Low-intensity pulsed ultrasound promotes osteoarthritic cartilage regeneration by BMSC-derived exosomes via modulating the NF-κB signaling pathway.

Osteoarthritis is the most common disabling joint disease throughout the world, and the effect of therapy on its course is still unsatisfactory in clinical practice. Recent studies have shown that mesenchymal stem cell (MSC)-derived exosomes can promote cartilage repair and regeneration in osteoarthritis, indicating that these exosomes could be a novel and promising strategy for treating osteoarthritis. This study investigated whether low-intensity pulsed ultrasound (LIPUS) enhances the effects of bone marrow MSC (BMSC)-derived exosomes on cartilage regeneration in osteoarthritis and examined the underlying mechanism. Our results revealed that BMSC-derived exosomes display the typical morphological features of exosomes. LIPUS-mediated BMSC-derived exosomes promoted cartilage regeneration, increased chondrocyte proliferation and extracellular matrix synthesis, suppressed inflammation, and inhibited the interleukin (IL)-1ß-induced activation of the nuclear factor kappa B (NF-κB) pathway. In brief, LIPUS enhances the promoting effects of BMSC-derived exosomes on osteoarthritic cartilage regeneration, mainly by strengthening the inhibition of inflammation and further enhancing chondrocyte proliferation and cartilage matrix synthesis. The underlying mechanism could be related to the inhibition of the IL-1ß-induced activation of the NF-κB pathway.

Downregulation of AATK mediates microRNA-558-induced resistance of A549 cells to radiotherapy.

The deregulation of microRNAs (miRNAs) is often implicated in the control of sensitivity to radiotherapy. The objective of the present study was to identify the association between miR­558 and apoptosis­associated tyrosine kinase (AATK), and their importance in regulating the development of resistance to radiotherapy. The current study demonstrated that AATK, a radiosensitization-associated gene, is a target of miR­558 in lung cancer cells, using in silico analysis and a luciferase reporter system. Furthermore, it was determined that transfection of 30 or 50 nM miR­558 mimics and AATK specific siRNA markedly suppressed the mRNA and protein expression of AATK. To determine whether miR­558 was required for lung cancer cell radioresistance, A549 cells were treated with different doses of ionizing radiation, from 0 to 10 Gy, following transfection with miR­558 mimics or AATK specific siRNA. It was determined that the administration of miR­558 mimics or AATK specific siRNA alone did not significantly alter the survival rate of the cells. By contrast, in the cells exposed to 4, 6 or 8 Gy, the administration of miR­558 mimics or AATK specific siRNA significantly promoted cell survival rate and overexpression of AATK reversed this effect. In conclusion, these data demonstrate that the miR­558/AATK cascade is important for the radiosensitization of lung cancer cells and may be a potential radiotherapy target.

Serum miR-152, miR-148a, miR-148b, and miR-21 as novel biomarkers in non-small cell lung cancer screening.

Lung cancer, predominantly by non-small cell lung cancer (NSCLC), is the leading cause of cancer-related deaths over the world. Late diagnosis is one of important reasons for high mortality rate in lung cancer. Current diagnostic approaches have disadvantages such as low accuracy, high cost, invasive procedure, etc. MicroRNAs were previously proposed as promising novel biomarkers in cancer screening. In this study, we evaluated the predictive power of four candidate miRNAs in NSCLC detection. Our study involved 152 NSCLC patients and 300 healthy controls. Blood samples were obtained from the total 452 subjects. After miRNA extraction from serum, the expression of miRNAs in cases and controls were quantified by qRT-PCR and normalized to the level of U6 small RNA. Statistical analyses were performed to compare miRNA levels between cases and controls. Stratified analyses were employed to compare miRNA levels in NSCLC patients with different clinical characteristics. Serum miR-148a, miR-148b, and miR-152 were significantly downregulated in NSCLC patients. However, overexpression of serum miR-21 was observed in NSCLC patients. The combination of four candidate miRNAs exhibited the highest predictive accuracy in NSCLC screening compared with individual miRNAs (AUC = 0.97). Low level of miRNA-148/152 members may associate with advanced stage, large tumor size, malignant cell differentiation, and metastasis. High expression of miR-21 was possibly correlated with large size tumor and advanced cancer stage. Our results showed the dysregulation of miR-148/152 family and miR-21 in NSCLC patients. Hence, the four candidate miRNAs have great potential to serve as promising novel biomarkers in NSCLC screening. Further large-scale studies are needed to validate our results.

Using siRNA in prophylactic and therapeutic regimens against SARS coronavirus in Rhesus macaque.

Development of therapeutic agents for severe acute respiratory syndrome (SARS) viral infection using short interfering RNA (siRNA) inhibitors exemplifies a powerful new means to combat emerging infectious diseases. Potent siRNA inhibitors of SARS coronavirus (SCV) in vitro were further evaluated for efficacy and safety in a rhesus macaque (Macaca mulatta) SARS model using clinically viable delivery while comparing three dosing regimens. Observations of SARS-like symptoms, measurements of SCV RNA presence and lung histopathology and immunohistochemistry consistently showed siRNA-mediated anti-SARS efficacy by either prophylactic or therapeutic regimens. The siRNAs used provided relief from SCV infection-induced fever, diminished SCV viral levels and reduced acute diffuse alveoli damage. The 10-40 mg/kg accumulated dosages of siRNA did not show any sign of siRNA-induced toxicity. These results suggest that a clinical investigation is warranted and illustrate the prospects for siRNA to enable a massive reduction in development time for new targeted therapeutic agents.

Simultaneous detection of seven mutations with seven forward primers and one common reverse primer in a single PCR step.

A new PCR method is described here to amplify simultaneously several fragments sharing the same template in a single PCR step with a hemi-primer. Using this method, we successfully detected various point mutations in tissue plasminogen activator (tPA) mutants.

[A method for preparation of genomic DNA from Grifola frondosa and construction of a genomic library].

Grifola frondosa, is a valuable medicinal fungus. High quality total genomic DNA is difficult to prepare due to its high polysaccharide content. A method for the preparation of Grifola frondosa total genomic DNA and construction of Grifola frondosa, genomic library is described. Genomic DNA prepared by this method is digested by Sau3A I restriction enzyme. Constructed genomic library give a titer of 2 x 10(5) transformants/50mg , with a average insert size of 14kb. This has paved way for the cloning of other Grifola frondosa genes and molecular biology studies.

[Transferring of multiple insect-resisting genes into two-line genetic male sterile rice Pei'ai 64S by particle bombardment].

Transformation of two-line genetic male sterile indica rice variety Pei'ai 64S was conducted by particle bombardment. Three insect-resisting genes ligased into plasmid vector pKC-3 was introduced into calli derived from mature embryo and a total of 33 transgenic plants had been obtained. The PCR and Southern blot analysis of each different gene in R0 plants and also PCR analysis of each different gene in R1 plants showed the integration of three insect-resisting genes into the genome of transgenic plants which could stably pass down.

[An efficient approach to gene discovery--normalization of cDNA library by reassociation].

The cDNA library normalized by reassociation is a newly-developed, effective platform for EST acquisition and gene discovery. This paper presents the principle, procedure, comparison, deficiencies, application and future of the technique of the normalization.