Mesenchymal stem cells (MSCs) were induced into a nucleus pulposus-like phenotype utilizing simulated microgravity in vitro in order to establish a new cell-based tissue engineering treatment for intervertebral disc degeneration. For induction of a nucleus pulposus-like phenotype, MSCs were cultured in simulated microgravity in a chemically defined medium supplemented with 0 (experimental group) and 10 ng/mL (positive control group) of transforming growth factor β1 (TGF-β1). MSCs cultured under conventional condition without TGF-β1 served as blank control group. On the day 3 of culture, cellular proliferation was determined by WST-8 assay. Differentiation markers were evaluated by histology and reverse transcriptase-polymerase chain reaction (RT-PCR). TGF-β1 slightly promoted the proliferation of MSCs. The collagen and proteoglycans were detected in both groups after culture for 7 days. The accumulation of proteoglycans was markedly increased. The RT-PCR revealed that the gene expression of Sox-9, aggrecan and type II collagen, which were chondrocyte specific, was increased in MSCs cultured under simulated microgravity for 3 days. The ratio of proteoglycans/collagen in blank control group was 3.4-fold higher than positive control group, which denoted a nucleus pulposus-like phenotype differentiation. Independent, spontaneous differentiation of MSCs towards a nucleus pulposus-like phenotype in simulated microgravity occurred without addition of any external bioactive stimulators, namely factors from TGF-β family, which were previously considered necessary.

OBJECTIVETo observe the law of the meridian abnormal changes and the correlation with acupuncture efficacy based on the effectiveness study of electroacupuncture (EA) in treatment of severe functional constipation.

METHODSSeventy patients of severe functional constipation were randomized into an EA group and a sham-EA group, 35 cases in each one. In the EA group, Tianshu (ST 25) and Fujie (SP 14) were punctured deeply and stimulated with EA (dense-disperse wave, 2Hz/15 Hz, 0. 1 to 1. 0 mA), and Shangjuxu (ST 37) was needled. In the sham-EA group, the sites lateral to Tianshu (ST 25) and Fujie (SP 14) were punctured shallowly and stimulated with electricity. The site lateral to Shangjuxu (ST 37) was punctured shallowly. The treatment was given continuously for 8 weeks in the two groups, 5 times weekly in the first 2 weeks and 3 times weekly in the rest 6 weeks. WANG Juyi's meridian examination method was applied to detect the abnormalities of the spleen, stomach and large intestine meridians before treatment, and in 2, 4, 6 and 8 weeks among 70 patients separately. The frequency of complete spontaneous bowel movement (CSBM) every two weeks, meridian abnormal value and the relativity with CSBM change rate were compared in the patients between two groups.

RESULTSRegarding the increase of CSBM frequency, the effect started since the 2nd week in the EA group, with the treatment going on, CSBM frequency was increased apparently (all P<0. 05). In the sham-EA group, after 6 and 8-weeks of treatment, CSBM frequency was increased apparently as compared with that before treatment (all P<0. 05). The increase of CSBM frequency in the EA group was remarkably as compared with the sham-EA group at every time point (all P<0. 05). The abnormal value of the large intestine meridian in 2 weeks of treatment and the values of the stomach and spleen meridians and the relevant meridians in 4 weeks of treatment were all reduced apparently as compared with those in the baseline in the EA group (all P<0. 05). With the treatment time going on, the abnormal reflections on the large intestine and stomach meridians were reduced gradually (all P<0. 05), and the total change rate of abnormalities on the large intestine, stomach and spleen meridians presented the negative correlation with the total change rate of CSBM frequency (P<0. 01, P<0. 05).

CONCLUSIONIn the EA group, the efficacy on CSBM frequency in severe functional constipation is advantageous and stable as compared with the sham-EA group. Acupuncture at the relevant acupoints of the spleen, stomach and large intestine meridians achieves the definite regulation to the meridian abnormalities. It is discovered that the abnormal changes in the spleen, stomach

Acupuncture Points ; Adult ; Aged ; Constipation ; physiopathology ; therapy ; Defecation ; Electroacupuncture ; Female ; Humans ; Male ; Meridians ; Middle Aged ; Treatment Outcome ; Young Adult

Mesenchymal stern cells (MSCs) were induced into a nucleus pulposus-like phenotype utilizing simulated microgravity in vitro in order to establish a new cell-based tissue engineering treatment for intervertebral disc degeneration.For induction of a nucleus pulposus-like phenotype,MSCs were cultured in simulated microgravity in a chemically defined medium supplemented with 0 (experimental group) and 10 ng/mL (positive control group) of transforming growth factor β1 (TGF-β1).MSCs cultured under conventional condition without TGF-β1 served as blank control group.On the day 3 of culture,cellular proliferation was determined by WST-8 assay.Differentiation markers were evaluated by histology and reverse transcriptase-polymerase chain reaction (RT-PCR).TGF-β1 slightly promoted the proliferation of MSCs.The collagen and proteoglycans were detected in both groups after culture for 7 days.The accumulation of proteoglycans was markedly increased.The RT-PCR revealed that the gene expression of Sox-9,aggrecan and type Ⅱ collagen,which were chondrocyte specific,was increased in MSCs cultured under simulated microgravity for 3 days.The ratio of proteoglycans/collagen in blank control group was 3.4-fold higher than positive control group,which denoted a nucleus pulposus-like phenotype differentiation.Independent,spontaneous differentiation of MSCs towards a nucleus pulposus-like phenotype in simulated microgravity occurred without addition of any external bioactive stimulators,namely factors from TGF-β family,which were previously considered necessary.