Different Intermolecular Interactions Drive Nonpathogenic Liquid-Liquid Phase Separation and Potentially Pathogenic Fibril Formation by TDP-43.

The liquid-liquid phase separation (LLPS) of proteins has been found ubiquitously in eukaryotic cells, and is critical in the control of many biological processes by forming a temporary condensed phase with different bimolecular components. TDP-43 is recruited to stress granules in cells and is the main component of TDP-43 granules and proteinaceous amyloid inclusions in patients with amyotrophic lateral sclerosis (ALS). TDP-43 low complexity domain (LCD) is able to de-mix in solution, forming the protein condensed droplets, and amyloid aggregates would form from the droplets after incubation. The molecular interactions regulating TDP-43 LCD LLPS were investigated at the protein fusion equilibrium stage, when the droplets stopped growing after incubation. We found the molecules in the droplet were still liquid-like, but with enhanced intermolecular helix-helix interactions. The protein would only start to aggregate after a lag time and aggregate slower than at the condition when the protein does not phase separately into the droplets, or the molecules have a reduced intermolecular helix-helix interaction. In the protein condensed droplets, a structural transition intermediate toward protein aggregation was discovered involving a decrease in the intermolecular helix-helix interaction and a reduction in the helicity. Our results therefore indicate that different intermolecular interactions drive LLPS and fibril formation. The discovery that TDP-43 LCD aggregation was faster through the pathway without the first protein phase separation supports that LLPS and the intermolecular helical interaction could help maintain the stability of TDP-43 LCD.

[Influence of Land Use Change on Ecosystem Service Value Based on GEE in the Beijing-Tianjin-Hebei Region from 1998 to 2018].

In sustainable development assessment of the Beijing-Tianjin-Hebei region, the ability to dynamically estimate the value of ecosystem services is of great significance. This study considers the Beijing-Tianjin-Hebei region as the research area, based on the google earth engine (GEE); the classification and decision tree (CART) classification algorithm was adopted to supervise and classify the Landsat Thematic Mapper/Operational Land Imager (TM/OLI) images in the study area in 1998, 2003, 2008, 2013, and 2018, and land use types in these five periods were obtained. Quantitative analysis of the dynamic changes of land use in the Beijing-Tianjin-Hebei region from 1998 to 2018 was carried out. Then, the ecosystem service value (ESV) equivalent estimation method was used to quantitatively estimate the ESV in the Beijing-Tianjin-Hebei region and combine it with a 15 km×15 km scale grid to detect its temporal and spatial dynamics. The main results were as follows. ① From 1998 to 2018, the area of construction land (increased by 16.67%) and grassland (reduced by 13.73%) in the six land use types in the Beijing-Tianjin-Hebei region was the largest, and the change in the proportion of water bodies (0.2%) was the smallest. ② The total value of ESV in the Beijing-Tianjin-Hebei region experienced a short-term increase from 1998 to 2003 (an increase of 91.97×108 yuan), and continued to decrease from 2003 to 2018 (a decrease of 239.07×108 yuan), mainly related to the expansion of construction land area in the other three time periods excluding 1998 and 2003. Among the six land use types, the forest provides the highest value of ecosystem services, and the construction land and unused land provide the lowest value of ecosystem services. ③ The ESV time-space analysis based on the 15 km×15 km scale grid showed that the ESV medium area in the Beijing-Tianjin-Hebei region gradually decreased from 1998 to 2018, the ESV lower area and the higher area gradually increased, and the ESV lower-area growth rate was higher than for the higher area. ④ The revised value of the Beijing-Tianjin-Hebei region (sensitivity coefficient range 0-0.83) has good significance and reliability. In future economic development, the Beijing-Tianjin-Hebei region should rationally optimize the land use pattern and strengthen the protection of forest land, grassland, water bodies and cultivated land.

Comparisons of Emu Necrotic Femoral Head Micro Structure Repaired in Two Different Methods / 中国医学科学院学报

<p><b>OBJECTIVE</b>To compare emu necrotic femoral head micro structure repaired in two different methods.</p><p><b>METHODS</b>Fifteen adult emus were divided into 3 groups (all n=5), and the right femoral head was selected to research. The first group was the control group; in the second group, femoral head necrosis was made by cryogen with liquid nitrogen; and in the third group, femoral head necrosis was made by local pure ethanol injection. Right femurs were taken for micro CT examination,then femoral head micro structures were compared among these three groups.</p><p><b>RESULTS</b>No infection or unexpected death was found in all groups. Compared with normal group, necrotic femoral heads in cryogen group showed that bone mineral density significantly reduced after repaire (P=0.015), trabecular space significantly reduced (P=0.001), bone volume fraction significantly enlarged (P=0.036), bone surface/volume fraction (P=0.032) and trabecular numbers (P=0.002) significantly enlarged; trabecular thickness showed no significant difference (P=0.060). Compared with control group, necrotic femoral heads in ethanol group showed that bone mineral density significantly enlarged after repaire (P=0.001), trabecular thickness (P=0.003) and bone surface/volume fraction (P=0.022) significantly enlarged, trabecular space (P=0.001) and bone volume fraction (P=0.001) significantly reduced; the trabecular numbers showed no significant difference (P=0.143). Compared with ethanol group, necrotic femoral heads in cryogen group showed significant lower bone mineral density after repair (P=0.001), significantly lower bone volume fraction (P=0.001), significantly lower trabecular thickness (P=0.001), significantly higher bone surface/volume fraction (P=0.022) and higher trabecular numbers (P=0.003); the trabecular space showed no significant difference (P=0.398).</p><p><b>CONCLUSION</b>Different repair methods make reconstructed femoral head weight bearing area have different bone structure and bone mineral density, along with different bone trabecular quality.</p>

Research progress on epithelial-mesenchymal transition in cancer recurrence and metastasis / 浙江大学学报·医学版

Epithelial-mesenchymal transition (EMT) is a process in which epithelial cells lose their morphology and function and gradually transformed into mesenchymal-like cells. It is considered that EMT is the main cause for tumor recurrence and metastasis. Many factors are involved in the regulation of EMT, such as E-cadherin, transforming growth factor-β, Wnt signaling pathway, microRNA and EMT-related transcription factors. This article reviews the research progress on EMT and the involved mechanisms, and thus to provide a new perspective on cancer therapy in the future.

Antioxidative effects of cysteinyl leukotriene receptor antagonists montelukast and HAMI 3379 on ischemic injury in rat cortical neurons in vitro / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To investigate the antioxidative effects of two cysteinyl leukotriene receptors antagonists (CysLT1R and CysLT2R) montelukast and HAMI 3379 on ischemic injury of rat cortical neurons in vitro.</p><p><b>METHODS</b>Cultured rat cortical neurons were pretreated with CysLT1R antagonist montelukast and CysLT2R antagonist HAMI 3379, and then exposed to oxygen-glucose deprivation/recovery (OGD/R）or H2O2. Reactive oxygen species (ROS) mitochondrial membrane potential (MMP) depolarization, neuronal viability and lactate dehydrogenase (LDH) release were determined. Meanwhile, RNA interference was used to inhibit the expression of CysLT1R and CysLT2R，and the effects were observed.</p><p><b>RESULTS</b>ROS production in neurons was significantly increased after 1 h OGD, which reached the peak at 30 min and lasted for 1.5 h after recovery. Montelukast and HAMI 3379 at 0.01-1μmol/L moderately decreased OGD/R-induced ROS production (P<0.05). Montelukast mildly attenuated OGD/R-induced MMP depolarization (P<0.05)，but HAMI 3379 had no effect. H2O2 reduced neuronal viability and increased LDH release, namely inducing neuronal injury. Montelukast and HAMI 3379 at 0.1-1μmol/L moderately attenuated H2O2-induced neuronal injury (P<0.05). However, both CysLT1R siRNA and CysLT2R shRNA did not significantly affect the responses mentioned above.</p><p><b>CONCLUSION</b>In ischemic neuronal injury, montelukast and HAMI 3379 exert a moderate antioxidative effect, and this effect may be receptor-independent.</p>

Effect of histone deacetylase inhibitor NL101 on rat neurons / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To investigate the protective effect of histone deacetylase inhibitor NL101 on L-homocysteine (HCA)-induced toxicity in rat neurons, and the toxic effect on normal rat neurons.</p><p><b>METHODS</b>In the presence of NL101 at various concentrations, HCA (5 mmol/L)-induced changes in cell density, necrosis, and viability were determined in the mixed cultures of rat cortical cells and the primary cultures of rat neurons. The direct effect of NL101 on primary neurons was also observed in the absence of HCA. Histone deacetylase inhibitor suberoylanilide hydroxamic acid (SAHA) was used as the control. After the treatments, cell viability, the density, and morphology of neurons and glial cells, and cell necrosis were determined.</p><p><b>RESULTS</b>In the mixed cultures of cortical cells, NL101 had no effect on HCA (5 mmol/L)-induced cell number reduction at 0.001-10μmol/L; however, it significantly attenuated necrosis at 1-10 μmol/L, and increased neuronal number at 1 μmol/L. NL101 had no effect on the mixed cortical cells in the absence of HCA. In the primary neurons, NL101 reduced neuronal viability and mildly increased necrosis at 1-10 μmol/L in the absence of HCA, while it significantly attenuated HCA-induced neuronal viability reduction at 0.01-10 μmol/L and reduced neuronal necrosis at 1-10 μmol/L. The effects of NL101 were apparently similar to those of SAHA.</p><p><b>CONCLUSION</b>NL101 has protective effect on HCA-induced neuronal injury but it is neurotoxic at high concentrations, which is similar to the typical histone deacetylase inhibitor SAHA.</p>

Effects of the water channel aquaporin 4 deficiency on bleomycin-induced lung fibrosis in mice / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To evaluate the effect of water channel aquaporin 4 (AQP4) on bleomycin-induced lung fibrosis in mice.</p><p><b>METHODS</b>In wild type and AQP4 gene knockout (AQP4-/-) mice, lung fibrosis was induced by injection of bleomycin (3 mg/kg) into the trachea and saline injection was used as a control. At d3, 7, 14, 28 after bleomycin-treatment, mice were randomly sacrificed in batch and the lung coefficient was determined. Serum levels of TGF-β1 and TNF-α were measured by ELISA and hydroxyproline contents in lung tissue were determined by Alkaline hydrolysis method. H-E staining and Masson's staining were performed to examine the pathological changes of lung tissues after bleomycin-treatment.</p><p><b>RESULTS</b>On d14 after bleomycin-treatment, the lung coefficients in wild type mice and AQP4-/- mice were 1.9-fold (12.69 ± 6.05 vs 6.80 ± 0.82, q=4.204, P<0.05) and 2.3-fold (14.05 ± 5.82 vs 6.05± 0.58, q=5.172, P<0.01) of that in control, respectively, but no significant difference was found between wild type and AQP4-/- mice in the lung coefficient value (P>0.05). The hydroxyproline contents in the lung increased after bleomycin-treatment; on d28, the lung hydroxyproline contents in wild type and in AQP4-/- mice were 1.55-fold (0.85 ± 0.22 g/mg vs 0.55 ± 0.14 μg/mg, q=4.313, P<0.05) and 1.4-fold (0.84 ± 0.13 μg/mg vs 0.60 ± 0.14μg/mg, q=4.595，P<0.05) of that in control, respectively, but no significant difference was noticed between wild type and AQP4-/- mice in lung hydroxyproline contents. There was a tendency that serum TGF-β1 and TNF-α levels increased in bleomycin-treated mice, but no significant difference was found between wild type and AQP4-/- mice. AQP4-knockout showed no effects on pathological changes of lung tissues with H-E staining and Masson's staining in mice with bleomycin-induced lung fibrosis.</p><p><b>CONCLUSION</b>AQP4 might not be involved in bleomycin-induced lung fibrosis in mice.</p>

Effects of leukotriene D4 on proliferation and migration of lung epithelial A549 cells in vitro / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To investigate the effects of cysteinyl leukotriene (CysLT) receptor agonist leukotriene D4 (LTD4) on proliferation and migration in lung epithelial A549 cells.</p><p><b>METHODS</b>The expression of CysLT1 receptor and CysLT2 receptor was determined by immunofluoresence staining in A549 cells. A549 cells were treated with LTD4 (0.01-100 nmol/L) for 24-72 h. Cell viability was detected by MTT reduction assay. Cell migration was determined by modified scratch and healing model.</p><p><b>RESULTS</b>In A549 cells, CysLT1 receptor and CysLT2 receptor were mainly expressed in the cytoplasm, membrane and few in the nuclei. The treatment of LTD4 (0.01-100 nmol/L) for 24-72 h caused no effect on cell viability (Ps>0.05); when A549 cells were treated with 100 nmol/L LTD4 for 24, 48 and 72 h the cell viability was (103.00±4.46)%，(107.00±9.45)% and (105.00±9.02)% of control, respectively (Ps>0.05). The migration rate of A549 cells after scratching during the first 24 h was markedly greater than that during the second and third 24 h in the same concentration groups; however, no significant difference in migration rate was noticed when the cells were treated with different concentrations of LTD4 (0.01-100 nmol/L)(Ps>0.05). The migration of A549 cells was 1.15-fold, 1.21-fold and 1.06-fold of that of control when the cells were treated with 100 nmol/L LTD4 for 24, 48 and 72 h, respectively (Ps>0.05).</p><p><b>CONCLUSION</b>The proliferation and migration of A549 cells are not changed when treated with 0.01-100 nmol LTD4 for up to 72h.</p>

Research advances on the roles of nicotinamide phosphoribosyltransferase in inflammation / 浙江大学学报·医学版

Nicotinamide phosphoribosyltransferase (Nampt) is also called visfatin or pre-B-cell colony-enhancing factor. The functions of Nampt have been reported as a cytokine, an adipokine and the rate-limiting enzyme in nicotinamide adenine dinucleotide biosynthesis. As a pleiotropic multifunctional protein, Nampt is involved in a variety of physiological and pathological conditions including innate immunity, metabolic disorders, and stress; and Nampt also participates in inflammatory disorders such as acute lung injury, atherosclerosis, myocardial infarct, obesity, type 2 diabetes, and rheumatoid arthritis. The studies indicate that Nampt might be a potential target for pharmacological intervention against inflammatory diseases. We review research advances on the roles of Nampt in inflammation.

Study of animal model of osteonecrosis induced by local ethanol injection in emu / 中国医学科学院学报

<p><b>OBJECTIVE</b>To establish a new animal model of osteonecrosis of the femoral head by local ethanol injection in emu.</p><p><b>METHODS</b>Eight milliliter ethanol was injected slowly to the operated femoral head with customized probe in twenty adult male emus. Postoperatively, hip magnetic resonance imaging was performed at 1, 4, 8, 12 weeks. After emus were sacrificed, the femurs were collected for micro-computed tomography and histological analysis.</p><p><b>RESULTS</b>No emu demonstrated signs of infection or died unexpectedly. Magnetic resonance imaging examination showed broad edema at proximal femur at 1(th) week, and the edema decreased with time, till local edema at femoral head at the 12(th) week. Histological images showed human-like osteonecrotic changes with active bone repair. There were significant differences in trabecular structure and bone mineral density between the operated and intact femoral heads. No collapse was found 6 months after the operation.</p><p><b>CONCLUSIONS</b>This emu model of femoral head osteonecrosis by local ethanol injection can progress to early stage osteonecrosis. The different repair methods may have certain correlation with the results of osteonecrosis of the femoral heads.</p>

In vitro cartilage production using an extracellular matrix-derived scaffold and bone marrow-derived mesenchymal stem cells / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Cartilage repair is a challenging research area because of the limited healing capacity of adult articular cartilage. We had previously developed a natural, human cartilage extracellular matrix (ECM)-derived scaffold for in vivo cartilage tissue engineering in nude mice. However, before these scaffolds can be used in clinical applications in vivo, the in vitro effects should be further explored.</p><p><b>METHODS</b>We produced cartilage in vitro using a natural cartilage ECM-derived scaffold. The scaffolds were fabricated by combining a decellularization procedure with a freeze-drying technique and were characterized by scanning electron microscopy (SEM), micro-computed tomography (micro-CT), histological staining, cytotoxicity assay, biochemical and biomechanical analysis. After being chondrogenically induced, the induction results of BMSCs were analyzed by histology and Immunohisto-chemistry. The attachment and viability assessment of the cells on scaffolds were analyzed using SEM and LIVE/DEAD staining. Cell-scaffold constructs cultured in vitro for 1 week and 3 weeks were analyzed using histological and immunohistochemical methods.</p><p><b>RESULTS</b>SEM and micro-CT revealed a 3-D interconnected porous structure. The majority of the cartilage ECM was found in the scaffold following the removal of cellular debris, and stained positive for safranin O and collagen II. Viability staining indicated no cytotoxic effects of the scaffold. Biochemical analysis showed that collagen content was (708.2-44.7) µg/mg, with GAG (254.7 ± 25.9) µg/mg. Mechanical testing showed the compression moduli (E) were (1.226 ± 0.288) and (0.052 ± 0.007) MPa in dry and wet conditions, respectively. Isolated canine bone marrow-derived stem cells (BMSCs) were induced down a chondrogenic pathway, labeled with PKH26, and seeded onto the scaffold. Immunofluorescent staining of the cell-scaffold constructs indicated that chondrocyte-like cells were derived from seeded BMSCs and excreted ECM. The cell-scaffold constructs contained pink, smooth and translucent cartilage-like tissue after 3 weeks of culture. We observed evenly distributed cartilage ECM proteoglycans and collagen type II around seeded BMSCs on the surface and inside the pores throughout the scaffold.</p><p><b>CONCLUSION</b>This study suggests that a cartilage ECM scaffold holds much promise for in vitro cartilage tissue engineering.</p>

Combined effects of NEL-like type 1 gene and zoledronate in preventing collapse of the femoral head / 中国医学科学院学报

<p><b>OBJECTIVE</b>To determine if combined therapy consisting of NEL-like type 1 gene (NELL-1) and zoledronate can prevent the collapse of the femoral head and stimulate the new bone formation in an animal model of osteonecrosis.</p><p><b>METHODS</b>Ischemic osteonecrosis was surgically induced in 24 SD rats, whicih were equally randomly divided into three groups: combination group, treated with both NELL-1 and zoledronate; sham operation group; and placebo group, treated with normal saline solution. The animals were killed 5 weeks after surgery. Radiography, MicroCT, histology, and immunohistochemistry were performed to analyze the results.</p><p><b>RESULTS</b>Morphologically, the femoral head was at good shape in the combination group, while mildly flattened femoral head was seen in the placebo group. No heterotopic ossifications were observed in each group. MicroCT assessment showed significantly higher total and bone mineral volume in the combination group than in the placebo group (P<0.01), whereas no such significant difference was found when compared with the sham operation group(P>0.05). Histological assessment showed more active osteoblast activity and reduced osteoclast activity in the combination group compared with placebo group.</p><p><b>CONCLUSION</b>A combination of NELL-1 and zoledronate can decrease the femoral head deformity while stimulating bone formation in a traumatic rat osteonecrois model, showing a potential to reverse the osteonecrosis.</p>

A phase I study on pharmacokinetics and pharmacodynamics of higenamine in healthy Chinese subjects.

AIM: To investigate the pharmacokinetics, pharmacodynamics, and safety of higenamine, an active ingredient of Aconite root, in healthy Chinese volunteers. METHODS: Ten subjects received continuous, intravenous infusion of higenamine at gradually escalating doses from 0.5 to 4.0 µg·kg(-1)·min(-1), each dose was given for 3 min. Blood and urine samples were collected at designated time points to measure the concentrations of higenamine. Pharmacodynamics was assessed by measuring the subject's heart rate. A nonlinear mixed-effect modeling approach, using the software Phoenix NLME, was used to model the plasma concentration-time profiles and heart rate. RESULTS: Peak concentrations (C(max)) of higenamine ranged from 15.1 to 44.0 ng/mL. The half-life of higenamine was 0.133 h (range, 0.107-0.166 h), while the area under concentration-time curve (AUC), extrapolated to infinity, was 5.39 ng·h·mL(-1) (range, 3.2-6.8 ng·h·mL(-1)). The volume of distribution (V) was 48 L (range, 30.8-80.6 L). The total clearance (CL) was 249 L/h (range, 199-336 L/h). Within 8 h, 9.3% (range, 4.6%-12.4%) of higenamine was recovered in the urine. The pharmacokinetics of higenamine was successfully described using a two-compartment model with nonlinear clearance. In the pharmacodynamic model, heart rates were related to the plasma drug concentrations using a simple direct effect model with baseline. The E(0), E(max), and EC(50) were 68 bpm, 73 bpm and 8.1 µg/L, respectively. CONCLUSION: Higenamine has desirable pharmacokinetic and pharmacodynamic characteristics. The results provide important information for future clinical studies on higenamine.

Three-dimensional deformation field measurement system based on micro-CT images / 医用生物力学

Objective To develop a novel measurement system composed of micro-CT, mechanical loading device and digital volume correlation (DVC) technique, so as to measure the three-dimensional microstructural deformation field in bone tissue. Methods Uniaxial compression was applied on the specimen with the micromechanical loading device, and CT scans were also conducted while maintaining the same loads; then sequential CT images were matched and searched accordingly by DVC method to calculate the micro-displacement in the specimen along three directions before and after loading; repeated scanning of zero-displacement and rigid body translation were used to evaluate the accuracy and precision of the system. The three-dimensional distribution of displacement field in bovine cancellous bone was measured by the system. Results The result from repeated scanning of zero-displacement showed that the highest accuracy of measurement was performed in the loading direction and the precision was less than tenth of the CT resolution. The result of rigid body translation showed that the standard deviation was 0.001~0.002 μm. For cancellous bone specimen under the load of 600 N, the range of micro-displacement was 100.35~110.25 μm, with multilayer field distribution. Conclusions The accuracy and precision of this measurement system can meet the requirement of DVC method. It is proved that this system can be used for measuring the three-dimensional micro-deformation field in the cancellous bone and as a measurement platform for investigating the relationship between deformation distribution and structural response in bone tissue for the future research.

Cysteinyl leukotriene receptor 1 is involved in rotenone-induced injury of PC12 cells / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To investigate whether cysteinyl leukotriene receptor 1 (CysLT₁ receptor) is involved in rotenone-induced injury of PC12 cells.</p><p><b>METHODS</b>After 24 h treatment with rotenone or with rotenone and the CysLT₁ receptor antagonist montelukast, PC12 cell viability was determined by the colorimetric MTT reduction assay. After PC12 cells were treated with various concentrations of rotenone for 24 h or with 3 μmol/L rotenone for various durations, the expression of CysLT(1) receptor was determined by Western blotting, and its intracellular distribution was detected by immunocytochemistry.</p><p><b>RESULTS</b>Rotenone (0.3-30 μmol/L) induced PC12 cell injury; this injury was significantly attenuated by montelukast at 1 and 5 μmol/L.The expression of CysLT(1) receptor increased after rotenone treatment at 1-10 μmol/L, or at 3 μmol/L for 3 and 24 h. Rotenone caused concentration-and time-dependent translocation of CysLT₁ receptor from the nucleus to the cytosol.</p><p><b>CONCLUSION</b>Cysteinyl leukotriene receptor 1 is involved in rotenone-induced injury of PC12 cells.</p>

Effect of montelukast on morphological changes in neurons after ischemic injury / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To determine the effect of montelukast, a cysteinyl leukotriene receptor 1 antagonist, on morphological changes in rat neurons after ischemic injury.</p><p><b>METHODS</b>The in vivo ischemia injury was induced by oxygen-glucose deprivation (OGD) for 2 h and reperfusion (R) for 24 h (OGD/R) in rat neurons primary culture and mixed cortex culture. In the presence or absence of various concentrations of montelukast, neuron number, area of neuron, number of neuritis per neuron, branch number of primary neuritis and primary neurite length were determined for evaluating morphological changes in neurons.</p><p><b>RESULTS</b>OGD/R significantly reduced neuron number, and altered neuron morphology. In cortical neuron cultures, montelukast (0.0001-1 μmol/L) attenuated OGD/R-induced reduction in neuron number, and inhibited OGD/R-induced increase in branch number of primary neuritis. In the mixed cultures, montelukast (0.0001-0.1 μmol/L) increased the primary neurite length, and reduced number of neuritis and branch number of primary neurite after OGD/R.</p><p><b>CONCLUSION</b>Montelukast has a protective effect on ischemic injury in neurons.</p>

Research advances in experimental animal models of osteonecrosis / 中国医学科学院学报

Osteonecrosis is a common disease, mainly affecting femoral head. Good animal models are helpful in research on the pathologic mechanism of osteonecrosis and the exploration of effective treatment. Although it is relatively easy to establish animal models of early osteonecrosis of femoral head using various approaches, it is difficult to develop an animal model that mimics the full range of osteonecrosis of femoral head. In this paper, we reviewed the current researches on experimental animal models of osteonecrosis, with an attempt to provide evidences for choosing the appropriate animal models and find the way of future development.

Effect and mechanism of zoledronate on prevention of collapse in osteonecrosis of the femoral head / 中国医学科学院学报

<p><b>OBJECTIVE</b>To observe the effect and mechanism of zoledronate on prevention of collapse in an animal model of osteonecrosis.</p><p><b>METHODS</b>Ischemic osteonecrosis was surgically induced in 16 SD rats (which were further divided into zoledronate group and placebo group); another 8 rats were used as sham surgery group (n=8). The animals were killed 5 weeks after surgery. Radiographic, Micro-CT, histological, and immunohistochemical assessments were performed.</p><p><b>RESULTS</b>Radiographic assessment showed better preservation of the femoral head shape in the zoledronate group than in the placebo group but not significantly different from the sham surgery group. Micro-CT assessment showed higher total volume, bone volume, and total mineralized content in the zoledronate group(all P0.05). Compared with the placebo group, the zoledronate group had reduced osteoclast and osteoblast activity, as confirmed by histological examinations.</p><p><b>CONCLUSION</b>Zoledronate can decrease the femoral head deformity by reducing the osteoclast activity while suppressing new bone and vessels formation in a rat model of traumatic osteonecrosis, and therefore may delay the collapse of femoral head.</p>

[Influence on pubertal reproductive function in female rats by immune challenge in early life].

OBJECTIVE: To investigate the long-term programming effects on pubertal reproductive function by immunological challenge in early life. METHODS: Female Sprague-Dawley rats were administered by endotoxin (lipopolysaccharide, LPS) at a dosage of 50 µg/kg and saline intraperitoneally on postnatal day 3 and 5. Body weight was measured weekly. Puberty onset (vaginal opening) and oestrous cyclicity were monitored from postnatal day 30. At the age of 6 weeks, bilateral ovariectomy was performed. The histological and morphological change of the ovaries (the thickness of the theca interna and the number of different kinds of follicles) were observed and the immunoreactivity of the ovarian sympathetic nerve markers (low affinity receptor of nerve growth factor, p75NGFR) was evaluated by immune staining. RESULTS: Immunological challenge (exposed to LPS) in early life delayed vaginal opening significantly [LPS-treated (40.6 ± 0.7) days versus controls (38.6 ± 0.5) days, P < 0.05], decreased the percentage of normal oestrous cyclicity (LPS-treated 26.1% versus controls 66.8%, P < 0.05), decreased the total number of different types of follicles (primordial follicles: LPS-treated 610 ± 47 versus controls 1181 ± 57, P < 0.05; primary follicles: LPS-treated 624 ± 41 versus controls 960 ± 30, P < 0.05; preantral follicles: LPS-treated 183 ± 16 versus controls 260 ± 14, P < 0.05; antral follicles: LPS-treated 32 ± 4 versus controls 79 ± 7, P < 0.05) and increased the thickness of the theca interna [LPS-treated (15.8 ± 0.4) µm versus controls (11.4 ± 0.3) µm, P < 0.05]. The immunostaining of p75NGFR was obviously enhanced in the LPS-treated ovaries when compared with that of controls (P < 0.05). CONCLUSIONS: Immunological stress during early critical developmental windows could have long dysfunctional effects on the pubertal reproductive function. It delayed puberty onset, reduced the percentage of the normal oestrous cycles, decreased follicles reserve and increased the thickness of the theca interna which might involve the up-regulation of the local ovarian sympathetic nerve activity.

Biocompatibility evaluation of electrospun aligned poly (propylene carbonate) nanofibrous scaffolds with peripheral nerve tissues and cells in vitro / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Peripheral nerve regeneration across large gaps is clinically challenging. Scaffold design plays a pivotal role in nerve tissue engineering. Recently, nanofibrous scaffolds have proven a suitable environment for cell attachment and proliferation due to similarities of their physical properties to natural extracellular matrix. Poly(propylene carbonate) (PPC) nanofibrous scaffolds have been investigated for vascular tissue engineering. However, no reports exist of PPC nanofibrous scaffolds for nerve tissue engineering. This study aimed to evaluate the potential role of aligned and random PPC nanofibrous scaffolds as substrates for peripheral nerve tissue and cells in nerve tissue engineering.</p><p><b>METHODS</b>Aligned and random PPC nanofibrous scaffolds were fabricated by electrospinning and their chemical characterization were carried out using scanning electron microscopy (SEM). Dorsal root ganglia (DRG) from Sprague-Dawley rats were cultured on the nanofibrous substrates for 7 days. Neurite outgrowth and Schwann-cell migration from DRG were observed and quantified using immunocytochemistry and SEM. Schwann cells derived from rat sciatic nerves were cultured in electrospun PPC scaffold-extract fluid for 24, 48, 72 hours and 7 days. The viability of Schwann cells was evaluated by 3-[4,5-dimethyl(thiazol-2-yl)-2,5-diphenyl] tetrazolium bromide (MTT) assay.</p><p><b>RESULTS</b>The diameter of aligned and random fibers ranged between 800 nm and 1200 nm, and the thickness of the films was approximately 10 - 20 µm. Quantification of aligned fiber films revealed approximately 90% alignment of all fibers along the longitudinal axis. However, with random fiber films, the alignment of fibers was random through all angle bins. Rat DRG explants were grown on PPC nanofiber films for up to 1 week. On the aligned fiber films, the majority of neurite outgrowth and Schwann cell migration from the DRG extended unidirectionally, parallel to the aligned fibers. However, on the random fiber films, neurite outgrowth and Schwann cell migration were randomly distributed. A comparison of cumulative neurite lengths from cultured DRGs indicated that neurites grew faster on aligned PPC films ((2537.6 ± 987.3) µm) than randomly-distributed fibers ((493.5 ± 50.6) µm). The average distance of Schwann cell migration on aligned PPC nanofibrous films ((2803.5 ± 943.6) µm) were significantly greater than those on random fibers ((625.3 ± 47.8) µm). The viability of Schwann cells cultured in aligned PPC scaffold extract fluid was not significantly different from that in the plain DMEM/F12 medium at all time points after seeding.</p><p><b>CONCLUSIONS</b>The aligned PPC nanofibrous film, but not the randomly-oriented fibers, significantly enhanced peripheral nerve regeneration in vitro, indicating the substantial role of topographical cues in stimulating endogenous nerve repair mechanisms. Aligned PPC nanofibrous scaffolds may be a promising biomaterial for nerve regeneration.</p>