Hypophosphatemia is a common metabolism disease in humans. Fibroblast growth factor 23 (FGF23) inhibits phosphate reabsorption by targeting on the renal tubules. FGF23^C-tail contains 73 amino acids from C-terminus of FGF23, serves as an inhibitor of FGF23, and can increase phosphate reabsorption. Therefore, FGF23^C-tail is an important drug for hypophosphatemia. In this paper, we constructed a fusion protein of FGF23^C-tail with HSA, and investigated the expression of the fusion protein in the Pichia pastoris system. The recombinant gene was constructed by fusion PCR. A high-yield strain was selected by G418 resistance and fermentation yield, and the expression yield was 43.7 mg·L^-1 in flask. In 5 L fermenters, the highest expression yield could reach 265.6 mg·L^-1. FGF23^C-tail-HSA could be used as an inhibitor for FGF23, and could significantly increase blood phosphorus levels in rats. The procedures for care and use of animals were approved by the Ethics Committee of YiChun University. This paper provided a basis research for further studying physiological activity of FGF23^C-tail-HSA.

γ-aminobutyric acid can be produced by a one-step enzymatic reaction catalyzed by glutamic acid decarboxylase. The reaction system is simple and environmentally friendly. However, the majority of GAD enzymes catalyze the reaction under acidic pH at a relatively narrow range. Thus, inorganic salts are usually needed to maintain the optimal catalytic environment, which adds additional components to the reaction system. In addition, the pH of solution will gradually rise along with the production of γ-aminobutyric acid, which is not conducive for GAD to function continuously. In this study, we cloned the glutamate decarboxylase LpGAD from a Lactobacillus plantarum capable of efficiently producing γ-aminobutyric acid, and rationally engineered the catalytic pH range of LpGAD based on surface charge. A triple point mutant LpGAD^{S24R/D88R/Y309K} was obtained from different combinations of 9 point mutations. The enzyme activity at pH 6.0 was 1.68 times of that of the wild type, suggesting the catalytic pH range of the mutant was widened, and the possible mechanism underpinning this increase was discussed through kinetic simulation. Furthermore, we overexpressed the Lpgad and Lpgad^{S24R/D88R/Y309K} genes in Corynebacterium glutamicum E01 and optimized the transformation conditions. An optimized whole cell transformation process was conducted under 40 ℃, cell mass (OD₆₀₀) 20, 100 g/L l-glutamic acid substrate and 100 μmol/L pyridoxal 5-phosphate. The γ-aminobutyric acid titer of the recombinant strain reached 402.8 g/L in a fed-batch reaction carried out in a 5 L fermenter without adjusting pH, which was 1.63 times higher than that of the control. This study expanded the catalytic pH range of and increased the enzyme activity of LpGAD. The improved production efficiency of γ-aminobutyric acid may facilitate its large-scale production.
Glutamate Decarboxylase/genetics* ; Lactobacillus plantarum/genetics* ; Catalysis ; gamma-Aminobutyric Acid ; Hydrogen-Ion Concentration ; Glutamic Acid

In recent years, continuous manufacturing technology has attracted considerable attention in the pharmaceutical industry. This technology is highly sought after for its significant advantages in cost reduction, increased efficiency, and improved productivity, making it a growing trend in the future of the pharmaceutical industry. Compared to traditional batch production methods, continuous manufacturing technology features real-time control and environmentally friendly intelligence, enabling pharmaceutical companies to produce drugs more efficiently. However, the adoption of continuous manufacturing technology has been slow in the field of traditional Chinese medicine(TCM) pharmaceuticals. On the one hand, there is insufficient research on continuous manufacturing equipment and technology that align with the characteristics of TCM preparations. On the other hand, the scarcity of talent with diverse expertise hampers its development. Therefore, in order to promote the modernization and upgrading of the TCM pharmaceutical industry, this article combined the current development status of the TCM industry to outline the development status and regulatory requirements of continuous manufacturing technology. At the same time, it analyzed the problems with existing TCM manufacturing models and explored the prospects and challenges of applying continuous manufacturing technology in the field of TCM pharmaceuticals. The analysis focused on continuous manufacturing control strategies, technical tools, and pharmaceutical equipment, aiming to provide targeted recommendations to drive the development of the TCM pharmaceutical industry.
Medicine, Chinese Traditional ; Quality Control ; Drug Industry ; Technology, Pharmaceutical/methods* ; Drugs, Chinese Herbal ; Pharmaceutical Preparations

Objective     To investigate the early safety and efficacy of transapical transcatheter aortic valve implantation (TAVI) for high-risk elderly patients with pure aortic valve insufficiency. Methods     A prospective multicenter clinical study of domestic J-valveTM TAVI for high-risk native non-calcified aortic valve insufficiency was conducted from April 2014 to May 2018, and the early postoperative results were analyzed. A total of 82 patients were enrolled, including 62 patients from West China Hospital, Sichuan University, 16 patients from Zhongshan Hospital, Fudan University, and 4 patients from Beijing Fuwai Hospital, National Center for Cardiovascular Diseases. There were 55 males and 27 females. The age was 61-90 (73.8±6.3) years. The logistic EuroSCORE was 10.0%-44.4% (17.5%±8.1%). All patients underwent TAVI using J-ValveTM system. Clinical evaluation and echocardiography were performed preoperatively and 1 month postoperatively. Multislice spiral CT was reviewed before discharge. Results     Three patients were transferred to thoracotomy for cardiopulmonary bypass operation, and 1 patient had decreased cardiac function due to leakage of the valve 1 week after surgery. The overall technical and procedural success rate was 95.1% and 93.9%, respectively. During hospitalization, 1 patient died of moderate pericyclosis complicated with multiple organ failure, and 1 patient died of pulmonary infection. Six (7.6%) patients received pacemaker implantation due to new onset Ⅲ° atrioventricular block. Echocardiographic follow-up showed paravalvular leak was observed in the few of patients, mild paravalvular leak was in 13 patients on the 30th day. Two patients showed moderate paravalvular leak. Left ventricular end-diastolic volume decreased from 197.7±66.8 mL (pre-TAVI) to 147.2±53.3 mL (30-day post-TAVI) (P<0.05). Mean pressure gradient was 9.5±4.1 mm Hg (30-day post-TAVI). Conclusion     This multicenter study demonstrates that TAVI with the J-Valve system for the treatment of pure aortic regurgitation is associated with sustained clinical and functional cardiovascular benefits in high-risk patients with symptomatic aortic regurgitation early-term follow-up. Our results further support that TAVI with the specific designed J-Valve system is an acceptable alternative therapy for high-risk patients with pure AR. Our result demonstrates good early-term durability and preserved hemodynamic function. The procedure appears to offer an adequate and lasting resolution for selected patients with pure aortic regurgitation.

Objective: To evaluate the effects of staining with carbon nanoparticles on the identification of parathyroid glands and lymph nodes during thyroid carcinoma surgery combined with lymphadenectomy. Methods: A total of 194 patients with papillary thyroid carcinoma who underwent thyroidectomy combined with lymphadenectomy from April 2016 to January 2018 were reviewed. Of them 104 cases were injected with carbon nanoparticles in operation area (nanocarbon group) and other 90 cases without the injection of carbon nanoparticles were as control group. The incidence of mistakenly dissection of parathyroid glands and the levels of serum calcium and parathyroid hormone in 1 day, 3 days, 1 month and 6 months after surgery were compared between two groups of patients. Chisquare and ranksum test were used to analyze 2 data. Results: There were no significant differences in age, gender, tumor size, operation time, extrathyroidal invasion and multifocality between two groups. Compared with control groups, nanocarbon group showed a significantly lower incidence of mistakenly dissection of parathyroid glands (8 cases vs 2 cases, 8.9% vs 1.9%,χ²=4.9, P=0.026) and a significantly lower incidence of hypoparathyroidism (41 cases vs 28 cases, 45.5% vs 26.9%, χ²=7.3, P=0.007). The number of lymph nodes dissected from central compartment was 791 in nanocarbon group and 536 in control groups, with a statistically significant difference (Z=-2.2, P=0.028). There was a significant difference in the number of lymph nodes removed from the right neck Ⅵb level between nanocarbon group and control group (41 nodes vs 93 nodes, Z=-2.1, P=0.034). Conclusion Treatment with nanocarbon can significantly facilitate the identification of the parathyroid during total thyroidectomy combined with central compartment lymphadenectomy, reduce the incidence of postoperative hypoparathyroidism, and improve the dissection of lymph node in the central compartment.