Objective:To investigate the reparative efficacy and mechanism of pressure-adjustable macroporous antibacterial hydrogel in the treatment of infected wounds.Methods:Staphylococcus aureus was used to establish wound infection models in healthy C57BL/6 mice. The models were divided into 3 groups subjected to 3 different treatments: a negative control group with no hydrogel treatment (group A), a control group treated by common medical hydrogel (group B) and an experiment group treated by pressure-adjustable macroporous antibacterial hydrogel (group C). On days 1, 3, 6, 9 and 12, the effects of 3 treatments were compared on the wound area and the number of bacterial colonies under scab, on the apoptosis of fibroblasts based on the changes of type Ⅰ procollagen, and on the inhibition of inflammation during wound repair by detecting the expression of interleukin-6 (IL-6) and tumor necrosis factor (TNF-α).Results:On days 1 and 3, there was no significant difference between the 3 groups in the wound area ( P>0.05), but on days 6, 9 and 12, there were significant differences between the 3 groups in the wound area ( P<0.05). On day 6, the wound areas in group B (1.23 cm 2 ± 0.16 cm 2) and in group C (1.14 cm 2 ± 0.12 cm 2) were significantly smaller than that in group A (1.56 cm 2 ± 0.16 cm 2) ( P<0.05), but there was no significant difference between groups B and C ( P>0.05). On days 9 and 12, the wound areas in group B (0.97 cm 2 ± 0.13 cm 2 and 0.76 cm 2 ± 0.10 cm 2) and in group C (0.66 cm 2 ± 0.06 cm 2 and 0.48 cm 2 ± 0.07 cm 2) were significantly smaller than those in group A (1.49 cm 2 ± 0.11 cm 2 and 1.39 cm 2 ± 0.13 cm 2), and those in group C were significantly smaller than those in group B (all P<0.05). On day 1, there was no significant difference between the 3 groups in the number of bacterial colonies under scab ( P>0.05). On days 3, 6, 9 and 12, the numbers of bacterial colonies under scab in groups B and C were significantly smaller than that in group A ( P<0.05), and that in group C was significantly smaller than that in group B ( P< 0.05). The nucleic acid electrophoresis showed that the grayscale bands in group C were significantly darker than those in groups A and B. The early apoptosis rate of the fibroblasts in group C[low-right positive fluorescence (LR%): 9.72%] was significantly lower than that in group A (43.99%) and that in group B (38.43%), and that in group B was significantly lower than that in group A ( P<0.05). On day 12, the ratio of the gray values of IL-6 and β-actin (0.64 ± 0.10) and the ratio of the gray values of TNF-α and β-actin (0.34 ± 0.05) in the fibroblasts in group C were significantly higher than those in group A (1.22 ± 0.21 and 0.60 ± 0.14) and in group B (0.88 ± 0.02 and 0.41 ± 0.06) ( P<0.01). Conclusion:The pressure-adjustable macroporous antibacterial hydrogel is an effective treatment of infected wounds and its mechanism may be related to the reduced apoptosis of fibroblasts.

In order to improve the expression of recombinant human atrial natriuretic peptide (ANP), a new plasmid (pET28a(+)/ANP₃) containing 3 tandem ANP genes with lysine codon as the interval linker, was constructed. Target gene was transformed into Escherichia coli BL21 (DE3) and induced by IPTG, about 60% of the total-cell-protein was the target protein, His₆-ANP₃. After denaturation and refolding, it was digested by Endoproteinase Lys-C and Carboxypeptidase B (CPB) and then purified by a series of purification processes, about 16 mg purified ANP monomer could be obtained from one liter bacteria broth of shaking culture. Ultimately, the purity of protein was above 90% determined by UPLC and Tricine SDS-PAGE, its molecular weight was 3 080 Da according to LC-MS identification and it was proved to be equivalent to the reference product by ELISA. The use of tandem gene expression can provide a new possible model for the expression of other peptide drugs.
Atrial Natriuretic Factor ; biosynthesis ; Electrophoresis, Polyacrylamide Gel ; Escherichia coli ; metabolism ; Gene Expression ; Humans ; Metalloendopeptidases ; Peptides ; Plasmids ; genetics ; Recombinant Fusion Proteins ; biosynthesis