Objective To establish an HPLC method for concentration determination of emodin in plasma and colon tissues after gavage withHuchang Qingdu Pellets;To explore pharmacokinetic properties and colon tissue distribution ofHuchang Qingdu Pellets in rats.Methods Wistar rats were randomly divided into Pellets group and ordinary capsule group. Blood was taken after gavage withHuchang QingduPellets or capsule content in given time points (1, 2, 3, 4, 5, 6, 8, 12, 24 h) from rat venous sinus and analyzed. Rats were put to death, and gastric and colon tissues were isolated and extracted for analysis. Chromatographic conditions:Phenomenex C18 column (4.6 mm × 150 mm, 10μm), the mobile phase of methanol-0.1% acetic acid solution (80∶20), flow rate of 1 mL/min, the detection wavelength of 254 nm.Results Emodin was in good linear range of 0.102-20.40μg/mL. The regression equations were:plasma solutionA=76 825C + 3567.8,r=0.999 4 (n=6);intestinal tissue solutionA=75 931C + 3384.2,r=0.999 0 (n=6). Day-to-day precision and within-day precision were less than 4%. The recovery rate was more than 90%. Pharmacokinetic parameters of T1/2 and Tmax of emodin in rat blood and intestinal tissues increased compared with the ordinary capsule group;Cmax and AUC0-24 were obviously improved compared with the ordinary capsule group.ConclusionHuchang Qingdu Pellets do have controlled release and colon targeting effect.

Bacterial antitumor therapy has great application potential given its unique characteristics, including genetic manipulation, tumor targeting specificity and immune system modulation. However, the nonnegligible side effects and limited efficacy of clinical treatment limit their biomedical applications. Engineered bacteria for therapeutic applications ideally need to avoid their accumulation in normal organs and possess potent antitumor activity. Here, we show that macrophage-mediated tumor-targeted delivery of Salmonella typhimurium VNP20009 can effectively reduce the toxicity caused by administrating VNP20009 alone in a melanoma mouse model. This benefits from tumor-induced chemotaxis for macrophages combined with their slow release of loaded strains. Inspired by changes in the tumor microenvironment, including a decrease in intratumoral dysfunctional CD8⁺ T cells and an increase in PDL1 on the tumor cell surface, macrophages were loaded with the engineered strain VNP-PD1nb, which can express and secrete anti-PD1 nanoantibodies after they are released from macrophages. This novel triple-combined immunotherapy significantly inhibited melanoma tumors by reactivating the tumor microenvironment by increasing immune cell infiltration, inhibiting tumor cell proliferation, remodeling TAMs to an M1-like phenotype and prominently activating CD8⁺ T cells. These data suggest that novel combination immunotherapy is expected to be a breakthrough relative to single immunotherapy.

ObjectiveTo study the effect of Xiaoyusan new formula on the articular cartilage of knee osteoarthritis (KOA) rabbits and its mechanism. MethodsA total of 42 New Zealand white rabbits aged 6 months were randomly divided into normal group, model group, ointment of Xiaoyusan group, and ointment of Xiaoyusan new formula group, with 10 rabbits in each group (the other 2 rabbits were used for model validation). Except for the normal group, the right knee joints of all rabbits in the other groups were prepared as KOA models according to the modified Hulth method. After 5 weeks of molding, the rabbits in ointment of Xiaoyusan group, ointment of Xiaoyusan New Formula group were given corresponding ointments for knee arthritis treatment, once a day, each time for 10 hours. After 2-week continuous administration and treatment, the knee joint cartilage of the four groups of rabbits was taken and the cartilage damage of each group was evaluated by Outerbridge grading method. The pathological changes of the cartilage, calcified layer and subchondral bone of the knee joint of rabbits in each group were observed by HE staining method under the light microscope, and the degree of cartilage degeneration was evaluated by Mankin's method. The expression of matrix metalloproteinase-13 (MMP-13) in the cartilage of rabbit knee joint in each group was deteced by immunohistochemistry. Results After the general observation of articular cartilage, the Outerbridge grading showed that the number of high-grade animals in ointment of Xiaoyusan group was reduced compared with the model group (P<0.05), and the number of high-grade animals in ointment of Xiaoyusan new formula group was also reduced (P<0.05) compared with ointment of Xiaoyusan group. HE staining showed that Mankin's scores of articular cartilage in the four groups ranked from high to low: model group (10.82±1.76), ointment of Xiaoyusan group (6.19±1.23), ointment of Xiaoyusan new formula group (2.64±1.18) and normal group (0.28±0.17). The difference among four groups was statistically significant (P<0.05). Immunohistochemical detection showed that the positive rates of MMP-13 expression in rabbit articular cartilage tissues in each group were (67.90±13.94)% of model group, (37.10±19.16)% of ointment of Xiaoyusan group, (13.60±3.10)% of ointment of Xiaoyusan new formula group and (3.20±2.39) % of normal group, ranking from high to low, and the difference among four groups was statistically significant (P<0.05). ConclusionXiaoyusan new formula can repair articular cartilage degeneration in KOA rabbits and decrease the expression of MMP-13 in cartilage, which may be one of the mechanisms of the treatment.