Conventional and multi-omics assessments of subacute inhalation toxicity due to propylene glycol and vegetable glycerin aerosol produced by electronic cigarettes.

Propylene glycol (PG) and vegetable glycerin (VG) are the most common solvents used in electronic cigarette liquids. No long-term inhalation toxicity assessments have been performed combining conventional and multi-omics approaches on the potential respiratory effects of the solvents in vivo. In this study, the systemic toxicity of aerosol generated from a ceramic heating coil-based e-cigarette was evaluated. First, the aerosol properties were characterized, including carbonyl emissions, the particle size distribution, and aerosol temperatures. To determine toxicological effects, rats were exposed, through their nose only, to filtered air or a propylene glycol (PG)/ glycerin (VG) (50:50, %W/W) aerosol mixture at the target concentration of 3 mg/L for six hours daily over a continuous 28-day period. Compared with the air group, female rats in the PG/VG group exhibited significantly lower body weights during both the exposure period and recovery period, and this was linked to a reduced food intake. Male rats in the PG/VG group also experienced a significant decline in body weight during the exposure period. Importantly, rats exposed to the PG/VG aerosol showed only minimal biological effects compared to those with only air exposure, with no signs of toxicity. Moreover, the transcriptomic, proteomic, and metabolomic analyses of the rat lung tissues following aerosol exposure revealed a series of candidate pathways linking aerosol inhalation to altered lung functions, especially the inflammatory response and disease. Dysregulated pathways of arachidonic acids, the neuroactive ligand-receptor interaction, and the hematopoietic cell lineage were revealed through integrated multi-omics analysis. Therefore, our integrated multi-omics approach offers novel systemic insights and early evidence of environmental-related health hazards associated with an e-cigarette aerosol using two carrier solvents in a rat model.

Constitutively activated AMPKα1 protects against skeletal aging in mice by promoting bone-derived IGF-1 secretion.

Senile osteoporosis is characterized by age-related bone loss and bone microarchitecture deterioration. However, little is known to date about the mechanism that maintains bone homeostasis during aging. In this study, we identify adenosine monophosphate-activated protein kinase alpha 1 (AMPKα1) as a critical factor regulating the senescence and lineage commitment of mesenchymal stem cells (MSCs). A phospho-mutant mouse model shows that constitutive AMPKα1 activation prevents age-related bone loss and promoted MSC osteogenic commitment with increased bone-derived insulin-like growth factor 1 (IGF-1) secretion. Mechanistically, upregulation of IGF-1 signalling by AMPKα1 depends on cAMP-response element binding protein (CREB)-mediated transcriptional regulation. Furthermore, the essential role of the AMPKα1/IGF-1/CREB axis in promoting aged MSC osteogenic potential is confirmed using three-dimensional (3D) culture systems. Taken together, these results can provide mechanistic insight into the protective effect of AMPKα1 against skeletal aging by promoting bone-derived IGF-1 secretion.

CCL3 in the bone marrow microenvironment causes bone loss and bone marrow adiposity in aged mice.

The central physiological role of the bone marrow renders bone marrow stromal cells (BMSCs) particularly sensitive to aging. With bone aging, BMSCs acquire a differentiation potential bias in favor of adipogenesis over osteogenesis, and the underlying molecular mechanisms remain unclear. Herein, we investigated the factors underlying age-related changes in the bone marrow and their roles in BMSCs' differentiation. Antibody array revealed that CC chemokine ligand 3 (CCL3) accumulation occurred in the serum of naturally aged mice along with bone aging phenotypes, including bone loss, bone marrow adiposity, and imbalanced BMSC differentiation. In vivo Ccl3 deletion could rescue these phenotypes in aged mice. CCL3 improved the adipogenic differentiation potential of BMSCs, with a positive feedback loop between CCL3 and C/EBPα. CCL3 activated C/EBPα expression via STAT3, while C/EBPα activated CCL3 expression through direct promoter binding, facilitated by DNA hypomethylation. Moreover, CCL3 inhibited BMSCs' osteogenic differentiation potential by blocking ß-catenin activity mediated by ERK-activated Dickkopf-related protein 1 upregulation. Blocking CCL3 in vivo via neutralizing antibodies ameliorated trabecular bone loss and bone marrow adiposity in aged mice. This study provides insights regarding age-related bone loss and bone marrow adiposity pathogenesis and lays a foundation for the identification of new targets for senile osteoporosis treatment.

MAPKAPK5-AS1 drives the progression of hepatocellular carcinoma via regulating miR-429/ZEB1 axis.

BACKGROUND: Hepatocellular carcinoma (HCC) is a common malignancy. Long non-coding RNAs (lncRNAs) partake in the progression of HCC. However, the role of lncRNA MAPKAPK5-AS1 in the development of HCC has not been fully clarified. METHODS: RNA sequencing data and quantitative real-time polymerase chain reaction (qRT-PCR) were adopted to analyze MAPKAPK5-AS1, miR-429 and ZEB1 mRNA expressions in HCC tissues and cell lines. Western blot was used to detect ZEB1, E-cadherin and N-cadherin protein expressions. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), Transwell and flow cytometry assays were adopted to analyze the effects of MAPKAPK5-AS1 on cell proliferation, migration, invasion and apoptosis. Besides, luciferase reporter assay was used to detect the targeting relationship between miR-429 and MAPKAPK5-AS1 or ZEB1 3'UTR. The xenograft tumor mouse models were used to explore the effect of MAPKAPK5-AS1 on lung metastasis of HCC cells. RESULTS: MAPKAPK5-AS1 and ZEB1 expressions were up-regulated in HCC tissues, and miR-429 expression is down-regulated in HCC tissues. MAPKAPK5-AS1 knockdown could significantly impede HCC cell proliferation, migration, invasion and epithelial-mesenchymal transition (EMT), as well as promote cell apoptosis. MAPKAPK5-AS1 overexpression could enhance L02 cell proliferation, migration, invasion and EMT, and inhibit cell apoptosis. MiR-429 was validated to be the target of MAPKAPK5-AS1, and miR-429 inhibitors could partially offset the effects of knocking down MAPKAPK5-AS1 on HCC cells. MAPKAPK5-AS1 could positively regulate ZEB1 expression through repressing miR-429. Moreover, fewer lung metastatic nodules were observed in the lung tissues of nude mice when the MAPKAPK5-AS1 was knocked down in HCC cells. CONCLUSION: MAPKAPK5-AS1 can adsorb miR-429 to promote ZEB1 expression to participate in the development of HCC.

Author Correction: Plumbagin attenuates cancer cell growth and osteoclast formation in the bone microenvironment of mice.

During re-read of our previously article Plumbagin attenuates cancer cell growth and osteoclast formation in the bone microenvironment of micepublished in Acta Pharmacologica Sinica, we were regretted to point out a mistake shown in Fig. 2a. The representative figure chosen to indicate the inhibitory effect of 4 mg/kg of plumbagin treatment at 1 week against MDA-MB-231SArfp cells localization within bone environment was incorrect due to the mishandling in manuscript preparation. Although this correction does not affect the results and conclusion of the paper, all the authors agree on the correction of our negligence as providing the corrected Fig. 2a presented below. We feel sorry and apologize for all the inconvenience it caused.An amendment to this paper has been published and can be accessed via a link at the top of the paper.

LncRNA-241 inhibits 1,2-Dichloroethane-induced hepatic apoptosis.

Chlorinated organic chemical 1,2-dichloroethane (1,2-DCE) is used widely in industrial production processes, and excessive exposure may lead to liver damage. The mechanisms underlying 1,2-DCE-induced hepatotoxicity are not fully understood. Numerous studies have demonstrated that long-non-coding RNAs (lncRNAs) play a pivotal role in the chemical-induced toxicity. To explore whether aberrant lncRNA expression is involved in hepatotoxicity mediated by 1,2-DCE exposure, we detected alterations of lncRNA expression profiling in a mouse model of 1,2-DCE-induced hepatotoxicity by microarray chip. Bioinformatic analysis indicated that a down-regulated lncRNA (lncRNA241) after 1,2-DCE exposure might be involved in 1,2-DCE-induced hepatotoxicity. We treated AML12 cells with 1,2-DCE and its metabolite 2-chloroacetic acid (2-CA) for 48 h, and the results revealed that it was 2-CA rather than primary form (1,2-DCE) that resulted in the decline of lncRNA241 expression in hepatocytes. In vitro intervention studies revealed that the repression of lncRNA241 expression after 2-CA exposure led to the down-regulation of anti-apoptosis-associated factor insulin growth factor-1 (Igf1) at mRNA and protein levels through modulation of their common target mmu-miR-451a, which promoted hepatic apoptosis. This study provides valuable insight into the role of lncRNAs in response to hepatocyte apoptosis induced by 1,2-DCE.

Author Correction: Synergistic suppression of human breast cancer cells by combination of plumbagin and zoledronic acid in vitro.

The authors regretted to find the mis-representative images in Fig. 3a, c and Fig. 4a, c when re-read our previously published article Synergistic suppression of human breast cancer cells by combination of plumbagin and zoledronic acid In vitro (DOI: 10.1038/aps.2015.42) in the journal of Acta Pharmacologica Sinica. This mistake occurred due to the careless compilation when the authors tried to show the synergistic effect against tumor apoptosis during figure presentation process. The right Fig. 3a, c and Fig. 4a, c were provided below. Despite that this correction does not affect the results and conclusions of the aforementioned paper, all the authors still consent on the correction of this negligence. We apologize to the Editor and the readership of the journal for any inconvenience caused. Your thoughtful understanding is highly appreciated.

[Corrigendum] Inhibition of MDA-MB-231 breast cancer cell migration and invasion activity by andrographolide via suppression of nuclear factor-κB-dependent matrix metalloproteinase-9 expression.

Subsequent to the publication of the above article, the authors have realized that Fig. 2 in their paper contained an error. The image selected to represent the experiment showing the migration of cells in the presence of andrographolide (30 µM) was chosen incorrectly during the figure compilation process. A corrected version of Fig. 2 is presented here. Note that this change does not affect the results or the conclusions reported in this paper, and all the authors agree to this correction. The authors apologize to the Editor and to the readership of the Journal for any inconvenience caused. [the original article was published in the Molecular Medicine Reports 11: 1139­1145, 2015; DOI: 10.3892/mmr.2014.2872].

Improved antibacterial properties of collagen I/hyaluronic acid/quaternized chitosan multilayer modified titanium coatings with both contact-killing and release-killing functions.

Implant infection is one of the most severe complications after orthopedic surgery. The construction of an antibacterial coating on orthopedic implants with release-killing or contact-killing is one of the most efficient strategies to prevent implant-related infections. Here we reported a hydroxypropyltrimethyl ammonium chloride chitosan (HACC) based multilayer modified plasma-sprayed porous titanium coating generated via the layer-by-layer covalent-immobilized method. We demonstrated that the multilayer coating inhibited the colonization and biofilm formation of several bacterial strains, including Staphylococcus aureus (ATCC 25923), methicillin-resistant Staphylococcus aureus (MSRA, ATCC 43300) and clinical isolates of methicillin-resistant Staphylococcus epidermidis (MRSE 287), in vitro. HACC in the multilayer was released slowly with the degradation of the coating under the action of collagenase, further killing the planktonic bacteria, while the remaining HACC could kill the colonized bacteria. In a rat model of femur implants, the HACC-based multilayer-modified TCs effectively controlled the infection caused by MRSA and prevented bone destruction. Therefore, the HACC-based multilayer modified TCs with multiple antimicrobial properties could be a new potential ideal surface modification strategy to prevent implant associated infections.

CXCR1/Akt signaling activation induced by mesenchymal stem cell-derived IL-8 promotes osteosarcoma cell anoikis resistance and pulmonary metastasis.

The loss of appropriate cell adhesion normally induces apoptosis via a process termed anoikis. The aim of this study was to investigate the effects of mesenchymal stem cells (MSCs) in the cancer microenvironment on the anoikis resistance and pulmonary metastasis of osteosarcoma (OS) cells, and to evaluate the critical role of the interleukin (IL)-8/C-X-C chemokine receptor (CXCR) 1/Akt-signaling pathway in these processes. Metastatic OS subtype cells, which did or did not interact with MSC-conditioned medium (MSC-CM) in vitro, were isolated from the pulmonary site and named Saos2-lung-M. Both MSC-CM and IL-8 treatment increased the anoikis resistance of Saos2 cells in vitro. Moreover, exogenous MSC-CM promoted the survival and metastasis of Saos2 cells in nude mice. Saos2-lung-M cells were more malignant and resistant to anoikis than parental cells. MSCs secreted IL-8, thereby protecting OS cells from anoikis. Blocking the IL-8/CXCR1/Akt pathway via CXCR1 knockdown inhibited the pulmonary metastasis of Saos2-lung-MSCs and prolonged the survival of tumor-bearing mice. In conclusion, MSCs enhanced OS cell resistance to anoikis and pulmonary metastasis via regulation of the IL-8/CXCR1/Akt pathway. These findings suggest that MSCs can "select for" OS cells with high metastatic potential in vivo, and highlight CXCR1 as a key target in the regulation of pulmonary metastasis of OS cells.

TIMP3 Overexpression Improves the Sensitivity of Osteosarcoma to Cisplatin by Reducing IL-6 Production.

Osteosarcoma is the most common bone cancer in children and adolescents. Tissue inhibitors of metalloproteinases (TIMPs)-3 inhibit matrix metalloproteinases to limit extracellular matrix degradation. Cisplatin is a widely used chemotherapeutic drug used to cure osteosarcoma. Interleukin (IL)-6 and TIMP3 play important roles in the drug resistance of osteosarcoma; however, their relationship in this process remains unclear. This study aimed to explore the role of TIMP3 in the cisplatin sensitivity of osteosarcoma and its underlying molecular mechanisms in vitro and in vivo. We compared TIMP3 expression levels between patients with cisplatin-sensitive and -insensitive osteosarcoma. TIMP3 was overexpressed or knocked down in the Saos2-lung cell line, which is a Saos2 subtype isolated from pulmonary metastases that has higher cisplatin chemoresistance than Saos2 cells. IL-6 expression, cell proliferation, sensitivity to cisplatin, migration, and invasion after TIMP3 overexpression or knockdown were determined. The same experiments were performed using MG63 and U2OS cells. Subsequently, luciferase-labeled Saos2-lung cells overexpressing TIMP3 were injected into the tibiae of nude mice treated with cisplatin. The results showed that IL-6 inhibited TIMP3 expression in Saos2 and Saos2-lung cells via signal transducer and activator of transcription 3 (STAT3) activation. STAT3 knockdown reversed the effect of IL-6. The expression of TIMP3 was higher in patients with cisplatin-sensitive osteosarcoma than in those with insensitive osteosarcoma. IL-6 expression was downregulated upon TIMP3 overexpression, and upregulated by TIMP3 knockdown. TIMP3 overexpression suppressed cell proliferation and enhanced cisplatin sensitivity by activating apoptosis-related signal pathways and inhibiting IL-6 expression in vitro and in vivo. In conclusion, cisplatin sensitivity correlated positively with TIMP3 expression, which is regulated by the IL-6/TIMP3/caspase pathway. The TIMP3 pathway could represent a target for new therapies to treat osteosarcoma.

Aberrant expression of miR-451a contributes to 1,2-dichloroethane-induced hepatic glycerol gluconeogenesis disorder by inhibiting glycerol kinase expression in NIH Swiss mice.

The identification of aberrant microRNA (miRNA) expression during chemical-induced hepatic dysfunction will lead to a better understanding of the substantial role of miRNAs in liver diseases. 1,2-Dichloroethane (1,2-DCE), a chlorinated organic toxicant, can lead to hepatic abnormalities in occupationally exposed populations. To explore whether aberrant miRNA expression is involved in liver abnormalities mediated by 1,2-DCE exposure, we examined alterations in miRNA expression patterns in the livers of NIH Swiss mice after dynamic inhalation exposure to 350 or 700 mg m-3 1,2-DCE for 28 days. Using a microarray chip, we discovered that only mmumiR-451a was significantly upregulated in the liver tissue of mice exposed to 700 mg m-3 1,2-DCE; this finding was validated by quantitative real-time polymerase chain reaction. In vitro study revealed that it was metabolite 2-chloroacetic acid, not 1,2-DCE that resulted in the upregulation of mmu-miR-451a in the mouse AML12 cell line. Furthermore, our data showed that the upregulation of mmu-miR-451a induced by 2-chloroacetic acid could suppress the expression of glycerol kinase and lead to the inhibition of glycerol gluconeogenesis in mouse liver tissue and AML12 cells. These observations provide evidence that hepatic mmu-miR-451a responds to 1,2-DCE exposure and might induce glucose metabolism disorders by suppressing the glycerol gluconeogenesis process.

1,2-Dichloroethane Induces Reproductive Toxicity Mediated by the CREM/CREB Signaling Pathway in Male NIH Swiss Mice.

1,2-Dichloroethane (1,2-DCE) is a widely used chlorinated organic toxicant but little is known about the reproductive disorders induced by its excessive exposure. To reveal 1,2-DCE-induced male reproductive toxicity and to elucidate the underlying mechanisms, we exposed male National Institutes of Health Swiss mice to 1,2-DCE by inhalation at 0, 100, 350, and 700 mg/m3 for 6 h/day, for 1 and 4 weeks. Our findings showed a significant decrease in body weight with increased testis/body weight ratio, reduced sperm concentration and induced malformation of spermatozoa, and vacuolar degeneration of germ cells in the seminiferous tubules of testes in mice exposed to 1,2-DCE. Cyclic adenosine monophosphate (cAMP)-response element binding protein (CREB) and cAMP-response element modulator (CREM) were significantly inhibited by 1,2-DCE. This is consistent with the declines in the transducer of regulated CREB activity 1 and activator of CREM in testis, which results in the decrease in lactate dehydrogenase C and testis-specific kinase 1 in the testes. Moreover, the activation of p53 and Bax with the inhibition of Bcl-2 might be the reason for the upregulation of caspase-3 in the apoptosis, as detected by TdT-mediated dUTP nick-end labeling assay in the testes induced by 1,2-DCE. Finally, elevated testosterone levels were found along with increased levels of gonadotropin-releasing hormone, cAMP, luteinizing hormone (LH), and LH receptors in the testes. These findings suggest that 1,2-DCE inhibits CREM/CREB signaling cascade and subsequently induces apoptosis associated with p53 activation and mitochondrial dysfunction. This also results in induced malformation of spermatozoa, reduced sperm concentration, and pathological impairment of the testes.

Uridine diphosphate-glucuronosyltransferase 2B15 D85Y gene polymorphism is associated with lower prostate cancer risk: a systematic review and meta-analysis.

UGT2B15 (uridine diphosphate-glucuronosyltransferase 2B15) catalyzes the conversion of lipophilic C19 steroid androgens such as dihydrotestosterone (DHT) into water-soluble metabolites that can be excreted. Studies of the association between the UGT2B15 gene D85Y polymorphism and prostate cancer have yielded contradictory results. We therefore systematically searched in the PubMed, EMBASE, Science Direct/Elsevier, CNKI, and Cochrane Library databases, and identified six relevant studies with which to perform a meta-analysis of the relation between UGT2B15 D85Y polymorphism and prostate cancer risk. Our meta-analysis revealed a significant association between UGT2B15 D85Y gene polymorphism and prostate cancer in all genetic models (P<0.05). The combined odds ratios and 95% confidence intervals were as follows: additive model, 0.53 and 0.32-0.88; dominant model, 0.51 and 0.33-0.79; recessive model, 0.76 and 0.60-0.96; co-dominant model, 0.55 and 0.35-0.86; and allele model, 0.70 and 0.55-0.89. These results are consistent with the idea that the UGT2B15 D85Y enzyme variant reduces the risk of prostate cancer by efficiently metabolizing dihydrotestosterone (DHT), which is associated with prostate cancer progression.

Upregulation of Akt signaling enhances femoral fracture healing by accelerating atrophic quadriceps recovery.

Muscle damage and disuse muscular atrophy are detrimental for fracture healing. It has been reported that the Akt signaling pathway plays a role in skeletal muscle hypertrophy and atrophy. The aim of this study was to further investigate whether promoting local muscle function through regulating Akt signaling affects fracture healing. For this purpose, we combined a rat model of short-term atrophy of the quadriceps with a femoral fracture model. In brief, botulinum toxin-A (BTX) were administered locally into the quadriceps one week before femur osteotomy to induce muscle atrophy. For the following weeks after BTX treatment, animals received injection of the Akt activator SC79 (20mg/kg/week) or the Akt inhibitor MK2206 (100mg/kg/week). We found that SC79 significantly accelerated the recovery of quadriceps weight and fiber size after BTX treatment. Moreover, animals that received SC79 injection showed greater bone callus volumes and superior femur mechanical properties. Immunological analysis revealed that the expression levels of the muscle-specific marker myosin heavy chain (MHC) were increased while expression of a negative regulator of muscle mass and function, myostatin, was decreased after SC79 treatment. Furthermore, SC79 increased the mRNA levels of the myogenic regulatory factors MyoD, MRF4 and Myf5 and promoted myotube formation in vitro. Taken together, these findings reveal that SC79 could accelerate the recovery of reversible muscular atrophy induced by BTX and subsequently promote fracture healing through activation of the Akt signaling pathway, which suggests its therapeutic potential in orthopedics.

Targeting Osteocytes to Attenuate Early Breast Cancer Bone Metastasis by Theranostic Upconversion Nanoparticles with Responsive Plumbagin Release.

The early detection and thus treatment of breast cancer bone metastasis remain a big challenge clinically. As the most abundant cells within bone tissue, osteocytes have been found to manipulate the activity of early cancer bone metastasis by its crosstalk with cancer cells and osteoclasts. However, conventional bone-targeting nanomedicine has limited bone-lesion specificity and ignores the vital role of osteocytes during breast cancer bone metastasis. Also, it lacks detailed insight into the therapeutic mechanisms, which hinders the following translational practice. Previously, we have shown that a combination of zoledronic acid (ZA) and plumbagin (PL) synergistically alleviates cancer-induced bone destruction. Herein, we further develop a pH-responsive bone-targeting drug delivery system, i.e., the ZA-anchored bimodal mesoporous slica covered gadolinium(III) upconversion nanoparticles loaded with PL, to detect and treat bone metastasis sensitively and specifically at an early stage. This multifunctional nanosystem can target osteocytes to release PL as controlled by pH, decreasing osteocytic RANKL expression synergistically through the structural simulation of adenosine phosphate, which competitively inhibits the phosphorylation of osteocytic protein kinase-a, cAMP-response element binding protein, extracellular regulated protein kinase, and c-Jun N-terminal kinase. More importantly, by establishing a breast cancer bone metastasis mice model via intracardiac injection, we show that tumoriogenesis and osteoclastogenesis can both be attenuated significantly. We thereby realize the effective theranostics of tiny bone metastasis in breast cancer bone metastasis. Our work highlights the significance of theranostic nanomedicine and osteocyte-targeting therapy in the treatment of early bone metastasis, which could be applied in achieving efficient theranostic effects for other bone diseases.

1,2-Dichloroethane impairs glucose and lipid homeostasis in the livers of NIH Swiss mice.

Excessive exposure to 1,2-Dichloroethane (1,2-DCE), a chlorinated organic toxicant, can lead to liver dysfunction. To fully explore the mechanism of 1,2-DCE-induced hepatic abnormalities, 30 male National Institutes of Health (NIH) Swiss mice were exposed to 0, 350, or 700mg/m3 of 1,2-DCE, via inhalation, 6h/day for 28days. Increased liver/body weight ratios, as well as serum AST and serum ALT activity were observed in the 350 and 700mg/m3 1,2-DCE exposure group mice, compared with the control group mice. In addition, decreased body weights were observed in mice exposed to 700mg/m3 1,2-DCE, compared with control mice. Exposure to 350 and 700mg/m3 1,2-DCE also led to significant accumulation of hepatic glycogen, free fatty acids (FFA) and triglycerides, elevation of blood triglyceride and FFA levels, and decreases in blood glucose levels. Results from microarray analysis indicated that the decreases in glucose-6-phosphatase catalytic subunit (G6PC) and liver glycogen phosphorylase (PYGL) expression, mediated by the activation of AKT serine/threonine kinase 1 (Akt1), might be responsible for the hepatic glycogen accumulation and steatosis. Further in vitro study demonstrated that 2-chloroacetic acid (1,2-DCE metabolite), rather than 1,2-DCE, up-regulated Akt1 phosphorylation and suppressed G6PC and PYGL expression, resulting in hepatocellular glycogen accumulation. These results suggest that hepatic glucose and lipid homeostasis are impaired by 1,2-DCE exposure via down-regulation of PYGL and G6PC expression, which may be primarily mediated by the 2-chloroacetic acid-activated Akt1 pathway.

YAP-mediated mechanotransduction regulates osteogenic and adipogenic differentiation of BMSCs on hierarchical structure.

Hierarchical structure mimicking the natural bone microenvironment has been considered as a promising platform to regulate cell functions. We have previously fabricated hierarchical macropore/nanowire structure and evidence has shown that it can better manipulate the cytoskeleton status and osteogenic performance of osteoblasts. However, how cues of hierarchical structure are translated and ultimately linked to BMSC lineage commitment have still remained elusive, which hinders the accurate knowledge and further development of the hierarchical structure. In this study, bone marrow-derived mesenchymal stem cells (BMSCs) fate on hierarchical structure was investigated as well as the detailed mechanisms. It was shown that well-developed cytoskeleton and focal adhesion were observed for BMSCs on hierarchical structure, which was accompanied by enhanced osteogenic and depressed adipogenic potential. Evidence of increased YAP activity and nuclear translocation were exhibited on hierarchical structure and YAP knockdown inhibited osteogenic differentiation and promoted adipogenic differentiation induced by hierarchical structure. Further remove of cytoskeleton tension inhibited YAP function, which confirmed the key role of YAP-mediated mechanotransduction in the BMSC differentiation. These results together provide information of the stem cell fate commitment on hierarchical structure and a promising approach to design advanced biomaterials by focusing on specific mechanotransduction process.

Functional differences between AMPK α1 and α2 subunits in osteogenesis, osteoblast-associated induction of osteoclastogenesis, and adipogenesis.

The endocrine role of the skeleton-which is impaired in human diseases including osteoporosis, obesity and diabetes-has been highlighted previously. In these diseases, the role of AMPK, a sensor and regulator of energy metabolism, is of biological and clinical importance. Since AMPK's main catalytic subunit α has two isoforms, it is unclear whether functional differences between them exist in the skeletal system. The current study overexpressed AMPKα1 and α2 in MC3T3-E1 cells, primary osteoblasts and mouse BMSCs by lentiviral transduction. Cells overexpressing AMPKα2 showed higher osteogenesis potential than AMPKα1, wherein androgen receptor (AR) and osteoactivin played important roles. RANKL and M-CSF were secreted at lower levels from cells overexpressing α2 than α1, resulting in decreased osteoblast-associated osteoclastogenesis. Adipogenesis was inhibited to a greater degree in 3T3-L1 cells overexpressing α2 than α1, which was modulated by AR. An abnormal downregulation of AMPKα2 was observed in human BMSCs exhibiting the fibrous dysplasia (FD) phenotype. Overexpression of AMPKα2 in these cells rescued the defect in osteogenesis, suggesting that AMPKα2 plays a role in FD pathogenesis. These findings highlight functional differences between AMPKα1 and α2, and provide a basis for investigating the molecular mechanisms of diseases associated with impaired functioning of the skeletal system.