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Defining drivers of tumour initiation can provide opportunities to control cancer progression. Here we report that lysophosphatidic acid receptor 4 (LPAR4) becomes transiently upregulated on pancreatic cancer cells exposed to environmental stress or chemotherapy where it promotes stress tolerance, drug resistance, self-renewal and tumour initiation. Pancreatic cancer cells gain LPAR4 expression in response to stress by downregulating a tumour suppressor, miR-139-5p. Even in the absence of exogenous lysophosphatidic acid, LPAR4-expressing tumour cells display an enrichment of extracellular matrix genes that are established drivers of cancer stemness. Mechanistically, upregulation of fibronectin via an LPAR4/AKT/CREB axis is indispensable for LPAR4-induced tumour initiation and stress tolerance. Moreover, ligation of this fibronectin-containing matrix via integrins α5β1 or αVβ3 can transfer stress tolerance to LPAR4-negative cells. Therefore, stress- or drug-induced LPAR4 enhances cell-autonomous production of a fibronectin-rich extracellular matrix, allowing cells to survive 'isolation stress' and compensate for the absence of stromal-derived factors by creating their own tumour-initiating niche.

While mesenchymal stem cells (MSCs) have been widely used to repair radiation-induced bone damage, the molecular mechanism underlying the effects of MSCs in the maintenance of bone homeostasis under radiation stress remains largely unknown. In this study, the role and mechanisms of R-spondin 1 (Rspo1)-leucine-rich repeat-containing G protein-coupled receptor 4 (LGR4) axis on the initiation of self-defense of bone mesenchymal stem cells (BMSCs) and maintenance of bone homeostasis under radiation stress were investigated. Interestingly, radiation increased levels of Rspo1 and LGR4 in BMSCs. siRNA knockdown of Rspo1 or LGR4 aggravated radiation-induced impairment of self-renewal ability and osteogenic differentiation potential of BMSCs. However, exogenous Rspo1 significantly attenuated radiation-induced depletion of BMSCs, and promoted the lineage shift towards osteoblasts. This alteration was associated with the reversal of mammalian target of rapamycin (mTOR) activation and autophagy decrement. Pharmacological and genetic blockade of autophagy attenuated the radio-protective effects of Rspo1, rendering BMSCs more vulnerable to radiation-induced injury. Then bone radiation injury was induced in C57BL6J mice to further determine the radio-protective effects of Rspo1. In mice, administration of Rspo1 recombinant protein alleviated radiation-induced bone loss. Our results uncover that Rspo1-LGR4-mTOR-autophagy axis are key mechanisms by which BMSCs initiate self-defense against radiation and maintain bone homeostasis. Targeting Rspo1-LGR4 may provide a novel strategy for the intervention of radiation-induced bone damage.

Viral entry inhibitors are absent in hepatitis C virus (HCV) treatment regimens although a dozen direct-acting antiviral (DAA) drugs are available now. Based on a previously identified HCV entry inhibitor

Straw return plays an important role in the improvement of farmland ecological environments, increasing soil fertility, crop quality, and yield. However, in the early growth of rice (Oryza sativa L.), straw decomposition and rice growth may be inhibited by suboptimal carbon/nitrogen (C/N) ratios. We, therefore, explored methods that could alleviate the inhibition of straw decomposition using different combinations of ammonium bicarbonate and compound fertilizer. No fertilizer (CK) and compound fertilizer (S0) were used as the control. The ratios of added ammonium bicarbonate were 10% (S1), 20% (S2), 30% (S3), and 40% (S4), respectively. Compared with the S0 treatment, the S2 treatment accelerated straw decomposition, promoted the early growth of tillers and roots, and improved the leaf area index, dry matter accumulation, and N use efficiency (apparent N utilization, agronomic N utilization, and partial factor productivity increased by 4.05‒19.48% in two years). Moreover, panicle number, filled‐grain percentage, and 1000‐grain weight were significantly higher in the S2 treatment than other treatments. The grain yield of S2 treatment was significantly higher than that of S0 treatment by 4.06% (2019) and 5.66% (2020). Regression analysis revealed that the optimal ratios of ammonium bicarbonate for increasing straw decomposition and grain yield were relatively close, 16.27% and 14.60%, respectively. These results suggest that compared with conventional fertilization method (S0), the 20% treatment is a more effective fertilization method that prevents inhibition of straw decomposition, promoting rice growth and yield, and improving nitrogen (N) use efficiency.

Academic performance of college students is the main concern of educational institutions. Effective and timely predicting performance is not only conducive to the school’s Ministry to improve the efficiency of supervision, but also helps students to develop good study habits. With the rapid construction of digital campus, university as the main range for students’ life can not only serve convenience but also record daily life. The popular using of smart card makes it easy to outline students behavior pattern with rich data. The purpose of our work is to predict students performance based on their behavior pattern and analyze the correlation between them. In this paper, we propose a general framework to model students performance. Firstly, we describe students behavior pattern and extract behavior features in two perspectives including statistics and relevance. Then we employ a multi-task model to learn performance of every course simultaneously. Our experiments on a real world data set of college students show a good outcome. We do a further analysis on relation between students behavior and academic performance. Moreover, our experiments indicate that our framework is feasible for early warning.

New evidence of the interactions between the immune system and bone has accumulated in bone diseases, including osteoporosis, periodontitis and rheumatoid arthritis. A marked imbalance between bone resorption and formation is central to the onset of pathological bone loss. Osteoimmunology has revealed that the immune system, including T cells, B cells and inflammatory cytokines, is a key regulator of both osteoclasts and osteoblasts. Th1 cells, which differentiate from CD4+T cells, are thought to play a major function during bone loss. Moreover, the correlated expression of Th1 cytokines (interleukin-12 (IL-12), interferon-γ (IFN-γ)) and bone-resorbing cytokines (tumor necrosis factor-α (TNF-α), IL-1) also plays a key role during inflammatory induced bone resorption. Furthermore, a relatively new member of the CD4+T cell family Th17 displays the ability to promote osteoclast activity. The effect of IFN-γ and IL-17 released by Th 17 cells on pre-osteoclast proliferation, differentiation and apoptosis provides the preliminary basis for the immune mechanism of pathological bone loss. The role of B cells in osteoimmunological interactions has long been suspected based on findings of B cells as active regulators of the RANK/RANKL/OPG axis. Pathological bone loss, including osteoporosis and human immunodeficiency virus-associated bone loss, are related to the altered RANKL/OPG through modified production by B cells, supporting this assumption. All of the above evidence may provide new theoretical explanations for the relationship between bone metabolism and the immune system as well as offer perspectives for the prevention and treatment of pathological bone loss.

Background Previous clinical trials have already demonstrated that combinations of two or more drugs were more effective in the cancer treatment, especially sequential photodynamic design combing with sequential chemotherapy. In our study, we propose a ternary cocktail NP delivery system based on self-decomposable NPs, which could realize synergistic chemo-photodynamic therapy through double loading chemo-drugs and multi-level programmable PDT treatment. Methods PS drug methylene blue (MB) was encapsulated into the center of the NPsmall, NPbig&thin, and NPbig&thick carriers through “grown-in” loading mechanism, which was released based on the drug concentration difference of the drug release environment. NPsmall, NPbig&thin, and NPbig&thick carriers have three different drug release profiles, which could realize multi-level programmable PDT treatment. At the same time, antitumor drug gemcitabine hydrochloride (GM) and Docetaxel (DTX), were chosen as the double loading chemo-drugs that absorbed onto the NPbig&thin and NPbig&thick surface, respectively. In specific, various particle configurations were used for modulating the inner MB sequential release with three pulse Tmax. Also, by adjusting the NPbig&thin and NPbig&thick configuration, the release interval lag time between absorbed GM and DTX can be successfully modulated to achieve maximized chemotherapeutic efficacy. Results In vitro and in vivo results demonstrated that these three pulses Tmax and the sustained release of MB could maximize the multi-level programmable PDT treatment. And the absorbed GM and DTX also have a release time lag of 12 h, which has been proved as the most effectiveness synergistic interval lag time in the cancer treatment. Conclusion Such a precise sequential release manner ternary cocktail NPs provided a promising platform for efficient and safe chemo-photodynamic therapy, which serves as a promising drug delivery system to cure cancer in the future. Electronic supplementary material The online version of this article (10.1186/s13046-017-0586-1) contains supplementary material, which is available to authorized users.

There are interactions between immune response and destruction of articular cartilage/synovial tissue in osteoarthritis (OA), which leads to chronic inflammation and systemic failure of joints. However, the role of immunological factors in the pathogenesis of OA has not been fully elucidated. In this study, expressions of 47 cytokines and chemokines were tested in the peripheral bloods and synovial fluids from 13 normal controls (NCs) and 31 OA patients. The primary chondrocytes, which were isolated from cartilages of OA patients, were stimulated by recombinant CXCL8 and CXCL11 to analyze the proliferation, cytokine secretion, and signaling pathways. The levels of IL-17A, CXCL8, CXCL9, and CXCL11 were elevated in the serum and synovial fluids of OA patients. Moreover, expressions of CXCL8 and CXCL11 were remarkably increased in the synovial fluids of late stage OA. Stimulation of CXCL8/11 resulted in the reduction of primary chondrocytes proliferation with downregulation of G2-M stage but elevation of S stage and apoptosis cells. The secretions of proinflammatory cytokines and MMPs were also increased upon stimulation. Furthermore, CXCL8/11 stimulation induced the higher expressions of phosphorylated STAT3, NF-kB p50 and JNK, but not p38MAPK or ERK1/2. Our findings suggested that CXCL8 and CXCL11 promoted the apoptosis and suppressed the proliferation of chondrocytes probably via influencing JAK-STAT, NF-kB and JNK MAPK signaling pathway and enhancing the expressions of other proinflammatory cytokines. CXCL8/11 may aggravate the disease progression of OA, and may also be served as new therapeutic targets for treatment of OA.

The present study aimed to evaluate the effects of Enterococcus faecalis, inactivated by the common intracanal irrigants sodium hypochlorite (NaOCl) and chlorhexidine (CHX), on osteoblasts. E. faecalis was inactivated with 2% CHX or 5.25% NaOCl. Subsequently, the Cell Counting kit‑8 assay was used to examine the effects of CHX‑ and NaOCl-inactivated E. faecalis on MC3T3‑E1 osteoblast cell proliferation. Alizarin red staining was used to determine osteoblast mineralization, and osteogenic induction was quantified by determining the optical density of the dye solution. The relative expression levels of osteogenic genes were detected after 1, 4, 7 and 14 days of stimulation with CHX‑ and NaOCl-inactivated E. faecalis by reverse transcription‑quantitative polymerase chain reaction. The results indicated that CHX‑inactivated E. faecalis inhibited osteoblast proliferation, whereas NaOCl‑inactivated E. faecalis did not suppress cell proliferation. Various concentrations of CHX‑ and NaOCl‑inactivated E. faecalis induced different degrees of osteoblast mineralization. The expression levels of osteocalcin, alkaline phosphatase, runt‑related transcription factor 2, osteopontin and osterix were upregulated in cells following stimulation with 107 and 105 colony‑forming units/ml E. faecalis inactivated by CHX and NaOCl; the upregulation of these osteogenic genes occurred at various time points. In conclusion, the present study demonstrated that CHX‑inactivated E. faecalis exerted more of an effect on osteoblast proliferation compared with NaOCl‑inactivated E. faecalis. In addition, CHX‑ and NaOCl‑inactivated E. faecalis was able to induce mineralization and relevant osteogenic gene expression in osteoblast cells.

Intact and stable bone reconstruction is ideal for the treatment of periodontal bone destruction but remains challenging. In research, biomaterials are used to encapsulate stem cells or bioactive factors for periodontal bone regeneration, but, to the best of our knowledge, using a supramolecular hydrogel to encapsulate bioactive factors for their sustained release in bone defect areas to promote periodontal bone regeneration has not been reported. Herein, we used a well-studied hydrogelator, NapFFY, to coassemble with SDF-1 and BMP-2 to prepare a supramolecular hydrogel, SDF-1/BMP-2/NapFFY. In vitro and in vivo results indicated that these two bioactive factors were ideally, synchronously, and continuously released from the hydrogel to effectively promote the regeneration and reconstruction of periodontal bone tissues. Specifically, after the bone defect areas were treated with our SDF-1/BMP-2/NapFFY hydrogel for 8 weeks using maxillary critical-sized periodontal bone defect model rats, a superior bone regeneration rate of 56.7% bone volume fraction was achieved in these rats. We anticipate that our SDF-1/BMP-2/NapFFY hydrogel could replace bone transplantation in the clinic for the repair of periodontal bone defects and periodontally accelerated osteogenic orthodontics in the near future.

Evidence indicates that microRNAs (miRNAs) play vital roles in regulating osteogenic differentiation and bone formation. Methods: Here, we show that a polyethyleneimine (PEI)-functionalized graphene oxide (GO) complex efficiently loaded with the miR-214 inhibitor is assembled into silk fibroin/hydroxyapatite (SF/HAP) scaffolds that spatially control the release of the miR-214 inhibitor. Results: SF/HAP/GO scaffolds with nanosized GO show high mechanical strength, and their hierarchical microporous structures promote cell adhesion and growth. The SF/HAP/GO-PEI scaffolds loaded with mir-214 inhibitor (SF/HAP/GPM) were tested for their ability to enhance osteogenic differentiation by inhibiting the expression of miR-214 while inversely increasing the expression of activating transcription factor 4 (ATF4) and activating the Akt and ERK1/2 signaling pathways in mouse osteoblastic cells (MC3T3-E1) in vitro. Similarly, the scaffolds activated the osteoblastic activity of endogenous osteoblast cells to repair critical-sized bone defects in rats without the need for loading osteoblast cells. Conclusion: This technology is used to increase osteogenic differentiation and mineralized bone formation in bone defects, which helps to achieve cell-free scaffold-based miRNA-inhibitor therapy for bone tissue engineering.

The aim of this study was to prepare a promising biomaterial for dental pulp tissue repair and regeneration. The ultrasmall superparamagnetic iron oxide (USPIO)-labeled hydroxyapatite (HA)/silk fibroin (SF) scaffold loaded dental pulp stem cells (DPSCs) was designed. The odontogenic differentiation of DPSCs was investigated, and USPIO was used as magnetic resonance imaging (MRI) contrast agent. The results indicated that scaffolds freeze-dried from SF solution with 0.025 mg/mL USPIO and 5 mg/mL HA showed stable physical properties, accurate MRI images, and low cytotoxicity in vitro. The composite scaffolds were characterized and implanted to the subcutaneous space under the nude mice with tooth fragment. After 2 W of implantation, the relaxation rate (R2) values increased because of incorporation of USPIO, then decreased with the degradation of scaffolds. The tissue regeneration and scaffolds degradation behaviors were monitored by MRI. The expression of dentin sialophosphoprotein (DSPP) and dentin matrix acidic phosphoprotein 1 (DMP1) indicated that DPSCs differentiated into odontoblast-like cells. Revascularization and mineralization evaluated by HE-stained images and alizarin red staining images showed good formation. The USPIO/HA/SF composite scaffold loaded DPSCs promoted the repair and regeneration of dental pulp tissue and provided the ability of imaging noninvasively.

Vip insecticidal proteins are produced by Bacillus thuringiensis (Bt) during its vegetative growth phase. In the present study, Vip3Aa11 and Vip3Aa39 proteins were investigated. These two proteins present 39 amino acid differential sites and they shared 95.06% amino acid sequence similarity. They are effective against some Lepidoptera insect larvae. In a previous study, using artificial diet bioassays, we estimated the LC50 of Vip3Aa11 and Vip3Aa39 strains against Agrotis ipsilon larvae were 73.41 μg/mL (with 95% confidence interval of 2.34–11.19) and 5.43 μg/mL (with 95% confidence interval of 43.20–115.03), respectively. To investigate the response of Agrotis ipsilon transcriptome in defending against Vip3Aa11 and Vip3Aa39 toxins, we performed high-throughput RNA-sequencing on cDNA generated from the midguts of Agrotis ipsilon larvae that consumed a control diet (CK-M-A), Vip3Aa11 (Vip3Aa11-M-A) and Vip3Aa39 (Vip3Aa39-M-A) proteins. We generated about 98.87 Gb bases in total on BGISEQ-500 sequencing platform. After assembling all samples together and filtering the abundance, we got 51,887 unigenes, the total length, average length, N50 and GC content of unigenes are 64,523,651 bp, 1243 bp, 2330 bp and 41.81% respectively. We revealed 558 midgut genes differential expressed in Vip3Aa11-M-A and 65 midgut genes differentially expressed in Vip3Aa39-M-A. The differentially expressed genes were enriched for serine proteases and potential Bt Vip toxin midgut receptor genes. Eleven serine proteases related genes and 13 Bt toxin potential receptor genes with differential expression were found. Based on transcriptome profiling, we focused on validation the sensitivity of these two Vip3Aa proteins to trypsin and their binding properties to Agrotis ipsilon midgut BBMV (Brush Border Membrane Vesicles). The results show that the sensitivity of the two proteins to trypsin is similar. Binding experiments revealed that both proteins can bind to Agrotis ipsilon midgut BBMV, and there is a competitive binding between them. This transcriptome dataset provided a comprehensive sequence resource of Agrotis ipsilon and provides a foundation for comparative studies with other species of insects.

Targeted delivery of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR‐associated protein 9 (Cas9) system to the receptor cells is essential for in vivo gene editing. Exosomes are intensively studied as a promising targeted drug delivery carrier recently, while limited by their low efficiency in encapsulating of large nucleic acids. Here, a kind of hybrid exosomes with liposomes is developed via simple incubation. Different from the original exosomes, the resultant hybrid nanoparticles efficiently encapsulate large plasmids, including the CRISPR–Cas9 expression vectors, similarly as the liposomes. Moreover, the resultant hybrid nanoparticles can be endocytosed by and express the encapsulated genes in the mesenchymal stem cells (MSCs), which cannot be transfected by the liposome alone. Taken together, the exosome–liposome hybrid nanoparticles can deliver CRISPR–Cas9 system in MSCs and thus be promising in in vivo gene manipulation.

Cumulative MB release of (a) NPsmall, (c) NPbig&thin, and (e) NPbig&thick and hour-by-hour MB release profiles of (b) NPsmall, (d) NPbig&thin, and (f) NPbig&thick, from PBS buffer at 37 °C. Figure S2. Silica content of Panc-1 cells presented in percentages after 6, 18, and 24 hrs incubation with cocktail nanoparticles. Figure S3. IC50 determination of free GM and DTX treatment groups in 4 different cell lines. Figure S4. Cytotoxicity evaluation of NPsmall, NPbig&thin and NPbig&thick in 4 different cells. Figure S5. Cytotoxicity evaluation of Cocktail NPs. Figure S6. The tumor size growth curves during the treatment in (a) AsPC-1 cells, (b) BxPC-3 cells, (c) MIA PaCa-2 cells and (d) Panc-1 cells. Figure S7. The bio-distribution of cocktail NPs presented by Silica percentage in (a) AsPC-1, (b) Panc-1, (c) MIA PaCa-2 and (d) BxPC-3 tumors bearing mice. Table S1-S4. Statistical analysis of tumor weight treated with Cocktail NPs upon different irradiation protocols in AsPC-1, BxPC-3, MIA PaCa-2 and Panc-1 tumor model. (DOCX 1397 kb)

Tooth loss occurs with age and significantly impacts the quality of elderly's life both physically and psychologically. It has been well known that odontogenesis is a complicated process with sequential and reciprocal interactions between epithelial and mesenchymal tissues and different types of dental tissue-derived stem cells involve in it. However, only a small portion of the intricate mechanisms has been defined nowadays. Among them, epigenetics has become an increasingly important mechanism for tooth development and regeneration.This review aims at illustrating the function of epigenetic regulation in odontogenesis, which plays an important role in dental tissue-derived stem cell self-renewal and differentiation nowadays and would be a new strategy for tooth regeneration.In this review, we introduced the natural process of tooth development and the functions of stem cells involved in. Furthermore, we summarized the current knowledge on epigenetic regulation including DNA methylation, histone modification, and non-coding RNAs during odontogenesis, providing the theoretical basis for tooth regeneration.Along with a deeper understanding of odontogenesis, the epigenetic mechanism involved in has become increasingly important. Therefore, it's necessary to further study the functions of epigenetic regulation in tooth development and regeneration, which may make tooth regeneration a reality in the future.

Oral squamous cell carcinoma (OSCC) is one of the most common types of the head and neck cancer. Chemo resistance of OSCC has been identified as a substantial therapeutic hurdle. In this study, we analyzed the role of miR-203 in the OSCC and its effects on cisplatin-induced cell death in an OSCC cell line, Tca8113. There was a significant decrease of miR-203 expression in OSCC samples, compared with the adjacent normal, non-cancerous tissue. After 3 days cisplatin treatment, the survived Tca8113 cells had a lower expression of miR-203 than that in the untreated control group. In contrast, PIK3CA showed an inverse expression in cancer and cisplatin survived Tca8113 cells. Transfection of Tca8113 cells with miR-203 mimics greatly reduced PIK3CA expression and Akt activation. Furthermore, miR-203 repressed PIK3CA expression through targeting the 3'UTR. Restoration of miR-203 not only suppressed cell proliferation, but also sensitized cells to cisplatin induced cell apoptosis. This effect was absent in cells that were simultaneously treated with PIK3CA RNAi. In summary, these findings suggest miR-203 plays an important role in cisplatin resistance in OSCC, and furthermore delivery of miR-203 analogs may serve as an adjuvant therapy for OSCC.

myo-Inositol phosphate synthase (MIPS) is the key enzyme of myo-inositol synthesis, which is a central molecule required for cell metabolism and plant growth as a precursor to a large variety of compounds. A full-length fragment of MfMIPS1 cDNA was cloned from Medicago falcata that is more cold-tolerant than Medicago sativa. While MfMIPS1 transcript was induced in response to cold, dehydration and salt stress, MIPS transcript and myo-inositol were maintained longer and at a higher level in M. falcata than in M. sativa during cold acclimation at 5 °C. MfMIPS1 transcript was induced by hydrogen peroxide (H(2) O(2)) and nitric oxide (NO), but was not responsive to abscisic acid (ABA). Pharmacological experiments revealed that H(2) O(2) and NO are involved in the regulation of MfMIPS1 expression by cold and dehydration, but not by salt. Overexpression of MfMIPS1 in tobacco increased the MIPS activity and levels of myo-inositol, galactinol and raffinose, resulting in enhanced resistance to chilling, drought and salt stresses in transgenic tobacco plants. It is suggested that MfMIPS1 is induced by diverse environmental factors and confers resistance to various abiotic stresses.

Upregulation of miR-34a by p53 is recently believed to be a key mediator in the pro-apoptotic effects of this tumor suppressor. We sought to determine whether restoration of miR-34a levels in p53 deficient cells could rescue the response to DNA damage. Compared with the p53 wildtype U2OS cells, miR-34a expression was much lower in p53 deficient Saos2 cells upon cisplatin treatment. Unexpectedly, delivery of miR-34a in Saos2 cells does not increase the cell sensitivity to apoptosis. This effect was mediated by direct downregulation of SirT1 expression by miR-34a, which in turn increased the NFκB activity. Inhibition of NFκB activity in Saos2 cells by Aspirin sensitized the miR-34a overexpressing cells to cell death. Thus, in tumors with p53 deficiency, miR-34a restoration alone confers drug resistance through Sirt1-NFκB pathway and combination of miR-34a and NFκB inhibitor could be considered as a promising therapeutic strategy.

From its inception, tissue engineering has had three tenets: cells, biomaterial scaffolds and signaling molecules. Among the triad, cells are the center piece, because cells are the building blocks of tissues. For decades, cell therapies have focused on the procurement, manipulation and delivery of healthy cells for the treatment of diseases or trauma. Given the complexity and potential high cost of cell delivery, there is recent and surging interest to orchestrate endogenous cells for tissue regeneration. Biomaterial scaffolds are vital for many but not all, tissue-engineering applications and serve to accommodate or promote multiple cellular functions. Signaling molecules can be produced by transplanted cells or endogenous cells, or delivered specifically to regulate cell functions. This review highlights recent work in tissue engineering and cell therapies, with a focus on harnessing the capacity of endogenous cells as an alternative or adjunctive approach for tissue regeneration.

Centipedegrass (Eremochloa ophiuroides [Munro] Hack.) is an important warm-season turfgrass and pasture grass. Due to availability of an efficient regeneration system, this study was undertaken to develop an Agrobacterium-mediated transformation system for centipedegrass for potential use in its genetic improvements. Embryogenic calli were co-cultivated with Agrobacterium strain EHA105 harboring pCAMBIA1301, followed by selection of hygromycin B (hyg B)-resistant callus and regenerated plantlets. The transformants were analyzed using PCR, DNA blot, RNA blot and semi-quantitative RT-PCR. Two transformant lines, lines 2 and 6, showed integration and expression of the hyg B resistance gene hpt. Line 2 revealed a single integration of the hpt transgene, while line 6 revealed two integration sites of hpt. Both lines exhibited resistance to hyg B, although line 2 showed higher level of expression of hpt.

Multipotent mesenchymal stem cell (MSC) therapies are being tested clinically for a variety of disorders. However, despite the remarkable clinical advancements in this field, most applications still use traditional culture media containing fetal bovine serum. Platelet-rich plasma (PRP) appears as a novel application for tissue engineering and its effect on bone healing is thought to be mainly dependent on the proliferation promoting function, with the molecular mechanisms largely unknown. In this study, mouse osteogenic progenitor mesenchymal stem cells (MSCs) were cultured in PRP or washed platelet (WPLT)-treated wells or in untreated wells, and analyzed on cycloxygenase 2 (COX2) expression (qRT-PCR), cell growth (MTT assay) and cell differentiation (alkaline phosphatase activity). The results showed that PRP and WPLT stimulated cell growth similarly in the first 6 days, together with the steady induction of COX2 and PGE2. 10 μmol/l celecoxib (an inhibitor of COX2) significantly inhibited the pro-proliferation effects. Interestingly, WPLT had stronger effects than PRP in proliferation at the later time points (6-9 days). ALP activity assay and collagen 1a expression revealed PRP had a mild but statistically significant pro-differentiation effect, while no obvious effects observed in WLPT group. In summary, PRP stimulates initial growth of MSCs in a COX2 partially dependent manner and the less obvious osteogenic differentiation promoting effects of WPLT strongly indicates WPLT rather than the PRP should be the optional choice for expanding MSCs in vitro for clinical use.

Within a tumor, cancer stem cells (CSCs) represent a small yet highly aggressive, metastatic population that enhances the progression of the surrounding non-CSCs. Here we define lysophosphatidic acid (LPA) as a primary factor produced by pancreatic CSCs that converts non-CSCs to a stem-like fate. LPA promotes increased anchorage independence, drug resistance, self-renewal and stem gene expression including Oct4 and Nanog. Cancer stem cells were found to express high levels of the LPA producing enzyme cPLA2 explaining their production of LPA. Pancreatic cancer cells exposed to the drug Gemcitabine showed a dramatic upregulation of the LPA receptor 4 enabling the cells to become drug resistant and tolerant to a range of microenvironmental stresses. These finding indicate that LPA is a bioactive lipid produced by CSCs that converts surround pancreatic tumor cells to a highly aggressive state. Therapeutic intervention and/or microenvironmental stress will induce non-CSCs to express LPA Receptor 4, thereby making them stress tolerant and drug resistant. Evidence is provided that targeting the LPA receptor pathway may reverse this paracrine effect and thereby reverse cancer progression and drug resistance. Citation Format: Taha Rakhshandehroo, Jiali Tan, Brandon Grosshart, David Cheresh. Cancer stem cell-produced lysophosphatidic acid reprograms surrounding tumor cells to a stem-like state [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-049.

Osteoporosis is a common disease that affects patient quality of life, especially among the elderly population. Although inflammation contributes significantly to osteoporosis, the underlying mechanism is unclear. In this study, we found that tumor necrosis factor (TNF)-α, an inflammatory environment mimic, inhibits osteogenesis of bone mesenchymal stem cells (BMSCs), induces miR-146a and decreases Smad4. Moreover, overexpression of miR-146a inhibited the osteogenic ability of BMSCs, whereas blocking miR-146a partially rescued the osteogenesis deficiency under TNF-α treatment. Molecularly, miR-146a decreased Smad4 expression at the protein level by binding to an element located in the Smad4 3′-untranslated region, and restoration of Smad4 reversed the inhibitory effects of miR-146a on osteogenesis. Together, our results showed that the inflammatory environment mimic TNF-α inhibits osteogenesis via upregulation of miR-146a and subsequent downregulation of Smad4, thus suggesting that therapeutic manipulation of miR-146a maybe a potential strategy to improve osteogenesis in the context of osteoporosis.