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Background: The aim of the present study was to investigate the function of novel circular RNA hsa_circ_0036683 (circ-36683) in non-small cell lung cancer (NSCLC)., Methods: RNA sequencing was used to screen out differentially expressed miRNAs. Expression levels of miR-4664-3p and circ-36683 were evaluated in lung carcinoma cells and tissues by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). The effects of miR-4664-3p and circ-36683 on proliferation and migration were assessed using cell counting kit-8 (CCK-8), wound healing and transwell migration assays and xenograft experiments. The targeting relationship of circ-36683/miR-4664-3p/CDK2AP2 was assessed by luciferase reporter assays, western blot, qRT-PCR and argonaute2-RNA immunoprecipitation (AGO2 RIP). Co-immunoprecipitation (Co-IP), 5-ethynyl-2'-deoxyuridine (EdU) staining and CCK-8 were used to validate the indispensable role of CDK2AP2 in suppressing cell proliferation as a result of CDK2AP1 overexpression., Results: By RNA sequencing, miR-4664-3p was screened out as an abnormally elevated miRNA in NSCLC tissues. Transfection of miR-4664-3p could promote cell proliferation, migration and xenograft tumor growth. As a target of miR-4664-3p, CDK2AP2 expression was downregulated by miR-4664-3p transfection and CDK2AP2 overexpression could abolish the proliferation promotion resulting from miR-4664-3p elevation. Circ-36683, derived from back splicing of ABHD2 pre-mRNA, was attenuated in NSCLC tissue and identified as a sponge of miR-4664-3p. The functional study revealed that circ-36683 overexpression suppressed cell proliferation, migration and resulted in G0/G1 phase arrest. More importantly, the antioncogenic function of circ-36683 was largely dependent on the miR-4664-3p/CDK2AP2 axis, through which circ-36683 could upregulate the expression of p53/p21/p27 and downregulate the expression of CDK2/cyclin E1., Conclusion: The present study revealed the antioncogenic role of circ-36683 in suppressing cell proliferation and migration and highlighted that targeting the circ-36683/miR-4664-3p/CDK2AP2 axis is a promising strategy for the intervention of NSCLC., (© 2024 The Author(s). Thoracic Cancer published by John Wiley & Sons Australia, Ltd.)

Regenerating family protein 2 (Reg2) is a trophic factor which stimulates β-cell replication and resists islet destruction. However, Reg2 also serves as an islet autoantigen, which makes it complicated to judge the effectiveness in treating diabetes. How Reg2 treatment behaves in non-obese diabetic (NOD) mice is to be investigated. NOD mice were treated with recombinant Reg2 protein, Complete Freund's adjuvant (CFA) + PBS and CFA+Reg2 vaccinations, CFA+PBS- and CFA+Reg2-immunized antisera, and single chain variable fragment (scFv)-Reg2 and mIgG2a-Reg2 antibodies. Glycemic level, bodyweight, serum Reg2 antibody titer, glucose tolerance, and insulin secretion were determined. Islet morphological characteristics, insulitis, cell apoptosis, islet cell components, and T cell infiltration were analyzed by histological examinations. The autoantigenicity of constructed Reg2C and Reg2X fragments was determined in healthy BALB/c mice, and the bioactivity in stimulating cell proliferation and survival was assessed in insulinoma MIN6 cells. Reg2 administration alleviated diabetes in NOD mice with improved glucose tolerance and insulin secretion but elevated serum Reg2 autoantibodies. Histomorphometry showed reduced inflammatory area, TUNEL signal and CD8 + T cell infiltration, and increased β-cell proportion in support of the islet-protective effect of Reg2 treatment. CFA+PBS and CFA+Reg2 immunizations prevented diabetic onset and alleviated insulitis while injections of the antisera offered mild protections. Antibody treatments accelerated diabetic onset without increasing the overall incidence. Reg2C fragment depletes antigenicity, but reserves protective activity in streptozotocin (STZ)-treated MIN6 cells. In conclusion, Reg2 treatment alleviates type 1 diabetes (T1D) by preserving islet β-cells, but induces Reg2 autoantibody production which poses a potential risk of accelerating diabetic progression., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

Cancer immunotherapy has emerged as a novel therapeutic approach against tumors, with immune checkpoint inhibitors (ICIs) making significant clinical practice. The traditional ICIs, PD-1 and PD-L1, augment the cytotoxic function of T cells through the inhibition of tumor immune evasion pathways, ultimately leading to the initiation of an antitumor immune response. However, the clinical implementation of ICIs encounters obstacles stemming from the existence of an immunosuppressive tumor microenvironment and inadequate infiltration of CD8 + T cells. Considerable attention has been directed towards advancing immunogenic cell death (ICD) as a potential solution to counteract tumor cell infiltration and the immunosuppressive tumor microenvironment. This approach holds promise in transforming "cold" tumors into "hot" tumors that exhibit responsiveness to antitumor. By combining ICD with ICIs, a synergistic immune response against tumors can be achieved. However, the combination of ICD inducers and PD-1/PD-L1 inhibitors is hindered by issues such as poor targeting and uncontrolled drug release. An advantageous solution presented by stimulus-responsive nanocarrier is integrating the physicochemical properties of ICD inducers and PD-1/PD-L1 inhibitors, facilitating precise delivery to specific tissues for optimal combination therapy. Moreover, these nanocarriers leverage the distinct features of the tumor microenvironment to accomplish controlled drug release and regulate the kinetics of drug delivery. This article aims to investigate the advancement of stimulus-responsive co-delivery nanocarriers utilizing ICD and PD-1/PD-L1 inhibitors. Special focus is dedicated to exploring the advantages and recent advancements of this system in enabling the combination of ICIs and ICD inducers. The molecular mechanisms of ICD and ICIs are concisely summarized. In conclusion, we examine the potential research prospects and challenges that could greatly enhance immunotherapeutic approaches for cancer treatment., Competing Interests: The authors report no conflicts of interest in this work., (© 2024 Zhang et al.)

Purpose: The drug resistance and low response rates of immunotherapy limit its application. This study aimed to construct a new nanoparticle (CaCO 3 -polydopamine-polyethylenimine, CPP) to effectively deliver interleukin-12 (IL-12) and suppress cancer progress through immunotherapy., Methods: The size distribution of CPP and its zeta potential were measured using a Malvern Zetasizer Nano-ZS90. The morphology and electrophoresis tentative delay of CPP were analyzed using a JEM-1400 transmission electron microscope and an ultraviolet spectrophotometer, respectively. Cell proliferation was analyzed by MTT assay. Proteins were analyzed by Western blot. IL-12 and HMGB1 levels were estimated by ELISA kits. Live/dead staining assay was performed using a Calcein-AM/PI kit. ATP production was detected using an ATP assay kit. The xenografts in vivo were estimated in C57BL/6 mice. The levels of CD80 + /CD86 + , CD3 + /CD4 + and CD3 + /CD8 + were analyzed by flow cytometry., Results: CPP could effectively express EGFP or IL-12 and increase ROS levels. Laser treatment promoted CPP-IL-12 induced the number of dead or apoptotic cell. CPP-IL-12 and laser could further enhance CALR levels and extracellular HMGB1 levels and decrease intracellular HMGB1 and ATP levels, indicating that it may induce immunogenic cell death (ICD). The tumors and weights of xenografts in CPP-IL-12 or laser-treated mice were significantly reduced than in controls. The IL-12 expression, the CD80 + /CD86 + expression of DC from lymph glands, and the number of CD3 + /CD8 + T or CD3 + /CD4 + T cells from the spleen increased in CPP-IL-12-treated or laser-treated xenografts compared with controls. The levels of granzyme B, IFN-γ, and TNF-α in the serum of CPP-IL-12-treated mice increased. Interestingly, CPP-IL-12 treatment in local xenografts in the back of mice could effectively inhibit the growth of the distant untreated tumor., Conclusion: The novel CPP-IL-12 could overexpress IL-12 in melanoma cells and achieve immunotherapy to melanoma through inducing ICD, activating CD4 + T cell, and enhancing the function of tumor-reactive CD8 + T cells., Competing Interests: The authors declare no conflicts of interest in this work., (© 2024 Shen et al.)

Blocking immune checkpoint CD47/SIRPα is a useful strategy to engineer macrophages for cancer immunotherapy. However, the roles of CD47-related noncoding RNA in regulating macrophage phagocytosis for lung cancer therapy remain unclear. This study aims to investigate the effects of long noncoding RNA (lncRNA) on the phagocytosis of macrophage via CD47 and the proliferation of non-small cell lung cancer (NSCLC) via TIPRL. Our results demonstrate that lncRNA KCTD21-AS1 increases in NSCLC tissues and is associated with poor survival of patients. KCTD21-AS1 and its m6A modification by Mettl14 promote NSCLC cell proliferation. miR-519d-5p gain suppresses the proliferation and metastasis of NSCLC cells by regulating CD47 and TIPRL. Through ceRNA with miR-519d-5p, KCTD21-AS1 regulates the expression of CD47 and TIPRL, which further regulates macrophage phagocytosis and cancer cell autophagy. Low miR-519d-5p in patients with NSCLC corresponds with poor survival. High TIPRL or CD47 levels in patients with NSCLC corresponds with poor survival. In conclusion, we demonstrate that KCTD21-AS1 and its m6A modification promote NSCLC cell proliferation, whereas miR-519d-5p inhibits this process by regulating CD47 and TIPRL expression, which further affects macrophage phagocytosis and cell autophagy. This study provides a strategy through miR-519-5p gain or KCTD21-AS1 depletion for NSCLC therapy by regulating CD47 and TIPRL., (© 2024. The Author(s).)

Introduction: As the special modality of cell death, immunogenic cell death (ICD) could activate immune response. Phototherapy in combination with chemotherapy (CT) is a particularly efficient tumor ICD inducing method that could overcome the defects of monotherapies., Methods: In this study, new dual stimuli-responsive micelles were designed and prepared for imaging-guided mitochondrion-targeted photothermal/photodynamic/CT combination therapy through inducing ICD. A dual-sensitive methoxy-polyethylene glycol-SS-poly(L-γ-glutamylglutamine)-SS-IR780 (mPEG-SS-PGG-SS-IR780) polymer was synthesized by grafting IR780 with biodegradable di-carboxyl PGG as the backbone, and mPEG-SS-PGG-SS-IR780/paclitaxel micelles (mPEG-SS-PGG-SS-IR780/PTXL MCs) were synthesized by encapsulating PTXL in the hydrophobic core., Results: In-vivo and -vitro results demonstrated that the three-mode combination micelles inhibited tumor growth and enhanced the therapeutic efficacy of immunotherapy. The dual stimuli-responsive mPEG-SS-PGG-SS-IR780/PTXL MCs were able to facilitate tumor cell endocytosis of nanoparticles. They were also capable of promoting micelles disintegration and accelerating PTXL release. The mPEG-SS-PGG-SS-IR780/PTXL MCs induced mitochondrial dysfunction by directly targeting the mitochondria, considering the thermo- and reactive oxygen species (ROS) sensitivity of the mitochondria. Furthermore, the mPEG-SS-PGG-SS-IR780/PTXL MCs could play the diagnostic and therapeutic roles via imaging capabilities., Conclusion: In summary, this study formulated a high-efficiency nanoscale platform with great potential in combined therapy for tumors through ICD., Competing Interests: The authors report no conflicts of interest in this work., (© 2023 Liang et al.)

Background: The present study aimed to investigate the function of miR-3529-3p in lung adenocarcinoma and MnO 2 -SiO 2 -APTES (MSA) as a promising multifunctional delivery agent for lung adenocarcinoma therapy., Methods: Expression levels of miR-3529-3p were evaluated in lung carcinoma cells and tissues by qRT-PCR. The effects of miR-3529-3p on apoptosis, proliferation, metastasis and neovascularization were assessed by CCK-8, FACS, transwell and wound healing assays, tube formation and xenografts experiments. Luciferase reporter assays, western blot, qRT-PCR and mitochondrial complex assay were used to determine the targeting relationship between miR-3529-3p and hypoxia-inducible gene domain family member 1A (HIGD1A). MSA was fabricated using MnO 2 nanoflowers, and its heating curves, temperature curves, IC50, and delivery efficiency were examined. The hypoxia and reactive oxygen species (ROS) production was investigated by nitro reductase probing, DCFH-DA staining and FACS., Results: MiR-3529-3p expression was reduced in lung carcinoma tissues and cells. Transfection of miR-3529-3p could promote apoptosis and suppress cell proliferation, migration and angiogenesis. As a target of miR-3529-3p, HIGD1A expression was downregulated, through which miR-3529-3p could disrupt the activities of complexes III and IV of the respiratory chain. The multifunctional nanoparticle MSA could not only efficiently deliver miR-3529-3p into cells, but also enhance the antitumor function of miR-3529-3p. The underlying mechanism may be that MSA alleviates hypoxia and has synergistic effects in cellular ROS promotion with miR-3529-3p., Conclusions: Our results establish the antioncogenic role of miR-3529-3p, and demonstrate that miR-3529-3p delivered by MSA has enhanced tumor suppressive effects, probably through elevating ROS production and thermogenesis., (© 2023 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.)

Reduced drug uptake and elevated drug efflux are two major mechanisms in cancer multidrug resistance (MDR). In the present study, a new multistage O 2 -producing liposome with NAG/R8-dual-ligand and stimuli-responsive dePEGylation was developed to address the abovementioned issues simultaneously. The designed C-NAG-R8-PTXL/MnO 2 -lip could also achieve magnetic resonance imaging (MRI)-guided synergistic chemodynamic/chemotherapy (CDT/CT). In vitro and in vivo studies showed that C-NAG-R8-PTXL/MnO 2 -lip enhanced circulation time by PEG and targeted the tumor site. After tumor accumulation, endogenous l-cysteine was administered, and the PEG-attached disulfide bond was broken, resulting in the dissociation of PEG shells. The previously hidden positively charged R8 by different lengths of PEG chains was exposed and mediated efficient internalization. In addition, the oxygen (O 2 ) generated by C-NAG-R8-PTXL/MnO 2 -lip relieved the hypoxic environment within the tumor, thus reducing the efflux of chemotherapeutic drug. O 2 was able to burst liposomes and triggered the release of PTXL. The toxic hydroxyl radical (·OH), which was produced by H 2 O 2 and Mn 2+ , strengthened CDT/CT. C-NAG-R8-PTXL/MnO 2 -lip was also used as MRI contrast agent, which blazed the trail to rationally design theranostic agents for tumor imaging., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier B.V.)

Long non-coding RNAs (lncRNAs) modulate cell proliferation, cycle, and apoptosis. However, the role of lncRNA-WFDC21P in the tumorigenesis of triple-negative breast cancer (TNBC) remains unclear. Results of this study demonstrated that WFDC21P levels significantly increased in TNBC, which was associated with the poor survival of patients. WFDC21P overexpression significantly promoted TNBC cell proliferation and metastasis. WFDC21P interacted with miR-628-5p, which further suppressed cell proliferation and metastasis by negatively regulating Smad3-related gene expression. Recovery of miR-628-5p weakened the roles of WFDC21P in promoting the growth and metastasis of TNBC cells. Moreover,N6-methyladenosine (m6A) modification upregulated WFDC21P expression in the TNBC cells. WFDC21P and its m6A levels were increased after methyltransferase like 3 (METTL3) overexpression but reduced after METTL3 silencing. The proliferation and metastasis of TNBC cells were promoted by METTL3 overexpression but suppressed by METTL3 silencing. This study demonstrated the vital roles of WFDC21P and its m6A in regulating the proliferation and metastasis of TNBC cells via the WFDC21P/miR-628/SMAD3 axis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Wei, Liang, Zhang, Li, Sun, Wang, Liang, Li, Wang, Yang, Wang and Xie.)

PKM2 is an important regulator of the aerobic glycolysis that plays a vital role in cancer cell metabolic reprogramming. In general, Trib2 is considered as a "pseudokinase", contributing to different kinds of cancer. However, the detailed roles of TRIB2 in regulating cancer metabolism by PKM2 remain unclear. This study demonstrated that TRIB2, not a "pseudokinase", has the kinase activity to directly phosphorylate PKM2 at serine 37 in cancer cells. The elevated pSer37-PKM2 would subsequently promote the PKM2 dimers to enter into nucleus and increase the expression of LDHA, GLUT1, and PTBP1. The aerobic glycolysis is then elevated to promote cancer cell proliferation and migration in TRIB2- or PKM2-overexpressed cultures. The glucose uptake and lactate production increased, but the ATP content decreased in TRIB2- or PKM2-treated cultures. Experiments of TRIB2 -/- mice further supported that TRIB2 could regulate aerobic glycolysis by PKM2. Thus, these results reveal the new kinase activity of TRIB2 and its mechanism in cancer metabolism may be related to regulating PKM2 to promote lung cancer cell proliferation in vitro and in vivo, suggesting promising therapeutic targets for cancer therapy by controlling cancer metabolism., (© 2022. The Author(s).)

Although the development of safe and efficient cancer therapeutic agents is essential, this process remains challenging. In this study, a mitochondria-targeted degradable nanoplatform (PDA-MnO₂-IR780) for synergistic photothermal, photodynamic, and sonodynamic tumor treatment was investigated. PDA-MnO₂-IR780 exhibits superior photothermal properties owing to the integration of polydopamine, MnO₂, and IR780. IR780, a photosensitizer and sonosensitizer, was used for photodynamic therapy and sonodynamic therapy. When PDA-MnO₂-IR780 was delivered to the tumor site, MnO₂ was decomposed by hydrogen peroxide, producing Mn 2+ and oxygen. Meanwhile, alleviating tumor hypoxia promoted the production of reactive oxygen species during photodynamic therapy and sonodynamic therapy. Moreover, large amounts of reactive oxygen species could reduce the expression of heat shock proteins and increase the heat sensitivity of tumor cells, thereby improving the photothermal treatment effect. In turn, hyperthermia caused by photothermal therapy accelerated the production of reactive oxygen species in photodynamic therapy. IR780 selectively accumulation in mitochondria also promoted tumor apoptosis. In this system, the mutual promotion of photothermal therapy and photodynamic therapy/sonodynamic therapy had an enhanced therapeutic effect. Moreover, the responsive degradable characteristic of PDA-MnO₂-IR780 in the tumor microenvironment ensured excellent biological safety. These results reveal a great potential of PDA-MnO₂-IR780 for safe and highly-efficiency synergistic therapy for cancer.

The construction of high-efficiency tumor theranostic platform will be of great interest in the treatment of cancer patients; however, significant challenges are associated with developing such a platform. In this study, we developed high-efficiency nanotheranostic agent based on ferroferric oxide, manganese dioxide, hyaluronic acid and doxorubicin (FMDH-D NPs) for dual targeting and imaging guided synergetic photothermal-enhanced chemodynamic/chemotherapy for cancer, which improved the specific uptake of drugs at tumor site by the dual action of CD44 ligand hyaluronic acid and magnetic nanoparticles guided by magnetic force. Under the acidic microenvironment of cancer cells, FMDH-D could be decomposed into Mn 2+ and Fe 2+ to generate •OH radicals by triggering a Fenton-like reaction and responsively releasing doxorubicin to kill cancer cells. Meanwhile, alleviating tumor hypoxia improved the efficacy of chemotherapy in tumors. The photothermal properties of FMDH generated high temperatures, which further accelerated the generation of reactive oxygen species, and enhanced effects of chemodynamic therapy. Furthermore, FMDH-D NPs proved to be excellent T 1 /T₂-weighted magnetic resonance imaging contrast agents for monitoring the tumor location. These results confirmed the considerable potential of FMDH-D NPs in a highly efficient synergistic therapy platform for cancer treatment.

RFWD2, an E3 ubiquitin ligase, is overexpressed in numerous human cancers, including leukemia, lung cancer, breast cancer, renal cell carcinoma, and colorectal cancer. The roles of RFWD2 in cancer are related to the targeting of its substrates for ubiquitination and degradation. This study aimed to investigate the role of TRIB2 in relation to the regulation of protein degradation through RFWD2. inBio Discover™ results demonstrated that TRIB2 can perform its functions by interacting with RFWD2 or other factors. TRIB2 can interact with and regulate RFWD2, which further attends the proteasome-mediated degradation of the RFWD2 substrate p-IκB-α. TRIB2 colocalizes with RFWD2-related IκB-α to form a ternary complex and further affects the IκB-α degradation by regulating its phosphorylation. Specific domain analysis showed that TRIB2 may bind to RFWD2 via its C-terminus, whereas it binds to IκB via its pseudokinase domain. TRIB2 acts as an oncogene and promotes cancer cell proliferation and migration, whereas RFWD2 knockdown reversed the role of TRIB2 in promoting cancer cell growth and colony formation in vitro and in vivo . In summary, this study reveals that TRIB2 promotes the progression of cancer by affecting the proteasome-mediated degradation of proteins through the interaction with RFWD2., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Hao, Hu, Liu, Liang, Li, Wang, Zhang, Wang, Li and Xie.)

The emergence of multidrug resistance (MDR) in malignant tumors is the primary reason for invalid chemotherapy. Antitumor drugs are often adversely affected by the MDR of tumor cells. Treatments using conventional drugs, which have specific drug targets, hardly regulate the complex signaling pathway of MDR cells because of the complex formation mechanism of MDR. However, natural products have positive advantages, such as high efficiency, low toxicity, and ability to target multiple mechanism pathways associated with MDR. Natural products, as MDR reversal agents, synergize with chemotherapeutics and enhance the sensitivity of tumor cells to chemotherapeutics, and the co-delivery of natural products and antitumor drugs with nanocarriers maximizes the synergistic effects against MDR in tumor cells. This review summarizes the molecular mechanisms of MDR, the advantages of natural products combined with chemotherapeutics in offsetting complicated MDR mechanisms, and the types and mechanisms of natural products that are potential MDR reversal modulators. Meanwhile, aiming at the low bioavailability of cocktail combined natural products and chemotherapeutic in vivo, the advantages of nanoplatform-based co-delivery system and recent research developments are illustrated on the basis of our previous research. Finally, prospective horizons are analyzed, which are expected to considerably improve the nano-co-delivery of natural products and chemotherapeutic systems for MDR reversal in cancer., (Copyright © 2021. Published by Elsevier B.V.)

Non-coding RNAs (ncRNAs) involve in diverse biological processes by post-transcriptional regulation of gene expression. Emerging evidence shows that miRNA-4293 plays a significant role in the development of non-small cell lung cancer. However, the oncogenic functions of miR-4293 have not been studied. Our results demonstrated that miR-4293 expression is markedly enhanced in lung carcinoma tissue and cells. Moreover, miR-4293 promotes tumor cell proliferation and metastasis but suppresses apoptosis. Mechanistic investigations identified mRNA-decapping enzyme 2 (DCP2) as a target of miR-4293 and its expression is suppressed by miR-4293. DCP2 can directly or indirectly bind to WFDC21P and downregulates its expression. Consequently, miR-4293 can further promote WFDC21P expression by regulating DCP2. With a positive correlation to miR-4293 expression, WFDC21P also plays an oncogenic role in lung carcinoma. Furthermore, knockdown of WFDC21P results in functional attenuation of miR-4293 on tumor promotion. In vivo xenograft growth is also promoted by both miR-4293 and WFDC21P. Overall, our results establish oncogenic roles for both miR-4293 and WFDC21P and demonstrate that interactions between miRNAs and lncRNAs through DCP2 are important in the regulation of carcinoma pathogenesis. These results provided a valuable theoretical basis for the discovery of lung carcinoma therapeutic targets and diagnostic markers based on miR-4293 and WFDC21P., (© 2021. The Author(s).)

Peritrophic membrane (PM) refers to a vital physical barrier enabling shrimp to resist pathogen invasion. It primarily consists of chitin and proteins, mostly chitin-binding protein (CBP). CBPs have been identified from microorganisms to higher organisms. In the present study, a CBP, designated MjCBP, was reported from Marsupenaeus japonicus. The open reading frame of MjCBP was 1854 bp, encoding a protein with 618 amino acids (MH544098). To be specific, the theoretical pI and molecular mass of mature MjCBP reached 5.43 and 66064.00 Da, respectively. MjCBP consisted of seven type Ⅱ chitin-binding domains (ChtB D2), which was up-regulated after being challenged with Vibrio anguillarum and then agglutinating several bacteria. In addition, MjCBP and the first chitin-binding domain (CBD1) could bind to several Gram-positive and Gram-negative bacteria via the binding process to lipopolysaccharides and peptidoglycans, whereas CBD1 was not capable of agglutinating bacteria. Moreover, the anterior and posterior segments of CBD1 were synthesized in vitro, and the posterior segment could bind to lipopolysaccharides. However, both segments fail to agglutinate bacteria. Furthermore, MjCBP and CBD1 facilitated the clearance of V. anguillarum in vivo, and the silencing of MjCBP via RNA interference reduced the ability of bacterial clearance. As revealed from the mentioned results, MjCBP acts as an opsonin or pattern recognition receptor to achieve antibacterial immune response in shrimp., (Copyright © 2020. Published by Elsevier Ltd.)

Background: Long noncoding (lncRNA) single-nucleotide polymorphisms (SNPs) are associated with the susceptibility to the development of various malignant tumors. The aim of this study was to investigate the roles of HOX transcript antisense intergenic RNA (HOTAIR) and its SNPs in lung cancer., Methods: Initially, the expression of HOTAIR in different tumors was investigated using the online Gene Expression Profiling Interactive Analysis (GEPIA) resource. Three SNPs (rs920778, rs1899663, and rs4759314) of HOTAIR were identified using the MassArray system. Following this, the relationship between these SNPs and susceptibility to lung cancer was investigated., Results: Expression of HOTAIR was found to increase in a variety of cancers, including nonsmall cell lung cancer (NSCLC). We found that the genotypes of these SNPs (rs920778, rs1899663, and rs4759314) were not significantly associated with lung cancer type, family history, lymph node metastasis, or lung cancer stage. In gender stratification, the results of rs920778 genotypes showed that, compared to genotype AA, the AG (OR = 0.344, 95% CI: 0.133-0.893, p = .028) and AG + GG (OR = 0.378, 95% CI: 0.153-0.932, p = .035) genotypes of rs920778 are protective factors against NSCLC in females. In smoking stratification, compared with AA of rs920778, the genotype AG + GG (OR = 0.507, 95% CI: 0.263-0.975, p = .042) was a protective factor against NSCLC in nonsmoking people. No statistical differences were observed in the classifications of rs1899663 and rs4759314 genotypes. Linkage disequilibrium analysis revealed a high linkage disequilibrium between the rs920778 and rs1899663 (D' = 0.99, r 2  = .74), rs920778 and rs4759314 (D' = 0.85, r 2  = .13), and rs1899663 and rs4759314 (D' = 0.79, r 2  = .00)., Conclusion: Our study demonstrated that HOTAIR expression increased in NSCLC, and that the genotypes of rs920778 could be useful in the diagnosis and prognosis of lung cancer., (© 2020 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals LLC.)

L-type lectins (LTLs) belong to the lectin family and are characterized by a conserved structural motif in their carbohydrate recognition domain. LTLs are homologous to leguminous lectins. In this study, we identified and functionally characterized an LTL from kuruma shrimp Marsupenaeus japonicus. We designated this LTL as MjLTL2. MjLTL2 contains a signal peptide, a Lectin_leg domain, a coiled coil, and transmembrane domain. MjLTL2 is distributed in hemocytes, heart, hepatopancreas, gill, stomach, and intestine; higher expression levels are seen in hemocytes and the hepatopancreas than in other tissues. MjLTL2 was upregulated following challenge of shrimp with Vibrio anguillarum and white spot syndrome virus (WSSV). MjLTL2 can agglutinate several bacteria without Ca 2+ . In addition, MjLTL2 could bind to several Gram-positive and -negative bacteria by binding to their lipopolysaccharide and peptidoglycan. However, MjLTL2 could not enhance the clearance of V. anguillarum in vivo. In the presence of WSSV infection, MjLTL2 knockdown by RNA interference resulted in a 7-day lower cumulative mortality of M. japonicus. Moreover, less VP19, VP24, VP26, and VP28 mRNAs were extracted from the hemocytes of MjLTL2 knockdown shrimp than from the control. These results suggest that MjLTL2 is involved in immune responses in shrimp., (Copyright © 2020. Published by Elsevier Ltd.)

Endometrial cancer (EC) is a common form of cancer in women. Metastasis is the main cause of EC treatment failure. Eukaryotic translation initiation factor 4E (eIF4E) is an oncogene that is overexpressed in a variety of malignancies and their distant metastases. The present study analyzed microarray data from the Oncomine database and revealed that high eIF4E expression was associated with poor prognosis and high pathological grade of EC. The expression of eIF4E was higher in EC tissues compared with in adjacent normal tissues. In addition, microRNA (miR)‑320a and miR‑340‑5p expression levels were downregulated in EC tissues compared with those in adjacent normal tissues, which suggested that these microRNAs may serve as EC tumor suppressor genes. miR‑320a and miR‑340‑5p could bind to the 3'‑UTR of eIF4E mRNA, thus downregulating the expression of eIF4E and phosphorylated (p)‑eIF4E in EC cells. Overexpression of miR‑320a or miR‑340‑5p effectively suppressed HEC‑1A cell migration and invasion. The downregulation of eIF4E and p‑eIF4E following miR‑320a or miR‑340‑5p transfection reduced the invasiveness and metastatic capability of EC cells in a manner associated with decreased expression of matrix metallopeptidase (MMP)‑3 and MMP‑9. In addition, one of the effects of transforming growth factor β1 (TGF‑β1), which is to induce the phosphorylation of eIF4E, was suppressed by miR‑320a and miR‑340‑5p overexpression. These two microRNAs also attenuated the features of TGF‑β1‑induced epithelial‑mesenchymal transition (EMT). In conclusion, the results of the present study demonstrated that eIF4E was upregulated in EC, whereas miR‑320a and miR‑340‑5p were downregulated in EC compared with adjacent normal tissues. In vitro, miR‑320a and miR‑340‑5p inhibited the migratory capability of EC cells by downregulating MMP‑3 and MMP‑9 and prevented TGF‑β1‑induced EMT through p‑eIF4E.

MicroRNAs (miRNAs/miRs) serve important roles in the chemotherapeutic effect of anticancer drugs. To investigate the roles of miRNAs in cisplatin‑induced suppression of lung adenocarcinoma cell proliferation, A549 cells were treated with different concentrations of cisplatin. An MTT assay demonstrated that cisplatin inhibited A549 cell proliferation in a dose‑dependent manner. Cisplatin induced cell apoptosis and inhibited cell migration by increasing the levels of miR‑93, miR‑26a and miR‑26b. Furthermore, as an upstream factor, miR‑93 was proposed to regulate cyclin D2 expression in miR‑93‑transfected A549 cells. Cisplatin also induced Bcl‑2‑associated X protein expression, and decreased that of Bcl‑2 and c‑Myc in lung adenocarcinoma cells. In vivo analysis further supported that cisplatin inhibited lung adenocarcinoma cell growth by regulating cyclin D2 and miR‑93 expression. In conclusion, our findings demonstrated that cisplatin could effectively inhibit lung adenocarcinoma cell proliferation by decreasing cyclin D2 expression via miR‑93.

Background: Regenerating islet-derived gene family member 4 (Reg4), a well-investigated growth factor in the regenerative pancreas, has recently been reported to be highly associated with a majority of gastrointestinal cancers. Pathological hyper-expression or artificial over-expression of Reg4 causes acceleration of tumor growth, migration, and resistance to chemotherapeutic 5-Fluorouracil (5-FU). Until now, no method has been successfully established for eliminating the effects of Reg4 protein., Methods: This study reports the production of an engineered immunoglobin, a single-chain variable fragment (scFv-Reg4), to specifically bind Reg4 and block the bioactivity. The complementary-determining regions (CDRs) against Reg4 were assigned using MOE and ZDOCK servers. The binding affinity (KD) was determined by bio-layer interferometry (BLI). MKN45 and AGS cell proliferation was determined by Thiazolyl blue tetrazolium bromide (MTT) method and the cell apoptosis was detected by flow cytometry assay., Results: The KD of scFv-Reg4 to Reg4 was determined to be 1.91×10-8. In MKN45 and AGS cell lines, scFv- Reg4 depressed Reg4-stimulated cell proliferation and the inhibitory rates were 27.7±1.5% and 17.3±2.6%, respectively. Furthermore, scFv significantly enhanced 5-FU-induced cell death, from 23.0±1.0% to 28.4±1.2% in MKN45 and 28.2±0.7% to 36.6±0.6% in AGS cells. Treatment with scFv alone could lyse cancer cells to a certain extent, but no significance has been observed., Conclusion: The single-chain antibody (scFv-Reg4) significantly inhibited gastric cancer cell proliferation and synergistically enhanced the lethal effect of 5-FU. Thus, traditional chemo-/radio- therapeutics supplemented with scFv-Reg4 may provide advances in the strategy for gastrointestinal cancer treatment., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Lung cancer is the most common cause of cancer‑associated mortality. MicroRNAs (miRNAs), as oncogenes or tumor suppressor genes, serve crucial roles not only in tumorigenesis, but also in tumor invasion and metastasis. Although miRNA‑let‑7a (let‑7a) has been reported to suppress cell growth in multiple cancer types, the biological mechanisms of let‑7a in lung adenocarcinoma are yet to be fully elucidated. In the present study, the molecular roles of let‑7a in lung adenocarcinoma were investigated by detecting its expression in lung adenocarcinoma tissues and exploring its roles in the regulation of lung cancer cell proliferation. Let‑7a expression was identified to be downregulated in lung adenocarcinoma tissues compared with normal tissues. Overexpression of let‑7a effectively suppressed cancer cell proliferation, migration and invasion in H1299 and A549 cells. Let‑7a also induced cell apoptosis and cell cycle arrest. Furthermore, let‑7a significantly inhibited cell growth by directly regulating cyclin D1 signals. This novel regulatory mechanism of let‑7a in lung adenocarcinoma provides possible avenues for future targeted therapies of lung cancer.

Multidrug resistance (MDR) in leukemia cells is a major obstacle to chemotherapeutic treatment. High expression and constitutive activation of multidrug resistance protein 1 (MRP1) has been associated with the development of resistance to anticancer drugs in a number of tumor types. The activity of c-Jun N-terminal kinase 1 (JNK1) is associated with the occurrence of MDR and MRP1 expression. The present study aimed to investigate the ability of solanine to resensitize the Adriamycin ® (ADR)-resistant human myelogenous leukemia cell line K562/ADM to ADR. Results of the Cell Counting Kit-8 assay demonstrated that solanine inhibited K562/ADM cell proliferation. K562/ADM cell sensitivity to ADR was increased following treatment with solanine, indicated by increased intracellular accumulation of ADR. Western blotting demonstrated that treatment with solanine led to reduced MRP1 protein expression, suggesting that solanine-induced ADR accumulation is due to the downregulation of MRP1 expression. Solanine-mediated MRP1 downregulation was observed to be dependent on the JNK signaling pathway. In conclusion, the results of the present study suggest that solanine reverses MDR in K562/ADM cells and may represent a novel therapeutic agent for the treatment of human myelogenous leukemia.

Solanine is an alkaloid and is the main extract of the traditional Chinese herb, Solanum nigrum Linn . It has been reported that Solanine has anti-inflammatory and antitumor properties. The present study aimed to investigate the antitumor effect of Solanine in Jurkat cells and demonstrate the molecular mechanism of antitumor activity of Solanine. A Cell Counting Kit-8 assay demonstrated that Solanine inhibited the proliferation of Jurkat cells in a dose-and time-dependent manner. Cell apoptosis was measured by flow cytometry. Flow cytometry revealed that Solanine induced apoptosis in a dose-dependent manner in Jurkat cells. Reverse transcription-quantitative polymerase chain reaction demonstrated that Solanine modulated the mRNA levels of B-cell lymphoma-2 (Bcl-2) and Bcl-2-associated X protein (Bax). Additionally, Bcl-2 and Bax expression was measured using western blot analysis. Western blot analysis revealed a significant increase in the expression of Bax and decrease in the expression of Bcl-2. Solanine increased the chemosensitivity of Jurkat cells to Adriamycin. In summary, the present results indicated that the antitumor activity of Solanine was associated with inhibition of cell proliferation, induction of apoptosis and increasing cytotoxicity of Adriamycin. Therefore, Solanine may have potential as a novel agent for the treatment of acute lymphocytic leukemia.

MicroRNAs (miRNAs or miRs) have recently become a popular focus of cancer research due to their ability to act as oncogenes or tumor suppressors. In the present study, miR‑33a‑5p expression was identified to be downregulated in lung adenocarcinoma samples compared with normal, which suggested that miR‑33a‑5p may serve as a tumor suppressor gene. Transfection with miR‑33a‑5p mimics inhibited the proliferation and migration of A549 and LTEP‑a‑2 cells and increased cellular apoptosis. A luciferase reporter assay confirmed that miR‑33a‑5p targets the 3'‑untranslated region of the mechanistic target of rapamycin (mTOR) gene. mTOR expression was decreased in A549 and LTEP‑a‑2 cells treated with miR‑33a‑5p mimics, as well as the expression of its downstream effectors phosphorylated (p)‑p70 ribosomal protein S6 kinase (p70S6K) and p‑eukaryotic translation initiation factor 4E binding protein 1 (4EBP1). Following treatment with celastrol, miR‑33a‑5p expression was upregulated, and miR‑33a‑5p could enhance cellular sensitivity to celastrol. Western blot analysis revealed that the expression of mTOR, p‑p70S6K and p‑4EBP1 decreased following celastrol treatment. These results suggested that mTOR was involved in the mechanism by which miR‑33a‑5p enhanced the sensitivity of lung adenocarcinoma cells to celastrol. Furthermore, LTEP‑a‑2 cells were xenografted subcutaneously into nude mice, to examine the effect of celastrol and miR‑33a‑5p on the growth of LTEP‑a‑2 cells in vivo. The results demonstrated that tumor growth in the celastrol‑treated or miR‑33a‑5p‑treated group was attenuated compared with the control group. Notably, tumor growth in the combination treatment group was almost arrested after 2 weeks. In addition, celastrol upregulated the expression of miR‑33a‑5p, and high expression of miR‑33a‑5p inhibited mTOR and its downstream effectors. In summary, miR‑33a‑5p inhibited the proliferation of lung adenocarcinoma cells, enhanced the antitumor effect of celastrol, and improved sensitivity to celastrol by targeting mTOR in lung adenocarcinoma in vitro and in vivo.

Activating transcription factor 6 (ATF6) pathway is the key branch of unfolded protein response (UPR). In this study, a homolog of ATFα from Marsupenaeus japonicus (MjATF6) was identified using genome sequencing and characterized, so as to investigate the role of ATF6 pathway in anti-viral immunity of M. japonicus. The cDNA of MjATF6 obtained was 1008 bp in length, with an open reading frame (ORF) of 849bp, which had encoded a putative of 283 amino acid proteins. Results of qRT-PCR showed that MjATF6 was distributed in all the six tested tissues, with the higher expression level being seen in hemocytes and hepatopancreas. Furthermore, MjATF6 expression would be up-regulated from 1 day to 7 day under white spot syndrome virus (WSSV) challenge. In comparison, RNA interference-induced MjATF6 knockdown had resulted in a lower 7-day cumulative mortality of M. japonicus in the presence of WSSV infection. Additionally, our results also revealed that less VP28 mRNA was extracted from hemocytes or hepatopancreas of MjATF6 knockdown shrimp than that from the control. Taken together, these results have confirmed that ATF6 pathway is vital for WSSV replication, and that UPR in M. japonicus may facilitate WSSV infection., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Cumulative evidence has indicated that celastrol may suppress cancer growth; however, the underlying mechanism requires further investigation. In the present study, A549 cells were treated with various concentrations of celastrol. Lung cancer cell proliferation was evaluated using an MTT assay and observed under a microscope. Cell apoptosis was detected by Annexin V fluorescein isothiocyanate/propidium iodide double-labeled flow cytometry. The results demonstrated that celastrol suppressed proliferation and induced apoptosis in a dose-independent manner. Celastrol may also decrease the phosphorylation levels of signal transducer and activator of transcription 3 (STAT3) and the B cell lymphoma-2 (Bcl-2)/Bcl-2 associated C protein (Bax) ratio. As microRNA (miR-24 and miR-181b) were predicated to target STAT3, STAT3 activation was inhibited in miR-24-or miR-181b-treated A549 cells compared with the control treatment. The ratio of Bcl-2/Bax was further reduced in miR-24 or miR-181b-treated A549 cells. The results were further confirmed by detecting in another lung adenocarcinoma cell line, LTEP-a-2. In summary, the results of the present study demonstrated that celastrol treatment suppressed the proliferation and induced apoptosis by regulating the expression levels of miR-24 and miR-181b.

Chemotherapy is the main treatment method for patients with chronic myeloid leukemia (CML) and has achieved marked results. However, the acquisition of multidrug resistance (MDR) has seriously affected the quality of life and survival rate of patients. The overexpression of the inhibitors of apoptosis proteins (IAPs) and the adenosine triphosphate (ATP)-dependent binding cassette (ABC) transporters are the two main causes of MDR. Apollon and MDR1 are the most important and representative members, respectively, among the IAPs and ABC transporters. In the present study, we investigated the role of Apollon and MDR1 in chemotherapy resistance and their mechanism of interaction. We respectively knocked down the expression of Apollon and MDR1 using short hairpin RNA (shRNA) in adriamycin (ADM) resistant human CML K562 cells and examined the drug sensitivity, the consequences with regard to ADM accumulation and the alterations in the expression of Apollon and MDR1. The expression levels of Apollon and MDR1 mRNA were higher in the K562/ADM cells compared with the parental K562 cells as determined by reverse transcription‑polymerase chain reaction (RT-PCR). The plasmids of Apollon and MDR1 shRNA were respectively stably transfected into K562/ADM cells using Lipofectamine 2000. The transfection efficiency was detected by fluorescence microscopy. Cell Counting Kit-8 (CCK-8) assay revealed that Apollon or MDR1 knockdown significantly increased the chemosensitivity of the K562/ADM cells to ADM. Flow cytometric assay revealed that K562/ADM/shMDR1 cells exhibited a significantly increased intracellular accumulation of ADM, and that changes were not found in the K562/ADM/shApollon cells. Compared with the parental K562/ADM cells, a significantly decreased expression of Apollon mRNA and protein was determined in the K562/ADM/shApollon cells without affecting the expression of MDR1 as determined by RT-PCR and western blotting. Likewise, the expression levels of MDR1 mRNA and protein also markedly downregulated in the K562/ADM/shMDR1 cells had no effect on Apollon expression. Collectively, our findings demonstrated, for the first time, that downregulation of Apollon or MDR1 through stable transfection with the Apollon- or MDR1-targeting shRNA induced MDR reversal through respective inhibition of Apollon or MDR1 expression and function. However, the reversal mechanism of Apollon and MDR1 revealed no direct interaction with each other.

MicroRNAs play important roles in tumorigenesis of various types of cancers. MiR-320a can inhibits cell proliferation of some cancers, but the biologic roles of miR-320a in lung cancer need to be further studied. Here, we investigated the roles of miR-320a in suppressing the proliferation of lung adenocarcinoma cells. MiR-320a treatment was found to effectively suppress LTEP-a-2 and A549 cell proliferation, and induce more apoptotic cells with irradiation treatment compared with control treatment. Our results also showed that miR-320a, as a novel miRNA, directly regulated signal transducer and activator of transcription 3 (STAT3) and its signals, such as Bcl - 2, Bax, and Caspase 3. The siRNA-inhibited STAT3 levels further proved its roles in regulating STAT3 signals. Moreover, miR-320a treatment effectively suppressed cancer cell growth in mice xenografts compared with controls, and significantly inhibited cell migration in vitro and in vivo. Our findings collectively demonstrated that miR-320a, by directly regulating STAT3 signals, not only suppressed cell proliferation and metastasis, but also enhanced irradiation-induced apoptosis of adenocarcinomia cells.

MicroRNAs (miRNAs) and Smad3, as key transcription factors in transforming growth factor-β1 (TGF-β1) signaling, help regulate various physiological and pathological processes. We investigated the roles of Smad3-regulated miRNAs with respect to lung adenocarcinoma cell apoptosis, proliferation, and metastasis. We observed that Smad3 and phospho-SMAD3 (p-Smad3) were decreased in miR-206- (or miR-140)-treated cells and there might be a feedback loop between miR-206 (or miR-140) and TGF-β1 expression. Smad3-related miRNAs affected tribbles homolog 2 (TRIB2) expression by regulating trib2 promoter activity through the CAGACA box. MiR-206 and miR-140 inhibited lung adenocarcinoma cell proliferation in vitro and in vivo by suppressing p-Smad3/Smad3 and TRIB2. Moreover, lung adenocarcinoma data supported a suppressive role for miR-206/miR-140 and an oncogenic role for TRIB2-patients with higher TRIB2 levels had poorer survival. In summary, miR-206 and miR-140, as tumor suppressors, induced lung adenocarcinoma cell death and inhibited cell proliferation by modifying oncogenic TRIB2 promoter activity through p-Smad3. MiR-206 and miR-140 also suppressed lung adenocarcinoma cell metastasis in vitro and in vivo by regulating EMT-related factors.

Multidrug resistance (MDR) plays a pivotal role in human chronic myelogenous leukemia (CML) chemotherapy failure. MDR is mainly associated with the overexpression of drug efflux transporters of the ATP-binding cassette (ABC) proteins. Phosphoinositide 3-kinase (PI3K)/Akt signaling cascade is involved in the MDR phenotype and is correlated with multidrug resistance 1 (MDR1)/P-glycoprotein (P-gp), multidrug resistance-associated protein 1 (MRP1) and breast cancer resistance protein (BCRP) expression in many human malignancies. Homeobox (HOX) B4, a member of the HOX gene family, has been reported to be correlated with occurrence, development, poor prognosis and drug resistance of human leukemia. In the present study, HOXB4 expression was analyzed in K562 cell line and its MDR subline K562/ADM. Compared with K562 cells, drug-resistant K562/ADM cells demonstrated evidently higher HOXB4 expression. In addition, we firstly investigated the reversal effect of HOXB4 deletion on K562/ADM cells and the underlying mechanism. The Cell Counting kit-8 (CCK-8) and flow cytometry assays showed that knockdown of HOXB4 enhanced chemosensitivity and decreased drug efflux in K562/ADM cells. Moreover, HOXB4 knockout led to downregulation of P-gp, MRP1 and BCRP expression and PI3K/Akt signaling activity, suggesting that repression of HOXB4 might be a key point to reverse MDR of K562/ADM cells.

Cyclin D2 is involved in the pathology of vascular complications of type 2 diabetes mellitus (T2DM). This study investigated the role of cyclin-D2-regulated miRNAs in endothelial cell proliferation of T2DM. Results showed that higher glucose concentration (4.5 g/l) significantly promoted the proliferation of rat aortic endothelial cells (RAOECs), and significantly increased the expression of cyclin D2 and phosphorylation of retinoblastoma 1 (p-RB1) in RAOECs compared with those under low glucose concentration. The cyclin D2-3' untranslated region is targeted by miR-98, as demonstrated by miRNA analysis software. Western blot also confirmed that cyclin D2 and p-RB1 expression was regulated by miR-98. The results indicated that miR-98 treatment can induce RAOEC apoptosis. The suppression of RAOEC growth by miR-98 might be related to regulation of Bcl-2, Bax and Caspase 9 expression. Furthermore, the expression levels of miR-98 decreased in 4.5 g/l glucose-treated cells compared with those treated by low glucose concentration. Similarly, the expression of miR-98 significantly decreased in aortas of established streptozotocin (STZ)-induced diabetic rat model compared with that in control rats; but cyclin D2 and p-RB1 levels remarkably increased in aortas of STZ-induced diabetic rats compared with those in healthy control rats. In conclusion, this study demonstrated that high glucose concentration induces cyclin D2 up-regulation and miR-98 down-regulation in the RAOECs. By regulating cyclin D2, miR-98 can inhibit human endothelial cell growth, thereby providing novel therapeutic targets for vascular complication of T2DM., (© 2016 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

P-glycoprotein (P-gp)-mediated multidrug resistance (MDR) has been reported to play a pivotal role in tumor chemotherapy failure. Study after study has illustrated that the phosphoinositide 3-kinase (PI3K)/Akt signaling cascade is involved in the MDR phenotype and is correlated with P-gp expression in many human malignancies. In the present study, osthole, an O-methylated coumarin, exhibited potent reversal capability of MDR in myelogenous leukemia K562/ADM cells. Simultaneously, the uptake and efflux of Rhodamine-123 (Rh-123) and the accumulation of doxorubicin assays combined with flow cytometric analysis suggested that osthole could increase intracellular drug accumulation. Furthermore, osthole decreased the expression of multidrug resistance gene 1 (MDR1) at both the mRNA and protein levels. Further experiments elucidated that osthole could suppress P-gp expression by inhibiting the PI3K/Akt signaling pathway which might be the main mechanism accounting for the reversal potential of osthole in the MDR in K562/ADM cells. In conclusion, osthole combats MDR and could be a promising candidate for the development of novel MDR reversal modulators.

Multidrug resistance (MDR) of leukemia cells is a major obstacle in chemotherapeutic treatment. The high expression and constitutive activation of P-glycoprotein (P-gp) and multidrug resistance protein-1 (MRP-1) have been reported to play a vital role in enhancing cell resistance to anticancer drugs in many tumors. The present study aimed to investigate the reversal of MDR by silencing homeobox A10 (HOXA10) in adriamycin (ADR)-resistant human chronic myelogenous leukemia (CML) K562/ADM cells by modulating the expression of P-gp and MRP-1. K562/ADM cells were stably transfected with HOXA10-targeted short hairpin RNA (shRNA). The results of reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analysis showed that the mRNA and protein expression of HOXA10 was markedly suppressed following transfection with a shRNA-containing vector. The sensitivity of the K562/ADM cells to ADR was enhanced by the silencing of HOXA10, due to the increased intracellular accumulation of ADR. The accumulation of ADR induced by the silencing of HOXA10 may be due to the downregulation of P-gp and MRP-1. Western blot analysis revealed that downregulating HOXA10 inhibited the protein expression of P-gp and MRP-1. Taken together, these results suggest that knockdown of HOXA10 combats resistance and that HOXA10 is a potential target for resistant human CML.

One of the major causes of failure in chemotherapy for patients with human chronic myelogenous leukemia (CML) is the acquisition of multidrug resistance (MDR). MDR is often associated with the overexpression of drug efflux transporters of the ATP-binding cassette (ABC) protein family. Timosaponin A-III (TAIII), a saponin isolated from the rhizome of Anemarrhena asphodeloides, has previously demonstrated the ability to suppress certain human tumor processes and the potential to be developed as an anticancer agent. Nevertheless, the ability of TAIII to reverse MDR has not yet been explored. In this study, the adriamycin (ADM) resistance reversal effect of TAIII in human CML K562/ADM cells and the underlying mechanism was investigated. The Cell Counting Kit-8 (CCK-8) assay showed that TAIII had a reversal effect on the drug resistance of K562/ADM cells. Flow cytometry assay showed increased intracellular accumulation of ADM after cells were pretreated with TAIII, and the changes in the accumulation of rhodamine-123 (Rho-123) and 5(6)-carboxyfluorescein diacetate (CFDA) dye in K562/ADM cells were determined to be similar to the changes of intracellular accumulation of ADM. After pretreatment of cells with TAIII, the decreasing expression of P-gp and MRP1 mRNA was examined by reverse transcription polymerase chain reaction (RT-PCR). Western blotting showed TAIII inhibiting P-gp and MRP1 expression depended on the PI3K/Akt signaling pathway by decreasing the activity of p-Akt. Moreover, wortmannin an inhibitor of PI3K/Akt signaling pathway has a strong inhibitory effect on the expression of p-Akt, P-gp and MRP1. Besides, the combined treatment with TAIII did not have an affect on wortmannin downregulation of p-Akt, P-gp and MRP1. Taken together, our findings demonstrate, for the first time, that TAIII induced MDR reversal through inhibition of P-gp and MRP1 expression and function with regained adriamycin sensitivity which might mainly correlate to the regulation of PI3K/Akt signaling pathway.

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive type of blood malignancy, deriving from T-cell progenitors in the thymus, and comprises 10-15% of pediatric and 25% of adult primary ALL cases. Despite advances, 20% of pediatric and the majority of adult patients with T-ALL succumb to mortality from resistant or relapsed disease, and the survival rate for patients with resistant or relapsed T-ALL remains poor. Alterations in the expression of Forkhead box protein M1 (FoxM1) have been detected in several types of cancer, and the inhibition of FoxM1 has been investigated as therapeutic strategy in cancer. The present study investigated the effects of the inhibition of FoxM1 by thiostrepton in human T-ALL Jurkat cells. The cells were treated with different concentrations of thiostrepton, either alone or in combination with doxorubicin. Cell viability was measured using CCK-8 assays and the cell cycle distribution, apoptosis and cell-associated mean fluorescence intensity of intracellular doxorubicin were assessed using flow cytometric analysis. The mRNA and protein expression levels were detected by reverse transcription-quantitative polymerase chain reaction and western blot analyses. The inhibition of FoxM1 by thiostrepton significantly decreased the proliferation of the Jurkat cells proliferation in a time- and dose-dependent manner. Cell arrest at the G2/M phase, and apoptosis was significantly increased in the thiostrepton-treated Jurkat cells. Thiostrepton reduced the half maximal inhibitory concentration of doxorubicin in the Jurkat cells, and significantly enhanced the cytotoxicity of doxorubicin within the Jurkat cells by enhancing doxorubicin-induced apoptosis and increasing the accumulation of intracellular doxorubicin. Furthermore, the inhibition of FoxM1 by thiostrepton enhanced doxorubicin-induced apoptosis, possibly through a caspase-3-dependent pathway, and increased the accumulation of intracellular doxorubicin, possibly through downregulating the expression of glutathione S-transferase pi. Collectively, the results of the present study suggested that targeting FoxM1 with thiostrepton resulted in potent antileukemia activity and chemosensitizing effects in human T-ALL Jurkat cells.

Pulmonary fibrosis (PF) is a disease with an unknown cause and a poor prognosis. In this study, we aimed to explore the pathogenesis of PF and the mechanism of sulindac in attenuating bleomycin (BLM)-induced PF. The rat PF model was induced by BLM and verified through histological studies and hydroxyproline assay. The severity of BLM-induced PF in rats and other effects, such as the extent of the wet lung to bw ratios, thickening of alveolar interval or collagen deposition, was obviously ameliorated in sulindac-treated rat lungs compared with BLM-induced lungs. Sulindac also reversed the epithelial mesenchymal transition (EMT) and inhibited the PF process by restoring the levels of E-cadherin and α-smooth muscle actin (SMA) in A549 cells. Our results further demonstrated that the above effects of sulindac might be related to regulating of interferon gamma (IFN-γ) expression, which further affects signal transducers and activators of transcription 3 (STAT3) and phosphorylated STAT3 (p-STAT3) levels. Moreover, higher miR-21 levels with the decreased E-cadherin and increased α-SMA expressions were found in transforming growth factor-β1-treated A549 cells, which can be reversed by sulindac. Collectively, our results demonstrate that by decreasing IFN-γ-induced STAT3/p-STAT3 expression to down-regulate miR-21, sulindac could significantly reverse EMT in A549 cells and prevent BLM-induced PF., (© 2015 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

Circulating microRNAs (miRNAs) are important in the diagnosis of a number of diseases, since serum or plasma miRNAs are more stable compared with miRNA isolated from blood samples. The aim of the present study was to investigate the association between the expression levels of serum let-7c miRNA and the clinical diagnosis of breast cancer (BC). The circulating let-7c levels of 90 BC patients and 64 healthy controls were determined by performing a reverse transcription-quantitative polymerase chain reaction assay. The results demonstrated that let-7c expression was downregulated in the BC tissues compared with the paracarcinoma control tissues. In addition, the let-7c expression in the serum of BC patients was significantly lower compared with the healthy controls (P<0.01). Using a cutoff value of 0.374×10 3 copies/ml, the serum expression levels of let-7c exhibited 87.5% sensitivity and 78.9% specificity for distinguishing BC patients from healthy controls (area under the receiver operating characteristic curve, 0.848; 95% confidence interval, 0.785-0.911). Furthermore, the results demonstrated that the serum expression levels of let-7c were significantly higher in premenopausal compared with postmenopausal patients (P<0.05), supporting the hypothesis that postmenopausal status may affect the serum expression levels of let-7c. However, no statistically significant differences were detected in the serum levels of let-7c between ER (or PR)-positive and -negative patients. Therefore, the current study hypothesized that serum let-7c may be used as a novel and valuable biomarker for the diagnosis of BC.

Aims: This study aimed to investigate the pathogenesis mechanisms of bleomycin (BLM)-induced pulmonary fibrosis (PF) in Sprague-Dawley rats and explore the anti-fibrotic role of arsenic trioxide (As2O3) in preventing PF., Main Methods: Intratracheal instillation of BLM was performed to establish PF rat models. The treatment group was treated with As2O3 (0.4 mg/kg/day). Morphological changes were observed by hematoxylin-eosin and Masson staining. Related proteins were determined by immunohistochemistry, immunofluorescence, and Western blot. MicroRNA detection was performed by quantitative real-time polymerase chain reaction., Key Findings: As a novel miRNA in PF, miR-98 decreased in the fibrotic lung tissues. Based on microRNA analysis software, we found that Stat3-3'-UTR is targeted by miR-98. Then, we found that Stat3 was activated with PF development and the expression of Stat3 and p-Stat3 was significantly increased in BLM-induced PF at day 28 compared with saline-treated rats. Our results showed that both Stat3 and p-Stat3 were significantly decreased in miR-98-treated A549 cells compared with that in mu-98-treated cultures or untreated controls. The fibrotic marker α-SMA was upregulated, whereas E-cadherin was inhibited in fibrotic lung tissues. The ratio of apoptotic factors Bax/Bcl-2 increased with the development of fibrosis. Furthermore, As2O3 treatment prevented lung interstitial thickening and inhibited the collagen type I and hydroxyproline, thereby preventing the development of PF. As2O3 also significantly down-regulated α-SMA but increased E-cadherin and miR-98 levels., Significance: The study revealed that arsenic trioxide prevented rat PF by up-regulation of miR-98 and inhibition of its downstream Stat3 signals., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Radioresistance is one of the main reasons for the failure of radiotherapy in lung cancer. The present study was conducted to identify the role of miR-511 in suppressing the growth of radioresistant lung adenocarcinoma cells. First, a radioresistant A549/R cell line was generated after prolonged exposure to X-rays for 68 Gy (2 Gy/day, 5 days/week) and the radioresistance was confirmed by wound healing assay. Next, oncogenic TRIB2 was found to be upregulated in the radioresistant A549/R cells when compared to that of the control A549 cells as determined by western blot analysis. As the upstream miRNA, quantitative PCR showed that miR-511 expression was decreased in the radioresistant A549/R cells. Overexpression of miR-511 in miR-511-transfected A549/R cells inhibited cell growth and increased the number of apoptotic cells when compared with the control treatment. Flow cytometric analysis further demonstrated that the growth suppressive effect of miR-511 on A549/R cells was mediated by regulation of the cell cycle, most likely due to a block in the G1-S transition. Finally, our results showed that the expression of BAX was lower in the radioresistant A549/R cells when compared with that in the control A549 cells. After downregulation of TRIB2 by miR-511 treatment, BAX expression was obviously increased in the miR-511-transfected apoptotic A549/R cells when compared to that in the NC-treated or control cultures. In summary, our results revealed that miR-511 regulates the growth of radioresistant A549/R cells by increasing BAX expression through TRIB2, which suggests that miR-511 may be a potential therapeutic molecule for the treatment of radioresistant lung adenocarcinoma.

Brain-derived neurotropic factor (BDNF) is a member of the neurotropin family. High levels of BDNF are associated with more aggressive malignant behavior in human cancer. In the present study, we observed the effect of cisplatin on BDNF expression in SH-SY5Y cells and investigated the mechanism of cisplatin in inducing the apoptosis of SH-SY5Y cells. Our results revealed that the expression of BDNF was obviously decreased in cisplatin-treated SH-SY5Y cells. In addition, the 3'-untranslated region of BDNF was found to be targeted by miR-16 using microRNA analysis software. After miR-16 was synthesized chemically, SH-SY5Y cells were transfected with miR-16 to investigate the regulatory role of miR-16 in regards to BDNF. The results showed that the expression of BDNF was markedly decreased in the miR‑16-transfected cells when compared with that in the control cultures as determined by western blotting. Moreover, miR-16 expression was obviously upregulated in the cisplatin-treated cells when compared with the untreated controls. Furthermore, SH-SY5Y cells were xenografted subcutaneously in nude mice to study the effect of cisplatin on the growth of SH-SY5Y cells in vivo. The results further showed that cisplatin inhibited the proliferation of SH-SY5Y cells in the cisplatin-treated mice when compared with the saline-treated control. The expression of miR-16 was increased, while the expression of BDNF was decreased in the cisplatin-treated mice. Our results demonstrated that cisplatin downregulated the expression of BDNF through miR-16 to inhibit SH-SY5Y cell proliferation in vitro and in vivo. These findings provide the basis for new targets for drug design or cancer therapy.

MicroRNAs (miRNAs) have significant roles in cell processes, including proliferation, apoptosis and stress responses. To investigate the involvement of miR-99 in the inhibition of HeLa cell proliferation, an miR-99 gene expression vector (pU6.1/miR-99), which overexpressed miR-99 in HeLa cells after transient transfection, was constructed. The expression of miR-99 was detected by qPCR. Cell proliferation and apoptosis were analyzed by cell viability, proliferation and apoptosis assays, as well as by electron microscopy. The results showed that overexpression of miR-99 in HeLa cells increased the HeLa cell mortality rate. Moreover, miR-99 overexpression was able to markedly inhibit HeLa cell proliferation according to the 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The cell apoptosis rate was significantly higher in pU6.1/miR-99-treated cells compared with that in the control cultures. Increases in intracellular electron density, as well as the proportion of nuclear plasma, blebbing phenomena and apoptotic bodies were observed in pU6.1/miR-99-treated cells compared with control cultures according to electron microscopy analysis. The Tribbles 2 (TRIB2) 3'-untranslated region was also observed to be targeted by miR-99 and the results further demonstrated that miR-99 was able to negatively regulate TRIB2 expression in HeLa cells The results indicate that miR-99 acts as a tumor suppressor gene in HeLa cells, establishing a theoretical basis for its application in cancer therapeutics.

Depression is a serious and potentially life-threatening mental disorder with unknown etiology. Emerging evidence shows that brain-derived neurotrophic factor (BDNF) and microRNAs (miRNAs) play critical roles in the etiology of depression. Here this study was aimed to identify and characterize the roles of BDNF and its putative regulatory miRNAs in depression. First, we identified that miR-182 may be a putative miRNA that regulates BDNF levels by bioinformatic studies, and characterized the effects of miR-182 on the BDNF levels using cell-based studies, side by side with miR-132 (a known miRNA that regulates BDNF expression). We showed that treatment of miR-132 and miR-182 respectively decreased the BDNF protein levels in a human neuronal cell model, supporting the regulatory roles of miR-132 and miR-182 on the BDNF expression. Furthermore, we explored the roles of miR-132 and miR-182 on the BDNF levels in depression using human subjects by assessing their serum levels. Compared with the healthy controls, patients with depression showed lower serum BDNF levels (via the enzyme-linked immunosorbent assays) and higher serum miR-132 and miR-182 levels (via the real-time PCR). Finally, the Pearson's (or Spearman's) correlation coefficient was calculated to study whether there was a relationship among the Self-Rating Depression Scale score, the serum BDNF levels, and serum BDNF-related miRNA levels. Our results revealed that there was a significant negative correlation between the SDS scores and the serum BDNF levels, and a positive correlation between the SDS scores and miR-132 levels. In addition, we found a reverse relationship between the serum BDNF levels and the miR-132/miR-182 levels in depression. Collectively, we provided evidence supporting that miR-182 is a putative BDNF-regulatory miRNA, and suggested that the serum BDNF and its related miRNAs may be utilized as important biomarkers in the diagnosis or as therapeutic targets of depression.

miR-21 can act as an oncogene. MSH2 has been reported that it involved in the DNA mismatch repair (MMR) system and overexpression of MSH2 can induce cell apoptosis. We predicted that MSH2-3'-untranslated region (3'-UTR) was targeted by miR-21 using microRNA analysis softwares. To further explore the roles of miR-21 and MSH2 in A549 cells, we constructed pcDNA-GFP-msh-UTR vector (including MSH2-3'-UTR) to transfect A549 cells with miR-21, GFP positive cells were estimated under a fluorescence microscopy and by flow cytometry. We found miR-21 could obviously downregulate the expression of MSH2, which was further proved by western blotting. Moreover, we treated A549 cells with cisplatin and found that cisplatin could inhibit A549 cell growth in vitro and in vivo. We also found that cisplatin could downregulate miR-21 expression, while increase MSH2 expression in A549 cells. Our results demonstrated that cisplatin could upregulate the expression of MSH2 through downregulating miR-21 to inhibit A549 cell proliferation, which provides new gene targets for drug design or cancer therary., (Copyright © 2013 Elsevier Masson SAS. All rights reserved.)

microRNAs (miRNAs) have been shown to play a role in cancer. Antisense oligonucleotides can bind directly to miRNAs and block their activity, which are generally named anti-miRNAs. To suppress A549 cell proliferation in vitro and in vivo by anti-miRNAs, an anti-miR-150 expression vector (PR-ASO-150), regulated by the H1 promoter and containing a 'TTTTT' sequence following a hairpin to stop transcription, was constructed. A549 cell proliferation in vitro or in nude mice was observed after PR-ASO-150 treatment. Our results showed that miR-150 expression was inhibited and the growth inhibition rate of A549 cells was higher in the PR-ASO-150-treated group compared with the control, which indicated that PR-ASO-150 could inhibit A549 cell proliferation by regulating miR-150 expression. Following establishment of A549 cancer cell xenografts in nude mice, PR-ASO-150 was delivered intratumorally to investigate the suppressive action to tumor proliferation by regulating miR-150 expression. The results indicate that the tumor volume and weight were lower compared to the control group. Our results further showed that p53 expression was higher after tumor tissue was treated with PR-ASO-150, indicating that up-regulation of p53 contributed to the suppression to tumor growth. Our study provides a novel strategy for cancer therapy through the development of anti-miRNAs.

To explore the mechanism of apoptosis induced by cisplatin, the expression of microRNAs (miRNAs) and regulating genes in K562 cells was analyzed using reverse transcription PCR, quantitative real-time PCR and enzyme-linked immunosorbent assays. Our results showed that miR-16, miR-34a-c, miR-17-5p and miR-125 were up-regulated, and their associated oncogenes (BCL2, E2F1 and E2F3, respectively) were down-regulated after cisplatin treatment. We also showed that miR-106 and miR-150 were down-regulated while their target genes (RB1 and P53, respectively) were up-regulated after cisplatin treatment. Moreover, miR-16, miR-34a-c and miR-17-5p proved to be upstream factors, regulating the expression of BCL2, E2F1 and E2F3, respectively. The oncogene E2F3 was down-regulated when RB1 expression was increased after treatment with antisense oligonucleotides (ASO). Similarly, BCL2 and E2F3 were down-regulated when P53 expression was elevated by ASO treatment. The study demonstrated that cisplatin induces K562 cells to apoptosis by reducing miR-106 which up-regulates RB1 or by inhibiting miR-150 which increases P53 expression.

MicroRNAs (miRNAs) have a profound impact on cell processes, including proliferation, apoptosis, and stress responses. We aimed to explore the role of antisense oligonucleotide (ASO) to induce proliferation or apoptosis of A549 cancer cells by inhibiting the expression of miRNAs. After A549/HBE/293T cells were treated with ASO, cells proliferation/apoptosis, and their relevant oncogenes/tumor suppressor genes were detected by light and electron microscopy, real-time PCR, enzyme-linked immunosorbent assay, etc. The results showed that ASO could inhibit the expression of miRNAs effectively. miR-16, miR-17, miR-34a-c, and miR-125 served as tumor suppressor miRNAs, while miR-20, miR-106, and miR-150 acted as oncogenic miRNAs. Our results also indicated that miR-16/34a-c, miR-17-5p, miR-125, miR-106, and miR-150 were the upstream factors, which could regulate the expression of BCL-2, E2F1, E2F3, RB1, and P53, respectively. After A549 cells treated with ASO for 24 h and different concentrations of anti-cancer drug (cisplatin or demethylcantharidin) were added into culture medium, the results indicated the percentage of alive cells in group treated with both ASO-106 (or ASO-150) and anti-cancer drug was lower than that in group treated with ASO, or anti-cancer drug, or both ASO-16 (or ASO-34a) and anti-cancer drug. In conclusion, ASO (specific to oncogenic miRNAs) could induce A549 cells apoptosis by inhibiting oncogenic miRNAs, and could increase chemotherapy sensitivity of A549 cells to anti-cancer drug, which holds great promise to lung cancer therapy.