Downregulation of Long Noncoding RNA LINC00261 Attenuates Myocardial Infarction through the miR-522-3p/Trinucleotide Repeat-Containing Gene 6a (TNRC6A) Axis.

BACKGROUND: Myocardial infarction (MI) is cardiac tissue necrosis caused by acute and persistent ischemic hypoxia of the coronary arteries. This study is aimed at investigating the expression of long noncoding RNA (lncRNA) LINC00261 in MI and its effect on myocardial cells. METHODS: qRT-PCR was performed to detect the expression levels of LINC00261, miR-522-3p, and TNRC6A in normal and MI cells. Western blotting analysis was performed to detect the expression of TNRC6A protein. Viability and apoptosis of myocardial cells after MI with the knockout of LINC00261 or TNRC6A were detected. The relationships among miR-522-3p, LINC00261, and TNRC6A in cardiomyocytes were evaluated using a double luciferase reporter gene assay. Hypoxic preconditioning in normal cells was used to construct a simulated MI environment to investigate the effect of LINC00261 on apoptosis of cardiac cells. RESULTS: LINC00261 and TNRC6A were upregulated, while miR-522-3p was downregulated in coronary heart disease tissues with MI. Knockout of LINC00261 can increase the viability of cardiomyocytes and inhibit cell apoptosis. LINC00261 targets miR-522-3p in cardiomyocytes. In addition, miR-522-3p targets TNRC6A in cardiomyocytes. TNRC6A regulates cell viability and apoptosis of cardiomyocytes after MI, and TNRC6A-induced MI can be reversed by overexpression of miR-522-3p. CONCLUSIONS: LINC00261 downregulated miR-522-3p in cardiomyocytes after MI by directly targeting miR-522-3p. TNRC6A is the direct target of miR-522-3p. Our results indicated that LINC00261 might serve as a therapeutic target for the treatment of MI.

McH-lpr/lpr-RA1 mice: A novel spontaneous mouse model of autoimmune sialadenitis.

Many studies of the autoimmune disease Sjögren's syndrome have been performed using spontaneous mouse models. In the present study, we describe the characteristics of McH/lpr-RA1 mice and propose their use as a novel murine model of autoimmune sialadenitis. The McH/lpr-RA1 mouse is a recombinant congenic strain derived from generation F54 or more of MRL-Faslpr x (MRL- Faslpr x C3H- Faslpr) F1. We show for the first time that this mouse spontaneously develops autoimmune sialadenitis and vasculitis in submandibular gland tissues. Sialadenitis was accompanied by extensive inflammatory cell infiltration and tissue destruction. Immunohistochemical studies revealed that the salivary gland lesions strongly expressed four sialadenitis-related molecules: SSA and SSB (autoantigens of Sjögren's syndrome), gp91phox (an accelerator of reactive oxygen species production) and single strand DNA (a marker of apoptotic cells). In contrast, expression of aquaporin-5 (AQP5), which stimulates salivary secretion was weak or negligible. Statistical correlation analyses indicated that the apoptosis of salivary gland cells provoked by oxidative stress contributed to the severe sialadenitis and reduced expression of AQP5. Our study has demonstrated that McH/lpr-RA1 mice spontaneously develop the pathognomonic features of autoimmune sialadenitis and thus could be used as a new animal model of Sjögren's syndrome.

PP2A and E3 ubiquitin ligase deficiencies: Seminal biological drivers in endometrial cancer.

OBJECTIVE: PI3K-AKT pathway mutations initiate a kinase cascade that characterizes endometrial cancer (EC). As kinases seldom cause oncogenic transformation without dysregulation of antagonistic phosphatases, pivotal interactions governing this pathway were explored and correlated with clinical outcomes. METHODS: After exclusion of patients with POLE mutations from The Cancer Genome Atlas EC cohort with endometrioid or serous EC, the study population was 209 patients with DNA sequencing, quantitative gene-specific RNA expression, copy number variation (CNV), and surveillance data available. Extracted data were annotated and integrated. RESULTS: A PIK3CA, PTEN, or PIK3R1 mutant (-mu) was present in 83% of patients; 57% harbored more than 1 mutation without adversely impacting progression-free survival (PFS) (P = .10). PIK3CA CNV of at least 1.1 (CNV high [-H]) was detected in 26% and linked to TP53-mu and CIP2A expression (P < .001) but was not associated with PFS (P = .24). PIK3CA expression was significantly different between those with CIP2A-H and CIP2A low (-L) expression (the endogenous inhibitor of protein phosphatase 2A [PP2A]), when stratified by PIK3CA mutational status or by PIK3CA CNV-H and CNV-L (all P < .01). CIP2A-H or PPP2R1A-mu mitigates PP2A kinase dephosphorylation, and FBXW7-mu nullifies E3 ubiquitin ligase (E3UL) oncoprotein degradation. CIP2A-H and PPP2R1A-mu (PP2A impairment) and FBXW7-mu (E3UL impairment) were associated with compromised PFS (P < .001) and were prognostically discriminatory for PIK3CA-mu and PIK3CA CNV-H tumors (P < .001). Among documented recurrences, 84% were associated with impaired PP2A (75%) and/or E3UL (20%). CONCLUSION: PP2A and E3UL deficiencies are seminal biological drivers in EC independent of PIK3CA-mu, PTEN-mu, and PIK3R1-mu and PIK3CA CNV.

Chd4 choreographs self-antigen expression for central immune tolerance.

Autoreactive T cells are eliminated in the thymus to prevent autoimmunity by promiscuous expression of tissue-restricted self-antigens in medullary thymic epithelial cells. This expression is dependent on the transcription factor Fezf2, as well as the transcriptional regulator Aire, but the entire picture of the transcriptional program has been obscure. Here, we found that the chromatin remodeler Chd4, also called Mi-2ß, plays a key role in the self-antigen expression in medullary thymic epithelial cells. To maximize the diversity of self-antigen expression, Fezf2 and Aire utilized completely distinct transcriptional mechanisms, both of which were under the control of Chd4. Chd4 organized the promoter regions of Fezf2-dependent genes, while contributing to the Aire-mediated induction of self-antigens via super-enhancers. Mice deficient in Chd4 specifically in thymic epithelial cells exhibited autoimmune phenotypes, including T cell infiltration. Thus, Chd4 plays a critical role in integrating Fezf2- and Aire-mediated gene induction to establish central immune tolerance.

Involvement of collagen XVII in pluripotency gene expression and metabolic reprogramming of lung cancer stem cells.

BACKGROUND: Recent advancements in cancer biology field suggest that glucose metabolism is a potential target for cancer treatment. However, little if anything is known about the metabolic profile of cancer stem cells (CSCs) and the related underlying mechanisms. METHODS: The metabolic phenotype in lung CSC was first investigated. The role of collagen XVII, a putative stem cell or CSC candidate marker, in regulating metabolic reprogramming in lung CSC was subsequently studied. Through screening the genes involved in glycolysis, we identified the downstream targets of collagen XVII that were involved in metabolic reprogramming of lung CSCs. Collagen XVII and its downstream targets were then used to predict the prognosis of lung cancer patients. RESULTS: We showed that an aberrant upregulation of glycolysis and oxidative phosphorylation in lung CSCs is associated with the maintenance of CSC-like features, since blocking glycolysis and oxidative phosphorylation reduces sphere formation, chemoresistance, and tumorigenicity. We also showed that the Oct4-hexokinase 2 (HK2) pathway activated by collagen XVII-laminin-332 through FAK-PI3K/AKT-GSB3ß/ß-catenin activation induced the upregulation of glycolysis and maintenance of CSC-like features. Finally, we showed that collagen XVII, Oct4, and HK2 could be valuable markers to predict the prognosis of lung cancer patients. CONCULSIONS: These data suggest the Oct4-HK2 pathway regulated by collagen XVII plays an important role in metabolic reprogramming and maintenance of CSC-like features in lung CSCs, which may aid in the development of new strategies in cancer treatment.

Expression of anti-aging type-XVII collagen (COL17A1/BP180) in hair follicle-associated pluripotent (HAP) stem cells during differentiation.

Hair-follicle-associated pluripotent (HAP) stem cells reside in the upper part of the bulge area of the the hair follicle. HAP stem cells are nestin-positive and keratin 15-negative and have the capacity to differentiate into various types of cells in vitro. HAP stem cells are also involved in nerve and spinal cord regeneration in mouse models. Recently, it was shown that the DNA-damage response in non-HAP hair follicle stem cells induces proteolysis of type-XVII collagen (COL17A1/BP180), which is involved in hair-follicle stem-cell maintenance. COL17A1 proteolysis stimulated hair-follicle stem-cell aging, characterized by the loss of stemness signatures and hair-follicle miniaturization associated with androgenic alopecia. In the present study, we demonstrate that HAP stem cells co-express nestin and COL17A1 in vitro and in vivo. The expression of HAP stem cell markers (nestin and SSEA1) increased after HAP stem-cell colonies were formed, then decreased after differentiation to epidermal keratinocytes. In contrast COL17A1 increased after differentiation to epidermal keratinocytes. These results suggest that COL17A1 is important in differentiation of HAP stem cells.

The intercellular expression of type-XVII collagen, laminin-332, and integrin-ß1 promote contact following during the collective invasion of a cancer cell population.

Cancer cells can invade as a population in various cancer tissues. This phenomenon is called collective invasion, which is associated with the metastatic potential and prognosis of cancer patients. The collectiveness of cancer cells is necessary for collective invasion. However, the mechanism underlying the generation of collectiveness by cancer cells is not well known. In this study, the phenomenon of contact following, where neighboring cells move in the same direction via intercellular adhesion, was investigated. An experimental system was created to observe the two-dimensional invasion using a collagen gel overlay to study contact following in collective invasion. The role of integrin-ß1, one of the major extracellular matrix (ECM) receptors, in contact following was examined through the experimental system. Integrin-ß1 was localized to the intercellular site in squamous carcinoma cells. Moreover, the intercellular adhesion and contact following were suppressed by treatment of an integrin-ß1 inhibitory antibody. ECM proteins such as laminin-332 and type-XVII collagen were also localized to the intercellular site and critical for contact following. Collectively, it was demonstrated that the activity of integrin-ß1 and expression of ECM proteins in the intercellular site promote contact following in the collective invasion of a cancer cell population.

Technology for Discovery of Antifibrotic Drugs: Phenotypic Screening for LARP6 Inhibitors Using Inverted Yeast Three Hybrid System.

Fibrosis is defined by excessive production of type I collagen in various organs. Excessive type I collagen production in fibrosis is stimulated by binding of RNA protein LARP6 to the structural element of collagen mRNAs, the 5' stem loop (5'SL). The LARP6-dependent regulation is specific for type I collagen and critical for fibrosis development. Inhibitors of LARP6 binding have potential to be specific antifibrotic drugs, as evidenced by the discovery of one such inhibitor. To create technology for phenotypic screening of additional compounds we developed an inverted yeast three hybrid system. The system is based on expression of human LARP6 and a short RNA containing the 5'SL of human collagen α1(I) mRNA in Saccharomyces cerevisiae cells. The cells were engineered in such a way that when LARP6 is bound to 5'SL RNA they fail to grow in a specific synthetic medium. Dissociation of LARP6 from 5'SL RNA permits the cell growth, allowing identification of the inhibitors of LARP6 binding. The assay simply involves measuring optical density of cells growing in multiwall plates and is pertinent for high throughput applications. We describe the specificity of the system and its characteristics for high throughput screening. As a proof of principle, the result of one screen using collection of FDA approved drugs is also presented. This screen demonstrates that using this technology discovery of novel LARP6 inhibitors is possible.

Expression of recombinant glutamic acid decarboxylase in insect larvae and its application in an immunoassay for the diagnosis of autoimmune diabetes mellitus.

Autoimmune Diabetes Mellitus (DM) is a chronic disease caused by the selective destruction of insulin producing beta cells in human pancreas. DM is characterized by the presence of autoantibodies that bind a variety of islet-cell antigens. The 65 kDa isoform of glutamate decarboxylase (GAD65) is a major autoantigen recognized by these autoantibodies. Autoantibodies to GAD65 (GADA) are considered predictive markers of the disease when tested in combination with other specific autoantibodies. In order to produce reliable immunochemical tests for large scale screening of autoimmune DM, large amounts of properly folded GAD65 are needed. Herein, we report the production of human GAD65 using the baculovirus expression system in two species of larvae, Rachiplusia nu and Spodoptera frugiperda. GAD65 was identified at the expected molecular weight, properly expressed with high yield and purity in both larvae species and presenting appropriate enzymatic activity. The immunochemical ability of recombinant GAD65 obtained from both larvae to compete with [35S]GAD65 was assessed qualitatively by incubating GADA-positive patients' sera in the presence of 1 µM of the recombinant enzyme. All sera tested became virtually negative after incubation with antigen excess. Besides, radiometric quantitative competition assays with GADA-positive patients' sera were performed by adding recombinant GAD65 (0.62 nM-1.4 µM). All dose response curves showed immunochemical identity between proteins. In addition, a bridge-ELISA for the detection of GADA was developed using S. frugiperda-GAD65. This assay proved to have 77.3% sensitivity and 98.2% of specificity. GAD65 could be expressed in insect larvae, being S. frugiperda the best choice due to its high yield and purity. The development of a cost effective immunoassay for the detection of GADA was also afforded.

Optimized upstream and downstream process conditions for the improved production of recombinant human asparaginase (rhASP) from Escherichia coli and its characterization.

The present work elucidates the production of recombinant human asparaginase (rhASP) under optimized fermentation and downstream processes in Escherichia coli. The maximum biomass yield of 6.7 g/L was achieved with fed-batch fermentation. The highest rhASP inclusion bodies recovery yield (91%) was achieved with the optimized lysis conditions. The 8.0 M urea at pH 8.5 has shown efficient solubilization (94%) of rhASP inclusion bodies. The refolding efficiency of rhASP increased at pH 8.5 (84%) and temperature 25°C (86%). The diluted rhASP solution was concentrated and partially purified (92%) using cross flow filtration. A single step ion exchange chromatography is successfully achieved the maximum purity of ≥ 97%. The molecular mass of purified rhASP is confirmed as 34.1 kDa by mass spectrometry. The secondary structure of rhASP is characterized by FT-IR spectroscopy based on the structural elements. Finally, cell proliferative assay of purified rhASP is signifies the similar biological activity over the standard.

Perturbation of epithelial apicobasal polarity by rhomboid family-1 gene overexpression.

The human rhomboid family (RHBDF)1 gene is highly expressed in breast cancer under clinical conditions but not in normal mammary gland tissues. Silencing the RHBDF1 gene in breast cancer xenograft tumors leads to inhibition of tumor growth. We show in this study that artificially raising RHBDF1 protein levels in the mammary epithelial cells MCF-10A results in severe perturbations of the ability of the cells to form lumen-containing acini, either in 3-dimensional cell cultures or implanted in mouse mammary fat pads. Knocking down RHBDF1 with short hairpin (sh)RNA leads to restoration of acinus formation. Consistently, RHBDF1 overexpression gives rise to disordered distribution of polarity markers GM130 and laminin-5, which otherwise are located in apical and basal positions, respectively, in the acini. Further investigations reveal that RHBDF1 directly binds to Par6a, a component of a protein complex consisting of partitioning-defective scaffold protein (Par)6, Par3, renin-angiotensin system-related C3 botulinum toxin substrate (Rac)1, and cell-division cycle (Cdc)42, which is structurally critical to the formation of apicobasal polarity. RHBDF1 binding to Par6a results in collapse of the protein complex and thus disruption of polarity formation. Since early stages of breast cancer are characterized by the loss of mammary gland epithelial cell polarity, our findings indicate that perturbations of apicobasal polarity by high levels of RHBDF1 is a significant attribute in the development of breast neoplasia.-Peng, X.-M., Gao, S., Deng, H.-T., Cai, H.-X., Zhou, Z., Xiang, R., Zhang, Q.-Z., Li, L.-Y. Perturbation of epithelial apicobasal polarity by rhomboid family-1 gene overexpression.

RNAi­mediated downregulation of asparaginase­like protein 1 inhibits growth and promotes apoptosis of human cervical cancer line SiHa.

Asparaginase like 1 (ASRGL1) protein belongs to the N­terminal nucleophile group, cleaving the isoaspartyl­dipeptides and L­asparagine by adding water. It tends to be overexpressed in cancerous tumors including ovarian cancer and breast tumors. The present study assessed the potential ability of ASRGL1 as a molecular target in gene­based cervical cancer treatment. The protein expression level of ASRGL1 was determined in paraffin­embedded tumor specimen by immunohistochemistry. Additionally, in order to assess the activity of ASRGL1 during the process of cervical cancer cell multiplication, ASRGL1­short hairpin (sh) RNA­expressing lentivirus was established, which was used to infect SiHa cells. The Cellomics ArrayScan VT1 Reader identified the influence of downregulation on SiHa caused by RNA interference­intervened ASRGL1. Flow cytometric analysis was also performed to evaluate the influence. The cyclin dependent kinase (CDK2), cyclin A2, B­cell lymphoma 2 (Bcl­2) and Bcl­2­associated X protein (Bax) expression levels were assessed by western blot analysis. ASRGL1 was observed to be overexpressed in cervical cancer tissues when compared with the adjacent normal tissues. The knockdown of ASRGL1 in SiHa by ASRGL1­shRNA lentivirus infection significantly inhibited cell growth and enhanced cellular apoptosis; the cells were also captured during the S phase. The knockdown of ASRGL1 expression led to the increased expression of Bax and decreased expression of Bcl­2, CDK2 and cyclin A2. In conclusion, ASRGL1 was closely associated with growth and apoptosis in cervical cancer. Therefore, ASRGL1 may be a novel, potentially effective anti­cervical cancer therapy.

Secretory expression and scale-up production of recombinant human thyroid peroxidase via baculovirus/insect cell system in a wave-type bioreactor.

The human thyroid peroxidase (hTPO) is an essential enzyme for thyroid hormone biosynthesis and is expressed in thyroid cells. It is an autoantigen against which antibodies are found in the sera of patients with a number of autoimmune thyroid disorders. Overexpression of hTPO has been achieved using the baculovirus expression vector system (BEVS). However, it is produced largely in an aggregated form in the cell lysate fraction, which increases the complexity of protein extraction. In this study, to achieve improved secretory expression of hTPO via BEVS, a truncated recombinant hTPO protein (hTPOt) was engineered by replacing its original signal peptide (SP) in the N-terminal with five heterologous SPs. Our data showed that the SP from the peptidyl-glycine alpha-amidating monooxygenase (PAM), referred to as SPPAM, significantly promoted the secretion of SPPAM-fused hTPOt (p-hTPOt) in High Five cells. Subsequently, we established an optimized scale-up production procedure for p-hTPOt in a 5-L wave-type bioreactor. The secretory p-hTPOt was purified by immobilized metal-chelating affinity chromatography and ion-exchange chromatography, achieving a protein purity of >95%. Finally, the purified p-hTPOt showed high sensitivity and specificity in reactions with positive or negative human serum samples via the double-antigen sandwich method, suggesting potential applications in hTPO-based research and product development.

Combined ASRGL1 and p53 immunohistochemistry as an independent predictor of survival in endometrioid endometrial carcinoma.

OBJECTIVE: In clinical practise, prognostication of endometrial cancer is based on clinicopathological risk factors. The use of immunohistochemistry-based markers as prognostic tools is generally not recommended and a systematic analysis of their utility as a panel is lacking. We evaluated whether an immunohistochemical marker panel could reliably assess endometrioid endometrial cancer (EEC) outcome independent of clinicopathological information. METHODS: A cohort of 306 EEC specimens was profiled using tissue microarray (TMA). Cost- and time-efficient immunohistochemical analysis of well-established tissue biomarkers (ER, PR, HER2, Ki-67, MLH1 and p53) and two new biomarkers (L1CAM and ASRGL1) was carried out. Statistical modelling with embedded variable selection was applied on the staining results to identify minimal prognostic panels with maximal prognostic accuracy without compromising generalizability. RESULTS: A panel including p53 and ASRGL1 immunohistochemistry was identified as the most accurate predictor of relapse-free and disease-specific survival. Within this panel, patients were allocated into high- (5.9%), intermediate- (29.5%) and low- (64.6%) risk groups where high-risk patients had a 30-fold risk (P<0.001) of dying of EEC compared to the low-risk group. CONCLUSIONS: P53 and ASRGL1 immunoprofiling stratifies EEC patients into three risk groups with significantly different outcomes. This simple and easily applicable panel could provide a useful tool in EEC risk stratification and guiding the allocation of treatment modalities.

Age-Associated Decline in Thymic B Cell Expression of Aire and Aire-Dependent Self-Antigens.

Although autoimmune disorders are a significant source of morbidity and mortality in older individuals, the mechanisms governing age-associated increases in susceptibility remain incompletely understood. Central T cell tolerance is mediated through presentation of self-antigens by cells constituting the thymic microenvironment, including epithelial cells, dendritic cells, and B cells. Medullary thymic epithelial cells (mTECs) and B cells express distinct cohorts of self-antigens, including tissue-restricted self-antigens (TRAs), such that developing T cells are tolerized to antigens from peripheral tissues. We find that expression of the TRA transcriptional regulator Aire, as well as Aire-dependent genes, declines with age in thymic B cells in mice and humans and that cell-intrinsic and cell-extrinsic mechanisms contribute to the diminished capacity of peripheral B cells to express Aire within the thymus. Our findings indicate that aging may diminish the ability of thymic B cells to tolerize T cells, revealing a potential mechanistic link between aging and autoimmunity.

BPIFB1 (LPLUNC1) inhibits migration and invasion of nasopharyngeal carcinoma by interacting with VTN and VIM.

BACKGROUND: Bactericidal/Permeability-increasing-fold-containing family B member 1 (BPIFB1, previously termed LPLUNC1) is highly expressed in the nasopharynx, significantly downregulated in nasopharyngeal carcinoma (NPC), and associated with prognosis in NPC patients. Because metastasis represents the primary cause of NPC-related death, we explored the role of BPIFB1 in NPC migration and invasion. METHODS: The role of BPIFB1 in NPC metastasis was investigated in vitro and in vivo. A co-immunoprecipitation assay coupled with mass spectrometry was used to identify BPIFB1-binding proteins. Additionally, western blotting, immunofluorescence, and immunohistochemistry allowed assessment of the molecular mechanisms associated with BPIFB1-specific metastatic inhibition via vitronectin (VTN) and vimentin (VIM) interactions. RESULTS: Our results showed that BPIFB1 expression markedly inhibited NPC cell migration, invasion, and lung-metastatic abilities. Additionally, identification of two BPIFB1-interacting proteins, VTN and VIM, showed that BPIFB1 reduced VTN expression and the formation of a VTN-integrin αV complex in NPC cells, leading to inhibition of the FAK/Src/ERK signalling pathway. Moreover, BPIFB1 attenuated NPC cell migration and invasion by inhibiting VTN- or VIM-induced epithelial-mesenchymal transition. CONCLUSIONS: This study represents the first demonstration of BPIFB1 function in NPC migration, invasion, and lung metastasis. Our findings indicate that re-expression of BPIFB1 might represent a useful strategy for preventing and treating NPC.

Asparaginase-like protein 1 is an independent prognostic marker in primary endometrial cancer, and is frequently lost in metastatic lesions.

OBJECTIVE: Loss of Asparaginase-like protein 1 (ASRGL1) has been suggested as a prognostic biomarker in endometrial carcinoma. Our objective was to validate this in a prospectively collected, independent patient cohort, and evaluate ASRGL1 expression in endometrial carcinoma precursor lesion and metastases. METHODS: 782 primary endometrial carcinomas, 90 precursor lesions (complex atypical hyperplasia), and 179 metastases (from 87 patients) were evaluated for ASRGL1 expression by immunohistochemistry in relation to clinical and histopathological data. ASRGL1 mRNA level was investigated in 237 primary tumors and related to survival and ASRGL1 protein expression. RESULTS: Low expression of ASRGL1 protein and ASRGL1 mRNA predicted poor disease specific survival (P<0.001). In multivariate survival analyses ASRGL1 had independent prognostic value both in the whole patient cohort (Hazard ratio (HR): 1.53, 95% confidence interval (CI): 1.04-2.26, P=0.031) and within the endometrioid subgroup (HR: 2.64, CI: 1.47-4.74, P=0.001). Low ASRGL1 expression was less frequent in patients with low grade endometrioid primary tumors compared to high grade endometrioid and non-endometrioid primary tumors, and ASRGL1 was lost in the majority of metastatic lesions. CONCLUSIONS: In a prospective setting ASRGL1 validates as a strong prognostic biomarker in endometrial carcinoma. Loss of ASRGL1 is associated with aggressive disease and poor survival, and is demonstrated for the first time to have independent prognostic value in the entire endometrial carcinoma patient population.

Increase in CIP2A expression is associated with cisplatin chemoresistance in gastric cancer.

BACKGROUND: The cancerous inhibitor of protein phosphatase 2A (CIP2A) is an oncoprotein which involves in the progression of several human malignancies. Development of cisplatin (DDP) resistance is the obstacle to an effective control of gastric cancer (GC) clinically. OBJECTIVE: We thus assessed whether CIP2A expression is associated with sensitivity of GC to DDP. METHODS: Real-time quantitative PCR, immunohistochemical analysis, or western blotting was performed to detect CIP2A expression in GC patients' tissues. SGC7901/DDP cells were transfected with CIP2A siRNA. MTT assay was used to determine the DDP-sensitivity of cells. Flow cytometry was used to measure cell apoptosis. RESULTS: CIP2A has higher expression in DDP-resistant GC patients. DDP-resistant GC patients with high CIP2A expression presented with poorer overall survival rates than those with low CIP2A expression. CIP2A knockdown in DDP-resistant GC cells resulted in attenuated proliferative abilities and increased apoptosis level. CIP2A depletion sensitizes DDP-resistant cells to DDP and CIP2A overexpression antagonizes DDP-sensitive cells to DDP. CIP2A influences the expression of multidrug resistance-related proteins in GC cells. CONCLUSIONS: Our results suggested that CIP2A oncoprotein plays an important role in DDP resistance of GC and could serve as a novel therapeutic target for the treatment of GC patients with DDP resistance.

Mode of Tolerance Induction and Requirement for Aire Are Governed by the Cell Types That Express Self-Antigen and Those That Present Antigen.

Aire controls the fate of autoreactive thymocytes (i.e., clonal deletion or development into regulatory T cells [Tregs]) through transcriptional control of the expression of tissue-restricted self-antigens (TRAs) from medullary thymic epithelial cells (mTECs) and bone marrow (BM)-derived cells. Although TRAs expressed by mTECs and BM-derived cells are suggested to complement each other to generate a full spectrum of TRAs, little is known about the relative contribution of TRAs from each component for establishment of self-tolerance. Furthermore, the precise role of Aire in specific types of Aire-expressing APCs remains elusive. We have approached these issues by generating two different types of transgenic mouse (Tg) model, which express a prefixed model self-antigen driven by the insulin promoter or the Aire promoter. In the insulin-promoter Tg model, mTECs alone were insufficient for clonal deletion, and BM-derived APCs were required for this action by utilizing Ag transferred from mTECs. In contrast, mTECs alone were able to induce Tregs, although at a much lower efficiency in the absence of BM-derived APCs. Importantly, lack of Aire in mTECs, but not in BM-derived APCs, impaired both clonal deletion and production of Tregs. In the Aire-promoter Tg model, both mTECs and BM-derived APCs could independently induce clonal deletion without Aire, and production of Tregs was impaired by the lack of Aire in mTECs, but not in BM-derived APCs. These results suggest that the fate of autoreactive thymocytes together with the requirement for Aire depend on the cell types that express self-antigens and the types of APCs involved in tolerance induction.