10.3969/j.issn.2095-4344.2013.26.004

Objective To construct MIA3 psicheck2 wild-type and mutant vectors targeting miR-374b,and provide the previous guarantee for the dual luciferase reporter assay.Methods The amplification primer was firstly designed according to mice MIA3-3'UTR sequence information,mice whole blood genomic DNA was taken as the template for PCR amplification of MIA3-3'UTR sequence,and the PCR product was cloned into psicheck2 dual luciferase reporter vector.Then,mutant primer was designed to mutate the MiR-374b seed sequence target TATTATA into AAATTAT so as to construct mutant vector.At last,the vector enzyme digestion evaluation and sequencing method was used to evaluate the constructed vectors.Results It could be seen from the analysis of agarose electrophoresis that the PCR amplification size of vector was consistent with the theoretical size.DNA sequencing evaluation showed that the MIA3-3'UTR-WT vector had been constructed successfully.The construction of mutant vector has successfully mutated the MiR-374b seed sequence target TATTATA into AAATTAT.Conclusion The successful construction of the vector will lay a foundation for the further evaluation on whether there is an actual binding site between the miR-374b and the chondrogenic differentiation-related target gene MIA3.

BACKGROUND:The effects of advanced glycation end products (AGEs) on osteoclast-induced bone resorption is controversial and the underlying mechanisms remain unclear. Most of the studies indicate that AGEs can enhance bone resorption, while some othersshowthe opposite effects.
OBJECTIVE:To investigate the effects of AGEs on osteoclast-induced inorganicmatrixdissolution and organic componentdegradation and the underlying mechanisms.
METHODS:RAW 264.7 cels were induced to generate osteoclasts,and AGEs (50-400 μg/mL) or control-bovine serum albumin (100 μg/mL) was added since the beginning of the induction. The effect of AGEs on bone resorption was evaluated by analyzing the area of resorption pits on the Osteo Assay Surface plates and the expression of cathepsin K. Furthermore, the number of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cels, nuclei per osteoclasts and the expression of integrinανβ3were detected.
RESULTS AND CONCLUSION:The area of resorption pits and expression of cathepsin K in AGEs groups were significantly decreased compared withthecontrol group, and this inhibiting effect became more obvious with the increase of AGEs concentration. TRAP staining also showed that number of TRAP-positivemultinucleated celsand nuclei per osteoclast were significantly reduced in an AGE dose-dependent manner. Quantitative PCR revealed that the expression of integrin ανβ3decreased significantly with the extension of AGEs incubation time. These data indicate that AGEs can exert inhibitory effects on organic and inorganicmatrixdegradation induced by osteoclasts. The underlying mechanism may be involved in the inhibitory effects of AGEs on directed differentiation and cel fusion of osteoclast precursor cels, and migration and adhension of osteoclasts.

STUDY DESIGN: A prospective cross-sectional study. PURPOSE: To evaluate the risk factors associated with the severity of pain intensity in patients with non-specific low back pain (NSLBP) in Southern China. OVERVIEW OF LITERATURE: Low back pain (LBP) is the leading cause of activity limitation and work absence throughout the world, so a firm understanding of the risk factor associated with NSLBP can provide early and prompt interventions that are aimed at attaining long-term results. METHODS: Participants were recruited from January 2014 to January 2016 and were surveyed using a self-designed questionnaire. Anonymous assessments included Short Form 36-Item Health Survey (SF-36) and Visual Analogue Scale (VAS). The association between the severity of NSLBP and these potential risk factors were evaluated. RESULTS: A total of 1,046 NSLBP patients were enrolled. The patients with primary school education, high body mass index (BMI), those exposed to sustained durations of driving and sitting, smoking, recurrent LBP had increased VAS and Oswestry Disability Index (ODI) scores with lower SF-36 scores (p<0.01). Workers and drivers compared with waiters and patients who lifted >10 kg objects in a quarter of their work time for >10 years had higher VAS and ODI scores with lower SF-36 scores (p<0.01). Multiple logistic regression showed lower levels of education, LBP for 1–7 days, long-lasting LBP in last year, smoking, long duration driving, and higher BMI were associated with more severe VAS score. CONCLUSIONS: The severity of NSLBP is associated with lower levels of education, poor standards of living, heavy physical labor, long duration driving, and sedentary lifestyle. Patients with recurrent NSLBP have more severe pain. Reducing rates of obesity, the duration of heavy physical work, driving or riding, and attenuating the prevalence of sedentary lifestyles and smoking may reduce the prevalence of NSLBP.
Anonyms and Pseudonyms ; Body Mass Index ; China ; Cross-Sectional Studies ; Education ; Health Surveys ; Humans ; Logistic Models ; Low Back Pain ; Obesity ; Prevalence ; Prospective Studies ; Risk Factors ; Sedentary Lifestyle ; Smoke ; Smoking