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1.  The polymorphism of Insulin-like growth factor-I (IGF-I) is related to osteoporosis and bone mineral density in postmenopausal population 
Objective: It has been shown that Insulin-like growth factor-1 (IGF-1) may be related with bone mineral density (BMD) or osteoporosis. But there are few evidences on the role of genetic variation of IGF-1 on the BMD or osteoporosis. We observed the relationship between polymorphisms of IGF-1(rs35767, rs2288377 and rs5742612) with osteoporosis and BMD in the postmenopausal female population in our study.
Methods: A total of 216 postmenopausal women with a primary diagnosis of osteoporosis and 220 normal healthy women were included in the study. Genomic DNA of IGF-1 rs35767, rs2288377 and rs5742612 was extracted from the whole blood using QIAamp blood DNA mini kits (QIAGEN, Hilden, Germany) according to the methods recommended by the manufacturer.
Results: We found that T allele of rs35767 had higher increased risk of osteoporosis (OR=1.34, 95%CI=1.0-1.81). Those carrying T allele of rs35767 had a significant lower BMD at L1–L4 vertebrae, femoral neck, total hip and trochanter when compared with those carrying C allele (P < 0.05). In addition, the BMD of L1–L4 vertebrae, femoral neck, total hip and trochanter decreased by 2.09%, 3.74%, 3.52% and 2.54% in women carrying T alleles compared with those carrying C alleles.
Conclusion: Our study suggests that polymorphism in IGF-I rs35767 was significantly associated with BMD and osteoporosis in postmenopausal female population, and polymorphism of rs35767 could be a marker for lower BMD and risk of osteoporosis.
doi:10.12669/pjms.301.4264
PMCID: PMC3955557  PMID: 24639846
Bone mineral density; Insulin-like growth factor-I; Osteoporosis; Polymorphism
2.  Genetically Encoded Fluorescent Redox Probes 
Sensors (Basel, Switzerland)  2013;13(11):15422-15433.
Redox processes are involved in almost every cell of the body as a consequence of aerobic life. In the past decades, redox biology has been increasingly recognized as one of the key themes in cell signaling. The progress has been accelerated by development of fluorescent probes that can monitor redox conditions and dynamics in cells and cell compartments. This short paper focuses on fluorescent redox probes that are genetically encoded, and discusses their properties, molecular mechanism, advantages and pitfalls. Our recent work on reaction-based encoded probes that are responsive to particular redox signaling molecules is also reviewed. Future challenges and directions are also commented.
doi:10.3390/s131115422
PMCID: PMC3871076  PMID: 24225906
redox signaling; fluorescent protein (FP)-based sensors; reaction-based probes; unnatural amino acids (UAAs); redox probes
3.  Variability in the Correlation between Asian Dust Storms and Chlorophyll a Concentration from the North to Equatorial Pacific 
PLoS ONE  2013;8(2):e57656.
A long-term record of Asian dust storms showed seven high-occurrence-frequency centers in China. The intrusion of Asian dust into the downwind seas, including the China seas, the Sea of Japan, the subarctic North Pacific, the North Pacific subtropical gyre, and the western and eastern Equatorial Pacific, has been shown to add nutrients to ocean ecosystems and enhance their biological activities. To explore the relationship between the transported dust from various sources to the six seas and oceanic biological activities with different nutrient conditions, the correlation between monthly chlorophyll a concentration in each sea and monthly dust storm occurrence frequencies reaching the sea during 1997–2007 was examined in this study. No correlations were observed between dust and chlorophyll a concentration in the <50 m China seas because atmospheric deposition is commonly believed to exert less impact on coastal seas. Significant correlations existed between dust sources and many sea areas, suggesting a link between dust and chlorophyll a concentration in those seas. However, the correlation coefficients were highly variable. In general, the correlation coefficients (0.54–0.63) for the Sea of Japan were highest, except for that between the subarctic Pacific and the Taklimakan Desert, where it was as high as 0.7. For the >50 m China seas and the North Pacific subtropical gyre, the correlation coefficients were in the range 0.32–0.57. The correlation coefficients for the western and eastern Equatorial Pacific were relatively low (<0.36). These correlation coefficients were further interpreted in terms of the geographical distributions of dust sources, the transport pathways, the dust deposition, the nutrient conditions of oceans, and the probability of dust storms reaching the seas.
doi:10.1371/journal.pone.0057656
PMCID: PMC3584023  PMID: 23460892
4.  Förster Resonance Energy Transfer-Based Biosensors for Multiparameter Ratiometric Imaging of Ca2+ Dynamics and Caspase-3 Activity in Single Cells 
Analytical chemistry  2011;83(24):9687-9693.
As one of the principal cytoplasmic second messengers, the calcium ion (Ca2+) is central to a variety of intracellular signal transduction pathways. Accordingly, there is a sustained interest in methods for spatially- and temporally resolved imaging of the concentration of Ca2+ in live cells using noninvasive methods such as genetically encoded biosensors based on Forster resonance energy transfer (FRET) between fluorescent proteins (FPs). In recent years, protein-engineering efforts have provided the research community with FRET-based Ca2+ biosensors that are dramatically improved in terms of enhanced emission ratio change and optimized Ca2+ affinity for various applications. We now report the development and systematic optimization of a pair of spectrally distinct FRET-based biosensors that enable the simultaneous imaging of Ca2+ in two compartments of a single cell without substantial spectral crosstalk between emission channels. Furthermore, we demonstrate that these new biosensors can be used in conjunction with previously reported caspase-3 substrates based on the same set of FRET pairs.
doi:10.1021/ac202595g
PMCID: PMC3560285  PMID: 22080726 CAMSID: cams2610
5.  MicroRNA-195 Inhibits the Proliferation of Human Glioma Cells by Directly Targeting Cyclin D1 and Cyclin E1 
PLoS ONE  2013;8(1):e54932.
Glioma proliferation is a multistep process during which a sequence of genetic and epigenetic alterations randomly occur to affect the genes controlling cell proliferation, cell death and genetic stability. microRNAs are emerging as important epigenetic modulators of multiple target genes, leading to abnormal cellular signaling involving cellular proliferation in cancers.In the present study, we found that expression of miR-195 was markedly downregulated in glioma cell lines and human primary glioma tissues, compared to normal human astrocytes and matched non-tumor associated tissues. Upregulation of miR-195 dramatically reduced the proliferation of glioma cells. Flow cytometry analysis showed that ectopic expression of miR-195 significantly decreased the percentage of S phase cells and increased the percentage of G1/G0 phase cells. Overexpression of miR-195 dramatically reduced the anchorage-independent growth ability of glioma cells. Furthermore, overexpression of miR-195 downregulated the levels of phosphorylated retinoblastoma (pRb) and proliferating cell nuclear antigen (PCNA) in glioma cells. Conversely, inhibition of miR-195 promoted cell proliferation, increased the percentage of S phase cells, reduced the percentage of G1/G0 phase cells, enhanced anchorage-independent growth ability, upregulated the phosphorylation of pRb and PCNA in glioma cells. Moreover, we show that miR-195 inhibited glioma cell proliferation by downregulating expression of cyclin D1 and cyclin E1, via directly targeting the 3′-untranslated regions (3′-UTR) of cyclin D1 and cyclin E1 mRNA. Taken together, our results suggest that miR-195 plays an important role to inhibit the proliferation of glioma cells, and present a novel mechanism for direct miRNA-mediated suppression of cyclin D1 and cyclin E1 in glioma.
doi:10.1371/journal.pone.0054932
PMCID: PMC3557299  PMID: 23383003
6.  Papulonecrotic Tuberculid with Positive Acid-fast Bacilli 
Two patients suffered from ‘boils’ on their arms and/or thighs for several months. A diagnosis of papulonecrotic tuberculid (PNT) was made based on clinical, laboratory parameters, histopathology, and a prompt response to multi-drug anti-tuberculosis treatment. We checked the pathological sections carefully and finally found a small amount of positive acid-fast bacilli. We analyzed the clinical and histopathological features of PNT in order to offer reference of preventing and controlling the disease.
doi:10.4103/0019-5154.105324
PMCID: PMC3555389  PMID: 23372228
Acid-fast bacilli; multi-drug therapy; papulonecrotic tuberculid; real-time polymerase chain reaction
7.  A Qualitative Exploration of Barriers to Condom Use among Female Sex Workers in China 
PLoS ONE  2012;7(10):e46786.
Background
Sex workers in China continue to engage in unprotected sex acts that put them at risk for contracting HIV (Human Immunodeficiency Virus) and other STIs (Sexually Transmitted Infections). The purpose of this study was to explore women’s work history, the context of sex work, condom use, HIV testing services, and potential barriers to condom use in a sample of FSWs (female sex workers) in Guangzhou, China.
Methodology/Principal Findings
In-depth, semi-structured, face-to-face interviews were conducted with 24 FSWs in Guangzhou, China. Informants were recruited using a purposive sampling technique. Qualitative data were coded and analyzed using NVivo 8.0. The majority of respondents were internal economic migrants who had entered the sex industry in pursuit of greater financial reward. Most women in the study were married or had steady boyfriends, and were young, with secondary education and limited knowledge about HIV and STIs. Most were not satisfied with their current living conditions and expressed a desire to leave the sex industry. Women reported that they were more likely to use condoms during sex acts with commercial partners than with non-commercial partners. The potential stigma of being seen as a sex worker prevented many from accessing HIV testing. Three key factors put these FSWs at risk for HIV and STIs: unreasonable trust toward clients, stereotypes and assumptions about customers, and financial incentives.
Conclusions/Significance
These findings suggest that social and economic factors play an important role in shaping sexual decision-making among female sex workers in Guangzhou. We argue that greater insight into and attention to these factors could enhance the success of HIV prevention efforts.
doi:10.1371/journal.pone.0046786
PMCID: PMC3466319  PMID: 23056452
8.  A Syndemic of Psychosocial Problems Places the MSM (Men Who Have Sex with Men) Population at Greater Risk of HIV Infection 
PLoS ONE  2012;7(3):e32312.
Background
The MSM (Men who have sex with men) population suffers from very high rates of concurrent psychosocial problems. Together, these problems comprise a syndemic that increases the risk of HIV infection for this community. The precise mechanisms through which this syndemic can raise the likelihood of HIV infection warrant further exploration.
Methodology/Principal Findings
A total of 522 MSM were enrolled via a multiframe sampling approach and were asked to report psychosocial problems, risky sexual behaviors and HIV test results. A count of psychosocial health problems was calculated to test the additive relationship of these factors on HIV risk. Adjusting analysis and restriction analysis were used to determine a proposed intermediate pathway. Psychosocial health problems are highly concurrent and intercorrelated among urban MSM. Greater numbers of health problems are significantly and positively associated with HIV infection, which is mediated, at least partially, by risky sexual behaviors.
Conclusions/Significance
MSM experience concurrent psychosocial health problems that correlate with HIV infection in this community. We recommend the development of coping strategies for this population to deal with these psychosocial problems, both in prevention research and health policy.
doi:10.1371/journal.pone.0032312
PMCID: PMC3316524  PMID: 22479319
9.  Association between sleep quality and arterial blood pressure among Chinese nonagenarians/centenarians 
Summary
Background
There is association between sleep quality and arterial blood pressure, but it is still unclear if the association also exists in the very elderly. We examined the individual association between sleep quality and arterial blood pressure among the very elderly.
Material/Methods
The present study analyzed data from a survey that was conducted on all residents aged 90 years or older in a district with 2,311,709 inhabitants in 2005. Sleep quality was measured using The Pittsburgh Sleep Quality Index (PSQI).
Results
The subjects included in the statistical analysis were 216 men and 444 women. There were no significant differences in sleep quality scores, sleep latency, and sleep efficiency percentage and prevalence of poor sleep quality between subjects with and without hypertension. None of the differences in systolic blood pressure, diastolic blood pressure, and prevalence of hypertension, systolic hypertension and diastolic hypertension among subjects with well, fairly and poor sleep quality were significant. Multiple logistic regressions showed that unadjusted and adjusted Odds Ratio (ORs) of poor sleep quality for increased risk for hypertension were significant.
Conclusions
Among very elderly subjects, there was no association between sleep quality and arterial blood pressure.
doi:10.12659/MSM.882512
PMCID: PMC3560755  PMID: 22367137
arterial blood pressure; sleep quality; nonagenarians/centenarians
10.  Apoptosis of human cholangiocarcinoma cells induced by ESC-3 from Crocodylus siamensis bile 
AIM: To investigate the effects of ESC-3 isolated from crocodile bile on the growth and apoptosis induction of human cholangiocarcinoma cells.
METHODS: ESC-3 was isolated from crocodile bile by Sephadex LH-20 and RP-18 reversed-phase column. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay was conducted to determine the effects of ESC-3 on the proliferation of human cholangiocarcinoma cell lines (QBC939, Sk-ChA-1 and MZ-ChA-1). Giemsa staining, Hoechst 33258 and acridine orange/ethidium bromide staining showed the morphological changes of Mz-ChA-1 cells exposed to ESC-3 at different concentrations. Flow cytometry with regular propidium iodide (PI) staining was performed to analyze the cell cycle distribution of Mz-ChA-1 cells and to assess apoptosis by annexin v-fluorescein isothiocyanate (V-FITC)/PI staining. Rh123 staining was used to detect the alteration of mitochondrial membrane potential (ΔΨm). The protein levels of Bax, Bcl-2, Cdk2, cytochrome c and caspase-3 were further confirmed by Western blotting.
RESULTS: ESC-3 significantly inhibited the growth of three human cholangiocarcinoma cell lines and arrested Mz-ChA-1 cell cycle at G0/G1 phase. Mz-ChA-1 cells showed typical apoptotic morphological changes after treated with ESC-3 (10 μg/mL) for 48 h. Cell death assay indicated that Mz-ChA-1 cells underwent apoptosis in a dose-dependent manner induced by ESC-3. In addition, ESC-3 treatment could downregulate the protein level of Bcl-2 and upregulate the Bax, leading to the increase in the ratio of Bax to Bcl-2 in Mz-ChA-1 cells. Meanwhile, cytochrome c was released from the mitochondria into the cytosol, which subsequently initiated the activation of caspase-3. All these events were associated with the collapse of the mitochondrial membrane potential.
CONCLUSION: ESC-3, the active ingredient of crocodile bile, induced apoptosis in Mz-ChA-1 cells through the mitochondria-dependent pathway and may be a potential chemotherapeutic drug for the treatment of cholangiocarcinoma.
doi:10.3748/wjg.v18.i7.704
PMCID: PMC3281230  PMID: 22363144
Crocodylus siamensis bile; Cholangiocarcinoma; Antiproliferation; Apoptosis; Mitochondria
11.  A method to site-specifically introduce methyllysine into proteins in E. coli 
A mutant pyrrolysyl-tRNA synthetase/tRNA pair was used to genetically encode allylcarbamoyl methyllysine in bacteria. This amino acid can be converted to methyllysine with a ruthenium catalyst, providing a straightforward approach for site-specifically introducing methyllysine residues into proteins.
doi:10.1039/c0cc00108b
PMCID: PMC2928331  PMID: 20571694
12.  The Effects of Exercise-induced Fatigue on Acetylcholinesterase Expression and Activity at Rat Neuromuscular Junctions 
Acta Histochemica et Cytochemica  2009;42(5):137-142.
Acetylcholinesterase is the enzyme that terminates neurotransmission by hydrolyzing the acetylcholine released by the motoneurons at the neuromuscular junctions. Although acetylcholinesterase has been studied for almost a century, the underlying relationship between exercise-induced fatigue and acetylcholinesterase activity at the synaptic cleft is not clear. The purpose of this study was to assess the effects of exercise-induced fatigue on the expression and activity of acetylcholinesterase at the neuromuscular junctions. The expression and activity of acetylcholinesterase at the gastrocnemius neuromuscular junctions was decreased transiently by exercise-induced fatigue and then gradually increased over 24 hr. The expression of acetylcholinesterase in the 24 hr recovery group returned to the level of the control (non-exercised) group, but the activity of acetylcholinesterase remained significantly lower. These data suggest that the decrease of acetylcholinesterase expression and activity may be involved in the production and/or maintenance of exercise-induced fatigue.
doi:10.1267/ahc.09019
PMCID: PMC2775104  PMID: 19918322
exercise; central fatigue; peripheral fatigue; acetylcholinesterase; neuromuscular junctions
13.  Synaptophysin Expression in Rat Retina Following Acute High Intraocular Pressure 
Acta Histochemica et Cytochemica  2008;41(6):173-178.
In response to injury, synapse alteration may occur earlier than the changes in the cell body of neurons. Although retinal ganglion cell death and thinning of the inner part of retina were found after acute high intraocular pressure (HIOP), the structural and functional changes of synapses in the retina remain unknown. In the present study, we investigated the protein and mRNA expression of synaptophysin (SYN), an important molecule closely related to synaptic activities, synaptogenesis and synaptic plasticity. In addition, we also studied the ultrastructural changes of the retinal synapses. We found that (1) synaptophysin was upregulated transiently at both protein and mRNA level following HIOP; (2) broadened distribution of synaptophysin protein was present within the outer nuclear layer at the early stage following HIOP; (3) in the outer nuclear layer bouton-like vesicle-containing structures were observed by electron microscopy. This data suggested that, besides degeneration, synapses in rat retina may undergo regenerative events following HIOP.
doi:10.1267/ahc.08034
PMCID: PMC2629489  PMID: 19180202
synaptophysin; synapse; plasticity; degeneration; acute high intraocular pressure
14.  Improved Method of Ink-Gelatin Perfusion for Visualising Rat Retinal Microvessels 
Acta Histochemica et Cytochemica  2008;41(5):127-133.
To visualize completely rat retinal microvessels, the gelatin-ink perfusion condition was systematically optimized using von Willebrand factor (vWf) immunostaining as control. Whether the vessel showed by the new perfusion condition can be used for double label with neurons or glial cells in the same retina was also tested. Our results showed that infusing rats first with 20 ml of 37°C ink plus 3% gelatin at 140% rat mean arterial pressure (MAP), and subsequently with 20 ml of 37°C ink plus 5% gelatin at 180% rat MAP allowed the ink to completely fill the rat retinal microvessels. Rat retinal microvessels labeled by the perfusion method were more in number than that by vWf immunostaining. Moreover, our data, for the first time, displayed that the improved gelatin-ink perfusion had no effect on and caused no contamination to the following fluorogold labeling or immunostaining of retinal neurons or glial cells in the same tissue. These data suggest that the improved gelatin-ink perfusion technique is a superior method for morphological characterization of rat retinal microvessels, compatible to the double labeling of glial cells and neurons, and it extends the practical scale of the classic method.
doi:10.1267/ahc.08015
PMCID: PMC2576503  PMID: 18989466
ink perfusion; von Willebrand factor; microvessel; retina; rat
15.  Hue-shifted monomeric variants of Clavularia cyan fluorescent protein: identification of the molecular determinants of color and applications in fluorescence imaging 
BMC Biology  2008;6:13.
Background
In the 15 years that have passed since the cloning of Aequorea victoria green fluorescent protein (avGFP), the expanding set of fluorescent protein (FP) variants has become entrenched as an indispensable toolkit for cell biology research. One of the latest additions to the toolkit is monomeric teal FP (mTFP1), a bright and photostable FP derived from Clavularia cyan FP. To gain insight into the molecular basis for the blue-shifted fluorescence emission we undertook a mutagenesis-based study of residues in the immediate environment of the chromophore. We also employed site-directed and random mutagenesis in combination with library screening to create new hues of mTFP1-derived variants with wavelength-shifted excitation and emission spectra.
Results
Our results demonstrate that the protein-chromophore interactions responsible for blue-shifting the absorbance and emission maxima of mTFP1 operate independently of the chromophore structure. This conclusion is supported by the observation that the Tyr67Trp and Tyr67His mutants of mTFP1 retain a blue-shifted fluorescence emission relative to their avGFP counterparts (that is, Tyr66Trp and Tyr66His). Based on previous work with close homologs, His197 and His163 are likely to be the residues with the greatest contribution towards blue-shifting the fluorescence emission. Indeed we have identified the substitutions His163Met and Thr73Ala that abolish or disrupt the interactions of these residues with the chromophore. The mTFP1-Thr73Ala/His163Met double mutant has an emission peak that is 23 nm red-shifted from that of mTFP1 itself. Directed evolution of this double mutant resulted in the development of mWasabi, a new green fluorescing protein that offers certain advantages over enhanced avGFP (EGFP). To assess the usefulness of mTFP1 and mWasabi in live cell imaging applications, we constructed and imaged more than 20 different fusion proteins.
Conclusion
Based on the results of our mutagenesis study, we conclude that the two histidine residues in close proximity to the chromophore are approximately equal determinants of the blue-shifted fluorescence emission of mTFP1. With respect to live cell imaging applications, the mTFP1-derived mWasabi should be particularly useful in two-color imaging in conjunction with a Sapphire-type variant or as a fluorescence resonance energy transfer acceptor with a blue FP donor. In all fusions attempted, both mTFP1 and mWasabi give patterns of fluorescent localization indistinguishable from that of well-established avGFP variants.
doi:10.1186/1741-7007-6-13
PMCID: PMC2292683  PMID: 18325109
17.  A model of inflammatory arthritis highlights a role for oncostatin M in pro-inflammatory cytokine-induced bone destruction via RANK/RANKL 
Arthritis Research & Therapy  2004;7(1):R57-R64.
Oncostatin M is a pro-inflammatory cytokine previously shown to promote marked cartilage destruction both in vitro and in vivo when in combination with IL-1 or tumour necrosis factor alpha. However, the in vivo effects of these potent cytokine combinations on bone catabolism are unknown. Using adenoviral gene transfer, we have overexpressed oncostatin M in combination with either IL-1 or tumour necrosis factor alpha intra-articularly in the knees of C57BL/6 mice. Both of these combinations induced marked bone damage and markedly increased tartrate-resistant acid phosphatase-positive multinucleate cell staining in the synovium and at the front of bone erosions. Furthermore, there was increased expression of RANK and its ligand RANKL in the inflammatory cells, in inflamed synovium and in articular cartilage of knee joints treated with the cytokine combinations compared with expression in joints treated with the cytokines alone or the control. This model of inflammatory arthritis demonstrates that, in vivo, oncostatin M in combination with either IL-1 or tumour necrosis factor alpha represents cytokine combinations that promote bone destruction. The model also provides further evidence that increased osteoclast-like, tartrate-resistant acid phosphatase-positive staining multinucleate cells and upregulation of RANK/RANKL in joint tissues are key factors in pathological bone destruction.
doi:10.1186/ar1460
PMCID: PMC1064887  PMID: 15642143
bone; IL-1; oncostatin M; RANK; tumour necrosis factor alpha

Results 1-17 (17)