Effects of short-term inflammatory and/or hypoxic pretreatments on periodontal ligament stem cells: in vitro and in vivo studies.

In this study, we extensively screened the in vitro and in vivo effects of PDLSCs following short-term inflammatory and/or hypoxic pretreatments. We found that the 24-h hypoxic pretreatment of PDLSCs significantly enhanced cell migration and improved cell surface CXCR4 expression. In addition, hypoxia-pretreated PDLSCs exhibited improved cell colony formation and proliferation. Cells that were dually stimulated also formed more colonies compared to untreated cells but their proliferation did not increase. Importantly, the hypoxic pretreatment of PDLSCs enhanced cell differentiation as determined by elevated RUNX-2 and ALP protein expression. In this context, the inflammatory stimulus impaired cell OCN protein expression, while dual stimuli led to decreased RUNX-2 and OCN mRNA levels. Although preconditioning PDLSCs with inflammatory and/or hypoxic pretreatments resulted in no differences in the production of matrix proteins, hypoxic pretreatment led to the generation of thicker cell sheets; the inflammatory stimulus weakened the ability of cells to form sheets. All the resultant cell sheets exhibited clear bone regeneration following ectopic transplantation as well as in periodontal defect models; the amount of new bone formed by hypoxia-preconditioned cells was significantly greater than that formed by inflammatory stimulus- or dual-stimuli-treated cells or by nonpreconditioned cells. The regeneration of new cementum and periodontal ligaments was only identified in the hypoxia-stimulus and no-stimulus cell groups. Our findings suggest that PDLSCs that undergo short-term hypoxic pretreatment show improved cellular behavior in vitro and enhanced regenerative potential in vivo. The preconditioning of PDLSCs via combined treatments or an inflammatory stimulus requires further investigation.

Preliminary Application of Precision Genomic Medicine Detecting Gene Variation in Patients with Multifocal Osteosarcoma.

OBJECTIVES: The purpose of this study was to present our clinical experience of treating multifocal osteosarcoma (MFOS) in our center and gain more insight into the biology of this rare condition; in particular, to address with the help of precision genomic medicine the issue of whether the multiple osteosarcoma (OS) lesions in such patients are multi-centric or originate from one primary lesion and metastasize to other sites. Finally, we aimed to identify particular gene phenotypes and mutations that differentiate MFOS from OS with only one tumor. METHODS: Clinical data of patients with MFOS treated at our center between June 2007 and October 2014 were collected and analyzed retrospectively. High throughput sequencing of the whole exome of normal tissue and multiple lesions had been performed on samples from two patients (HJF and JZ) diagnosed in 2014. To explore the particular gene phenotype and clinical significance of MFOS, these sequencing results were analyzed and compared with those from patients with osteosarcoma in a single site. Seven patients with MFOS (three male and four female; average age 19.71 ± 3.35 years were enrolled in this study. Two of these patients declined treatment and died after 4 and 6 months, respectively. The remaining patients received standard treatment comprising neoadjuvant chemotherapy, surgery and chemotherapy. The chemotherapy regimen was lobaplatin (45 mg/m(2) ), doxorubicin (60 mg/m(2) ) and ifosfamide (12 g/m(2) ). Patients were followed up every 3 months after completing treatment and evaluated by the Enneking and Response Evaluation Criteria in Solid Tumors scoring systems. RESULTS: Up to the last follow-up on 1 December 2015, three patients were still alive. The event-free survival ranged from 4 to 144 weeks (median, 50.14 weeks), the mean (±SD) being 55.45 ± 45.47 weeks. Overall survival ranged from 16 to 388 weeks (median, 89 weeks; mean ± SD, 118.7 ± 147.7 weeks). The rates of mutation of the targeted drug-related genes were 133.5% ± 3.0% in the proximal tibia lesion and 113.1% ± 1.9% in the distal femur of patient HJF (P < 0.01) and 136.1% ± 10.8% in the proximal tibial lesion and 122.3% ± 5.5% in the proximal humerus of patient JZ (P = 0.0335). Furthermore, there were several anti-oncogenes in the somatic copy number variation lists analyzed from the two patients, especially TP53. However, no kataegis was found. CONCLUSIONS: Early and radical surgery accompanied by appropriate chemotherapy is the optimal means of treating MFOS. These patients may benefit from precision genomic medicine.

Landscape of RNAs in human lumbar disc degeneration.

Accumulating evidence indicates noncoding RNAs (ncRNAs) fine-tune gene expression with mysterious machinery. We conducted a combination of mRNA, miRNA, circRNA, LncRNA microarray analyses on 10 adults' lumbar discs. Moreover, we performed additional global exploration on RNA interacting machinery in terms of in silico computational pipeline. Here we show the landscape of RNAs in human lumbar discs. In general, the RNA-abundant landscape comprises 14,635 mRNAs (37.93%), 2,059 miRNAs (5.34%), 18,995 LncRNAs (49.23%) and 2,894 (7.5%) circRNAs. Chromosome 1 contributes for RNA transcription at most (10%). Bi-directional transcription contributes evenly for RNA biogenesis, in terms of 5' to 3' and 3' to 5'. Despite the majority of circRNAs are exonic, antisense (1.49%), intergenic (0.035%), intragenic (1.69%), and intronic (6.29%) circRNAs should not be ignored. A single miRNA could interact with a multitude of circRNAs. Notably, CDR1as or ciRS-7 harbors 66 consecutive binding sites for miR-7-5p (previous miR-7), evidencing our pipeline. The majority of binding sites are perfect-matched (78.95%). Collectively, global landscape of RNAs sheds novel insights on RNA interacting mechanisms in human intervertebral disc degeneration.