Anti-osteoporosis activity of red yeast rice extract on ovariectomy-induced bone loss in rats.

Osteoporosis is the most common bone disease, affecting millions of people worldwide and leading to significant morbidity and high costs. Monacolin K, an extract of red yeast rice (RYR, Hongqu), plays important roles in the management of dyslipidemia, coronary heart disease, and diabetes. Our study aimed to investigate the protective effect of monacolin K on ovariectomy-induced bone loss in rats. Fifty female Sprague-Dawley rats were randomly divided into a sham-operated and five ovariectomized (OVX) groups: OVX with vehicle, OVX with fluvastatin, and OVX with RYR extract of three graded doses. Bone mineral density (BMD), biochemical markers, and cell viability were analyzed by dual energy X-ray absorptiometry, enzyme-linked immunosorbent assay, and 3(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assays. Gene expression was evaluated by real-time polymerase chain reaction amplification and western blot. Our results showed that administration of RYR extract markedly increased the bone mineral density in OVX rats. Moreover, RYR extract decreased the levels of bone turnover markers, including osteocalcin and tartrate resistant acid phosphatase activity. The MMT assay revealed that RYR extract treatment significantly improved the osteoblast viabilities in a dose-dependent manner (P < 0.05). At the molecular level, we further demonstrated that RYR extract enhanced the expression of Bmp2 and Bmp4 both at the mRNA and protein levels. Collectively, these data suggested RYR extract could protect against osteoporosis in ovariectomized rats, most likely through activation of BMP2/4 expression.

Hyperbaric oxygen therapy attenuates neuropathic hyperalgesia in rats and idiopathic trigeminal neuralgia in patients.

BACKGROUND: Neuropathic pain after nerve injury is severe and intractable, and current drug and non-drug therapies offer very limited pain relief. Hyperbaric oxygen (HBO 2) has been clinically used for protection of the nervous system after acute injury. We investigated whether HBO 2 treatment could prevent and/or attenuate neuropathic pain in animals and in patients. METHODS: Mechanical allodynia and thermal hyperalgesia and neurochemical alterations of neuropathic pain were analysed in male, adult, Sprague-Dawley rats with sciatic nerve injury. Clinical trials were conducted in patients with idiopathic trigeminal neuralgia. RESULTS: Repetitive HBO 2 treatment [a combination of pressure at 3 atmosphere absolute (ATA) and pure oxygen] greatly inhibited behavioural signs of neuropathic pain manifested as thermal hyperalgesia and mechanical allodynia. Such an HBO 2 treatment also inhibited nerve injury-induced induction of c-Fos and activation of astrocytes and increased phosphorylation of NR2B receptor and the subsequent Ca 2+-dependent signals in rats. Neither high pressure (up to 3 ATA) nor pure oxygen alone resulted in analgesic effect. In clinical trials, one course of HBO 2 therapy (10 consecutive days) produced a rapid-onset, dose-dependent and long-lasting analgesic effects evidenced by the decreased doses of carbamazepine required for keeping patient pain at a minimum and decreased scores of visual analogue scales, which was used for patient's self-evaluation. CONCLUSIONS: These findings support that HBO 2 therapy is an effective approach for treating neuropathic pain in both animals and human beings and suggest that neural protection, anti-inflammation and inhibition of nerve injury-induced altered neural activity may contribute to the analgesic effect of HBO 2 therapy.

Microbial succession of glycogen accumulating organisms in an anaerobic-aerobic membrane bioreactor with no phosphorus removal.

The succession of glycogen accumulating organisms (GAOs) has been observed in an acetate-fed, anaerobic-aerobic sequencing membrane bioreactor (MBR) operated for 260 days without enhanced biological phosphorus removal (EBPR) activity. Semi-quantitative fluorescence in situ hybridization results showed that a gammaproteobacterial lineage GB frequently observed in EBPR processes was initially the numerically dominant species (50-66% of total cells) of the GAO in the MBR from day 1 to day 38. During this period, succession of two different subgroups of group GB was also observed. On day 85 onward, a population shift from GB group to 'Defluvicoccus'-related tetrad-forming organisms (TFO) occurred. This microbial succession was suspected to be related to the applied operating conditions (long hydraulic retention time and long solid residence time) which favored the proliferation of 'Defluvicoccus'-related TFO rather than the GB group. Application of terminal restriction fragment length polymorphism on selected samples further revealed that the microbial diversity of the seeding sludge as determined by the number of terminal restriction fragments was higher than that of sludge samples taken after day 85.

Phylogenetic and physiological diversity of tetrad-forming organisms in deteriorated biological phosphorus removal systems.

Polyhydroxyalkanoate (PHA)- and polyphosphate-accumulating traits of different taxonomic tetrad-forming organisms (TFOs) in two anaerobic-aerobic sequential batch reactors (SBRs) were characterized by the simultaneous use of fluorescence in-situ hybridization, PHB stain and DAPI stain. The two SBRs with glucose as the main carbon source were operated under different P:total organic carbon feeding ratios for more than 300 days, but both exhibited no enhanced biological phosphorus removal (EBPR) activity. Microscopic observations on sludge samples taken at various times from those two SBRs revealed that TFOs consistently accounted for more than 50% of total cells, and were mostly affiliated with the beta- and gamma-subclasses of Proteobacteria and the high G+C phylum of gram-positive bacteria (HGC). Those TFOs from the beta-Proteobcateria exhibited PHB stain positive and DAPI stain negative, indicating that they could utilize compounds other than polyphosphate (i.e. glycogen) as reducing power for PHA synthesis from glucose. In contrast, two types of TFOs within the HGC group showed negative PHB stain and positive DAPI stain, indicating their capacity to accumulate polyphosphate without the synthesis and degradation of PHA. This metabolic trait was different from the widely accepted biochemical model of EBPR and non-EBPR metabolisms. Other TFOs within the HGC group and gamma-Proteobacteria showed negative responses to both PHA and DAPI stains, and their function in the deteriorated EBPR system need to be further clarified. Overall findings suggested that the phylogenic and physiological heterogeneity of TFOs in anaerobic-aerobic activated sludge systems were diverse and greatly exceeded the current understanding.

In situ identification of polyphosphate- and polyhydroxyalkanoate-accumulating traits for microbial populations in a biological phosphorus removal process.

Polyphosphate- and polyhydroxyalkanoate (PHA)-accumulating traits of predominant microorganisms in an efficient enhanced biological phosphorus removal (EBPR) process were investigated systematically using a suite of non-culture-dependent methods. Results of 16S rDNA clone library and fluorescence in situ hybridization (FISH) with rRNA-targeted, group-specific oligonucleotide probes indicated that the microbial community consisted mostly of the alpha- (9.5% of total cells), beta- (41.3%) and gamma- (6.8%) subclasses of the class Proteobacteria, Flexibacter-Cytophaga (4.5%) and the Gram-positive high G+C (HGC) group (17.9%). With individual phylogenetic groups or subgroups, members of Candidatus Accumulibacter phosphatis in the beta-2 subclass, a novel HGC group closely related to Tetrasphaera spp., and a novel gamma-proteobacterial group were the predominant populations. Furthermore, electron microscopy with energy-dispersive X-ray analysis was used to validate the staining specificity of 4,6-diamino-2-phenylindole (DAPI) for intracellular polyphosphate and revealed the composition of polyphosphate granules accumulated in predominant bacteria as mostly P, Ca and Na. As a result, DAPI and PHA staining procedures could be combined with FISH to identify directly the polyphosphate- and PHA-accumulating traits of different phylogenetic groups. Members of Accumulibacter phosphatis and the novel gamma-proteobacterial group were observed to accumulate both polyphosphate and PHA. In addition, one novel rod-shaped group, closely related to coccus-shaped Tetrasphaera, and one filamentous group resembling Candidatus Nostocoidia limicola in the HGC group were found to accumulate polyphosphate but not PHA. No cellular inclusions were detected in most members of the alpha-Proteobacteria and the Cytophaga-Flavobacterium group. The diversified functional traits observed suggested that different substrate metabolisms were used by predominant phylogenetic groups in EBPR processes.

Identification of a novel group of bacteria in sludge from a deteriorated biological phosphorus removal reactor.

The microbial diversity of a deteriorated biological phosphorus removal reactor was investigated by methods not requiring direct cultivation. The reactor was fed with media containing acetate and high levels of phosphate (P/C weight ratio, 8:100) but failed to completely remove phosphate in the effluent and showed very limited biological phosphorus removal activity. Denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S ribosomal DNA was used to investigate the bacterial diversity. Up to 11 DGGE bands representing at least 11 different sequence types were observed; DNA from the 6 most dominant of these bands was further isolated and sequenced. Comparative phylogenetic analysis of the partial 16S rRNA sequences suggested that one sequence type was affiliated with the alpha subclass of the Proteobacteria, one was associated with the Legionella group of the gamma subclass of the Proteobacteria, and the remaining four formed a novel group of the gamma subclass of the Proteobacteria with no close relationship to any previously described species. The novel group represented approximately 75% of the PCR-amplified DNA, based on the DGGE band intensities. Two oligonucleotide rRNA probes for this novel group were designed and used in a whole-cell hybridization analysis to investigate the abundance of this novel group in situ. The bacteria were coccoid and 3 to 4 microm in diameter and represented approximately 35% of the total population, suggesting a relatively close agreement with the results obtained by the PCR-based DGGE method. Further, based on electron microscopy and standard staining microscopic analysis, this novel group was able to accumulate granule inclusions, possibly consisting of polyhydroxyalkanoate, inside the cells.