Production and characterization of highly purified recombinant thymosin beta 4 in Escherichia coli.

Thymosin ß4 (Tß4) is a small peptide composed of 43 amino acids. It has many important biological functions, such as promoting cardiac repair and wound healing, and therefore has great potential in clinical applications. In this report, we describe a novel and efficient way to produce highly purified and active Tß4. It was expressed in a soluble form using a DsbA and hexahistindine tag in Escherichia coli (E. coli). Using high cell density cultivation, the final biomass concentration was about 50 g L(-1) dry cell weight with the expression level of the fusion protein being 40%. To obtain highly purified protein, a purification process involving a five-step column procedure was implemented. The purity of Tß4 was above 98% and all the host cell related impurities, such as endotoxin, host cell protein and residual DNA levels, were within the permissible range listed in the Chinese Pharmacopoeia. The E-rosette test demonstrated that the bioactivity of purified Tß4 was consistent with other published work. This is the first report producing highly purified Tß4 from genetically engineered sources.

[Thrombocytopoietic Efficacy of Pegylated Recombinant Human Interleukin-11 Mutein in Myelosuppressed Mice].

OBJECTIVE: To evaluate the effect of PEGylated IL-11 mutein (PEG-mIL 11) with different dose or injection frequency on thrombocytopenia in myelosuppressed mice and to compare its effect with mIL-11, so as to provide reference data for clinical use. METHODS: Myelosuppressive model with thrombocyopenia was produced in BALB/c mice by whole body 60Co γ-ray irradiation in dose of administration 2.5 Gy followed by i.p. injections of carboplatin at 50 mg/kg. In study of injection frequency, 30 thrombocytopenic BALB/c mice were randomly divided into 5 groups: vehicle control group (once daily on d1, 4, 7), mIL-11 group [200 µg/(kg·d)×9 d], PEG-mIL 11 A group [111800 µg/(kg·d)×1 d (d 1)], PEG-mIL 11 B group 900 µg/(kg·d)×2 d (d 1,5), and PEG-mIL 11 C group [600 µg/(kg·d)×3 d (d 1,4,7)]. The route of administration is subcutaneous injection. The platelet counts were monitored in the subsequant 5 weeks. In study of dose administration, 100 thrombocytopenic BALB/c mice were randomly divided into 5 groups: vehicle control group (once daily on d1 and 5), mIL-11 group 200 µg/(kg·d)×9 d, and PEG-mIL 11 low-, mid-, and high-dose groups (200, 420 and 900 µg/(kg·d)×2 d, once a day on d1 and 5). The route of administration is subcutaneous injection. The platelet counts were monitored every 2-3 days in the subsequant 5 weeks, and CFU-Meg was determined on d 8 of the bone marrow cells collection. RESULTS: After 60Co irradition and carboplatin injection, Plt level decreased with time, and a >80% reduction was noted at nadir when comparing with baseline. In frequency of administration study, the platelet nadir of the 3 PEG-mIL 11 groups were significantly higher than that of vehicle control and mIL-11 groups (P<0.05), while no significant difference was noted among the 3 groups of different administration frequences; In dose level study, the reduction of Plt count at the nadir in the 3 PEG-mIL 11 groups was significantly less than that of vehicle control and mIL-11 groups (P<0.05). And a rapid recovery of Plt count was found in the PEG-mIL 11 groups with a dose-dependent increase of Plt count on d 10. A lower reduction and a rapider recovery of RBC was also found in the PEG-mIL 11 groups. No significant effect on WBC was found for all the treatment groups. An increase in CFU-Meg was observed in PEG-mIL 11 and mIL-11 groups, with higher CFU-Meg in PEG-mIL 11 groups. CONCLUSION: A preventive effect of PEG-mIL 11 on thrombocytopenia in myelosuppressed mice has been confirmed. In comparison with mIL-11, a better effect of PEG-mIL 11 is obtained under lower dose frequency, indicating a better compliance of the treatment regimen, and providing a foundation for developing a long-acting preparation of rhIL-11.

Physiological control of intraaorta pump based on heart rate.

Because of the special structures of intraaorta pump, the pressure and blood flow sensors cannot be implanted in the blood pump. Moreover, the cardiovascular pump is a very complex system that has no accurate model but much uncertainty and disturbance. Hence, the conventional control algorithm cannot achieve good performance. To overcome this problem, on one hand, a cardiovascular pump model is established. The heart rate in this model is chosen as a controlled variable that is a nonlinear function of the mean arterial pressure. On the other hand, a fuzzy logic feedback control algorithm, which maintains the actual heart rate tracking the desired heart rate, is designed. Computer simulations are performed to verify the robustness and dynamic characters of the controller. The simulation results demonstrate that the controller can maintain the actual heart rate tracking the desired one without static error. When the desired heart rate changed from 100 to 80 bpm, the settling time is <10 seconds. When the peripheral resistance increases from 1.0 to 0.7 mm Hg/ml, the settling time is <10 seconds.