BACKGROUND: Psoralen is a plant estrogen, and a large number of studies have confirmed its role in promoting cell proliferation and differentiation. OBJECTIVE: To construct a cell-scaffold composite bone using psoralen, rabbit endosteal mesenchymal stem cells and polycaprolactone and to explore its effect on the treatment of rabbit nonunion. METHODS: (1) Rabbit endosteal mesenchymal stem cells were cultured and cultured until the third generation for each experiment. Passage 3 cells were seeded onto culture plates containing 50 mg/L bone morphogenetic protein 2 (positive control), 10-8, 10-7 and 10-6 mol/L psoralen (low-, middle-, and high-concentration psoralen groups) or the same volume of purified water (control group). The cell proliferation of each group was detected on the 3rd, 5th and 7th days after intervention using MTT method. (2) The three-dimensional polycaprolactone scaffold was added to the bottom of the cell culture plate, and rabbit endosteal mesenchymal stem cells were seeded into the scaffold at a density of 1×103 per well. Then, 10-6 mol/L of psoralen was added. Cell-scaffold composite bone was taken after 21 days of culture. (3) Animal models of radial nonunion were established in 27 New Zealand white rabbits, and were then randomized into experimental, scaffold and control groups followed by implantation of cell-scaffold composite bone, simple scaffold, and nothing, respectively. Pathological hematoxylin-eosin staining for observation of bone healing was performed at the 2nd, 4th, and 8th weeks after surgery. Healing of nonunion was observed on the X-ray films that were taken at the 4th week after surgery. RESULTS AND CONCLUSION: (1) Three concentrations of psoralen could induce the proliferation of rabbit endosteal mesenchymal stem cells. Compared with the control group, 10-6 mol/L psoralen exerted the strongest stimulation effect on rabbit endosteal mesenchymal stem cells (P < 0.05), whereas there was no significant difference between high-concentration psoralen group positive control group (P > 0.05). (2) Pathological hematoxylin-eosin staining of radial nonunion showed that the number of osteoblasts in the experimental group was higher than that in the scaffold and control groups (P < 0.05). (3) The X-ray films revealed bone healing in the experimental group, partial healing in the scaffold group and non-healing in the control group. Overall findings indicate that psoralen can promote the proliferation of rabbit endostealmesenchymal stem cells, and the effect is certainly related to the concentration of psoralen. Psoralen can be combined with rabbit endosteal mesenchymal stem cells and polycaprolactone scaffold to form composite bone, achieving good outcomes in the treatment of nonunion in animals.

BACKGROUND: Apigenin has been shown to hold the effects of antivirus, anti-inflammation, anti-oxidation and tranquilizer and sedative. OBJECTIVE: To study the effect of different doses of apigenin on osteoporosis in rats and to explore the underlying mechanism. METHODS: The study was approved by the Laboratory Animal Ethical Committee of Southern Medical University. Forty-two adult male Sprague-Dawley rats were selected, and the rat models of osteoporosis were established by ovariectomy. The rats were divided into seven groups (n=6/group): The sham operation group, the control group, the positive control group, and the 20, 40, and 60, and the 80 mg/kg apigenin groups according to the dose of apigenin. The sham operation group did not receive the removal of the ovaries. The positive control group was supplemented with diethylstilbestrol (0.02 mg/kg), vitamin D and calcium daily. The control group was given the subcutaneous injection of same volume of purified water. The apigenin groups were given the subcutaneous injection of various doses of apigenin, respectively, once daily, for 8 weeks. The femoral bone mineral density and serum levels of calcium, phosphorus, bone alkaline phosphatase, amino-terminal propeptide of type I procollagen, C-terminal telopeptide of type I collagen were measured at 4 and 8 weeks after intervention, respectively. RESULTS AND CONCLUSION: (1) Compared with the control group, when the dose of apigenin was 40, 60 and 80 mg/kg, the bone mineral density, and serum levels of calcium and phosphorus were increased significantly at 4 and 8 weeks (P < 0.05), and the levels of amino-terminal propeptide of type I procollagen, C-terminal telopeptide of type I collagen and bone alkaline phosphatase were decreased significantly (P < 0.05). (2) Compared with the positive control group, when the dose of apigenin was 60 and 80 mg/kg, the bone mineral density, serum levels of calcium and phosphorus, the levels of amino-terminal propeptide of type I procollagen, C-terminal telopeptide of type I collagen and bone alkaline phosphatase showed no significant difference at 4 and 8 weeks (P > 0.05). When the dose of apigenin was 20 and 40 mg/kg, compared with the positive control group, the bone mineral density, and serum levels of calcium and phosphorus were decreased significantly at 4 and 8 weeks (P < 0.05), and the levels of amino-terminal propeptide of type I procollagen, C-terminal telopeptide of type I collagen and bone alkaline phosphatase were increased significantly (P < 0.05). (3) Compared with the sham operation group, in the control group, the bone mineral density, and serum levels of calcium and phosphorus were decreased significantly at 4 and 8 weeks (P < 0.05), and the levels of amino-terminal propeptide of type I procollagen, C-terminal telopeptide of type I collagen and bone alkaline phosphatase were increased significantly (P < 0.05). (4) These results suggest that it is indeed effective to establish a rat osteoporosis model by ovariectomy. The therapeutic effect of apigenin on osteoporosis is dose-related, and a certain dose of apigenin has an anti-osteoporosis effect.

Objective: To establish a method for the simultaneous identification of Zika, Chikungunya and Mayaro viruses. Methods: The complete genome sequences of Zika, Chikungunya and Mayaro virus were retrieved from Global Shared Database for comparative analysis, estimate its conservative region and determine the target gene location, specific primers and probes were designed, then a triplex real-time RT-PCR assay was developed. The specificity, sensitivity and repeatability of the assay were assessed by viral nucleic acid of Zika virus, Chikungunya virus a, in vitro transcriptional RNA of Mayaro virus, normal human serum and related virus simulation sample. Results: The result showed that the established method could detect Zika virus, Chikungunya virus, as well as simulated Mayaro virus samples, the limit of detection (LOD) of Zika and Chikungunya virus was 16.22 Copy/PCR and 12.02 Copy/PCR, respectively, the LOD for simulated Mayaro virus RNA was 2.82 Copy/PCR, no significant difference was detected between the triplex and monoplex assays. No cross reaction was found in the detection of dengue virus, Hantavirus, severe fever with thrombocytopenia syndrome (SFTS) virus, yellow fever virus and influenza virus, and 100 healthy adults blood samples, the specificity of the method was 100%. The repeatability result showed that the standard deviation of all three detections were blow 0.5 and the coefficient of variation was less than 2% by selecting viral nucleic acids or transcribed RNA with high, medium and low concentration gradients. Conclusions A triplex real-time RT-PCR assay for detection of Zika, Chikungunya and Mayaro virus has been established with an acceptable specificity, sensitivity and repeatability.