Rspo1 inhibited apoptosis of glucocorticoid-induced osteoblasts via Wnt/ß-catenin pathway in Legg-Calve-Perthes disease.

BACKGROUND: The pathogenesis of Legg-Calve-Perthes disease (LCPD), a juvenile form of avascular necrosis of the femoral head (ANFH), is not fully understood. OBJECTIVES: The purpose of this work was to study the regulatory effect of R-spondin 1 (Rspo1) on osteoblastic apoptosis and evaluate the pre-clinical efficacy of recombinant human protein Rspo1 (rhRspo1) in treatment of LCPD. MATERIAL AND METHODS: This is an experimental study. In vivo rabbit ANFH model was established. Human osteoblast cell line hFOB1.19 (hFOB) was used to overexpress and silence Rspo1 in vitro. Additionally, hFOB cells were induced with glucocorticoid (GC) and methylprednisolone (MP), and treated with rhRspo1. The expressions of Rspo1, ß-catenin, Dkk-1, Bcl-2, and caspase-3, and the apoptosis rate of hFOB cells were examined. RESULTS: The expressions of Rspo1 and ß-catenin were lower in ANFH rabbits. The expression of Rspo1 was decreased in GC-induced hFOB cells. Compared to the control group, after 1 µM MP induction for 72 h, the expressions of ß-catenin and Bcl-2 were higher, while Dkk-1, caspase-3 and cleaved caspase-3 expressions were lower in Rspo1 overexpression and rhRspo1-treated groups. The apoptosis rate of GC-induced hFOB cells was decreased in Rspo1 overexpression and rhRspo1-treated groups compared to the control group. CONCLUSIONS: R-spondin 1 inhibited GC-induced osteoblast apoptosis via Wnt/ß-catenin pathway, which might be associated with the development of ANFH. Moreover, rhRspo1 had a potential pre-clinical therapeutic effect on LCPD.

[Start-up and Optimization of Denitrifying Phosphorus Removal in ABR-MBR Coupling Process].

The feasibility of the denitrifying phosphorus removal process in the ABR-MBR system with no sludge reflux and high concentration of seeding activated sludge (25 g ·L-1, in MLSS) in the ABR was investigated. The characteristics of the microbial community in the denitrifying phosphorus removal compartment were also evaluated. The denitrifying phosphorus removal function was achieved by gradually increasing the reflux ratio (R) from 0% to 200%. During the stable operation, the average removal rates of COD, PO43--P, and TN in the system were 88.28%, 54.45%, and 61.93%, respectively. When the influent loading rate, NOx--N reflux ratio, and hydraulic retention time (HRT) of ABR and MBR were 0.8 kg ·(m3 ·d)-1, 150%, and 9 h and 3.3 h, respectively, the average VFA concentration of 80.58 mg ·L-1, ρ(NO2--N)/ρ(NO3--N) reflux ratio of 1.68, and PO43--P and TN removal rates of 64.94% and 62.95% were obtained. The short-cut nitrification denitrifying phosphorus removal was achieved in the ABR-MBR system. Batch tests showed that denitrifying phosphorus removal bacteria (DPAOs) were the main functional bacteria in the ABR, with anaerobic phosphorus release and anoxic phosphorus uptake of 3.73 mg ·L-1 and 10.22 mg ·L-1, respectively. High throughput sequencing results showed that Proteobacteria and Bacteroidetes were the dominant phyla in the phosphorus removal compartment, accounting for 23.49%-53.66% and 16.55%-21.78% of the total phyla, respectively. Thauera, Thiothrix, Pseudomonas, norank_ f_Rhodocyclaceae, and unclassification_ f_Rhodocyclaceae in Proteobacteria, and Sphingobacteriales in Bacteroidetes were the potential denitrifying phosphorus removal microorganisms.

[Performance and Microbial Characteristics of Ammonium-limited and Nitrite-limited ANAMMOX Systems].

The performance and microbial characteristics of ammonium-limited and nitrite-limited ANAMMOX reactors were studied in two continuously stirred tank reactors. The influent TN concentrations were controlled below 50 mg·L-1. The hydraulic retention time and water temperature were maintained at 2.0 h and 20℃, respectively. Results showed that though both ANAMMOX reactors demonstrated similar TN removal loading rates[0.45-0.5 kg·(m3·d)-1] and TN removal efficiencies (around 70%), the ΔNO3-/ΔNH4+ ratio of the ammonium-limited ANAMMOX reactor showed a faster upward trend. Batch tests and high-throughput sequencing results indicated that the ammonium-limited ANAMMOX reactor had more significant functional and population heterogeneity than the nitrite-limited ANAMMOX reactor. Candidatus_Brocadia was the predominant ANAMMOX bacteria in both reactors. The relative abundance of Candidatus_Brocadia in large granules (53.9%) was significantly higher than that in flocs (19.1%) under the ammonium-limited conditions, whereas only a small difference in relative abundance of Candidatus_Brocadia was observed between the granules (28.1%) and flocs (21.3%) in the nitrite-limited ANAMMOX reactor. Nitrospira-like NOB were detected in both ANAMMOX reactors, which primarily inhabited flocs, seemingly driven by the availability of oxygen. Moreover, the ammonium-limited (i.e., excess nitrite) conditions seemingly favored the growth of Nitrospira. Building upon these results, a control strategy for optimal operation of the ammonium-limited ANAMMOX reactor was proposed based on selective floc discharge.

[Realization Process of Nitritation and Changes in Sludge Characteristics in Granular Sludge Reactor for Low Strength Sewage Treatment].

The realization process of nitritation was studied in a CSTR reactor seeding with nitrification granular sludge to treat low ammonia sewage. During the operation period, the physical and chemical properties, the spatial distribution of functional microbes, and the activity of the granular sludge were also investigated to elaborate the main factors for the stability of nitritation. The results showed that nitritation can be successfully achieved and maintained by the cooperative controlling of nitrogen loading rate (NLR) and dissolved oxygen (DO) levels, and the nitrite accumulation rate was over 80%. The obtained nitritation granular sludge was brownish yellow, showing a smooth, full ellipsoid or sphere, and the microorganisms on the surface of the particles were mainly cocci; the average particle size was 1.3 mm, and the average sedimentation rate was 71.3 m·h-1. Batch tests showed that there was a significant stratified distribution structure in granular sludge (particle size >0.8 mm), the ammonia-oxidizing bacteria (AOB) mainly occupied the surface space of the particles, and the nitrite-oxidizing bacteria (NOB) were mainly distributed inside the particles. Flocs or small-size sludge (particle size<0.8 mm) and granular sludge (particle size >0.8 mm) exhibit different spatial distribution characteristics of microorganisms. In the granular sludge reactor, well stratification of the nitrifier guilds, high level of residual ammonia concentrations in effluent (15-33 mg·L-1), or low ratio between DO and NH4+-N concentrations (0.08-0.15) should be key influencing factors in the process of achieving nitritation.

[Characteristics of Organics Transformation and Sludge Morphology in an ABR for Sewage Treatment with Different HRTs].

The characteristics of organics transformation and sludge morphology of in an ABR(anaerobic baffled reactor) for sewage treatment with different HRTs were investigated based on reactor performance, particle size distribution, and scanning electron microscopy (SEM). Results showed that the COD removal rate was stably maintained above 90.0% when the HRT decreased from 15 h to 4 h. However, the first compartment of ABR contributed to 90%, 78.56%, 74.18%, and 58.91% of the total COD removal when the HRT was 10, 7.5, 5, and 4 h, respectively. When the HRT was reduced, the total amount of volatile fatty acids (VFAs) in the first compartment of ABR significantly increased, and the abundance of acetic acid, being the major constituent of VFAs, gradually increased from 51.36% to 58.77%; the concentrations of butyric acid and propionic acid were maintained and constituted a minority of the VFAs. The sludge morphology in ABR significantly changed in the wake of run time. On day 111, granulation of sludge was achieved. Additionally, the degree of sludge granulation showed a decreasing trend with the direction of water flow. SEM observations of granular sludge showed that the separation of biomass did occur in the ABR. Along the direction of water flow, filamentous bacteria, M. methane, monococci, and bacilli were the dominant microbes in each compartment of the ABR.