Two undescribed compounds from Aspergillus Niger, an endophytic fungus isolated from Camellia flavida.

A new naphtho-γ-pyrone dimer, asperosperma A, and a new methyl nicotinate derivative, asperosperma B, with 12 known compounds were isolated from the endophytic fungus Aspergillus niger from the stem of Camellia flavida. Their structure was elucidated by NMR, ECD spectrum, and HR-ESI-MS data. Asperosperma A exhibited a highly cytotoxicity against H460 and 4T1 cancer cells with the IC50 values were 0.37 ± 0.06 and 2.04 ± 0.79 µM, respectively. Moreover, it showed a highly sensitive against Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa and methicillin-resistant S. aureus.

Enantioselective adsorption of ibuprofen enantiomers using chiral-active carbon nanoparticles induced S-α-methylbenzylamine.

The chiral media is crucial to the chiral recognition and separation of enantiomers. In this study, we report the preparation of novel chiral carbon nanoparticles (CCNPs) via surface passivation using glucose as the carbon source and S-(-)-α-methylbenzylamine as the chiral ligand. The structures of the obtained CCNPs are characterized via FT-IR, Raman spectroscopy, DLS, XPS, XRD, TEM, and zeta potential analysis. These CCNPs could be employed as the chiral adsorbent and used for the enantioselective adsorption of the ibuprofen enantiomers. The results demonstrated that the CCNPs could selectively adsorb R-enantiomer from ibuprofen racemate solution and give an enantiomeric excess (e.e.) of about 50% under an optimal adsorption condition. Moreover, the regeneration efficiency of the CCNPs remained above e.e. of 43% after the fifth cycle. The present work confirmed that the prepared CCNPs show great potential in the enantioselective separation of ibuprofen racemate.

Triterpenes and Pheophorbides from Camellia ptilosperma and Their Cytotoxicity, Photocytotoxicity, and Photodynamic Antibacterial Activity.

Phytochemical investigation of the leaves of Camellia ptilosperma S. Y. Liang et Q. D. Chen led to the isolation of ten undescribed compounds, including six new triterpenes (1-6) and four new pheophorbide-related compounds (7-10). Meanwhile, the cytotoxic activity of the six triterpenes against six cancer cell lines was evaluated by MTT assay. Compound 2 showed potent cytotoxicity toward HepG2 cells with an IC50 value of 2.57 µM. Compounds 4 and 5 exhibited cytotoxicity against MDA-MB231 cells, with IC50 values of 11.31 and 5.52 µM, respectively. Additionally, the cytotoxicity of four new pheophorbides against these cancer cells was evaluated both in the presence and absence of light treatment. Compound 7 exhibited exceptional photocytotoxicity against Hela, MCF-7, and A549 cells, with IC50 values of 0.43 µM, 0.28 µM, and 0.92 µM, respectively. Compound 10 demonstrated significant photodynamic cytotoxic activity against BEL-7402 and HepG2 cells with IC50 values of 0.77 µM and 0.33 µM, respectively. The photodynamic antibacterial activity of 7-10 was also tested for S. aureus, E. coli, K. pneumoniae, and P. aeruginosa under direct illumination. Compounds 8 and 10 exhibited sensitivity to E. coli and demonstrated a photodynamic antibacterial effect, with a MIC value of 0.625 µM.

Ursane-Type Triterpenes with a Phenylpropanoid Unit from Camellia ptilosperma and Evaluation of Their Cytotoxic Activities.

Six new ursane-type triterpenes with a phenylpropanoid unit and five known oleanane-type triterpenes were isolated from the leaves of Camellia ptilosperma. The undescribed compounds were identified by analysis of 1D and 2D NMR and HRESIMS spectroscopic data as ptilospermanols A-F. The cytotoxicity of new compounds against six human cancer cell lines and three mouse tumor cell lines was evaluated by MTT assay.

Distribution of bone voids in the thoracolumbar spine in Chinese adults with and without osteoporosis: A cross-sectional multi-center study based on 464 vertebrae.

Bone void is a novel intuitive morphological indicator to assess bone quality but its use in vertebrae has not been described. This cross-sectional and multi-center study aimed to investigate the distribution of bone voids in the thoracolumbar spine in Chinese adults based on quantitative computed tomography (QCT). A bone void was defined as a trabecular net region with extremely low bone mineral density (BMD) (<40 mg/cm3), detected by an algorithm based on phantom-less technology. A total of 464 vertebrae from 152 patients (51.8 ± 13.4 years old) were included. The vertebral trabecular bone was divided into eight sections based on the middle sagittal, coronal, and horizontal planes. Bone void of the whole vertebra and each section were compared between healthy, osteopenia, and osteoporosis groups and between spine levels. Receiver operator characteristic (ROC) curves were plotted and optimum cutoff points of void volume between the groups were obtained. The total void volumes of the whole vertebra were 124.3 ± 221.5 mm3, 1256.7 ± 928.7 mm3, and 5624.6 ± 3217.7 mm3 in healthy, osteopenia, and osteoporosis groups, respectively. The detection rate of vertebrae with bone voids was higher and the normalized void volume was larger in the lumbar than in thoracic vertebrae. L3 presented the largest void (2165.0 ± 3396.0 mm3), while T12 had the smallest void (448.9 ± 699.4 mm3). The bone void was mainly located in the superior-posterior-right section (40.8 %). Additionally, bone void correlated positively with age and increased rapidly after 55 years. The most significant void volume increase was found in the inferior-anterior-right section whereas the least increase was found in the inferior-posterior-left section with aging. The cutoff points were 345.1 mm3 between healthy and osteopenia groups (sensitivity = 0.923, specificity = 0.932) and 1693.4 mm3 between osteopenia and osteoporosis groups (sensitivity = 1.000, specificity = 0.897). In conclusion, this study demonstrated the bone void distribution in vertebrae using clinical QCT data. The findings provide a new perspective for the description of bone quality and showed that bone void could guide clinical practice such as osteoporosis screening.

Two new isoflavones from the barks of Dalbergia hancei Benth.

Two new isoflavone compounds, Dalhancei A (1) and Dalhancei B (2), along with a known compound epicatechin (3) were isolated from 80% methanol extract of the barks of Dalbergia hancei Benth. The structures of compounds 1-3 were elucidated by comparison with the literature and physical data analysis, including optical rotation, MS, 1D and 2D NMR spectra. Compounds 1 and 2 showed weak inhibitory activity against tyrosinase at 16.22 mmol/L, with inhibition rates of 42.23 ± 0.18% and 45.68 ± 0.17%, respectively; compound 1 exhibited weak inhibitory activity against α-glucosidase with the inhibition rate of 43.72 ± 0.22% at 5.41 mmol/L, compounds 2 and 3 had better α-glucosidase inhibitory activity than compound 1 with IC50 values of 0.90 ± 0.18 and 0.41 ± 0.17 mmol/L, respectively.

Bisphenol A analogues in associations with serum hormone levels among reproductive-aged Chinese men.

BACKGROUND: Bisphenol A (BPA) as an endocrine disrupting chemical has been shown to alter reproductive endocrine function, but little is known on its analogues such as bisphenol F (BPF) and bisphenol S (BPS) with increasing usage and exposure. OBJECTIVE: To explore the associations between exposures to BPA, BPF and BPS and serum reproductive hormones among reproductive-aged Chinese men. METHODS: We measured BPA, BPF and BPS concentrations in repeated urine samples and multiple reproductive hormones in the serum samples collected from 462 men attending an infertility clinic in Wuhan, China. Linear regression models were applied to assess the associations between averaged urinary BPA, BPF and BPS levels and serum hormone concentrations, and restricted cubic spline (RCS) models were further utilized to explore potential non-linear associations. We also examined potential modifying effects by age and body mass index (BMI). RESULTS: There was little evidence of associations between BPA exposure and altered reproductive hormones. However, we found that elevated BPF and BPS exposures were in negative associations with estrogen (E2) levels and E2/T (total testosterone) ratio (all P for trends < 0.05), and that elevated BPS exposure was negatively associated with SHBG levels (P for trend = 0.09). Based on the RCS models, these linear negative associations except that between BPS exposure and E2/T ratio were further confirmed. In stratified analyses, BPF and BPS exposures in relation to reduced E2 and E2/T ratio were more pronounced among men aged > 30 years, whereas their associations with reduced SHBG levels were more pronounced among men aged ≤ 30. Also, BPS exposure in negative association with FSH only emerged among men with BMI ≥ 24 kg/m2 (P for interaction = 0.03). CONCLUSION: BPF and BPS exposures were negatively associated with male serum E2, E2/T ratio and SHBG levels, and these associations varied by age and BMI.

Organochlorine pesticide exposures, metabolic enzyme genetic polymorphisms and semen quality parameters among men attending an infertility clinic.

The associations of organochlorine pesticides (OCPs) with semen quality from human studies are conflicting, and also it is largely unknown whether the associations are modified by genetic polymorphisms. We aimed to evaluate the associations between serum concentrations of 18 OCPs and semen quality among 387 Chinese men, and further to examine the modifying effects by genetic polymorphisms in cytochrome P450 (CYP2E1) and glutathione S-transferase (GSTT1). Multivariable linear regressions were used to evaluate the relationships between serum OCP concentrations and semen quality, and the role of CYP2E1 and GSTT1 polymorphisms in modifying the associations were assessed. Multiple testing was adjusted using the false discovery rate (FDR). We observed that men with detectable concentrations of serum ɤ-HCH had a decrease in sperm motility of 7.07% (95% CI: -10.9%, -3.24%) compared to those with undetectable concentrations (FDR-P value = 0.02). Men with TT of CYP2E1 rs 915906 genotypes had higher median concentrations of serum dieldrin compared with those with CT/CC of CYP2E1 rs 915906 genotypes. There were interactions between CYP2E1 and GSTT1 polymorphisms and certain OCPs namely ɤ-HCH, δ-HCH, dieldrin, endosulfan I, and endrin aldehyde on semen quality. For example, elevated dieldrin levels in relation to decreased sperm concentration, sperm count, and sperm motility were only observed among men with CC of CYP2E1 rs2031920 genotypes (all Pinteraction < 0.05). However, these interactions were not statistically significant after the FDR adjustment. Our results suggested that CYP2E1 and GSTT1 polymorphisms may modify the effects of OCP exposures on semen quality. Due to the relatively small size samples, further investigation is warranted to confirm the findings.

A novel surgical planning system using an AI model to optimize planning of pedicle screw trajectories with highest bone mineral density and strongest pull-out force.

OBJECTIVE: The purpose of this study was to evaluate the ability of a novel artificial intelligence (AI) model in identifying optimized transpedicular screw trajectories with higher bone mineral density (BMD) as well as higher pull-out force (POF) in osteoporotic patients. METHODS: An innovative pedicle screw trajectory planning system called Bone's Trajectory was developed using a 3D graphic search and an AI-based finite element analysis model. The preoperative CT scans of 21 elderly osteoporotic patients were analyzed retrospectively. The AI model automatically calculated the number of alternative transpedicular trajectories, the trajectory BMD, and the estimated POF of L3-5. The highest BMD and highest POF of optimized trajectories were recorded and compared with AO standard trajectories. RESULTS: The average patient age and average BMD of the vertebral bodies were 69.6 ± 7.8 years and 55.9 ± 17.1 mg/ml, respectively. On both sides of L3-5, the optimized trajectories showed significantly higher BMD and POF than the AO standard trajectories (p < 0.05). On average, the POF of optimized trajectory screws showed at least a 2.0-fold increase compared with AO trajectory screws. CONCLUSIONS: The novel AI model performs well in enabling the selection of optimized transpedicular trajectories with higher BMD and POF than the AO standard trajectories.

Automatic phantom-less QCT system with high precision of BMD measurement for osteoporosis screening: Technique optimisation and clinical validation.

BACKGROUND: Currently dual-energy X-ray absorptiometry (DXA) and phantom-based quantitative computed tomography (PB-QCT) have been utilized to diagnose osteoporosis widely in clinical practice. While traditional phantom-less QCT (PL-QCT) is limited by the precision of manual calibration using body tissues, such as fat and muscle. OBJECTIVE: The aim of this study is to validate the accuracy and precision of one newly-developed automatic PL-QCT system to measure spinal bone mineral density (BMD) and diagnose osteoporosis. METHODS: A total of 36 patients were enrolled for comparison of BMD measurement between DXA and QCT. CT images of 63 patients were analyzed by both PB-QCT and newly developed automatic PL-QCT system, then the BMD results generated by the automatic PL-QCT were utilized to diagnose osteoporosis. The diagnostic outcomes were compared with that of DXA and PB-QCT to assess the performance of the new system. RESULTS: BMD test results showed that the automatic PL-QCT system had higher precision than previous studies performed with QCT, while maintaining similar capability to diagnose osteoporosis as DXA and PB-QCT. Area under curve (AUC) result of PL-QCT was larger than 0.8 for predicting spine DXA T-score in receiver operating characteristic (ROC) analysis. Pearson correlation analysis (r â€‹= â€‹0.99) showed strong linear correlation and Bland-Altman analysis (bias â€‹= â€‹3.0mg/cc) indicated little difference between the two methods. The precision result (CV â€‹= â€‹0.89%) represented good reproducibility of the new system. CONCLUSION: The traditional PL-QCT system has relatively low reproducibility due to the manual selection of the region of interest (ROI) of body tissues. Automatic selection of ROI in this new system makes the BMD testing more convenient and improves precision significantly. Compared with traditional BMD measurement methods, the automatic PL-QCT system had higher precision in accurate diagnosis of osteoporosis with great potential in translational research and wide clinical application. TRANSLATIONAL POTENTIAL STATEMENT: With high accuracy and precision, the automatic PL-QCT system could serve as an opportunistic screening tool for osteoporosis patients in the future. It could also facilitate related researches by providing more reliable data collection, both retrospectively and longitudinally.

Structural Characterization and Assessment of Anti-Inflammatory and Anti-Tyrosinase Activities of Polyphenols from Melastoma normale.

Polyphenols, widely distributed in the genus Melastoma plants, possess extensive cellular protective effects such as anti-inflammatory, anti-tyrosinase, and anti-obesity, which makes it a potential anti-inflammatory drug or enzyme inhibitor. Therefore, the aim of this study is to screen for the anti-inflammatory and enzyme inhibitory activities of compounds from title plant. Using silica gel, MCI, ODS C18, and Sephadex LH-20 column chromatography, as well as semipreparative HPLC, the extract of Melastoma normale roots was separated. Four new ellagitannins, Whiskey tannin C (1), 1-O-(4-methoxygalloyl)-6-O-galloyl-2,3-O-(S)-hexahydroxydiphenoyl-ß-d-glucose (2), 1-O-galloyl-6-O-(3-methoxygalloyl)-2,3-O-(S)-hexahydroxydiphenoyl-ß-d-glucose (3), and 1-O-galloyl-6-O-vanilloyl-2,3-O-(S)-hexahydroxydiphenoyl-ß-d-glucose (4), along with eight known polyphenols were firstly obtained from this plant. The structures of all isolates were elucidated by HRMS, NMR, and CD analyses. Using lipopolysaccharide (LPS)-stimulated RAW2 64.7 cells, we investigated the anti-inflammatory activities of compounds 1-4, unfortunately, none of them exhibit inhibit nitric oxide (NO) production, their IC50 values are all > 50 µM. Anti-tyrosinase activity assays was done by tyrosinase inhibition activity screening model. Compound 1 showed weak tyrosinase inhibitory activity with IC50 values of 426.02 ± 11.31 µM. Compounds 2-4 displayed moderate tyrosinase inhibitory activities with IC50 values in the range of 124.74 ± 3.12-241.41 ± 6.23 µM. The structure-activity relationships indicate that hydroxylation at C-3', C-4', and C-3 in the flavones were key to their anti-tyrosinase activities. The successful isolation and structure identification of ellagitannin provide materials for the screening of anti-inflammatory drugs and enzyme inhibitors, and also contribute to the development and utilization of M. normale.

Magnetic Molecularly Imprinted Polymers Based on Dehydroabietylamine as Chiral Monomers for the Enantioseparation of RS-Mandelic Acid.

Stereoselective adsorption of the enantiomers shows potential in the resolution of a racemate. In this work, we synthesized novel magnetic surface molecularly imprinted polymers (MIPs) on the surface of the γ-methacryloxypropyltrimethoxysilane (MPS)-modified Fe3O4@SiO2 particles to utilize chiral dehydroabietylamine (DHA) as a functional monomer and R-mandelic acid as a template molecule (DHA-MIPs). We performed the resolution of mandelic acid racemate (RS-MA) via adsorption on the as-prepared MIPs. The results revealed that the MIPs have good affinity and high adsorption capacity for R-MA and show better enantioselective adsorption ability for R-MA than that for S-MA. One-stage adsorption of RS-MA on the MIPs can achieve up to 53.7% enantiomeric excess (ee) for R-MA. These help us to improve the chiral separation ability of the traditional MIPs using a chiral rather than an achiral monomer in MIP preparation. The MIPs can be employed as an economic and efficient adsorbent for chiral separation of MA racemate.

Prevent action of magnoflorine with hyaluronic acid gel from cartilage degeneration in anterior cruciate ligament transection induced osteoarthritis.

According to the Chinese medicine, magnoflorine exerted significant anti-inflammatory effects and potentially promoted synthesis of proteoglycans in chondrocytes to reverse the progression of rheumatoid arthritis. However, the latent beneficial effect of magnoflorine for the treatment of traumatic osteoarthritis (OA) is still unknown. Therefore, we aim to demonstrate the efficacy of magnoflorine combined with HA-gel in attenuating cartilage degeneration in anterior cruciate ligament transection (ACLT) induced OA rat model. We found that the histological results showed the elevated cartilage matrix, chondrogenic signals and chondroprogenitor cells in HA-gel + magnoflorine treatment. HA-gel + magnoflorine treatment resulted in a decreased modified Mankin's score, and a higher volume ratio of hyaline cartilage (HC)/calcified cartilage (CC) and HC/Sum (whole cartilage), compared to ACLT and HA-gel groups. Furthermore, both the volume ratios of HC/Sum and HC/CC were negatively correlated with modified Mankin's scores. Finally, HA-gel + magnoflorine could significantly increase the BV/TV, Tb.Th, and decrease the Tb.Pf, Po(tot), Conn.Dn and Tb.Sp. In vitro, 50 µg/ml magnoflorine treatment could significantly increase the viability, S-phase, migration rate and chondrogenesis of chondroprogenitor cells. There were significant downregulations of MAPK/NF-κB signaling, and upregulations of chondrogenic signals in 50 µg/ml magnoflorine treatment. There were significant downregulations of proinflammatory cytokines and upregulation of IL-10 in HA-gel + magnoflorine treated group. Therefore, our study elucidated a protective effect of HA-gel + magnoflorine on attenuating cartilage degradation and maintaining SCB stabilization in ACLT induced OA.

Mangiferin exerts neuroprotective activity against lead-induced toxicity and oxidative stress via Nrf2 pathway.

Objective: The study was designed to assess the beneficial role of mangiferin (MGN) in lead (Pb)-induced neurological damages in the activation of Nrf2-governed enzymes, genes and proteins. Methods: A total of 96 weaned Wistar rats (48 males and 48 females, 26- to 27-day-old), weighing 50-80 g were used. The experiment was performed in six groups: normal group (control, n = 16), model group (chronic Pb exposed, n = 16), Dimercaptosuccinic acid (DMSA)-treated group (positive control, Pb + DMSA, n = 16), three MGN-treated groups with different doses (Pb + MGN, n = 48). Normal group freely had access to purified water. DMSA-treated group was given DMSA, which was clinically used as the standard treatment for moderate Pb poisoning, at 50 mg/kg (2 mL suspension with purified water) by intragastric gavage (ig) 4 continual days a week for 4 weeks, MGN-treated groups were given MGN at 50, 100, or 200 mg/kg (2 mL suspension with purified water) by ig daily for 4 weeks. At the end of the treatment, all rats were sacrificed and the brain samples were collected. The haematoxylin and eosin (H&E) staining was used for observation of histopathology. Commercial kit, real-time quantitative polymerase chain reaction (RT-qPCR), Western-blot and immunohistochemistry (IHC) detection were used to detect the mRNA and protein expression. Results: Eight weeks exposure to Pb-containing water resulted in pathological alterations, anti-oxidative system disorder in the brain, all of which were blocked by MGN in a Nrf2-dependent manner. Nrf2 downstream enzymes such as HO-1, NQO1, γ-GCS were activated. Nrf2, GCLC, GCLM, HO-1 mRNA and total Nrf2, Nuclear Nrf2, γ-GCS, HO-1 protein expression were affected too. Conclusion: MGN ameliorated morphological damage in the hippocampus. Its neuroprotective effects were achieved by the activation of the Nrf2 downstream genes. The data from this in vitro study indicates that MGN targeting Nrf2 activation is a feasible approach to reduce adverse health effects associated with Pb exposure. Thus, MGN could be an effective candidate agent for the Pb-induced oxidative stress and neurotoxicity in the human body.

Effects of long-term fluoride exposure on cognitive ability and the underlying mechanisms: Role of autophagy and its association with apoptosis.

Autophagy and apoptosis are two important cellular processes that are crucial for neurodevelopment. Evidence shows that apoptosis is implicated in fluoride neurotoxicity. However, the biological roles of autophagy, especially its interplay with apoptosis in the neurotoxicity induced by long-term fluoride exposure remain unclear. Here we present in vivo and in vitro evidence that fluoride-induced defective autophagy elicits excessive apoptosis, thus inducing neurotoxicity. Using Sprague-Dawley rats exposed to sodium fluoride from 60 days before pregnancy until 6 months post-delivery as in vivo model, we showed that fluoride impaired the learning and memory abilities of offspring rats, with decreased neuronal number, suppressed autophagy and enhanced apoptosis in hippocampus. These results were validated in human neuroblastoma SH-SY5Y cells in vitro. Mechanistically, mTOR signaling, responsible for autophagy induction, was activated in vivo and in vitro, and targeting inhibition of mTOR with rapamycin protected SH-SY5Y cells from defective autophagy and excessive apoptosis, thereby enhancing neuronal survival. Furthermore, circulating levels of autophagy markers were low in children with higher fluoride body burden and lower intelligence quotient scores. Collectively, our results suggest that defective autophagy plays a pivotal role in fluoride neurotoxicity, and mTOR might be a promising target for the prevention and treatment of fluoride neurotoxicity.

Enantioseparation of racemic bupivacaine via ultrasonic-assisted diastereomeric crystallization using 12,14-dinitrodehydroabietic acid.

12,14-Dinitrodehydroabietic acid (12,14-dinitroDHAA), a chiral acid obtained by the nitration of optical dehydroabietic acid (DHAA), was successfully employed as resolving agent. The resolution of racemic bupivacaine by ultrasonic-assisted diastereomeric crystallization in ethanol was investigated. The results indicated that ultrasonic-assist can well facilitate resolution of (R,S)-bupivacaine and a higher enantiomeric excess (ee) and yield was obtained for (S)-bupivacaine, and while without ultrasound, the ee value decreases by increasing the crystallization time. A Box-Behnken experimental design with four factors (amount of 12,14-dinitroDHAA, ethanol amount, ultrasonic power and crystallization temperature) combined with response surface methodology (RSM) was applied to explore resolution effects. A second-order polynomial equation was adequate to model the relationship between the ee (or yield) and the dependent variables. When maintaining a lower limit of 90% for the yield of (S)-bupivacaine, the optimal resolution conditions by RSM were 12,14-dinitroDHAA/bupivacaine molar ratio of 1.6, solvent/propranolol ratio of 16.5 mL/g, 63.2 W ultrasonic power and crystallization temperature of 0 °C, respectively. Under the optimal conditions, the experimental ee and yield of (S)-bupivacaine were 69.8% and 87.5%.

Multichromatic TTF staining characterizes cartilage matrix in osteoarthritis and bone development.

Various histological staining methods have been explored to detect the joint lesions in osteoarthritis (OA), but these histological stains cannot comprehensively present the comparatively complex structures of articular cartilage in knee OA. In addition, no integrated histological staining method can be used to evaluate efficiently both the subzone region and matrix composition in cartilage containing tissues. Therefore, in this study, a novel multichromatic staining method termed TTF staining, using Toluidine Blue (T), Tartrazine (T) and Fast Green (F) sequential combined staining for histological analysis, has been exploited to characterize the changes of matrix components and contents in cartilage during OA and in the bone development. This specific TTF staining profile can be used to differentiate the major compartments of knee joint region, including the synovium, meniscus, multiple subzones of cartilage and subchondral bone. An anterior cruciate ligament transection induced OA model in rat has been established to profoundly present the alterations of glycosaminoglycans in cartilage degeneration by TTF staining profile. The changes of TTF staining profile in the chondrification and ossification centers of the postnatal rat knee joint indicate the developmental features of cartilage matrix during the growth of bone. In summary, we have developed an effective histological staining method that enables us to identify the subzones of cartilage in detail and to define the matrix features of bone development. Therefore, finally using this new TTF staining method may help us to exploit a histopathological grading system to assess cartilage lesions in clinical disease.

Essential oils and ethanol extract from Camellia nitidissima and evaluation of their biological activity.

Camellia nitidissima, a well-known species of yellow Camellia, has undergone commercial cultivation as a new tea resource recently. Herein, the composition, antioxidant and antimicrobial activities of the essential oil and ethanol extract of C. nitidissima were investigated. The essential oils from the leaves and flowers of C. nitidissima were obtained by hydro-distillation. A total of 56 and 34 constituents accounting for 77.5 and 96.8% of the oils were identified by GC-MS. Linalool (35.8%), phytol (7.9%), cis-geranyl acetone (7.3%) and methyl salicylate (6.8%) were found to be the primary components in the leaf oil, while the flower oil was rich in α-eudesmol (34.3%), γ-eudesmol (31.5%) and linalool (11.1%). The ethanol extract of C. nitidissima leaves contained 281.04 mg gallic acid equivalent/g of total phenols. The antioxidant activities of the two oils and extract were evaluated by DPPH and ABTS radical-scavenging assays. The IC50 values varied from 17.4 (extract) to 720.3 µg/mL (flower oil) for DPPH and from 28.8(extract) to 889.6 µg/mL (flower oil) for ABTS. Both essential oils exhibited moderate antioxidant activities, and the extract possessed strong effects close to ascorbic acid. Additionally, the antimicrobial activities of the oils and extract against Staphylococcus aureus, Bacillus subtilis, Escherichia coli and Pseudomonas aeruginosa were evaluated by agar dilution assay. No considerable bactericidal activities were observed for either essential oil or extract compared with ampicillin and tobramycin standards. The results indicated the extract was more efficient than the two essential oils against S. aureus (MIC = 0.625 mg/mL) and B. subtilis (MIC = 1.25 mg/mL).

Mutual promotion of apoptosis and autophagy in prepubertal rat testes induced by joint exposure of bisphenol A and nonylphenol.

BPA and NP are both typical endocrine disruptors, the exposed populations are widespread, and the health risks mustn't be ignored. However, the interactions between them on spermatogenesis are rarely mentioned. And the underlying mechanism is unclear yet. In the present study, prepubertal SD rats were exposed to different low doses of BPA and NP separately or jointly for 4 weeks. The results indicate that the joint exposure induced excessive apoptosis and autophagy in the testes, as proved by a series of characteristics such as chromatin condensation and autophagosomes formation. Besides, endocrine disorders and oxidative stress were also caused by the exposure. Apoptosis was mediated by the mitochondrial apoptosis pathway, since the Bax and Caspase-3 gene expressions significantly increased with a prominent decrease of Bcl-2. While autophagy was caused by the inhibition of the Akt/mTOR pathway, as the expressions of the downstream genes Beclin-1, Atg5, Atg12 and the split of LC3 protein increased altogether. Worse yet, autophagy and apoptosis might reinforce each other and make the situation more severe in the joint group. What's more, remarkable histopathological changes such as spermatogenic epithelium atrophy, germ cell loss, and various ultrastructural modifications were strongly related to the apoptosis and autophagy. In aggregate, this study shows the enormous risk on male reproductive system brought by the interactions between BPA and NP. The findings provide a broader vision to understand the roles of apoptosis and autophagy induced by the joint exposure in the aggravation of spermatogenesis impairment, which could be a reference for the situation of complex EDCs exposure-induced male reproductive toxicity, and possibly inspire us to find new ideas for preventive and therapeutic treatments.

Magnoflorine with hyaluronic acid gel promotes subchondral bone regeneration and attenuates cartilage degeneration in early osteoarthritis.

OBJECTIVE: To investigate efficacy of Chinese medicine magnoflorine combined with hyaluronic acid (HA)-gel in promoting subchondral bone (SCB) regeneration and attenuating cartilage degeneration in early osteoarthritis (OA). METHODS: MC3T3-E1 under magnoflorine treatment was assayed by XTT to determine cell viability. Cell proliferation was reflected through cell cycle. Osteoblast mineralization was stained by Alizarin Red. Standardized bone canal of 1 mm in diameter and 4 mm in depth was made on tibial medial plateau of 4-month-old Dunkin-Hartley spontaneous knee OA guinea pigs. Guinea pigs (n = 5/group) were treated once intra-bone canal injection of 2 µl HA-gel, 2 µl HA-gel+50 ng magnoflorine and null (Defect) respectively, except sham group. The left hind limbs were harvested for µCT scan and histopathological staining 2-month post-surgery. RESULTS: 25 µg/ml magnoflorine treatment significantly increased cell viability, S-phase and mineralization of MC3T3-E1 cells. In vivo, HA-gel + magnoflorine treatment significantly altered SCB microstructure; changes included increase in bone volume fraction (BV/TV), trabecular number (Tb.N), connectivity density (Conn.Dn), and decrease in degree of anisotropy (DA), which implied trabecular bone regeneration. Treatment also resulted in a decrease in modified Mankin's scores, and an increase in volume ratio of hyaline cartilage (HC)/calcified cartilage (CC) and fractal dimension (FD, roughness indicator of osteochondral conjunction), compared to Defect and HA groups. Furthermore, FD was positively associated with volume ratio of HC/CC and negatively associated with modified Mankin's scores. Finally, histological results showed that due to a faster regeneration of SCB with the HA-gel + magnoflorine treatment, the reduction of cartilage matrix and the decreased expression of chondrogenic signals were attenuated. CONCLUSION: Our study elucidated the potential benefits of HA-gel + magnoflorine in promoting SCB regeneration and revealed a protective effect of stimulating recovery of the SCB integrity on attenuating cartilage degradation to prevent OA progression.