Objective To investigate the effects of aggrecanase-2 knockdown in chondrocytes from rheumatoid arthritis patient by shRNA infection.Methods Cartilage harvested from rheumatoid arthritis patients who underwent total knee arthroplasty was digested by pancreatin and type Ⅱ collagen enzyme to obtain chondrocytes.Then chondrocytes were cultured and passaged to second or third generation.After aggrecanase-2 shRNA5,aggrecanase-2 shRNA6,aggrecanase-2 shRNA7,aggrecanase-2 shRNA8 infection,growth and morphological changes of the chondrocytes were examined.To select the best target sequence,mRNA expression of aggrecanase-2 and aggrecan was detected by RT-qPCR assay on day 2,5,10,represented with Ct (Cycle threshold) value.Expression of aggrecan protein was detected by immunocytochemistry,represented with IOD (integral optical density) value.Results aggrecanase-2 knockdown had no obvious effects on the morphology and growth of the chondrocytes.MOI (multiplicity of infection) was 100,50,25,and infection efficiency was 90％,60％,30％ with the corresponding viral load of 200 μl,100 μl,50 μl after transfection.mRNA expression of aggrecanase-2 was suppressed significantly,especially the group of shRNA5 which suppressed aggrecanase-2 expression from 0.876 3±0.115 6 to 0.069 9±0.015 1 (P ＜ 0.05).mRNA and protein expression of aggrecan were significantly upregulated after infection.mRNA expression of aggrecan increased from 0.992 1±0.201 3 to 3.049 2±0.278 2 (P ＜ 0.05) and protein expression of aggrecan increased from 496.160 5± 225.673 7 to 4 525.433 0±1 131.813 0 (P ＜ 0.05).Conclusion aggrecanase-2 suppression in chondrocytes by lentivirius infection is an effective method to protect the expression of aggrecan.

Xuzhou Medical University, relying on the National Demonstration Center for Experimental Basic Medical Science Education and the scientific research platform of Jiangsu Key Laboratory of Brain Disease Bio-information, organizes clinical medical undergraduates to participate in the innovative training program of scientific research practice. Taking the research on signal transduction of ischemic nerve cell injury as an example, this paper studies the innovative practice training on experimental ability and scientific research thinking of medical undergraduates, and preliminarily discusses the promoting effect of innovative practice training on teaching reform of biochemistry and molecular biology. Innovative training in scientific research practice provides a new teaching method, which effectively promotes the cultivation of scientific research quality of medical undergraduates and the teaching reform of theoretical courses.

Objective To investigate the effects of low-frequency sound waves (16~160 Hz) of somatosensory music on healthy people’s microcirculation in kidney and bladder meridians;To discuss the collateral channels biophysical mechanisms in somatosensory music treatment. Methods Low-frequency sound waves of somatosensory music were played near the 30 healthy people’s kidney and bladder meridians orderly. At the same time, Laser Doppler Flowmetry was used to analyze microcirculation in meridian acupoints.Results The low-frequency acoustic waves with different frequencies had significantly different effects on kidney and bladder meridians. The effects of the same frequency on different acupoints of the same meridian were similar. The G1 (49.00 Hz) showed the most significant effects on the kidney meridian (P<0.01);G2 (98.00 Hz) showed the most significant impact on bladder meridian (P<0.01).Conclusion The kidney and bladder meridians have selective absorption characteristics to the low-frequency sound waves of different frequencies. Specific frequency sound waves can cause resonance in kidney and bladder meridians.

Objective To discuss different effects of low frequency sound waves of different timbres on microcirculation and transcutaneous oxygen partial pressure of Weizhong (BL40) of healthy people;To study the mechanism of somatosensory music therapy. Methods The same frequency (98.00 Hz), different timbres (sounds of guqin, flute, ocarina, bell, and drum were simulated) low frequency sound waves were played near Weizhong acupoint of 30 healthy persons. Laser Doppler flowmetry was used to analyze microcirculation and the changes of transcutaneous oxygen pressure of meridian acupoints, when each timbre was played for 60, 120, 180, 240 and 300 s. Results Sound waves of bell, drum, and flute made point microcirculation and transcutaneous oxygen pressure gradually increase, and the influence of drum>bell>flute;timbre waves of ocarina and guqin made the point microcirculation and transcutaneous oxygen pressure decrease gradually decrease, and the decrease of ocarina was more significant than that of guqin. In the same time point, sound waves of bell made point microcirculation and transcutaneous oxygen pressure increase more than the other sound waves (P<0.01). Conclusion Low frequency sounds of different timbres belong to different properties of five elements. They have different components in frequency spectrum, and can create different effects on acupoints.