Mesenchymal stromal/stem cells (MSC) may represent the cell-of-origin for sarcoma development. A collection of human MSCs sequentially mutated with an increasing number of oncogenic hits served to recreate a step-wise process of sarcomagenesis. To identify potential protein targets of interest in the MSC-sarcoma transformation process, quantitative mass spectrometry-based (LC–MS/MS) proteomics was performed. Among the protein hits identified as significantly regulated in the transformation process, ALDH1A3 and CD99 were selected and further studied. Both ALDH1A3 abundance levels and activity were significantly upregulated in early-phase (immortalized) and fully transformed (sarcoma forming) cells as compared to normal MSCs. Inversely, CD99 total protein and cell-surface abundance levels were downregulated in immortalized and transformed MSCs. Downregulated CD99 was also identified in several human bone and soft tissue sarcoma subtypes. Proteomics investigation of a MSC-transformation model of sarcoma has yielded ALDH1A3 and CD99 as potential targets for sarcomagenesis that may contribute to a greater understanding of the disease and the development of novel therapeutic approaches.

Disulfiram (DSF), which is an inhibitor of aldehyde dehydrogenase (ALDH) and approved by the FDA for the treatment of alcoholism previously, has been repurposed for use as a cancer treatment because of its potent effect in preclinical studies. In this study, we found that disulfiram forms potent complexes with copper (DSF/Cu) inhibited cell proliferation, induced apoptosis in human pancreatic cancer cells, which was detected by flow cytometry and western blotting. Meanwhile, autophagy and autophagic flux also clearly observed by transmission electron microscopy, confocal microscopy and flow cytometry. Our results also showed that DSF/Cu induced transcription factor p8 upregulation and PI3K/mTOR signaling pathway activation detected by real-time PCR and western blotting. Additionally, suppression of p8 inactivated the mTOR signaling pathway and autophagic flux maintained. Furthermore, mechanism study indicated that autophagy induced by DSF/Cu was regulated by p8 and was related to PI3K/mTOR/p70S6K signaling pathway in pancreatic cancer cells. Our findings provide insights into the role of p8 in regulating autophagy induced by DSF/Cu effects in pancreatic cancer cells.

Objective: Abnormal lipid metabolism has a close link to the pathophysiology of schizophrenia (SZ). This study mainly aimed to evaluate the association of variants at apolipoprotein A1 (APOA1) and APOA4 with SZ in a Chinese Han population. Methods: The rs5072 of APOA1 and rs1268354 of APOA4 were examined in a case–control study involving 2,680 patients with SZ from the hospital and 2,223 healthy controls screened by physical examination from the community population. The association was estimated with the odds ratio (OR) and 95% confidence intervals (95% CIs) by logistic regression. The APOA1 and APOA4 messenger RNA (mRNA) in peripheral blood leukocytes were measured by real-time PCR and compared between SZ cases and controls. Serum apoA1 levels were detected by turbidimetric inhibition immunoassay and high-density lipoprotein cholesterol (HDL-C) levels were detected by the homogeneous method. Results: Both of the rs5072 of APOA1 and rs1268354 of APOA4 had statistically significant associations with SZ. After adjustment for age and sex, ORs (95% CIs) of the additive model of rs5072 and rs1268354 were 0.82 (0.75–0.90) and 1.120 (1.03–1.23), and p-values were 3.22 × 10−5 and 0.011, respectively. The association of rs5072 with SZ still presented statistical significance even after Bonferroni correction (p-value×6). SZ patients during the episode presented lower levels of apoA1, HDL-C, mRNA of APOA1 common variants and transcript variant 4, and APOA4 mRNA than controls (p < 0.01) while SZ patients in remission showed a significantly decreased APOA1 transcript variant 3 expression level and increased APOA4 mRNA expression level (p < 0.01). mRNA expression levels of APOA1 transcript variant 4 significantly increased with the variations of rs5072 in SZ during the episode (p trend = 0.017). After the SZ patients received an average of 27.50 ± 9.90 days of antipsychotic treatment, the median (interquartile) of serum apoA1 in the SZ episode significantly increased from 1.03 (1.00.1.20) g/L to 1.08 (1.00.1.22) g/L with the p-value of 0.044. Conclusion: Our findings suggest that the genetic variations of APOA1 rs5072 and APOA4 rs1268354 contribute to the susceptibility of SZ, and the expression levels of APOA1 and APOA4 mRNA of peripheral blood leukocytes decreased in SZ patients during the episode while APOA4 increased after antipsychotic treatment.

Background Clinical applications have shown extracellular vesicles (EVs) to be a major paracrine effector in therapeutic responses produced by human mesenchymal stromal/stem cells (hMSCs). As the regenerative capacity of EVs is mainly ascribed to the transfer of proteins and RNA composing its cargo, and to the activity attributed by the protein surface markers, we sought to profile the protein composition of small EVs released from hMSCs to identify hMSC-EV biomarkers with potential clinical relevance. Methods Small EVs were produced and qualified from five human bone marrow MSC donors at low passage following a 48-h culture in exosome-depleted medium further processed by steps of centrifugation, filtration, and precipitation. Quantitative proteomic analysis comparing the protein profile of the EVs released from hMSCs and their parental cell was conducted using tandem mass tag labeling combined to mass spectrometry (LC-MS/MS) to identify enriched EV protein markers. Results Nanoparticle tracking analysis showed no differences in the EV concentration and size among the five hMSC donors (1.83 × 10 10 ± 3.23 × 10 9 /mL), with the mode particle size measuring at 109.3 ± 5.7 nm. Transmission electron microscopy confirmed the presence of nanovesicles with bilayer membranes. Flow cytometric analysis identified commonly found exosomal (CD63/CD81) and hMSC (CD105/CD44/CD146) markers from released EVs in addition to surface mediators of migration (CD29 and MCSP). Quantitative proteomic identified 270 proteins significantly enriched by at least twofold in EVs released from hMSCs as compared to parental hMSCs, where neuropilin 1 (NRP1) was identified among 21 membrane-bound proteins regulating the migration and invasion of cells, as well as chemotaxis and vasculogenesis. Validation by western blot of multiple batches of EVs confirmed consistent enrichment of NRP1 in the nanovesicles released from all five hMSC donors. Conclusion The identification and verification of NRP1 as a novel enriched surface marker from multiple batches of EVs derived from multiple hMSC donors may serve as a biomarker for the assessment and measurement of EVs for therapeutic uses.

Denervated-dependent skeletal muscle atrophy (DSMA) is considered to be the neuro-disconnection of skeletal muscle. This study aimed to investigate the protective effects of Buyang Huanwu Tang (BYHWT) on the DSMA and clarify associated molecular and genetic mechanisms. DSMA rat models were established according to the previously published study and divided into Model group and BYHWT group. Meanwhile, normal rats were assigned as Normal control (NC) group. Hematoxylin and eosin (HE) staining was used to examine inflammatory responses. Motor endplate activity was evaluated with wholemount acetylcholinesterase (AChE) staining. Mass-spectrometry analysis was conducted to compare differentially expressed proteins. RNAs were prepared and applied to gene functional analysis. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were employed to analyze biological functions. The results indicated that BYHWT remarkably alleviated inflammatory responses and significantly improved motor endplate function, compared to that in DSMA Model rats (P<0.05). In BYHWT group, there were 393 differentially up-regulated and 576 differentially down-regulated molecules compared to that in Model group. Comparing to Model group, the cellular response to interferon-gamma, integral component of plasma membrane and voltage-gated potassium channel activity genes in BYHWT group were the most biological process (BP), cellular component (CC) and molecular function (MF) differential genes, respectively. Fructose/mannose metabolism and glycerolipid metabolism KEGG signaling pathways illustrated the most significant enrichment of differentially expressed genes. In conclusion, BYHWT alleviated the inflammations and improved the motor endplate function of DSMA rats by activating cellular response to interferon-gamma, integral component of plasma membrane and voltage-gated potassium channel activity genes and associated signaling pathways.

Intracerebral hemorrhage (ICH) has high morbidity and mortality, with no effective treatment at present. One possible therapeutic strategy involves the use of mesenchymal stem cells (MSCs), which have shown promise in experimental models and have great potential for treating nervous illnesses in humans. However, many deficiencies in MSC treatment still need to be addressed, including their poor survival rate post-transplantation. Previously, we reported that the microRNA-21 (miR-21) is downregulated in ICH patients' blood and brain tissue. In this study, we aimed to examine its role and therapeutic efficacy in ICH using miR-21-overexpressing MSCs. We found that this microRNA can enhance MSC survival and recovery of neurological function in ICH rats. Its mechanism of action involves reduced neuronal apoptosis. In addition, we demonstrated that miR-21 can be transported to neurons through exosomes derived from MSCs and that it can target transient receptor potential melastatin 7 (TRPM7) to alleviate neuronal injury following ICH. We also observed that the NF-κB pathway is involved in the regulation of miR-21 in neural cells. In conclusion, miR-21 significantly enhances the survival of MSCs in ICH, and miR-21-overexpressing MSCs clearly improved neurological function in ICH rats. Transplantation of miR-21-overexpressing MSCs may, therefore, provide an effective strategy for neuroprotection and treatment of cerebrovascular diseases.

Background: MiR-320a has been found playing important role in tumor progression. However, the molecular mechanisms of miR-320a in oral tongue squamous cell carcinoma (OTSCC) remain unknown. Methods: MicroRNA expression was detected by miRNA microarray in healthy people and OTSCC patients. RT-PCR was further used to verify microarray data. The function of miR-320a was measured by wound healing assay, transwell assay and Western blot in OTSCC cells. The miR-320a target gene was tested by using a luciferase reporter assay, RT-PCR and Western blot. Results: A total of 18 miRNAs showed altered expression in OTSCC patients, and miR-320a was markedly decreased (P<0.05). In addition, migration and invasion abilities of OTSCC cells were decreased in the miR-320a group (P<0.05); the luciferase activity of the MRP2-3’-UTR plasmid was inhibited following miR-320a binding (P<0.05). Conclusion: MiR-320a expression was significantly decreased in patients with OTSCC, indicating that miR-320a would be a potential antioncogene partly through targeting MRP2, it would become a therapeutic target for OTSCC.

Multipotent mesenchymal stromal cell (MSC) transplantation is proposed as a novel therapy for treating diabetes by promoting the regeneration of damaged islets. The clinical promise of such treatments may be hampered by a high degree of donor-related variability in MSC function and a lack of standards for comparing potency. Here, we set out to identify markers of cultured human MSCs directly associated with islet regenerative function. Stromal cultures from nine separate bone marrow donors were demonstrated to have differing capacities to reduce hyperglycemia in the NOD/SCID streptozotocin-induced diabetic model. Regenerative (R) and non-regenerative (NR) MSC cultures were directly compared using isobaric tags for relative and absolute quantitation (iTRAQ)-based quantitative proteomics. A total of 1,410 proteins were quantified resulting in the identification of 612 upregulated proteins and 275 downregulated proteins by ± 1.2-fold in R-MSC cultures. Elastin microfibril interface 1 (EMILIN-1), integrin-linked protein kinase (ILK), and hepatoma-derived growth factor (HDGF) were differentially expressed in R-MSCs, and Ingenuity Pathway Analyses revealed each candidate as known regulators of integrin signaling. Western blot validation of EMILIN-1, ILK, and HDGF not only showed significantly higher abundance levels in R-MSCs, as compared with NR-MSCs, but also correlated with passage-induced loss of islet-regenerative potential. Generalized estimating equation modeling was applied to examine the association between each marker and blood glucose reduction. Both EMILIN-1 and ILK were significantly associated with blood glucose lowering function in vivo. Our study is the first to identify EMILIN-1 and ILK as prospective markers of islet regenerative function in human MSCs. Stem Cells 2016;34:2249-2255.

The present study was aimed to determine the topographic relationship between root apex of the mesially and horizontally impacted mandibular third molar and lingual plate of mandible. The original cone beam computed tomography (CBCT) data of 364 teeth from 223 patients were retrospectively collected and analyzed. The topographic relationship between root apex and lingual plate on cross-sectional CBCT images was classified as non-contact (99), contact (145) and perforation (120). The cross-sectional morphology of lingual plate at the level of root apex was defined as parallel (28), undercut (38), slanted (29) and round (4). The distribution of topographic relationship between root apex and lingual plate significantly associated with gender, impaction depth, root number and lingual plate morphology. Moreover, the average bone thickness of lingual cortex and distance between root apex and the outer surface of lingual plate were 1.02 and 1.39 mm, respectively. Furthermore, multivariate regression analyses identified impaction depth and lingual plate morphology as the risk factors for the contact and perforation subtypes between root apex and lingual plate. Collectively, our findings reveal the topographic proximity of root apex of impacted mandibular third molar to the lingual plate, which might be associated with intraoperative and postoperative complications during tooth extraction.