Effect and molecular mechanism of Sulforaphane alleviates brain damage caused by acute carbon monoxide poisoning:Network pharmacology analysis, molecular docking, and experimental evidence.

Sulforaphane (SFN) has attracted much attention due to its ability on antioxidant, anti-inflammatory, and anti-apoptotic properties, while its functional targets and underlying mechanism of action on brain injury caused by acute carbon monoxide poisoning (ACOP) have not been fully elucidated. Herein, we used a systematic network pharmacology approach to explore the mechanism of SFN in the treatment of brain damage after ACOP. In this study, the results of network pharmacology demonstrated that there were a total of 81 effective target genes of SFN and 36 drug-disease targets, which were strongly in connection with autophagy-animal signaling pathway, drug metabolism, and transcription disorders in cancer. Upon the further biological function and KEGG signaling pathway enrichment analysis, a large number of them were involved in neuronal death, reactive oxygen metabolic processes and immune functions. Moreover, based on the results of bioinformatics prediction associated with multiple potential targets and pathways, the AMP-activated protein kinase (AMPK) signaling pathway was selected to elucidate the molecular mechanism of SFN in the treatment of brain injury caused by ACOP. The following molecular docking analysis also confirmed that SFN can bind to AMPKα well through chemical bonds. In addition, an animal model of ACOP was established by exposure to carbon monoxide in a hyperbaric oxygen chamber to verify the predicted results of network pharmacology. We found that the mitochondrial ultrastructure of neurons in rats with ACOP was seriously damaged, and apoptotic cells increased significantly. The histopathological changes were obviously alleviated, apoptosis of cortical neurons was inhibited, and the number of Nissl bodies was increased in the SFN group as compared with the ACOP group (p < .05). Besides, the administration of SFN could increase the expressions of phosphorylated P-AMPK and MFN2 proteins and decrease the levels of DRP1, Caspase3, and Casapase9 proteins in the brain tissue of ACOP rats. These findings suggest that network pharmacology is a useful tool for traditional Chinese medicine (TCM) research, SFN can effectively inhibit apoptosis, protect cortical neurons from the toxicity of carbon monoxide through activating the AMPK pathway and may become a potential therapeutic strategy for brain injury after ACOP.

Early predictors of brain injury in patients with acute carbon monoxide poisoning and the neuroprotection of mild hypothermia.

INTRODUCTION: Carbon monoxide (CO) poisoning can cause serious neurological sequelae. However, there is neither effective treatment strategy nor reliable indicators to determine the prognosis of patients with CO poisoning. The present study aimed to observe the changes of neurological function score, disease severity score, cerebral oxygen utilization (O2UCc), bispectral (BIS) index and neuron-specific enolase (NSE) concentration, and to elucidate the clinical significance of these potential indicators and the neuroprotective effect of mild hypothermia on brain injury in patients with severe acute CO poisoning. MATERIALS AND METHODS: A total of 277 patients with acute severe CO poisoning from 2013 to 2018 were enrolled in our hospital. Patients were divided into three groups according to their body temperature on the day of admission and their willingness to treat: a fever group (n = 78), a normal temperature group (NT group, n = 113), and a mild hypothermia group (MH group, n = 86). All patients were given hyperbaric oxygen therapy, while those in the MH group received additional mild hypothermia treatment. The severity of the disease, the neurobehavioral status, the incidence of delayed encephalopathy after acute carbon monoxide poisoning (DEACMP), and other indicators including BIS, O2UCc, NSE were further evaluated in all patients at given time-points. RESULTS: Mild hypothermia therapy improved the prognosis of patients with CO poisoning, significantly decreased the value of O2UCc and NSE, and up-regulated BIS. The incidence of DEACMP at 6 months was 27% in the fever group, 23% in the NT group, and 8% in the MH group. The values of Glasgow-Pittsburgh coma scale (G-P score), BIS index and NSE were closely related to the occurrence of DEACMP, the cutoff values were 12.41, 52.17 and 35.20 ng/mL, and the sensitivity and specificity were 79.3%, 77.6%, 79.3% and 67.6%, 89.5%, 88.6% in the receiver operating characteristic curve (ROC), respectively. CONCLUSIONS: Early mild hypothermia treatment could significantly reduce the severity of brain injury after CO poisoning, and might be further popularized in clinic. G-P scores, NSE and BIS index can be regarded as the prediction indicators in the occurrence and development of DEACMP. CLINICAL TRIAL REGISTRATION: The study protocol was granted from Qingdao University Research Ethics Committee (Clinical trial registry and ethical approval number: QD81571283).

Therapeutic effect and molecular mechanism of Salvia Miltiorrhiza on rats with acute brain injury after carbon monoxide poisoning based on the strategy of internet pharmacology.

The pathogenesis of brain injury caused by carbon monoxide poisoning (COP) is very complex, and there is no exact and reliable treatment in clinic. In the present study, we screened the therapeutic target and related signal pathway of Salvia Miltiorrhiza for acute COP brain injury, and clarified the pharmacological mechanism of multicomponent, multitarget, and multisignal pathway in Salvia Miltiorrhiza by network pharmacology. To further verify the therapeutic effect of Salvia Miltiorrhiza on acute brain injury based on the results of network analysis, a total of 216 male healthy Sprague Dawley rats were collected in the present study and randomly assigned to a normal control group, a COP group and a Tanshinone IIA sulfonate treatment group (72 rats in each group). The rat model of acute severe COP was established by the secondary inhalation in a hyperbaric oxygen chamber. We found that Salvia Miltiorrhiza had multiple active components, and played a role in treating acute brain injury induced by COP through multiple targets and multiple pathways, among them, MAPK/ERK1/2 signaling pathway was one of the most important. COP can start apoptosis process, activate the MAPK/ERK1/2 signaling pathway, and promote the expression of VEGF-A protein and the formation of brain edema. Tanshinone IIA can effectively inhibit apoptosis, up-regulate the expressions of VEGF-A, P-MEK1/2 and P-ERK1/2 proteins, thereby protect endothelial cells, promote angiogenesis and microcirculation, and finally alleviate brain edema.

[Effects of activated coke on soil properties and growth of two plant species in saline alkali soil in northern Shanxi Province, China.] / 活性焦对晋北盐碱地土壤性质和两种植物生长的影响.

Solid waste-based improver is one of the effective means to improve properties of saline-alkali soil. As a kind of porous waste, activated coke is expected to improve soil properties and alleviate salt-alkali stress. In order to understand the improvement effect of activated coke on saline alkali land in northern Shanxi Province, we examined the effects of different addition rates of activated coke (CK, 0 g·kg-1; A10, 10 g·kg-1; A20, 20 g·kg-1; A50, 50 g·kg-1) on the properties of saline alkali soil and the growth of two plant species. The results showed that activated coke addition could increase the content of water soluble soil aggregates, reduce soil salt content, soil pH, and the electrical conductivity (EC). Compared with CK, the mean weight diameters of the aggregates for the saline-alkali soils grown with Puccinellia distans and maize were increased by 5.1%-32.2%, soil pH was decreased by 0.4%-4.1%, sodium adsorption ratio (SAR) was decreased by 4.8%-18.7%, and the EC was decreased by 7.4%-8.2%. Applying appropriate amount of activated coke could promote plant growth through reducing the plasma membrane damage of plant cells, increasing plant chlorophyll and Ca2+ contents. The biomass of Puccinellia distans and maize both reached the maximum under the A20 treatment. It suggested that the application of 20 g·kg-1 activated coke (A20) in saline alkali soil could improve soil quality in the rhizosphere soil, increase plant selective Ca2+ absorption, thereby reducing salt damage to plant cells and promote plant growth in saline-alkali habitat.

Strontium Ranelate Combined with Insulin Is as Beneficial as Insulin Alone in Treatment of Fracture Healing in Ovariectomized Diabetic Rats.

BACKGROUND Type 2 diabetes mellitus (T2DM) and estrogen deficiency both predispose fracture patients to increased risk of delayed union or nonunion. The present study investigated the effects of strontium ranelate (SR) on fracture healing in ovariectomized (OVX) diabetic rats. MATERIAL AND METHODS A mid-shaft fracture was established in female normal control (CF), diabetic (DF), and OVX diabetic (DOF) rats. Treated DOF rats received either insulin alone (DOFI) or combined with SR (DOFIS). All rats were euthanized at 2 or 3 weeks after fracture. Fracture healing was evaluated using radiological, histological, immunohistochemical, and micro-computed tomography analyses. RESULTS At 3 weeks after fracture, radiological and histological evaluations demonstrated delayed fracture healing in the DF group compared with the CF group, which was exacerbated by OVX, as indicated by the significantly lower X-ray score, BMD, BV/TV, and Md.Ar/Ps.Cl.Ar, and the markedly decreased OCN and Col I expression in the DOF group. All these changes were prevented by insulin alone or combined with SR treatment. In comparison with the DOFI group, DOFIS rats displayed markedly higher OCN expression at 2 weeks after fracture and Col I expression at 2 and 3 weeks after fracture. CONCLUSIONS These results demonstrated delayed fracture healing with preexisting estrogen deficiency and T2DM. While insulin alone and combined with SR were both effective in promoting bone fracture healing in this model, their combined treatment showed significant improvement in promoting osteogenic marker expression, but not of the radiological appearance, compared with insulin alone.

Effects of human parathyroid hormone 1-34 on bone loss and lumbar intervertebral disc degeneration in ovariectomized rats.

PURPOSE: Lumbar intervertebral disc degeneration is a common cause of lower back pain that affects the physical and mental health of patients and increases social burden. Parathyroid hormone has been reported to be effective at inhibiting disc degeneration; however, these effects have not been fully established in vivo in ovariectomized (OVX) rats. Thus, in this study, we aimed to address this issue and examine the effects of parathyroid hormone treatment in OVX rats. METHODS: Thirty female Sprague-Dawley rats, three months-old, were subjected to sham or ovariectomy surgery. Twelve weeks postsurgery, OVX rats were treated with either human parathyroid hormone [hPTH(1-34), 30 µg/kg/day] or vehicle (normal saline (NS)) treatment. The L3-6 spinal segments were harvested after 12 weeks treatment. Bone mineral density (BMD), micro-architectural parameters, and biomechanical assessment were measured at the lumbar vertebral bodies. Histology and immunohistochemistry were performed to analyze the characteristics of the lumbar intervertebral discs. RESULTS: OVX + PTH rats had significantly higher BMD, percentage bone volume density, trabecular thickness, and biomechanical strength compared with those in Sham and OVX + NS rats. Histology and immunostaining revealed that disc degeneration was not significantly different between the OVX + NS rats and the OVX + PTH rats, compared with the Sham group; the structure of nucleus pulposus was disordered, the expression of collagen I was increased, and collagen II and aggrecan were decreased. CONCLUSIONS: These findings confirmed that hPTH(1-34) treatment has substantial anabolic effects on bone mass and trabecular micro-architecture, while the excessively enhanced bone mass and strength were coupled with a non-significant effect on the disc degeneration in ovariectomized rats.

Orally administered simvastatin partially preserves lumbar vertebral bone mass but not integrity of intervertebral discs in ovariectomized rats.

The present study aimed to investigate the effect of orally administered simvastatin on lumbar vertebral bone mass and intervertebral disc (IVD) degeneration in ovariectomized (OVX) rats. A total of 30 female Sprague-Dawley (SD) rats were subjected to either bilateral ovariectomy (n=20) or sham surgery (n=10). After 12 weeks, the OVX rats were orally administered either saline vehicle (OVX + V group; n=10), or 5 mg/kg/day simvastatin (OVX + SIM group; n=10). Following 12 weeks of treatment, necropsy was conducted and bone mineral density (BMD) was determined in the L5-6 vertebrae. Furthermore, the microstructure and biomechanical properties of the L3 vertebrae were detected by micro-computed tomography and compression testing, respectively. The L5-6 vertebrae were analyzed by measurement of IVD height, observation of histological changes by van Gieson staining, and evaluation of collagen-II (col-II), aggrecan (AGG) and collagen I (col-I) expression by immunohistochemical analysis. Rats in the OVX+V group had lower BMD, bone volume/trabecular volume ratio, maximum load and elastic modulus than the sham group. Rats in the OVX + SIM group had higher BMD and biomechanical strength values than the rats in the OVX+V group. Histological analysis showed that the OVX + V and OVX + SIM groups exhibited significantly higher disc degeneration scores and significantly lower IVD height than the sham group. Immunohistochemical analysis revealed lower expression levels of col-II and AGG, but higher levels of col-I in the annulus fibrosis and endplate in OVX+V rats compared with the sham group. The OVX + SIM group exhibited levels of col-II, AGG and col-I expression comparable with those of OVX+V rats, with the exception of an upregulation of col-II expression in the annulus fibrosis. These data demonstrate that simvastatin treatment partially prevented bone loss and the deterioration of biomechanical properties of lumbar vertebrae, but not the progression of IVD degeneration in OVX rats.

Strontium ranelate causes osteophytes overgrowth in a model of early phase osteoarthritis.

BACKGROUND: Osteoarthritis (OA) involves cartilage changes as well as modifications of subchondral bone and synovial tissues. Strontium ranelate (SR), an anti-osteoporosis compound, which is currently in phase III clinical trial for treatment of OA. Evidences suggest that SR preferably deposited in osteophyte, other than in subchondral bone in early phase of OA. This phenomenon raises concern about its utility for OA treatment as a disease-modifying drug. To evaluate the effect of SR on cartilage, subchondral bone mass and subchondral trabecular bone structure in medial meniscectomized (MNX) guinea pigs. METHOD: Thirty-six 3-month-old male Dunkin Hartley albino guinea pigs received either sham or medial meniscectomy operations. One week after the procedure, meniscectomized animals began 12 weeks of SR (625 mg/kg, daily) treatment by oral gavage for MNX + SR group, or normal saline for MNX + V group. All animals were euthanized 12 weeks later, cartilage degeneration and subchondral bone micro-architecture was analyzed. RESULTS: Both OARSI scores (P = 0.523 for marcoscopic scores, P = 0.297 for histological scores) and Cartilage thickness (P = 0.335) in MNX + SR group were comparable to MNX + V group. However, osteophyte sizes were larger in MNX + SR group (P = 0.014), and collapsed osteophytes in MNX + SR group (7 by 12) were significantly more than in MNX + V group (1 by 12) (P = 0.027), while immunohistochemistry indicates catabolic changes in osteophyte/plateau junction. Micro-CT analysis showed bone mineral density (BMD) (P = 0.001), bone volume fraction (BV/TV) (P = 0.008), trabecular spacing (Tb.Sp) (P = 0.020), trabecular thickness (Tb.Th) (P = 0.012) and structure model index (SMI) (P = 0.005) levels to be significantly higher in the MNX + SR group than in the MNX + V group. CONCLUSIONS: SR (625 mg/kg/day) did not protect cartilage from degeneration in MNX guinea pigs but subchondral bone was significantly enhanced. In early phase OA, SR administration causes osteophyte overgrowth, which may be related to incorporation into mineralizing osteophytes. This adverse effect is important for future studies of SR in OA.

Enhancement of Lumbar Fusion and Alleviation of Adjacent Segment Disc Degeneration by Intermittent PTH(1-34) in Ovariectomized Rats.

Osteoporosis, which is prevalent in postmenopausal or aged populations, is thought to be a contributing factor to adjacent segment disc degeneration (ASDD), and the incidence and extent of ASDD may be augmented by osteopenia. Parathyroid hormone (PTH) (1-34) has already been shown to be beneficial in osteoporosis, lumbar fusion and matrix homeostasis of intervertebral discs. However, whether PTH(1-34) has a reversing or retarding effect on ASDD in osteopenia has not been confirmed. In the present study, we evaluated the effects of intermittent PTH(1-34) on ASDD in an ovariectomized (OVX) rat model. One hundred 3-month-old female Sprague-Dawley rats underwent L4 -L5 posterolateral lumbar fusion (PLF) with spinous-process wire fixation 4 weeks after OVX surgery. Control groups were established accordingly. PTH(1-34) was intermittently administered immediately after PLF surgery and lasted for 8 weeks using the following groups (n = 20) (V = vehicle): Sham+V, OVX+V, Sham+PLF+V, OVX+PLF+V, OVX+PLF+PTH. The fused segments showed clear evidence of eliminated motion on the fusion-segment based on manual palpation. Greater new bone formation in histology was observed in PTH-treated animals compared to the control group. The extent of ASDD was significantly increased by ovariotomy. Intermittent PTH(1-34) significantly alleviated ASDD by preserving disc height, microvessel density, relative area of vascular buds, endplate thickness and the relative area of endplate calcification. Moreover, protein expression results showed that PTH(1-34) not only inhibited matrix degradation by decreasing MMP-13, ADAMTS-4 and Col-I, but also promote matrix synthesis by increasing Col-II and Aggrecan. In conclusion, PTH(1-34), which effectively improves lumbar fusion and alleviates ASDD in ovariectomized rats, may be a potential candidate to ameliorate the prognosis of lumbar fusion in osteopenia.

The inhibitory effect of salmon calcitonin on intervertebral disc degeneration in an ovariectomized rat model.

PURPOSE: Intervertebral disc degeneration related to postmenopausal osteoporosis is an important issue in spinal disorder research. This study aimed to investigate the effects of salmon calcitonin (sCT), as an antiresorptive medication, on lumbar intervertebral disc degeneration using a rat ovariectomy (OVX) model. METHODS: Thirty 3-month-old female Sprague-Dawley rats were randomly divided into three groups: the sham-operated (Sham) group and two ovariectomized groups treated with vehicle (OVX+V) or sCT (OVX+CT; 16 IU/kg, sc) on alternate days for 6 months. Treatment began after OVX and continued for 6 months. At the end of the experiment, bone mineral density (BMD), micro-CT analysis, biomechanical testing, histology, and immunohistochemistry were performed for all groups. RESULTS: Salmon calcitonin significantly maintained vertebrae BMD, percent bone volume, and biomechanical strength, when compared with the OVX+V group. The changes of mucoid degeneration in the nucleus pulposus and calcification in the middle cartilage endplate were more moderate in the OVX+CT group compared with the OVX+V group, and immunohistochemistry revealed a significant increase in aggrecan and type II collagen expressions, but marked reductions in matrix metalloproteinase (MMP)-1, MMP-3, and MMP-13 expressions in the OVX+CT group. CONCLUSIONS: Salmon calcitonin treatment was effective in delaying the process of the disc degeneration in OVX rats. The underlying mechanisms may be related to preservation of structural integrity and function of vertebrae, and affecting extracellular matrix metabolism by modulating the expressions of MMPs, aggrecan and type II collagen to protect the disc from degeneration.

Alendronate Prevents Intervertebral Disc Degeneration Adjacent to a Lumbar Fusion in Ovariectomized Rats.

STUDY DESIGN: A model of disc degeneration adjacent to a lumbar fusion in osteoporotic rats. OBJECTIVE: We determined the effect of alendronate (ALN) on the disc degeneration adjacent to a lumbar fusion in ovariectomized rats. SUMMARY OF BACKGROUND DATA: Adjacent-segment disc degeneration (ASDD) is one of the negative sequelae of spinal fusion. Previous studies have shown that ALN can alleviate disc degeneration. However, no data have been documented regarding the effect of ALN on ASDD after posterolateral lumbar fusion (PLF) in osteoporosis. METHODS: 50 female Sprague-Dawley rats underwent either a sham operation (sham) (n = 20) or bilateral ovariectomy (OVX) (n = 30). 4 weeks later, all but 10 rats from each group underwent PLF consisting of an intertransverse process spinal fusion using autologous-iliac-bone grafts with spinous-process wire fixation at the L4-L5 segment. Animals were subcutaneously administered vehicle (V) or ALN (70 µg/kg/wk) for 12 weeks post-PLF as follows: Sham+V, OVX+V, PLF+V, OVX+PLF+V, and OVX+PLF+ALN. Fusion status was analyzed by manual palpation and radiography. Adjacent-segment disc was assessed by histological, histomorphometric, immunohistochemical, and mRNA analysis. L6 vertebrae microstructures were evaluated by microcomputed tomography. RESULTS: The fused segments showed clear evidence of fusion based on manual palpation and radiographs. The OVX+PLF+V group showed more severe degenerative alterations and higher histological scores in the disc than the Sham+V, OVX+V, and PLF+V groups (P < 0.05). Compared with the OVX+PLF+V group, the OVX+PLF+ALN group exhibited significantly improved bone mass and vertebrae microstructures (P < 0.05), an increased disc height, and a decreased endplate calcification area (P < 0.05). ALN also significantly decreased Col-I, MMP-13, and ADAMTS-4 expression and increased Col-II and Aggrecan expression in the disc matrix (P < 0.05). CONCLUSION: ALN effectively alleviated ASDD post-PLF in ovariectomized rats. These data indicate that ALN can be used as a potential therapeutic agent to attenuate ASDD progression in osteoporosis.

Age dependent changes in cartilage matrix, subchondral bone mass, and estradiol levels in blood serum, in naturally occurring osteoarthritis in Guinea pigs.

The Dunkin Hartley (DH) guinea pig is a widely used naturally occurring osteoarthritis model. The aim of this study was to provide detailed evidence of age-related changes in articular cartilage, subchondral bone mineral density, and estradiol levels. We studied the female Dunkin Hartley guinea pigs at 1, 3, 6, 9, and 12 months of age (eight animals in each group). Histological analysis were used to identify degenerative cartilage and electron microscopy was performed to further observe the ultrastructure. Estradiol expression levels in serum were assessed, and matrix metalloproteinase 3 and glycosaminoglycan expression in cartilage was performed by immunohistochemistry. Bone mineral density of the tibia subchondral bone was measured using dual X-ray absorptiometry. Histological analysis showed that the degeneration of articular cartilage grew more severe with increasing age starting at 3 months, coupled with the loss of normal cells and an increase in degenerated cells. Serum estradiol levels increased with age from 1 to 6 months and thereafter remained stable from 6 to 12 months. Matrix metalloproteinase 3 expression in cartilage increased with age, but no significant difference was found in glycosaminoglycan expression between 1- and 3-month old animals. The bone mineral density of the tibia subchondral bone increased with age before reaching a stable value at 9 months of age. Age-related articular cartilage degeneration occurred in Dunkin Hartley guinea pigs beginning at 3 months of age, while no directly positive or negative correlation between osteoarthritis progression and estradiol serum level or subchondral bone mineral density was discovered.

An increase in the incidence of hip fractures in Tangshan, China.

UNLABELLED: We determined the number and incidence of hip fractures in Tangshan, China, in 2010. Compared with data we reported in Tangshan from 1994, the crude and age-specific incidence increased significantly for both sexes, especially in women. Strategies are needed for effective fracture prevention in the future. INTRODUCTION: The aims of the study were to determine the incidence of cervical and trochanteric fractures of the proximal femur in Tangshan, China, in 2010 and to compare the incidence with data from 1994. METHODS: The orthopedic departments of 15 hospitals in Tangshan were visited in 2010; the medical records and radiographs of patients who had sustained cervical and trochanteric fractures were reviewed. The absolute number of admissions was collated and the incidence rate per 100,000 person years was calculated, adjusted by different age ranges, and gender. We then calculated the age-standardized incidence in 2010 as compared with those from 1994. RESULTS: The population of Tangshan in 2010 was determined to be 3,075,382 (1,558,173 males; 1,517,209 females); there were 1,509 cervical and trochanteric fractures (in 745 males and 764 females). The overall incidence was 47.8 and 50.4 fractures per 100,000 per year for men and women, respectively. Females showed a higher fracture incidence than males in those aged 55 years and over. Comparing the 2010 data with the 1994 findings, the incidence increased by 85% in men and by 306% in women; age-specific increases were observed in all female and male groups (except the 55-59 years age group). CONCLUSIONS: Compared with the results in 1994, the incidence of hip fracture has markedly increased in 2010 in Tangshan, China. It is necessary to implement a comprehensive policy for hip fracture prevention in our communities.

Alendronate retards the progression of lumbar intervertebral disc degeneration in ovariectomized rats.

OBJECTIVE: Increasing evidence has revealed a positive correlation between postmenopausal osteoporosis and intervertebral disc degeneration, the underlying mechanism of which might be associated with changes in the vertebral bone and endplate. Alendronate (ALN) can increase bone mass and improve the microstructure of osteoporotic vertebrae, which might be helpful in preserving disc morphology and mechanical properties. This study aims to investigate the effects of ALN on lumbar intervertebral disc degeneration related to osteoporosis using an ovariectomized (OVX) rat model. METHODS: Thirty female Sprague-Dawley rats aged 3 months were randomly divided into three groups (with 10 rats each) as follows: the Sham group underwent sham surgery; the OVX + ALN group had twice-a-week subcutaneous injections of ALN (15 µg/kg) for 6 months. The OVX + V group received an equivalent volume of saline solution as placebo post-OVX. After animals were sacrificed at 6 months post-OVX, the L3-6 spinal segments were harvested. Bone mineral density (BMD), micro-CT analysis and biomechanical testing were performed to evaluate the bone quality and microstructural changes in the lumbar vertebral bodies. Histological analysis with van Gieson stain and the histological score were used to identify the characteristics of the degenerative discs. The disc height and the thickness of the cartilage endplate were measured and compared. Immunohistochemistry and real-time PCR measurements for aggrecan, type I collagen, type II collagen, and matrix metalloprotease (MMP)-1, MMP-3 and MMP-13 expressions on the disc were performed to assess the underlying molecular signaling changes in matrix metabolism during intervertebral disc degeneration. RESULTS: The OVX + ALN group significantly maintained vertebrae BMD, percent bone volume and biomechanical strength, when compared with the OVX + V group. Histological evaluation suggests that there was no significant difference in disc height between the OVX + ALN and Sham groups, and ALN significantly prevented cartilage endplate thickening and the development of abnormal bony tissues within the cartilage endplate. The histological score in the OVX + ALN group was significantly lower than the OVX + V group, suggesting that ALN treatment was effective in delaying the process of the disc degeneration. The results of molecular analysis revealed a significant increase in aggrecan and type II collagen expressions, but marked reductions in MMP-1, MMP-3 and MMP-13 expressions at both the protein and mRNA levels in the OVX + ALN group. CONCLUSIONS: ALN can retard the progression of lumbar intervertebral disc degeneration in OVX rats. The underlying mechanisms might be related to preservation of the structural integrity and function of the adjacent structures, including the vertebrae and endplates, which further links with modulations in extracellular matrix metabolism to protect the disc from degeneration. These results suggest that ALN might be a promising drug agent for preventing lumbar intervertebral disc degeneration related to osteoporosis.

[Experimental study on tissue engineering platelet lysates in the promotion of bone reconstruction].

OBJECTIVE: To evaluate the effects of tissue engineered allogeneic platelet lysates (PL) upon bone reconstruction. METHODS: After preparation of recombinant material with PL, allogeneic decalcified bone granules (ADBG) and collagen type I (CG), 30 healthy Wistar rats were used to prepare the bilateral bone defects in femoral condyles. The defects were filled with equivalent PL/ADBG/CG, ADBG/CG and CG in different groups of A, B and C (with 10 rats each) respectively. At 4 weeks, the defect reconstruction was evaluated with radiology, histology, immunology and biomechanics. RESULTS: (1) The X-ray showed that bone density in group A (4.18 +/- 0.96) was close to that of normal bone and it was significantly higher than that in group B (2.36 +/- 0.87) and group C (1.09 +/- 0.55) (P < 0.01). (2) In comparisons with B and C, the histological assay revealed that there were markedly more activities of new bone formation and more implanted bone granules surviving without significant lymphocyte infiltration, as well as more osteoclastic bone resorption in group A. The bone histomorphometric assay showed the newly formed bone area in group A (286.73 +/- 17.22) was significantly higher than that in group B (94.34 +/- 33.56) and group C (19.12 +/- 14.53) (P < 0.01). (3) Anti-press mechanical measures showed that the destructive load in A, B, C and normal control group was 259.63 +/- 34.57, 187.90 +/- 21.07, 91.33 +/- 26.58 and 311.93 +/- 82.45 respectively. The destructive energy in A, B, C and normal control group was 10.82 +/- 1.44, 7.83 +/- 0.88, 3.81 +/- 1.11 and 12.97 +/- 3.43 respectively. These results showed either destructive load or destructive energy in group A was markedly higher than that in group B and group C with significant difference (P < 0.01), but still lower than that in normal controls (P < 0.01). (4) Three-color flow cytometry assay showed that the T lymphocyte subsets of CD3+CD4+CD8-, CD3+CD8+CD4- and the ratio of CD4/CD8 showed no significance difference within these three groups as well as normal controls. CONCLUSION: Tissue engineering PL (PL/ADBG/CG) is capable of accelerating the regenerative repair of bone defect and promoting the bone regeneration and osetointergretion of allograft bone after transplantation.

Application of a magnetotactic bacterium, Stenotrophomonas sp. to the removal of Au(III) from contaminated wastewater with a magnetic separator.

In this study, the feasibility of applying a magnetotactic bacterial isolate (MTB), Stenotrophomonas sp. to the removal of Au(III) was investigated. Biosorption experiments showed that Au(III) biosorption capacity exhibited no significant difference in the initial pH range of 1.0-5.5, while decreased more significantly in the initial pH range of 5.5-13.0. Langmuir isotherm indicated that the maximum Au(III) biosorption capacity of Stenotrophomonas sp. were 506, 369 and 308 mg g(-1) dry weight biomass at the initial pH values of 2.0, 7.0 and 12.0, respectively. Thiourea was proved to be an effective desorbent to recover Au from the MTB biomass and 91% Au adsorbed on the biomass could be recovered at equilibrium when the thiourea concentration was 0.8M. The magnetic separator developed by our research team used for separating Au loaded MTB biomass showed high separation efficiency, with 100% biomass removed at the magnetic intensity of 1200 Gs in 180 min. The analyses from FTIR and XRD further confirmed that the reduction of Au(III) to Au(0) by the reductants on the MTB biomass occurred, and the deposition of nano-crystal Au(0) particles, ranging from 24.7 to 31.4 nm, could be estimated on the biomass surface.

Effects of ovariectomy and 17beta-estradiol treatment on the renin-angiotensin system, blood pressure, and endothelial ultrastructure.

The purpose of this study was to determine whether the renin-angiotensin system (RAS), nitric oxide (NO), atrial natriuretic peptide (ANP), blood pressure (BP), ultrastructural characteristics, and endothelium-dependent relaxation of thoracic aorta were modulated by the estrogen level. Rats were divided into 3 groups: ovariectomized (OVX); not ovariectomized (sham); and ovariectomized and treated with subcutaneous 17beta-estradiol (15 microg/kg/day, OVX+E(2)) (n=15-17 per group). For 13 weeks after surgery, blood pressure, serum estrogen, NO, plasma angiotensin II (Ang II), ANP, and renin activity levels were monitored. Thirteen weeks after surgery, the vasodilator responses of the aortic rings to acetylcholine and the ultrastructural characteristics of the thoracic aorta were determined. In the 9th and 13th week, OVX rats had a significantly higher blood pressure than the other two groups (p<0.05). Ovariectomy led to a significant decrease in plasma Ang II level and a significant increase in renin activity in OVX rats compared to sham rats; this effect could be reversed by estrogen treatment. In the 5th, 9th, and 13th weeks, the serum NO level was significantly lower in the OVX group than in the sham group (p<0.05); this effect could be reversed by estrogen treatment. Plasma ANP levels in the 9th and 13th weeks were significantly lower in the OVX group (p<0.05), and plasma ANP levels could be completely restored by estrogen treatment. Ovariectomy markedly reduced endothelium-dependent relaxation in response to acetylcholine in isolated rat thoracic aortic rings; chronic estrogen treatment significantly restored endothelium-dependent relaxation in response to acetylcholine. Under electron microscopy, the endothelial cells in OVX rats were swollen, even necrosed; estrogen treatment inhibited these changes. These results strongly suggest that estradiol protects rats from the development of hypertension and has a protective effect on the endothelium by increasing NO and ANP levels while decreasing renin activity. However, there was a discordance between the effects that estradiol had on angiotensin II and on blood pressure. This might be the result of negative feedback that ultimately results in the overall suppression of the RAS.

17Beta-oestradiol regulates the expression of Na+/K+-ATPase beta1-subunit, sarcoplasmic reticulum Ca2+-ATPase and carbonic anhydrase iv in H9C2 cells.

1. It is necessary to improve our understanding of the effect of 17beta-oestradiol (E2) on the heart at a molecular and cellular level. In the present study, the effects of E2 on Na(+)/K(+)-ATPase, sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPase (SERCA) and carbonic anhydrase IV (CAIV) in H9C2 cells were investigated. To identify the mechanism of action of E2 on these proteins, the oestrogen receptor (ER) antagonist tamoxifen was used. 2. The results indicated that 1 and 100 nmol/L E2 can enhance the activity of Na(+)/K(+)-ATPase and SERCA and upregulate the expression of the Na(+)/K(+)-ATPase beta1-subunit, SERCA2a and CAIV at both the mRNA and protein level compared with 0 and 0.01 nmol/L E2. 17beta-Oestradiol had the greatest effect at 100 nmol/L; 1 micromol/L E2 did not further protein expression compared with 100 nmol/L E2. 3. Tamoxifen (10 nmol/L) significantly decreased the activity of SERCA, as well as the expression of the Na(+)/K(+)-ATPase beta1-subunit and SERCA at the mRNA and protein level, in H9C2 cells cultured with 1 nmol/L E2. Tamoxifen alone had no significant effect on these proteins in H9C2 cells. 4. It may be hypothesized that a suitable E2 concentration has a protective effect on the heart and that the actual dose of E2 used in hormone-replacement therapy is important in menopausal women.

[Application of allogeneic bone in surgical treatment of benign bone neoplasm].

OBJECTIVE: To evaluate the clinical outcomes of allogeneic bone grafting for bone defect resulting from benign neoplasm resection and discuss the clinical application and bone defect repair mechanisms of allogeneic bone. METHODS: A retrospective review was conducted of 135 patients with benign neoplasm resection who received bone defect filling with the allogeneic bone graft. RESULTS: In the 104 patients with complete clinical follow-up data, 96 achieved bone union, 7 experienced relapses to require surgical intervention and 1 had severe infection to lead to failure of the operation. The mean time for bone union was 9.7 months, and during the follow-up, no viral disease in relation to the graft was found after surgery. CONCLUSION: Bone defect filling with allogeneic bone graft can be simple and safe in comparison with that with autograft or other biomaterials, and the bone healing time, infection rate and local tumor recurrence can be comparable with the autograft.