Mechano-biomimetic hydrogel 3D cell cultivation as a strategy to improve mammalian cell protein expression.

Eukaryotic expression systems are frequently employed for the production of recombinant proteins as therapeutics as well as research tools. Among which mammalian cell protein expression approach is the most powerful one, which can express complex proteins or genetic engineered biological drugs, such as PD-1. However, the high expense, which partially derives from its low protein yielding efficiency, limited the further application of such approach in large scale production of target proteins. To address this issue, we proposed a novel technique to promote the protein production efficiency of mammal cells without using conventional genetic engineered approaches. By placing 293T cells in a hydrogel 3D cell culture platform and adjusting the stress relaxation of the matrix hydrogel, cells formed multicellular spheroids by self-organization. In particular, the multicellular spheroids have a significantly enhanced ability to transiently express multiple proteins (SHH-N, PD-1 and PDL-1). We also examined in detail the mechanism underlying this phenomenon, and found that the reorganization of cytoskeleton during spheroids formation enhances the translation process of protein by recruiting ribosomes. Overall, this finding provides a novel approach for subsequent improvement of large-scale mammalian protein expression cell systems.

[Clinical observation of herb-separated moxibustion on segmental conception vessel combined with low-frequency TEAS for asthenospermia and oligospermia].

OBJECTIVE: To observe the clinical effect of herb-separated moxibustion on segmental conception vessel combined with low-frequency transcutaneous electrical acupoint stimulation (TEAS) for asthenospermia and oligospermia. METHODS: A total of 105 patients with asthenospermia and oligospermia were randomly divided into a combination group, a TEAS group and a medication group, 35 cases in each one. In the medication group, vitamin E capsules, coenzyme Q10 capsules，Wuzi Yanzong pills were taken. In the TEAS group, TEAS was applied at Shenshu (BL 23) and Pishu (BL 20), 2 Hz in frequency, 10-15 mA in intensity, once every 2 days, 30 min each time.On the base of the treatment as the TEAS group, herb-separated moxibustion on segmental conception vessel [Shenque (CV 8) to Zhongji (CV 3)] was applied in the combination group, once every 10 days. A total of 3 months were required in the 3 groups.Before treatment and 2, 3 months into treatment, the semen routine indexes (concentration, normal morphology rate, sperm motility), seminal plasma contents of superoxide dismutase (SOD) and malondialdehyde (MDA) and TCM syndrome score were observed in the 3 groups, and the clinical efficacy was evaluated. RESULTS: Compared before treatment, except for the sperm morphology 2 months into treatment in the medication group, the semen routine indexes 2, 3 months into treatment were improved in the 3 groups (P<0.01), and the semen routine indexes 2, 3 months into treatment in the combination group were superior to the medication group and the TEAS group (P<0.01). Compared before treatment, the seminal plasma contents of SOD were increased and MDA were decreased 2, 3 months into treatment in the 3 groups (P<0.01), and the changes of the combination group were larger than the medication group and the TEAS group (P<0.01). Compared before treatment, the TCM syndrome scores 2, 3 months into treatment in the combination group, and 3 months into treatment in the medication group and the TEAS group were decreased (P<0.01). The TCM syndrome scores 2, 3 months into treatment in the combination group were lower than the medication group and the TEAS group (P<0.01). The total effective rates in the combination group, the medication group and the TEAS group were 85.7% (30/35), 68.6% (24/35) and 65.7% (23/35), the clinical efficacy of the combination group was higher than the medication group and the TEAS group (P<0.05). CONCLUSION: Herb-separated moxibustion on segmental conception vessel combined with low-frequency TEAS can improve semen routine, reduce sperm oxidative stress damage for patients with asthenospermia and oligospermia, and the clinical efficacy is better than the medication and TEAS.

[Scraping technique of stuck needle at Anmian point in the treatment of insomnia: a randomized controlled trial].

OBJECTIVE: To compare the efficacy difference in the treatment of insomnia between scraping technique of stuck needle and conventional acupuncture at Anmian (Extra). METHODS: One hundred and thirty one cases were randomized into an Anmian group (68 cases) and a conventional acupuncture group (63 cases). In the Anmian group, Anmian (Extra) was selected. After arrival of qi, the stuck needling was adopted by rotating the needle gently in single direction, 2-3 rounds till the needle body was stuck tightly. Afterwards, the needle tail was touched gently with the index finger to fix the needle body and the needle handle was scraped gently with the thumbnail from bottom to up. The needle was retained for 30 min. In the conventional acupuncture group, Sanyinjiao (SP 6), Shenmen (HT 7) and Baihui (GV 20) were selected and stimulated with reducing technique by rotating the needles. The needles were retained for 30 min. The treatment was given once every day, continuously for 2 weeks in both groups. The score of each factor and the total score in Pittsburgh sleep quality index (PSQI) were assessed before and after treatment in the two groups. Additionally, the efficacies of two groups were evaluated. RESULTS: For the patients in the conventional acupuncture group, the sleep quality and time of falling into spleen after treatment were improved as compared with those before treatment (all P < 0.01). The improvement of sleep time, sleep efficiency, sleep disturbance, hypnotic drug and daytime dysfunction, and PSQI total score did not present statistically significant difference as compared with those before treatment (all P > 0.05). After treatment, for the patients in the Anmian group, the factor score and total score in PSQI were apparently improved as compared with those before treatment (P < 0.01, P < 0.05). And the improvements of the above items were superior to the conventional acupuncture group (P < 0.01, P < 0.05). The total effective rate was 91.2% (62/68) in the Anmian group and was 74.6% (47/63) in the conventional acupuncture group. The clinical efficacy in the Anmian group was apparently superior to the conventional acupuncture group. CONCLUSION: The scraping technique of stuck needle at Anmian (Extra) achieves the superior effect on insomnia as compared with the conventional acupuncture.