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Tumour-associated macrophages (TAMs) are thought to contribute to oral squamous cell carcinoma (OSCC) initiation and progression. However, the underlying mechanism through which TAMs foster OSCC progression is still unclear. This study intended to determine whether there are exclusively exosomal miRNAs-derived macrophages that are functionally necessary for OSCC progression. The phenotype of TAM recruitment in OSCC tissue samples was assessed, subsequently identifying the influence of M2 macrophages and exosomes derived from M2 macrophages on OSCC proliferation and tumorigenesis in vitro and in vivo. CD68 and CD163, the specific markers of M2 type macrophages, were upregulated in TAMs presented in intra-cancer tissues. M2 macrophages and M2 macrophage-derived exosomes (M2 exos) both can promote OSCC growth and tumorigenicity. An exosomal RNA-seq analysis was conducted to predict regulatory exosomal miRNAs related to OSCC growth, which determined miR-31-5p and LATS2 for subsequent experiments. Mechanistically, miR-31-5p was delivered to recipient OSCC cells through M2 exos and complementary pairing with the large tumor suppressor 2 (LATS2) coding sequence, thus suppressing the expression of LATS2 and inactivation the Hippo signaling pathway to support OSCC growth. Collectively, our findings demonstrate that M2 macrophage-derived exosomal miR- 31-5p can make tumor suppressor LATS2 gene inhibited and facilitate the progression of OSCC via inhibiting the Hippo signaling pathway, which possibly provides new targets for the molecular therapy of OSCC.

Reasonable optimization of development well type is the prerequisite to realize the economic and effective development of coalbed methane (CBM). However, the existing CBM development well type optimization methods are disadvantageous in many aspects. CBM is a kind of typical unconventional natural gas resource, and its study focuses on single-well evaluation. In this paper, the assessment method and the evaluation index (EI) for CBM development well type optimization based on the long-run marginal cost (LRMC) were constructed on the basis of CBM characteristics from the perspective of economic feasibility. Then, the corresponding decision making criteria were established. Finally, this newly developed method was applied to the well-type optimization of different development blocks in two important CBM development basins in China. And the following research and application results were obtained. First, under the current technical and economic conditions, both directional well and horizontal well are economically feasible to the development of Block A in the Qinshui Basin and Block B in eastern margin of the Ordos Basin, and the economically optimal well type is a horizontal well in Block A and a directional well in Block B. Second, compared with single-well steady daily gas production and single-well investment, the evaluation results on the schemes of different well types present critical value, above which the economic benefit of different schemes are equivalent. Third, EI of different well types is affected by many factors, e.g. geological characteristic of gas reservoirs, single-well production profile, CBM selling price, investment and cost. Therefore, it is necessary to carry out dynamic analysis and update in time based on the change of various influential factors so as to ensure the rationality of decision-making. In conclusion, this newly developed method is intrinsically consistent with traditional evaluation methods, and it is more advantageous with strong operability, intuitive evaluation results and quick and accurate optimization of CBM development well types. Keywords: Coalbed methane (CBM), Long-run marginal cost (LRMC), Development well, Well-type optimization, Directional wells, Horizontal wells, Surface development, Decision making criteria, Critical point

Background Endoplasmic reticulum stress (ER stress) fosters cancer cell escape from immune surveillance and upregulate PD-L1 expression, but the mechanisms remain unclear. Methods We analyzed protein levels by immunofluorescence and Western blotting, RNA levels by qRT-PCR. Exosomes were injected intravenously through the tail vein into 6-week-old nude mice once every other day for a total of 10 injections Results Expression of some ER stress markers, including GRP78 (glucose-regulated protein 78), ATF6 (activating transcription factor 6) and PERK (PKR-like endoplasmic reticulum kinase), was upregulated in OSCC tissues and correlated with poor overall survival. The level of ER stress-related proteins positively correlated with a cluster of PD-L1 expression and macrophage infiltration in OSCC tissues. PD-L1 expression in OSCC tissues was negatively correlated with cumulative survival. Incubation with Exo-ER upregulated PD-L1 levels in macrophages in vitro and vivo, and upregulation of PD-L1 promoted macrophage polarisation towards the M2 subtype. Conclusions ER stress induced exosome secretion by OSCC cells and PD-L1 expression in macrophages to promote M2 macrophage differentiation. A novel exosome-modulated mechanism was delineated for OSCCs-macropahge crosstalk that drove tumor growth and should be explored for its therapeutic utility.

Aerobic vaginitis (AV) can occur if normal vaginal microflora are dominated by aerobic bacteria, seriously affects not only female health, but also fetal health while they are pregnant. Besides, pregnant status also aggravates the symptoms and consequences of the infection. Here, we infected pregnant BALB/c mice with Escherichia coli on embryonic day 4.5 (E4.5) (study group), and administered an equivalent volume of phosphate-buffered saline in another cohort of pregnant mice (control group). We recorded the weight of pregnant mice and their fetuses. The maternal and fetal weight of the study group decreased in comparison with that of the control group, whereas the weight of placenta increased in the study group. Then, five genes with significant upregulation and 15 genes with downregulation were screened. Expression of interleukin 4 (IL-4) mRNA in the study group decreased to 18.5%. Enzyme-linked immunosorbent assay results showed IL-4 expression in mouse plasma declined in the study group at E11.5 and E18.5. mRNA expression of chemokine (c-c motif) ligand (CCL)-17, CCL-22, CCL-24, IL-4, Janus Kinase (JAK)-1, signal transducer and activator of transcription (STAT)-6, and GATA-3 showed significant downregulation in placental and uterine tissues. Flow cytometry of primary decidual macrophages (DMs) revealed more M1-like macrophages in the study group. And after addition of IL-4 to DMs, more M1 macrophages polarized to M2 type macrophages. We did not discover bacteria existed in mouse placentas. Our study affords a feasible method for exploring and managing AV during pregnancy.

Clinical and investigational proof indicates that tumour-related macrophages stimulate tumour initiation and development. Consequently, macrophage-derived molecular factors controlling OSCC initiation are not yet completely understood. Here, we show that M2 macrophage–regulated OSCC cell growth and tumorigenicity depend on M2 macrophage-derived exosomes (M2 exo). M2 exo exhibited a high level of miR-31-5p, and the M2 exo-mediated increase in OSCC cell proliferation and growth depended on this miRNA. Mechanistically, miR-31-5p was delivered to OSCC cells through M2 exo and complementary pairing with the LATS2 coding sequence, thus suppressing the expression of LATS2, which has been considered to be a major aspect inhibiting OSCC growth. Collectively, our findings demonstrate that M2 exo participate in OSCC tumorigenesis, and M2 exo mediate their tumour-inducing effect via the miR-31-5p/LATS2 axis of the Hippo signalling pathway in OSCC. This active and mutual communication among OSCC cells and M2 macrophages provides a novel strategy for treating OSCC. Funding Statement: This work was supported by research grants from the National Natural Science Foundation of China (21872026), the Outstanding Young scholar Foundation of Jiangsu Province, China (BK20170106), the Major Projects of Science and Technology Development Fund of Nanjing Medical University (NMUD2019001), the Open Fund of State Key Laboratory of Pharmaceutical Biotechnology, Nan-jing University, China (KF-GN-201903), the Southeast University and Nanjing Medical University Cooperative Research Project, China (2019DN0003), the Open Research Fund of State Key Laboratory of Bioelectronics, Southeast University. Declaration of Interests: None. Ethics Approval Statement: The study procedure was under the approval of the Ethics Committee of Stomatology Hospital Affiliated to Nanjing Medical University, and written informed consent was obtained from every patient.

This study was designed to investigate the protective effect of curcumin againstd-galactose (d-gal)-induced premature ovarian failure (POF) in mice. A mouse POF model was induced by subcutaneous injection ofd-gal (200 mg/kg/day) daily for 42 days. Mice in the curcumin group received bothd-gal treatment and intraperitoneal injection of curcumin (100 mg/kg/day) for 42 days. Ovarian function, oxidative stress and apoptosis were evaluated. The P, E2 and SOD levels were higher, and the FSH, LH and MDA levels were significantly lower in the curcumin group than those in thed-gal group. The proportion of primordial follicles was also significantly higher in the curcumin group than that in thed-gal group. In addition, curcumin treatment afterd-gal administration resulted in significantly lowerSod2,Cat, 8-OhdG, 4-HNE, NTY and senescence-associated protein P16 expression levels, higherAmhexpression levels and less apoptosis in granulosa cells than was observed in thed-gal group. Moreover, the p-Akt, Nrf2 and HO-1 protein expression levels were significantly higher and the apoptosis-related cleaved caspase-3 and -9 protein expression levels were markedly lower in the curcumin group than in thed-gal group. In conclusion, curcumin effectively inhibitedd-gal-induced oxidative stress, apoptosis and ovarian injury via a mechanism involving the Nrf2/HO-1 and PI3K/Akt signaling pathways, suggesting that curcumin is a potential protective agent against POF.

The aim of this study was to investigate the protective role and underlying mechanisms of curcumin on glycerol-induced acute kidney injury (AKI) in rats. Glycerol (10 ml/kg BW, 50% v/v in sterile saline, i.m.) was used to induce AKI, followed by curcumin (200 mg/kg/day, p.o.) administration for 3 days. To confirm renal damage and the effects of curcumin on AKI, serum BUN, Scr, and CK as well as renal SOD, MDA, GSH-Px were measured. Additionally, morphological changes were identified by H&E staining and transmission electron microscopy. The expression of several factors including chemotactic factor MCP-1, proinflammatory cytokines including TNF-α and IL-6, as well as the kidney injury markers, as Kim-1 and Lipocalin-2 were also assessed using q-PCR. Finally, cell apoptosis in renal tissue was detected using in situ TUNEL apoptosis fluorescence staining and expression of proteins associated with apoptotic, oxidative stress and lipid oxidative related signaling pathways were detected using immunohistochemical staining and western blot. The results showed that curcumin exerts renoprotective effects by inhibiting oxidative stress in rhabdomyolysis-induced AKI through regulation of the AMPK and Nrf2/HO-1 signaling pathways, and also ameliorated RM-associated renal injury and cell apoptosis by activating the PI3K/Akt pathway.

To assess the effect of cluster of differentiation (CD47) downregulation on autophagy in hypoxia/reoxygenation (H/R)‑treated H9c2 cardiomyocytes. H9c2 cells were maintained in normoxic conditions (95% air, 5% CO2, 37˚C) without CD47 antibodies, Si‑CD47 or chloroquine (CQ) treatment; H9c2 cells in the H/R group were subjected to 24 h of hypoxia (1% O2, 94% N2, 5% CO2, 37˚C) followed by 12 h of reoxygenation (95% air, 5% CO2, 37˚C). All assays were controlled, triplicated and repeated on three separately initiated cultures. The biochemical parameters in the medium supernatant were measured to evaluate the oxidative stress in cardiomyocytes. The Annexin V‑fluorescein isothiocyanate assay was used to detect the apoptotic rate in the H9c2 cells. Transmission electron microscope, immunofluorescent staining and western blot analysis were performed to detect the effect of the CD47 antibody on autophagic flux in H/R‑treated H9c2 cardiomyocytes. The cardiomyocytic oxidative stress and apoptotic rate decreased and autophagic clearance increased after CD47 downregulation. H/R triggered cell autophagy, autophagosome accumulation and apoptosis in H9c2 cell lines. However, these effects can be attenuated by CD47 downregulation. This study demonstrates its clinical implications in ischemia/reperfusion injury treatment.

Background/Aims: This study investigated the effect of consecutive superovulation on the ovaries and established a premature ovarian failure (POF) model in mice. Methods: The mouse POF model was induced by 5-15 consecutive superovulation treatments with pregnant mare serum gonadotropin (PMSG), human chorionic gonadotropin (HCG) and prostaglandin F2α (PGF2α). Normal adult mice were compared with mice displaying natural ovarian aging. The following serum biochemical parameters were measured: including follicle-stimulating hormone (FSH), luteinizing hormone (LH), progesterone (P), estradiol (E2), inhibin B (INH B), malondialdehyde (MDA), total superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) levels. Follicles were counted using H&E staining. Levels of 8-hydroxyguanosine (8-OhdG), 4-hydroxynonenal (4-HNE), nitrotyrosine (NTY), anti-Mullerian hormone (AMH) and CDKN2A/ p16 (p16) were detected using immunohistochemical staining. Reactive oxygen species (ROS) levels were measured using dihydroethidium (DHE) staining. Cell apoptosis was detected using an in situ TUNEL fluorescence staining assay. Levels of proteins involved in ROS-related pathways and the p16 protein were detected using Western blotting. Sod1, Sod2 and Sod3 mRNA levels were detected using quantitative polymerase chain reaction (Q-PCR). Oocyte quality was evaluated using in vitro fertilization (IVF) and zygote culture. Results: Consecutive superovulation groups presented lower P, E2, SOD, GSH-Px and INH B levels, significantly higher FSH, LH, MDA and ROS levels, and significantly fewer primordial follicles compared with the control group. Consecutive superovulation groups presented significantly increased levels of Sod2, 8-OhdG, 4-HNE, NTY, significantly increased levels of the SIRT1 and FOXO1 proteins, significantly increased levels of the senescence-associated protein p16, as well as decreased AMH, Sod1 and Sod3 levels and increased granulosa cell apoptosis compared with the control group. Conclusion: Consecutive superovulation significantly decreased ovarian function and oocyte quality and increased oxidative stress and apoptosis in the ovary via a mechanism involving the p16 and SIRT1/FOXO1 signaling pathways. These findings suggest that consecutive superovulation may be used to establish a mouse model of ovarian aging.

Based on their respective antitumor and thrombolytic activities, the superantigen staphylococcal enterotoxin C2 (SEC2) and staphylokinase (Sak) were chosen for the construction of the novel chimeric proteins Sak-linker- SEC2 and SEC2-linker-Sak using a linker composed of nine Ala residues. Both chimeric proteins possessed nearly the same PBMC proliferation stimulating activity and antitumor activity as SEC2 and thrombolytic activity as Sak. Neither the SEC2 or Sak component of each chimeric protein affected the activity of the other component. The results presented in this study provide a possible strategy to prevent and cure tumor thrombus.