Fossil evidence sheds light on sexual selection during the early evolution of birds.

The impact of sexual selection on the evolution of birds has been widely acknowledged. Although sexual selection has been hypothesized as a driving force in the occurrences of numerous morphological features across theropod evolution, this hypothesis has yet to be comprehensively tested due to challenges in identifying the sex of fossils and by the limited sample size. Confuciusornis sanctus is arguably the best-known early avialan and is represented by thousands of well-preserved specimens from the Early Cretaceous Jehol lagerstätte, which provides us with a chance to decipher the strength of sexual selection on extinct vertebrates. Herein, we present a morphometric study of C. sanctus based on the largest sample size of this taxon collected up to now. Our results indicate that the characteristic elongated paired rectrices is a sexually dimorphic trait and statistically robust inferences of the sexual dimorphism in size, shape, and allometry that have been established, providing the earliest known sexual dimorphism in avian evolution. Our findings suggest that sexual selection, in conjunction with natural selection, does act upon body size and limb length ratio in early birds, thereby promoting a deeper understanding of the role of sexual selection in large-scale phylogenetic evolution.

Systematic optimization and evaluation of culture conditions for the construction of circulating tumor cell clusters using breast cancer cell lines.

BACKGROUND: Circulating tumor cell (CTC) clusters play a critical role in carcinoma metastasis. However, the rarity of CTC clusters and the limitations of capture techniques have retarded the research progress. In vitro CTC clusters model can help to further understand the biological properties of CTC clusters and their clinical significance. Therefore, it is necessary to establish reliable in vitro methodological models to form CTC clusters whose biological characteristics are very similar to clinical CTC clusters. METHODS: The assays of immunofluorescence, transmission electron microscopy, EdU incorporation, cell adhension and microfluidic chips were used. The experimental metastasis model in mice was used. RESULTS: We systematically optimized the culture methods to form in vitro CTC clusters model, and more importantly, evaluated it with reference to the biological capabilities of reported clinical CTC clusters. In vitro CTC clusters exhibited a high degree of similarity to the reported pathological characteristics of CTC clusters isolated from patients at different stages of tumor metastasis, including the appearance morphology, size, adhesive and tight junctions-associated proteins, and other indicators of CTC clusters. Furthermore, in vivo experiments also demonstrated that the CTC clusters had an enhanced ability to grow and metastasize compared to single CTC. CONCLUSIONS: The study provides a reliable model to help to obtain comparatively stable and qualified CTC clusters in vitro, propelling the studies on tumor metastasis.

Stress perception and associated factors among patients with Parkinson's disease: a cross-sectional study after the COVID-19 pandemic.

BACKGROUND: Parkinson's disease (PD) is a serious neurodegenerative disease that brings great stress to the physical and mental health of patients. At the same time, long-term treatment will also bring great economic losses and social burden to the family and society, especially after COVID-19 pandemic. The aim of this study is to analyze the current status of stress perception and anxiety in patients with PD and explore the influencing factors after the COVID-19 pandemic. METHODS: This study used the convenient sampling method to select the research objects of patients with PD who were outpatients or inpatients in a general public hospital in Hangzhou, Zhejiang Province, and the survey time was from February 2023 to March 2023. The measurements included the General information questionnaire, The Perceived Stress Scale (PSS) and The Self Rating Anxiety Scale (SAS). SPSS 21.0 software was used for data statistical analysis. RESULT: 394 out of 420 patients with PD completed the questionnaire. The stress perception score of PD was (16.41 ± 6.435) and the anxiety score was (54.77 ± 10.477). The stress perception scores of patients with PD were significantly different in gender, age, educational, occupation, nature of costs, time of sleep, quality of sleep, duration of disease, way of medical treatment and anxiety level (p < 0.05). Among them, age, duration of disease, public expenses, online remote therapy and anxiety level were the main influencing factors of stress perception in patients with PD (p < 0.05). Besides, there were significant differences in gender, educational, nature of costs, time of sleep, quality of sleep and duration of disease in anxiety among patients with PD (p < 0.05). CONCLUSION: After the COVID-19 pandemic, the level of stress perception and anxiety in patients with PD is high, and the influencing factors are complex.

Dysphagia Pattern in Early to Moderate Parkinson's Disease Caused by Abnormal Pharyngeal Kinematic Function.

Airway invasion is common in patients with Parkinson's disease (PD) and can cause serious complications. However, a PD-related dysphagic pattern has not been clearly elucidated. In this study, 53 patients with early to moderate PD were enrolled to undergo a videofluoroscopic study of swallowing evaluation (VFSS) and a battery of neuropsychological assessments. A set of VFSS variables (three visuoperceptual, nine temporal, and six spatial) were measured. The main effects of bolus viscosity and volume on airway invasion were calculated. Statistical analyses were performed to determine key kinematic factors of airway invasion for swallowing each bolus type. Airway invasion frequency was significantly higher for liquid boluses (liquid vs. pudding P < 0.001; liquid vs. honey P = 0.006). Laryngeal vestibule closure reaction time (LVCrt) was the key kinematic factor of airway invasion for 3 ml liquid swallow (P = 0.040), anterior displacement of hyoid bone was the key kinematic factor for both 5 ml and 10 ml liquid swallows (P = 0.010, 0.034, respectively). Male sex and advanced Hoehn and Yahr stage were significantly related to reduced anterior displacement of hyoid bone. These results reveal the dysphagic pattern related to PD, demonstrating that prolonged LVCrt and reduced anterior displacement of hyoid bone are two crucial kinematic factors contributing to airway invasion during the liquid swallow. In addition, hyoid bone dysfunction was correlated with disease severity and male sex. Our findings warrant further investigation of the pathophysiological mechanism of dysphagia in PD and would guide clinical intervention.

An evaluation of the effect of hydrofluoric acid (HF) treatment on keratins.

Hydrofluoric acid (HF) is commonly used in geological and paleontological research to extract organic fossils for morphological and chemical studies. However, during HF treatment, organic matter can also be altered, which raises concerns that HF-treated organic matter may not be representative of the original organic matter. To provide reference data for protein studies on fossils, herein, we use Fourier transform infrared (FTIR) spectroscopy to investigate the effect of HF (21.3 M) treatment on keratins, with treatment durations ranging from 2 to 48 h. Results show that the FTIR spectra of HF-treated samples are overall similar to that of the untreated sample, while curve fitting shows that HF treatment has led to alteration of the secondary structure in all the HF-treated samples and the effect is time-dependent. The 2- and 4-h treatment mainly reduced the content of the random coils, α-helix, and intermolecular ß-sheet. From 8h onwards, the content of random coils greatly increased at the expense of other structures. Our results imply that for protein detection in fossils using FTIR spectroscopy, the negative effect of HF treatment is not substantial, as the bands characteristic of proteins, that is, amide A, amide B, amide I, amide II, and amide III, are still present after the 48-h treatment. If the target is a secondary structure, the effect of HF treatment should be considered. When HF treatment is necessary, limiting the treatment duration to less than 4h may be a choice.

Potential Therapeutic Value of the STING Inhibitors.

The stimulator of interferon genes (STING) is a critical protein in the activation of the immune system in response to DNA. It can participate the inflammatory response process by modulating the inflammation-preferred translation program through the STING-PKR-like endoplasmic reticulum kinase (PERK)-eIF2α pathway or by inducing the secretion of type I interferons (IFNs) and a variety of proinflammatory factors through the recruitment of TANK-binding kinase 1 (TBK1) and interferon regulatory factor 3 (IRF3) or the regulation of the nuclear factor kappa-B (NF-κB) pathway. Based on the structure, location, function, genotype, and regulatory mechanism of STING, this review summarizes the potential value of STING inhibitors in the prevention and treatment of infectious diseases, psoriasis, systemic lupus erythematosus, non-alcoholic fatty liver disease, and other inflammatory and autoimmune diseases.

Biodegradable controlled-release polymer containing butylidenephthalide to treat a recurrent cervical spine glioblastoma with promising result: a compassionate trial report.

Intramedullary spinal glioblastoma multiforme (GBM) tends to recur within 11 months of surgical resection, even after adjuvant chemoradiation therapy. Treatment options for recurrent spinal GBM are often limited. (Z)-n-butylidenephthalide [(Z)-BP] is a natural compound that induces apoptosis, antiproliferation, anti-invasion and antistemness effects in GBM cells. The Cerebraca wafer consists of (Z)-BP within a biodegradable wafer that can be implanted in the parenchyma of the central nervous system to treat high-grade glioma. We present a 44-year-old woman with a recurrent spinal GBM who underwent microscopic surgical tumor excision under fluorescein sodium guidance and intraoperative neurophysiologic monitoring. Four Cerebraca wafers were implanted into the cord and intradural space during the operation. MRI revealed that both tumor volume and spinal cord edema had decreased 4 days after surgery; both had substantially decreased 16 months after surgery. Neurologic functions and quality of life were improved after salvage therapy. No adverse events were reported. Cerebraca wafer implantation during surgical re-excision of spinal GBM may be a novel therapeutic approach for reduction of the tumor size and subsequent spinal cord edema with no toxicity to the spinal cord.

The molecular evolution of feathers with direct evidence from fossils.

Dinosaur fossils possessing integumentary appendages of various morphologies, interpreted as feathers, have greatly enhanced our understanding of the evolutionary link between birds and dinosaurs, as well as the origins of feathers and avian flight. In extant birds, the unique expression and amino acid composition of proteins in mature feathers have been shown to determine their biomechanical properties, such as hardness, resilience, and plasticity. Here, we provide molecular and ultrastructural evidence that the pennaceous feathers of the Jurassic nonavian dinosaur Anchiornis were composed of both feather ß-keratins and α-keratins. This is significant, because mature feathers in extant birds are dominated by ß-keratins, particularly in the barbs and barbules forming the vane. We confirm here that feathers were modified at both molecular and morphological levels to obtain the biomechanical properties for flight during the dinosaur-bird transition, and we show that the patterns and timing of adaptive change at the molecular level can be directly addressed in exceptionally preserved fossils in deep time.

A bizarre Jurassic maniraptoran theropod with preserved evidence of membranous wings.

The wings of birds and their closest theropod relatives share a uniform fundamental architecture, with pinnate flight feathers as the key component. Here we report a new scansoriopterygid theropod, Yi qi gen. et sp. nov., based on a new specimen from the Middle-Upper Jurassic period Tiaojishan Formation of Hebei Province, China. Yi is nested phylogenetically among winged theropods but has large stiff filamentous feathers of an unusual type on both the forelimb and hindlimb. However, the filamentous feathers of Yi resemble pinnate feathers in bearing morphologically diverse melanosomes. Most surprisingly, Yi has a long rod-like bone extending from each wrist, and patches of membranous tissue preserved between the rod-like bones and the manual digits. Analogous features are unknown in any dinosaur but occur in various flying and gliding tetrapods, suggesting the intriguing possibility that Yi had membranous aerodynamic surfaces totally different from the archetypal feathered wings of birds and their closest relatives. Documentation of the unique forelimbs of Yi greatly increases the morphological disparity known to exist among dinosaurs, and highlights the extraordinary breadth and richness of the evolutionary experimentation that took place close to the origin of birds.

Archaeorhynchus preserving significant soft tissue including probable fossilized lungs.

We describe a specimen of the basal ornithuromorph Archaeorhynchus spathula from the Lower Cretaceous Jiufotang Formation with extensive soft tissue preservation. Although it is the fifth specimen to be described, unlike the others it preserves significant traces of the plumage, revealing a pintail morphology previously unrecognized among Mesozoic birds, but common in extant neornithines. In addition, this specimen preserves the probable remnants of the paired lungs, an identification supported by topographical and macro- and microscopic anatomical observations. The preserved morphology reveals a lung very similar to that of living birds. It indicates that pulmonary specializations such as exceedingly subdivided parenchyma that allow birds to achieve the oxygen acquisition capacity necessary to support powered flight were present in ornithuromorph birds 120 Mya. Among extant air breathing vertebrates, birds have structurally the most complex and functionally the most efficient respiratory system, which facilitates their highly energetically demanding form of locomotion, even in extremely oxygen-poor environments. Archaeorhynchus is commonly resolved as the most basal known ornithuromorph bird, capturing a stage of avian evolution in which skeletal indicators of respiration remain primitive yet the lung microstructure appears modern. This adds to growing evidence that many physiological modifications of soft tissue systems (e.g., digestive system and respiratory system) that characterize living birds and are key to their current success may have preceded the evolution of obvious skeletal adaptations traditionally tracked through the fossil record.

Direct Analysis of Rare Circulating Tumor Cells in Whole Blood Based on Their Controlled Capture and Release on Electrode Surface.

The development of a simple, sensitive, and effective method for the analysis of circulating tumor cells (CTCs) is essential for cancer diagnosis and metastasis prediction. In this work, we have proposed an enzyme-free electrochemical method for specific capture, sensitive quantification, and efficient release of CTCs. To achieve this, the specific interaction between CTCs and the corresponding aptamer designed to be located in the identification probe (IP) will unfold the hairpin structure of IP. Consequently, IP will initiate a hybridization reaction to produce a duplex, which will further trigger the hybridization chain reaction (HCR) process to form a composite product of CTCs and double-stranded DNA polymers. Therefore, a significantly amplified signal readout can be obtained. Moreover, the composite product can be brought to the electrode surface by tetrahedral DNA nanostructures to achieve the purpose of capturing and quantifying CTCs. More significantly, these captured CTCs can be controlled released without compromising cell viability via a simple strand displacement reaction. Taking the breast cancer cell MCF-7 as a representative, the newly developed approach led to an ultralow detection limit of 3 cells mL-1, which is superior to several studies previously reported. The current method has also been demonstrated to analyze CTCs in human whole blood and hence revealed a great potential in the future.

An Electrochemical Biosensor Designed by Using Zr-Based Metal-Organic Frameworks for the Detection of Glioblastoma-Derived Exosomes with Practical Application.

Glioblastoma (GBM) is one of the most fatal tumors in the brain, and its early diagnosis remains technically challenging due to the complex repertoires of oncogenic alterations and blood-brain barrier (BBB). GBM-derived specific exosomes can cross the BBB and circulate in body fluids, so they can be noninvasive biomarkers for the early diagnosis of GBM. Herein, we propose a sensitive and label-free electrochemical biosensor designed by using Zr-based metal-organic frameworks (Zr-MOFs) for the detection of GBM-derived exosomes with practical application. In the design, a peptide ligand can specifically bind with human epidermal growth factor receptor (EGFR) and EGFR variant (v) III mutation (EGFRvIII), which are overexpressed on the GBM-derived exosomes. Meanwhile, Zr-MOFs encapsulated with methylene blue can absorb on the surface of the exosomes due to the interaction between Zr4+ and the intrinsic phosphate groups outside of exosomes. Consequently, the concentration of exosomes can be directly quantified by monitoring the electroactive molecules inside MOFs, ranging from 9.5 × 103 to 1.9 × 107 particles/µL with the detection of limit of 7.83 × 103 particles/µL. Furthermore, this proposed biosensor can distinguish GBM patients from healthy groups, demonstrating the great prospect for early clinical diagnosis.

Structural colours in diverse Mesozoic insects.

Structural colours, nature's most pure and intense colours, originate when light is scattered via nanoscale modulations of the refractive index. Original colours in fossils illuminate the ecological interactions among extinct organisms and functional evolution of colours. Here, we report multiple examples of vivid metallic colours in diverse insects from mid-Cretaceous amber. Scanning and transmission electron microscopy revealed a smooth outer surface and five alternating electron-dense and electron-lucent layers in the epicuticle of a fossil wasp, suggesting that multilayer reflectors, the most common biophotonic nanostructure in animals and even plants, are responsible for the exceptional preservation of colour in amber fossils. Based on theoretical modelling of the reflectance spectra, a reflective peak of wavelength of 514 nm was calculated, corresponding to the bluish-green colour observed under white light. The green to blue structural colours in fossil wasps, beetles and a fly most likely functioned as camouflage, although other functions such as thermoregulation cannot be ruled out. This discovery not only provides critical evidence of evolution of structural colours in arthropods, but also sheds light on the preservation potential of nanostructures of ancient animals through geological time.

Beta-elemene inhibits breast cancer metastasis through blocking pyruvate kinase M2 dimerization and nuclear translocation.

Pyruvate kinase M2 (PKM2), playing a central role in regulating aerobic glycolysis, was considered as a promising target for cancer therapy. However, its role in cancer metastasis is rarely known. Here, we found a tight relationship between PKM2 and breast cancer metastasis, demonstrated by the findings that beta-elemene (ß-elemene), an approved drug for complementary cancer therapy, exerted distinct anti-metastatic activity dependent on PKM2. The results indicated that ß-elemene inhibited breast cancer cell migration, invasion in vitro as well as metastases in vivo. ß-Elemene further inhibited the process of aerobic glycolysis and decreased the utilization of glucose and the production of pyruvate and lactate through suppressing pyruvate kinase activity by modulating the transformation of dimeric and tetrameric forms of PKM2. Further analysis revealed that ß-elemene suppressed aerobic glycolysis by blocking PKM2 nuclear translocation and the expression of EGFR, GLUT1 and LDHA by influencing the expression of importin α5. Furthermore, the effect of ß-elemene on migration, invasion, PKM2 transformation, and nuclear translocation could be reversed in part by fructose-1,6-bisphosphate (FBP) and L-cysteine. Taken together, tetrameric transformation and nuclear translocation of PKM2 are essential for cancer metastasis, and ß-elemene inhibited breast cancer metastasis via blocking aerobic glycolysis mediated by dimeric PKM2 transformation and nuclear translocation, being a promising anti-metastatic agent from natural compounds.

Correction to: Cryptotanshinone activates AMPK-TSC2 axis leading to inhibition of mTORC1 signaling in cancer cells.

Following publication of the original article [1], the author noticed the following errors in the article.

Molecular evidence of keratin and melanosomes in feathers of the Early Cretaceous bird Eoconfuciusornis.

Microbodies associated with feathers of both nonavian dinosaurs and early birds were first identified as bacteria but have been reinterpreted as melanosomes. Whereas melanosomes in modern feathers are always surrounded by and embedded in keratin, melanosomes embedded in keratin in fossils has not been demonstrated. Here we provide multiple independent molecular analyses of both microbodies and the associated matrix recovered from feathers of a new specimen of the basal bird Eoconfuciusornis from the Early Cretaceous Jehol Biota of China. Our work represents the oldest ultrastructural and immunological recognition of avian beta-keratin from an Early Cretaceous (∼130-Ma) bird. We apply immunogold to identify protein epitopes at high resolution, by localizing antibody-antigen complexes to specific fossil ultrastructures. Retention of original keratinous proteins in the matrix surrounding electron-opaque microbodies supports their assignment as melanosomes and adds to the criteria employable to distinguish melanosomes from microbial bodies. Our work sheds new light on molecular preservation within normally labile tissues preserved in fossils.

Triassic-Jurassic climate in continental high-latitude Asia was dominated by obliquity-paced variations (Junggar Basin, Ürümqi, China).

Empirical constraints on orbital gravitational solutions for the Solar System can be derived from the Earth's geological record of past climates. Lithologically based paleoclimate data from the thick, coal-bearing, fluvial-lacustrine sequences of the Junggar Basin of Northwestern China (paleolatitude ∼60°) show that climate variability of the warm and glacier-free high latitudes of the latest Triassic-Early Jurassic (∼198-202 Ma) Pangea was strongly paced by obliquity-dominated (∼40 ky) orbital cyclicity, based on an age model using the 405-ky cycle of eccentricity. In contrast, coeval low-latitude continental climate was much more strongly paced by climatic precession, with virtually no hint of obliquity. Although this previously unknown obliquity dominance at high latitude is not necessarily unexpected in a high CO2 world, these data deviate substantially from published orbital solutions in period and amplitude for eccentricity cycles greater than 405 ky, consistent with chaotic diffusion of the Solar System. In contrast, there are indications that the Earth-Mars orbital resonance was in today's 2-to-1 ratio of eccentricity to inclination. These empirical data underscore the need for temporally comprehensive, highly reliable data, as well as new gravitational solutions fitting those data.

Cryptotanshinone inhibition of mammalian target of rapamycin pathway is dependent on oestrogen receptor alpha in breast cancer.

Cryptotanshinone (CPT) has been demonstrated to inhibit proliferation and mammalian target of rapamycin (mTOR) pathway in MCF-7 breast cancer cells. However, the same results are unable to be repeated in MDA-MB-231 cells. Given the main difference of oestrogen receptor α (ERα) between two types of breast cancer cells, It is possibly suggested that CPT inhibits mTOR pathway dependent on ERα in breast cancer. CPT could significantly inhibit cell proliferation of ERα-positive cancer cells, whereas ERα-negative cancer cells are insensitive to CPT. The molecular docking results indicated that CPT has a high affinity with ERα, and the oestrogen receptor element luciferase reporter verified CPT distinct anti-oestrogen effect. Furthermore, CPT inhibits mTOR signalling in MCF-7 cells, but not in MDA-MB-231 cells, which is independent on binding to the FKBP12 and disrupting the mTOR complex. Meanwhile, increased expression of phosphorylation AKT and insulin receptor substrate (IRS1) induced by insulin-like growth factor 1 (IGF-1) was antagonized by CPT, but other molecules of IGF-1/AKT/mTOR signalling pathway such as phosphatase and tensin homolog (PTEN) and phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) were negatively affected. Finally, the MCF-7 cells transfected with shERα for silencing ERα show resistant to CPT, and p-AKT, phosphorylation of p70 S6 kinase 1 (p-S6K1) and eukaryotic initiation factor 4E binding protein 1 (4E-BP1) were partially recovered, suggesting ERα is required for CPT inhibition of mTOR signalling. Overall, CPT inhibition of mTOR is dependent on ERα in breast cancer and should be a potential anti-oestrogen agent and a natural adjuvant for application in endocrine resistance therapy.

Correlation between serum interleukin-6 level and type 1 diabetes mellitus: A systematic review and meta-analysis.

OBJECTIVE: This report aimed to explore the association between the change of circulating interleukin-6 (IL-6) in patients and the development of type 1 diabetes mellitus (T1DM). METHODS: Four databases (PubMed, CNKI, WanFang and Civip) were used to search and list all clinical case-control studies about serum IL-6 level in T1DM patients between Jan 1, 2000 and Aug 31, 2016. RESULTS: A total of 20 case-control studies with 1238 T1DM patients and 742 healthy controls were included in this study. Compared to healthy controls, the serum content of IL-6 in patients with T1DM was significantly greater (overall: SMD, 1.49; 95% CI, 1.04 to 1.93; p<0.001), and notably increased in all subgroup with different age, ethnic and disease duration (all p<0.001). Furthermore, the analysis in subgroup exhibited that serum levels of IL-6 in the age greater than 20-year old (SMD, 1.64; 95% CI, 0.57-2.71; p<0.001), the diseased duration among 0-10years (SMD, 2.43; 95% CI, 1.42-3.44; p<0.001) and the sorted American group (SMD, 1.68; 95% CI, 0.85-2.51; p<0.001) were higher than those in control groups. CONCLUSIONS: Patients with T1DM were found to be linked to elevated level of serum IL-6, which the age, ethnic and disease durations in T1DM patients had no effect on the serum IL-6 levels for promoting diabetes mellitus.

Cryptotanshinone activates AMPK-TSC2 axis leading to inhibition of mTORC1 signaling in cancer cells.

BACKGROUND: Cryptotanshinone (CPT), a fat-soluble phenanthraquinone from Salvia miltiorrhiza Bunge, has been demonstrated to inhibit phosphorylation of p70 S6 kinase 1 (S6K1) and eukaryotic initiation factor 4E binding protein 1 (4E-BP1), a couple of direct downstream effectors of the mammalian target of rapamycin complex 1 (mTORC1), resulting in cancer cell arrested in G0 phase and subsequent inhibition of proliferation. However, its concrete molecular mechanism about how CPT inhibits mTORC1 signaling pathway is unclear. METHODS: one solution was used to check cell viability and western blotting for determining expression of the indicated proteins. Molecular docking was performed to assess the binding of CPT with mTOR. The co-immunoprecipitation assay was to analyze whether CPT could disrupt the mTORC1 and TSC1/TSC2 complex. Recombinant adenoviral dominant-negative AMPKα was used to downregulate expression of AMPKα and lentiviral AMPK and TSC2 to silence the AMPK and TSC2 in Rh30 cells. RESULTS: Primarily, Rh30 cells expressing rapamycin-resistant mutant mTOR are also sensitive to CPT, while the molecular docking result for CPT binding to mTOR is negative, suggesting that CPT inhibition of mTORC1 is different from rapamycin. Then the related proteins of PTEN-PI3K pathway was proved not to be affected, but the phosphorylation of adenosine monophosphate-activated protein kinase (AMPK) was activated by a concentration- and time- dependent manner, meaning that it may be associated with AMPK. Further results indicated that compound C, inhibitor of AMPK, could clearly reversed CPT inhibitory effect on Rh30 cells, and dominant-negative AMPK in cancer cells conferred resistance to CPT inhibition of 4E-BP1 and phosphorylation of S6K1, as well as sh-AMPK. Furthermore, compared with AMPK-positive MEF cells, AMPK-negative MEF cells are less sensitive to CPT by the findings that 4E-BP1 and phosphorylation of S6K1 express comparatively more. Additionally, phosphorylation of tuberous sclerosis complex 2 (TSC2) was activated under the treatment of CPT, and down-expression of TSC2 by shRNA slightly recovered expression of 4E-BP1 and phosphorylation of S6K1, while co-immunoprecipitation of TSC2 did not alter expression of TSC1 by CPT. CONCLUSION: CPT inhibiting mTORC1 pathway was mostly due to activation of AMPK-TSC2 axis rather than specific binding to mTORC1. CPT is a potent anticancer agent targeting AMPK.