Hu'po Anshen Decoction Accelerated Fracture-Healing in a Rat Model of Traumatic Brain Injury Through Activation of PI3K/AKT Pathway.

Hu'po Anshen decoction (HPASD) is a traditional Chinese medicine formula comprising five herbal medicines for the treatment of concussion and fracture healing, but its pharmacological mechanism is still unclear. Ultra-performance liquid chromatography coupled with quadrupole time of flight mass spectrometry (UPLC/Q-TOF MS) was used to analyze the main active components of HPASD. Rats were randomly assigned to fracture group, fracture combined with traumatic brain injury (TBI) group (FBI) and FBI combined with HPASD treatment group (FBIH). Rats in the FBIH group were given oral doses of HPASD (2.4 g/kg, 4.8 g/kg and 9.6 g/kg) for 14 or 21 consecutive days. The fracture callus formation and fracture sites were determined by radiographic analysis and micron-scale computed tomography (micro-CT) analysis. Hematoxylin and eosin (H&E) staining and a three-point bending test were applied to assess histological lesions and biomechanical properties, respectively. The levels of cytokines-/protein-related to bone formation and differentiation as well as PI3K/AKT pathway-related proteins were determined by Enzyme-linked immunosorbent assay (ELISA), quantitative reverse transcription-polymerase chain reaction (qRT-PCR), or western blot assays, respectively. UPLC-Q/TOF-MS-based serum metabolomic analysis was also performed to investigate the therapeutic effects of HPASD in the treatment of FBI. UPLC/Q-TOF MS analysis showed the chemical components in HPASD, including flavonoids, amino acids, saponins, and phenylpropanoid constituents, etc. HPASD dose-dependently promoted callus formation, increased bone density, improved mechanical parameters and morphological scores, and facilitated the expressions of VEGF, PDGF, bFGF, VEGFA, CoL1A1, RUNX2, BMP2, and Aggrecan, inhibited the expression of MMP13, and activated PI3K/AKT pathway. Metabolomics analysis revealed abnormalities of malate-aspartate shuttle and glucose-alanine. HPASD accelerates fracture healing by promoting bone formation and regulating the malate-aspartate shuttle and glucose-alanine cycle, which might be associated with the activation of the PI3K/AKT pathway.

[Experimental study on invigorating kidney and activating blood on preventing and curing SD rats with knee osteoarthritis].

OBJECTIVE: To explore the effects and mechanism of invigorating kidney and activating blood, invigorating kidney and expelling wind on hemorheology, IL-1ß and TNF-α of SD rats with knee osteoarthritis, then definite the evolution of muscle certified turning into heumatism and compare the effect of Chinese herbal. METHODS: One hundred and eighty SD rats with 3-month-old (each weight was 185 to 215 g) received intra-articular injection of papain solution for establishing knee OA models. All rats were randomly divided into activating blood group, preventing group, expelling wind group, invigorating kidney group, invigorating kidney and activating blood group and model group. Laboratory indexes were obtained at the 30th, 60th, 90th days after gastric perfusion, which including state of mind, activity, fur, weight, joint swelling, largely image, hemorheology, inflammation and HE pathological appearance. RESULTS: After operation, rats appeared blood stasis and swelling and difficulty crawling. There was significant difference of hemorheology in invigorating kidney and activating blood group the content of IL-1ß and TNF-α was obviously lower than model group (P < 0.05 ). While the content of IL-1ß and TNF-α on the early stage was obviously higher than late stage (P < 0.05). CONCLUSION: Knee osteoarthritis mainly show synovial inflammation at the early stage, inflammation at early stage is more severe than late; invigorating kidney and activating blood decoction can inhibit the knee cartilage injury, improve blood circulation and prevent local inflammatory reaction. Activating blood decoction and invigorating kidney and activating blood Decoction have certain curative effect in early time, but the effects of invigorating kidney and activating blood Decoction is more effective than other on the late stage.

[Experimental study on treatment of femoral head necrosis with arterial perfusion of marrow stem cells].

OBJECTIVE: To explore the effect of arterial perfusion of marrow multifunctional stem cells (MFSC) in treating femoral head necrosis and its mechanism. METHODS: The rabbit model of femoral head necrosis was established by large dose of methyl-prednisone through Shwartzman response. Bone marrow was extracted from femoral bone of model rabbit and isolated in vitro for culturing and proliferating MFSC. The experimental rabbits were randomly divided into 4 groups, treated with normal saline (A), Salvia + urokinase (B), MFSC (C) and MFSC + Salvia + urokinase (D), respectively, they were sacrificed in batches at 2 and 4 weeks after treatment, and changes in various parameters, including molybdenum target roentgenogram, routine pathology with HE staining, tetracycline labeled fluorescent microscopy and ultrastructure alteration by scanning electron microscope (SEM), were observed. RESULTS: Typical appearance of femoral head necrosis was shown in the successfully modeled rabbits. Two and 4 weeks after treatment by high selective drug via medial and lateral femoral circumflex arterial perfusion, the X-ray examination showed significant improvement of bone density; pathohistologic manifestation showed decrease of empty bone lacuna, increase of osteoblast and new bone formation; tetracycline fluorescent labeled microscopic picture showed bright fluorescent band of increased osteoblasts in necrosis repairing region with widened border; SEM displayed irregularly arranged fibrosis in necrosis region, abundant organelles in osteoblasts with few empty bone lacuna. The above-mentioned improvement was more significant in rabbits treated by MFSC. CONCLUSION: High selective femoral drug arterial perfusion in treating femoral head necrosis could accelerate the process of revascularization and re-ossification in rabbits. As compared with Salvia, MFSC showed quicker and more potent effect.

Association of apolipoprotein E 4 polymorphism with cerebral infarction in Chinese Han population.

AIM: To study the association between APOE polymorphisms and cerebral infarction through a case-control study among the Chinese Han population. METHODS: First-ever cerebral infarction patients (n=226) whose ages ranged from 40 to 60 years old were recruited from Department of Neurology, Zhongshan Hospital, Shanghai, and Zhejiang Chinese Traditional Medicine Hospital, Zhejiang, China. Unrelated healthy controls (n=201) were selected from the general population in the same area with similar age and sex distribution. APOE was amplified by one-stage PCR using the forward primer: 5'-GGC ACG GCT GTC CAA GGA GCT-3' and reverse primer: 5'-GAT GGC GCT GAG GCC GCG CT-3'. The PCR product was digested directly with 5 U of CfoI and separated by a 20 % polyacrylamide (acrylamide: bis-acrylamide=29:1) nondenaturing gel. RESULTS: Both cerebral infarction patient and control groups were in Hardy-Weinberg equilibrium. The allele frequency of APOE*2, APOE*3, and APOE*4 was 4.6 %, 81.9 %, and 13.5 % respectively in the patients with cerebral infarction; 5.7 %, 87.3 %, and 7.0 % respectively in the healthy control group. Compared with APOE3/3 subjects, APOE4/4 carriers had a 2.1-fold risk of cerebral infarction (odds ratio 2.1, 95 % confidence limits 1.3 to 3.4). The allele frequency of APOE*4 in the cerebral infarction patient group was significantly higher than that in the control group (13.5 % vs 7.0 %; P=0.002). CONCLUSION: APOE 4 is a risk factor for cerebral infarction among the Chinese Han population.