Effects of Wenyangbushen formula on the expression of VEGF, OPG, RANK and RANKL in rabbits with steroid-induced femoral head avascular necrosis.

The present study aimed to investigate the effects of Wenyangbushen formula on the mRNA and protein expression levels of vascular endothelial growth factor (VEGF), osteoprotegerin (OPG), receptor activator of nuclear factor (NF)­κß ligand (RANK), and RANK ligand (RANKL) in a rabbit model of steroid­induced avascular necrosis of the femoral head (SANFH). The present study also aimed to examine the potential mechanism underlying the effect of this formula on the treatment of SANFH. A total of 136 New Zealand rabbits were randomly divided into five groups: Normal group, model group, and three groups treated with the traditional Chinese medicine (TCM), Wenyangbushen decoction, at a low, moderate and high dose, respectively. The normal group and positive control group were intragastrically administered with saline. The TCM groups were treated with Wenyangbushen decoction at the indicated dosage. Following treatment for 8 weeks, the mRNA and protein expression levels of VEGF, OPG, RANK and RANKL in the femoral head tissues were determined using reverse transcription­quantitative polymerase chain reaction and western blot analyses, respectively. The data revealed that Wenyangbushen decoction effectively promoted the growth of bone cells, osteoblasts and chondrocytes, and prevented cell apoptosis in the SANFH. The mRNA and protein expression levels of OPG and VEGF were increased, while the levels of RANK and RANKL were reduced in the necrotic tissue of the model group, compared with those in the normal rabbits. Wenyangbushen treatment prevented these changes, manifested by an upregulation in the expression levels of VEGF and OPG, and downregulation in the expression levels of RANK and RANKL in a dose­dependent manner. It was concluded that treatment with Wenyangbushen formula alleviated necrosis of the femoral head induced by steroids. It was observed to promote bone cell, osteoblast and chondrocyte growth, as well as prevent cell apoptosis. In addition, it upregulated the expression levels of OPG and VEGF, and inhibited the expression levels of RANK and RANKL. These results suggest the potential use of Wenyangbushen formula as a possible approach for the effective treatment of SANFH.

[Optimize extraction technology of Fufang Bajitian Shenggu Particles by central composite design and response surface method].

OBJECTIVE: To optimize extraction process of Fufang Bajitian Shenggu Particles by central composite design and response surface method. METHODS: The preparation of Fufang Bajitian Shenggu Particles was designed according to the test. With the overall desirability (OD) of Icariin content and the dry extract yield in medicinal materials extract concentrate as examining indexes, immersion time, material liquid ratio, extraction time and stress concentration temperature were investigated. Based on the single factor tests, material liquid ratio, extraction time and stress concentration temperature which impacted response values significantly were investigated by three factors and five levels of central composite design. RESULTS: Optimum extraction technology of Fufang Bajitian Shenggu Particles were as follows: material liquid ratio was 1:11.33, extraction time was 39 min, and stress concentration temperature was 66 °C. Bias of the lcariin content and the dry extract yield between observed and predicted values were 1.60% and 1.55% ,respectively. CONCLUSIONS: Using central composite design and response surface method to optimize extraction of Fufang Bajitian Shenggu particles has a good prediction.

Experimental research into using bone-strengthening pill to treat ischemic necrosis of femoral head.

OBJECTIVE: To study the use of a bone-strengthening pill to treat ischemic necrosis of the femoral head. METHODS: A model of castration in rats with osteoporosis and a model of ischemic necrosis in the bilateral femoral heads of young rats were used to detect bone density, bone formation measurements, lipid deposition in the femoral head, the distribution of capillaries in the femoral head and other relevant biochemical indices. RESULTS: In model rats, bone-strengthening pills were able to increase the bone density, the unit volume of bony trabeculae, the width of bony trabeculae, and the unit volume of the bony cortex. They could also enhance the mineral apposition rate in the femoral head and the seam width of osteoid deposition. Furthermore, there was a reduction in lipid deposition in the femoral head, and enriched distribution of arteries in the femoral head. Bone-related biochemical detections showed an increase in alkaline phosphatase staining and a decrease in tartaric acidic phosphatase staining. CONCLUSION: The results show that bone-strengthening pills can obviously promote bony growth, cause an increase in bone density, restore blood circulation, inhibit the occurrence and development of bony necrosis, and accelerate the repair of necrotic bones, with no toxic side-effects.

Effects of huogu I formula (I) on correlated factors of bone regeneration in chickens with steroid-induced necrosis of femoral head.

OBJECTIVE: To study the mechanism of Huogu I formula (I) in treating osteonecrosis of femoral head. METHODS: Forty-eight healthy female Leghorn chickens were randomly divided into control group, model group and Huogu I group, and each group consisted of 16 chickens. At the meantime of model establishment, chickens of the Huogu I group were administrated with decoction, while the model and control group with distilled water by gavage. At the 8th and 16th week after medication, blood samples were obtained for blood lipid detection while both sides of femoral head were harvested for the rest of examinations. Specifically, expressions of bone morphogenetic protein-2 (BMP2), transforming growth factor beta1 (TGFß(1)), Smad4 and Smad7 were evaluated by immunohistochemistry, while expression of osteoprotegerin/receptor activator of nuclear factor kappaB ligand (OPG/RANKL) mRNA was detected by in situ hybridization. RESULTS: Compared with the control group, serum levels of total cholesterol (TC), triglyceride (TG) and low-density lipoprotein cholesterol (LDL-C) in the model group rose significantly. Positive cell counting of BMP2, TGFß(1), Smad4 and OPG in femoral head of the model group dropped prominently. Positive cell counting of Smad7 and RANKL increased dramatically. In contrast with the model group, levels of TC, TG and LDL-C in Huogu I group reduced significantly. Positive cell counting of BMP2, TGFß(1), Smad4 and OPG in femoral head of the Huogu I group increased prominently. Indices of Smad7 and RANKL both decreased significantly. Especially at the 8th week, these variations were more significant. CONCLUSION: Huogu I formula is effective in promoting repair of necrotic femoral head by regulating the expressions of BMP2, TGFß(1), Smads and OPG/RANKL of osteoclast in femoral head.

[Effects of different concentrations of gubishu containing serum on the proliferation of rabbit articular chondrocytes in vitro culture].

OBJECTIVE: To investigate the effects of different concentrations of Gubishu containing serum on the proliferation of rabbit articular chondrocytes cultured in vitro. METHODS: Articular chondrocytes were obtained from the cartilage of 1-month rabbit and cultured in vitro. They were randomly divided into 8 groups,blank and Gubishu groups in different concentrations (5%, 10%,15%, 20%), MTT assay method was adopted to observe the influence of Gubishu containing serum with different concentrations to the proliferation of chondrocytes after incubated 1, 3, 5, 7 and 9 days. RESULTS: The proliferation of chondrocytes was dependent on the concentration in Gubishu groups. At same time point,there was significant value between every groups, 20% concentration was greatest (P<0.05); There was significant differences between 5%, 10% and 20% concentration of the blank groups at same time point (P<0.05), and was not between 15% and 20% concentration at the 1, 3, 5 and 7 days (P>0.05), 20% concentration of the blank group was greatest. 20% concentrations of Gubishu containing serum was significantly greater than 20% concentrations of blank group at the 1, 3, 5 and 7 days (P<0.05). CONCLUSION: 20% concentrations of Gubishu containing serum can significantly increase the proliferation of chondrocytes, and bring the logarithmic growth period forward to the 3 day.

[Experimental research progress of warming yang and reinforcing kidney of Chinese medicine to promote the differentiation of bone marrow stromal cells].

Bone marrow stromal cells (BMSCs), a kind of stem cells residing in bone marrow, have self-renewal, high proliferative capacity and the potential of multilineage differentiation. It has a good prospect in application of the cell replacement therapy, the gene therapy and the tissue engineering and so on. As the content of BMSCs is extremely low in bone marrow, BM-SCs must be amplified in vitro and induced to differentiation to meet the clinical needs. Researches of the recent years suggest there is a very promising way that Chinese medicine could induce BMSCs proliferation, differentiation. Based on the Chinese medicine theory, "the kidney generating marrow and dominating bone" and "kidney storing essence, essence and marrow", the TCM scholars have done some researches to explore the function of warming yang and reinforcing kidney of Chinese medicine to promote bone marrow stromal cells and found that these drugs can promote the BMSCs to proliferate and to differentiate into osteogenic, cartilage and nerve cells. This article elaborates and presents the researches on this aspect.

Progress on integrated Chinese and Western medicine in the treatment of osteoarthritis.

Osteoarthritis (OA) is a chronic degenerative disease of the joints caused by wide variety of factors. factors. This paper provides a review of the clinical and experimental research on integrated Chinese and Western medicine in the treatment of osteoarthritis. Western medicine in the treatment of osteoarthritis. (1) Clinical research: integrated Chinese and Western medicine therapies were used including physiotherapy, medications, acupuncture, functional training, intra-articular injection of sodium hyaluronate therapy, and arthroscopic debridement with Chinese medicine articular iontophoresis therapy. articular iontophoresis therapy. (2) Experimental research: modern methods were used in studying the mechanism of Chinese medicine in slowing down cartilage degeneration, promoting articular cartilage repair, inhibiting synovial inflammation, and blocking cartilage destruction. inflammation, and blocking cartilage destruction. In addition, this article also reviews the advantages, prospects, and problems of the therapies. and problems of the therapies.

[Experimental study on the effects of pharmaco-serum of diabetic rats fed with Chinese herbs qianggubao decoction ([Chinese characters: see text]) on osteoblast culture in vitro].

OBJECTIVE: To investigate the optimum phase and dose of pharmaco-serum of diabetic rats fed with Qianggubao decoction ([Chinese characters: see text]) on the differentiation and mineralization of osteoblast (OB). METHODS (OB) was isolated from the skull of 10 newly born SD rats aged 1 to 2 days by means of Trypsin-collagenase digestion. After the OB was identified, different kinds of pharmaco-serum of diabetic rats fed with inactive Qianggubao decoction ([Chinese characters: see text]) of different phase (rats were fed with medicine three days or five days after last fed with medicine one hour or three hours) and concentration (5%, 10%, 20%) were added to the OB and incubated. After 7 days and 18 days of culture,the effects of the differentiation and mineralization of osteoblast were detected. RESULTS: The secretion of ALP and formation of mineralized nodules of osteoblast in the different doses of pharmaco-serum groups were almost the same as that of normal control group, but were superior to that in the model control group. And the group with concentration of 20% pharmaco-serum was the best in the secretion of ALP and formation of mineralized nodules of osteoblast. As to the phases of pharmaco-serum, the best one on the differentiation and mineralization of osteoblast was the serums from diabetic rat-model fed with Qianggubao decoction ([Chinese characters: see text]) three days or five days, after one hour of last fed with medicine. CONCLUSION: The pharmaco-serum of diabetic rats fed with Qianggubao decoction ([Chinese characters: see text]) can promote the differentiation and mineralization of osteoblast. Allow for time and the cost of experiment,we presume that pharmaco-serum of diabetic rats fed with Qianggubao decoction ([Chinese characters: see text]) three days, after one hour of last fed, with concentration of 20% and not-inactivation is the optimum on the differentiation and mineralization of osteoblast.

[Effect of compound duzhong jiangu granule on knee joint osteoarthritis: a report of 400 cases].

OBJECTIVE: To study the clinical efficacy and adverse reaction of compound Duzhong Jiangu granule (CDJG) on knee joint osteoarthritis (KJO) of Gan-Shen deficiency with stasis in tendon and muscle syndrome. METHODS: The randomized controlled method was adopted in this study, comparative study was conducted in 400 patients in the treated group treated with CDJG and 200 patients in the control group treated with Zhuanggu Guanjie pill (ZGP). RESULTS: The total effective rate in the treated group was 92% and the curative-markedly effective rate was 47%, which were higher than those in the control group respectively. Moreover, CDJG showed superiority in improving symptoms and with shorter initiating time of action as compared with ZGP. However CDJG had the effect more favourable for patients of mild condition. CONCLUSION: CDJG is a kind of effective and safe medicine for treatment of KJO.

[Effect of Bugu Mixture on all-trans retinoic acid-induced apoptosis of bone marrow stromal cells].

OBJECTIVE: We used the SD rat's bone marrow stromal cells (BMSCs) cultured in vitro to observe the effects of Bugu Mixture on the apoptosis and to explore the molecular biologic mechanism of the treatment of osteoporosis with Bugu Mixture. METHODS: BMSCs were separated from the bones of the extremities of SD rats in vitro. The morphologic changes, the apoptosis cell cycles, the mitochondrion membrane potential changes, and the Bcl-2 and Bax gene expression were observed, and the effects of Bugu Mixture on the course of cell apoptosis were evaluated. RESULTS: The earlier use of Bugu Mixture could decrease the cells blocked in G0/G1 phase, and promote their synthesis of DNA in S phase. The expression of Bcl-2 was higher in the Bugu Mixture group than that in the all-trans retinoic acid (ATRA) induced group, and the expression of Bax was lower in the Bugu Mixture group than that in the ATRA induced group. The mitochondrion membrane potential descended significantly in the Bugu Mixture group than that in the ATRA induced group. CONCLUSION: The mechanism of the treatment of osteoporosis with Bugu Mixture is that the earlier use of Bugu Mixture can decrease the amount of apoptostic cells induced by ATRA, thus promoting the cell mitosis and restraining the apoptosis. It can also act as a protector to Bcl-2 located on the mitochondrion membrane. By preventing the transferring of the Bax protein from cell-plasma to mitochondrion membrane, it takes the advantage of Bcl-2 in forming Bcl-2/Bax homodimer so as to prevent the opening of the permeability transition pore to avoid the changing of mitochondrion membrane potential and the destruction of biosynthesis caused by the mitochondrion release of apoptosis inducing factors and to reach the objective of restraining apoptosis.