAM, and the clinical remission rate of cisplatin­based chemotherapy was significantly higher compared with other chemotherapy­based regimens in 122 patients with TAM resistant breast cancer. Taken together, the data of the present study demonstrated that although c­MYC was involved in TAM resistance, it increased the sensitivity of ER+ breast cancer to cisplatin. Thus, cisplatin may be a preferred chemotherapeutic agent for the treatment of patients with TAM­resistant breast cancer, particularly in patients where the rapid control of disease progression is required.

Sulfasalazine­induced ferroptosis in breast cancer cells is reduced by the inhibitory effect of estrogen receptor on the transferrin receptor.

The aim of the present study was to clarify the activation of ferroptosis in different breast cancer cells by sulfasalazine (SAS) and to explore the relationship between the estrogen receptor (ER) and the transferrin receptor (TFRC). MDA­MB­231 and T47D cells were treated with SAS for 24 h. Changes in cell morphology were observed under a microscope. CCK­8 was used to detect the proliferation inhibition rate and determine the IC50 values. Western blotting was used to detect the expression of glutathione peroxidase 4 (GPX4) and xCT. Flow cytometry was used to identify changes in the production of reactive oxygen species (ROS). Mitochondrial morphological changes in T47D were observed using transmission electron microscopy. Changes in the mitochondrial membrane potential (MMP) were observed using confocal fluorescence microscopy. RT­PCR was used to detect the mRNA expression levels of TFRC and divalent metal transporter 1 (DMT1). Bioinformatics analysis was performed on TFRC expression in 1,208 breast cancer samples and its relationship with ER. TFRC expression was detected in various breast cancer tissues using immunohistochemistry and in various breast cancer cells using western blotting. Small interfering RNA (siRNA) knocked down ER expression in T47D cells, and changes in the TFRC mRNA and protein levels were observed. RT­PCR was used to detect TFRC expression in 87 clinical specimens. The results of the present study revealed that SAS could inhibit breast cancer cell viability, which was accompanied by an abnormal increase in ROS and a depletion of GPX4 and system xc­. Liproxstatin­1 reversed the SAS­induced increase in ROS. The cells treated with SAS had shrunken mitochondria and decreased MMP. SAS upregulated TFRC and DMT1. Knockdown of the ER increased TFRC expression in breast cancer cells. Immunohistochemistry indicated that TFRC expression was lower in ER+ tissues than in ER­ tissues. After confirmation with RT­PCR in 87 clinical specimens, TFRC expression in ER­ tissue was revealed to be significantly higher than that of ER+ tissue. In conclusion SAS could trigger ferroptosis in breast cancer cells, especially in cells with low ER expression. Therefore, SAS is a potential agent for breast cancer treatment.