Development of a High-Titer Culture Medium for the Production of Cholesterol by Engineered Saccharomyces cerevisiae and Its Fed-Batch Cultivation Strategy.

Steroids are a class of compounds with cyclopentane polyhydrophenanthrene as the parent nucleus, and they usually have unique biological and pharmacological activities. Most of the biosynthesis of steroids is completed by a series of enzymatic reactions starting from cholesterol. Synthetic biology can be used to synthesize cholesterol in engineered microorganisms, but the production of cholesterol is too low to further produce other high-value steroids from cholesterol as the raw material and precursor. In this work, combinational strategies were established to increase the production of cholesterol in engineered Saccharomyces cerevisiae RH6829. The basic medium for high cholesterol production was selected by screening 8 kinds of culture media. Single-factor optimization of the carbon and nitrogen sources of the culture medium, and the addition of calcium ions, zinc ions and citric acid, further increased the cholesterol production to 192.53 mg/l. In the 5-L bioreactor, through the establishment of strategies for glucose and citric acid feeding and dissolved oxygen regulation, the cholesterol production was further increased to 339.87 mg/l, which was 734% higher than that in the original medium. This is the highest titer of cholesterol produced by microorganisms currently reported. The fermentation program has also been conducted in a 50-L bioreactor to prove its stability and feasibility.

Effects of vitamin D combined with pioglitazone hydrochloride on bone mineral density and bone metabolism in Type 2 diabetic nephropathy.

The study aims to investigate the effect of vitamin D (VD) combined with pioglitazone hydrochloride (PIO) on bone mineral density (BMD) and bone metabolism in patients with Type 2 diabetic nephropathy (T2DN). T2DN patients were selected and assigned into mild, moderate, and severe groups. In each group, three therapy regimens (VD, PIO, and VD plus PIO) were administered. X-ray absorptiometry was used to measure BMD. Intact parathyroid hormone (iPTH) and 25-hydroxyvitamin D3 (25-OH-VD3) were measured by chemiluminescence meter. ELISA was applied to detect levels of osteoprotegerin (OPG), bone gla protein (BGP), C-terminal telopeptides of type I collagen (ß-CTX), procollagen type I N-propeptide (PINP), pyridinoline (Pyr), and deoxypyridinoline (D-Pyr). Compared with the mild group, T2DN patients in the moderate and severe groups had longer course of disease and higher levels of total cholesterol (TC), triglyceride (TG), serum phosphorus, fasting plasma glucose (FPG), glycosylated hemoglobin (HbAlc) and creatine (Cr), and lower blood calcium. The BMD in different parts increased among the mild, moderate, and severe groups, and the highest BMD was found after VD plus PIO treatment. OPG, iPTH, BGP, ß-CTX, Pyr/Cr, and D-Pyr/Cr levels were reduced, while 25-OH-VD3 and PINP levels were elevated among three groups after different treatments, and the most obvious change was observed after VD plus PIO treatment. Our findings indicate that VD combined with PIO may be more effective in improving BMD and bone metabolism than VD or PIO alone in the treatment of T2DN patients, especially for T2DN patients with mild disease.