[Targeted AGEes and AGEs cross-link in drug discovery: preventing and reversing arterial sclerosis in aging and diabetes].
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Biomedical subjects
Publications and source records attributed to Song Li.
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OBJECTIVE: To study the prevalence rates of birth defects in high and low risk areas in China. METHODS: A population-based surveillance system on birth defects was used to obtain the prevalence rates of 24 kinds of major external birth defects from > or = 20 weeks of gestation to 7 days of life in selected areas in Shanxi and Jiangsu provinces. RESULTS: The birth prevalence of birth defects (232.4 per 10,000 births) and neural tube defects (NTDs) (138.7 per 10,000 births) in four counties of Shanxi province were significantly higher than that in Taiyuan city (75.3 and 28.2 per 10,000 births, respectively). There was no significant difference for all selected birth defects between Wuxi city and Xishan counties in low risk areas. There was a 6.1-fold of higher prevalence for NTDs in Taiyuan city compared with that in Wuxi areas (4.6 per 10,000 births). In four counties of Shanxi province, the prevalence rates of anencephaly, spina bifida, hydrocephaly, cleft palate alone and polydactyly were significantly higher than in Wuxi areas. The NTDs prevalence rate in four counties of Shanxi was 30.2 times higher than in Wuxi areas. When compared with previous surveillance data, the NTDs prevalence rate did not present obvious declining trend in high risk areas. The birth prevalence rate had a 31.8% decrease when births were calculated after 28 gestational weeks and compared with those from 20 gestational weeks. CONCLUSION: NTDs remained to be the most common birth defect seen in Shanxi province. The birth prevalence rate of NTDs in some areas of Shanxi province was among the highest that ever reported in the world in comparison with data from other countries and regions. The current prevalence rate in high risk areas in Shanxi province did not clearly show a declining trend. Programs on surveillance and prenatal diagnosis were proved to have made big impact on the rates of major external birth defects.
OBJECTIVE: To preliminarily investigate the relationship between the Lactate dehydrogenase (LDH) activity of Streptococcus mutans (S. mutans) and dental caries initiation. METHODS: 100 S. mutans strains derived from caries-active and caries-free individuals were cultured in BHI medium supplemented with glucose. Cells were extracted and ruptured, and the extracted liquid protein was quantified with Coomassie brilliant blue G250 staining methods. LDH activity was assayed using the pyruvate-dependent oxidation of NADH-with and without FDP. RESULTS: LDH activity of the two groups strains had no difference (P > 0.05), but the distribution of differ class enzyme activity strains in the two groups was different (P < 0.05). CONCLUSION: LDH activity of S. mutans is correlated to the initiation of dental caries to some extent. The measurement methods in this study can be applied in preliminary quantitation of LDH activity and the screening of S. mutans strains.
OBJECTIVE: The purpose of the study is to detect the expression of TGF-beta1 mRNA in the maxillary sutures of puberty rhesus during different periods of the loading of intermaxillary class III orthopedic force. METHODS: The animal model was established with 6 puberty female rhesus, which were randomly divided into experimental group (wearing class III twin-block magnet appliance, each 2 rhesus for 3 and 6 month respectively) and control group (not wearing any appliance, each 1 rhesus for 3 and 6 month respectively). Tissue sections were obtained perpendicular to the sutures. In situ hybridization was used to the expression of TGF-beta1 mRNA, and the expression intensity was measured and statistics was performed by SPSS 11.0. RESULTS: There were no statistic differences of cellular density between palatomaxillary suture in vertical group and pterygomaxillary suture in horizontal group, but statistic differences were found between other groups. The expression of TGF-beta1 mRNA was detected in the control group, but the expression intensity was obviously increased after the load of intermaxillary class III orthopedic force. Statistically significant differences were found among all groups except the six months experimental group and control group of temporozygomatic sutures and pterygomaxillary sutures. Experimental groups were more intensive than the control group and three months group was more intensive than the six months group. CONCLUSION: The active tissue remodeling happened in the circummaxillary sutures by the effect of class III intermaxillary orthopedic force. Cell proliferation, extracellular matrix synthesis and bone formation were accelerated. During the different remodeling periods, the expression intensities were different, which may be related to the different force loading manners, the different reaction of sutures to the orthopedic force and the different biological function of TGF-beta1 in the different periods.
To examine the self-association of CD4 molecules and preliminary studies on its biological function by several indirect methods. A series of CD4 chimeras were generated including truncated CD4 lacking the short cytoplasmic tail, deleted mutantsD1/D2 devoid of D3 and D4 and D3/D4 devoid of D1 and D2 by PCR techniques, as well as another three CD4 chimeric genes by fused human Fas cytoplasmic death domain to the downstream of the above chimeras respectively. All these molecules were subcloned into pEGFP-N1, forming the corresponding expression vectors. After introducing into HEK293 cells, gene-modified cell morphological changes and target protein subcellular localization were observed and analyzed by a confocal microscopy. Moreover, stable 293/CD4 clones were obtained by transfecting the truncated CD4 recombinant plasmid into the HEK293 cell line and selected by G418. The fluorescene intensity and rosette formation of different clones was each analyzed by a confocal microscopy and cell adhesive assays. It's seen that CD4-Fas fusion gene could induce approximately 80% cell apoptosis of transfected HEK293 cells, compared to FKBP12-Fas is about 30% and CD4 gene only is 7%. Furthermore, both D1/D2-Fas and D3/D4 Fas chimeras could trigger nearly all transfected HEK293 cells to death. Cell adhesion assays showed that neither the D1/D2 nor D3/D4 chimeras when expression in HEK293 cells binds to MHC class II + Raji B cells. Interestedly, there were two type stable clones among 293/CD4. Fluorescence intensity analysis displayed that one' mean fluorescence intensity value is about twice of the other while cell-cell binding examination showed that the former is capable of forming rosette with Raji cells but the latter. All these results suggest that CD4 molecules most likely could exist as a dimer or even an oligomer on transfected HEK293 cell surface, which constitute a functional form for stable binding to MHC class II molecules.
A reference material used for wavelength calibration of fluorescence spectrophotometer was found. The holmium doped oxide reference material GBW(E) 130112 is a kind of standard reference material for absorption spectrophotometer. It can emit 547.7 nm fluorescence when excited by xenon lamp light. The excitation spectrum of 547.7 nm fluorescence was measured. It was found that the measured peaks of excitation spectrum are positioned at 333.56, 360.43 and 418.39 nm, respectively, which are coincident with the true values 333.8, 360.9 and 418.5 nm of reference material certification. It was illustrated that the holmium doped oxide reference material GBW(E)130112 could be used as reference material for the excitation wavelength calibration of the fluorescence spectrophotometer. Its property could be enhanced very much if high luminescent efficiency material is selected as rare earth ion doped matrix, and the purity is enhanced to reduce the cross relaxation.
To screen NFAT antagonistic drugs and research signal transduction pathway related to NFAT. Four recombinant vectors were constructed. Each consists of three tandem copies of the human IL-2 distal NFAT-AP1 binding site in the context of the minimal IL-2 enhancer, either the sequence from -326 - +46 or the sequence from -89 - +46 (containing only the TATA box), driving a luciferase reporter gene or a destabilized enhanced green fluorescence protein (d2EGFP) reporter gene, respectively. Transient transfection of Jurkat cells was achieved by electroporation with 5 - 10 microg of the above plasmid and one pulse at 200V, 65ms. Plasmid pEFBos-mNFAT1 constitutively expressing murine full length NFAT1 protein was used for transient cotransfection. The results showed that neither of non-stimulation nor PMA or ionomycin stimulation alone could activate the reporter gene except PMA plus ionomycin costimulation. Furthermore, overexpressed murine NFAT1 augmented the activation of either IL-2 promoter or NFAT-AP1 enhancer drived reporter gene compared to the endogenous did. However, the reporter gene expression was nearly completely inhibited by pretreatment for 1h with FK506 at 5 microg/mL and then stimulation for 6-12h with PMA plus ionomycin in the presence of FK506. These findings indicated that such a transient Jurkat cell model offered a potential platform for preliminary screening of FK506 or CsA-like immunosuppressive agents.
The fluorescence spectroscopy of humen serum albumin (HSA) and the interaction of HSA and the Levofloxacin were studied. When the Levofloxacin was added into HSA solution gradually, an interesting new phenomenon emerged in emission spectrum peak. The intensity of 343.70 nm emission peak of HSA decreased obviously and moved towards long-wavelength, which is atypical quenching phenomenon. It was found that the excitation peak of HSA was positioned at 286.70 nm also. A new emission peak emerged at 503.96 nm, which resulted from the Levofloxacin. The excitation peaks of 503.96 nm fluorescence of Levofloxacin were positioned at 300.16 nm and 336.16 nm. When Levoflaxasin was added into HSA gradually, the 300.16 nm and 336.16 nm excitation peaks moved towards long wavelength. The dissociation constant of Levofloxacin from HSA is about Kd = 3.65 x 10(-5) (mol x L(-1)). The combination constant of Levofloxacin is about Ks = 2.742 x 10(4) (L x mol(-1)). The quenching process of Levofloxacin-HSA is not dynamic quenching, which resulted from the molecular diffusion and collision. It is caused by the static quenching process resulting from the chemical component between molecules. The energy transfer efficiency between Levofloxacin and HSA is E = 0.372. According to these calculation results, the combination position between the binding site of Levofloxacin and the tryptophane of HSA is about R = 1.933 nm.
PURPOSE: The purpose of this study is to quantitatively study the remodling changes of circummaxillary sutures of puberty rhesus loaded with Class III intermaxillary orthopedic force. METHODS: The animal model was established with 6 puberty female rhesus, which were randomly divided into experimental group (wore Class III Twin-block magnet appliance, each 2 rhesus for 3 and 6 months respectively) and control group(did not wear any appliance, each 1 rhesus for 3 and 6 months respectively ). The specimens were stained by hematoxylin and eosin, and image analysis system (IAS) was used in the quantatitive study of sutures width and cell density. One-way analysis of variance (ANOVA) was employed, and LSD analysis was used when interclass significance was found. RESULTS: Compared to the control group, different changes were shown in different sutures.New bone formation was observed in all experimental animal. There was no significance difference between palatomaxillary suture and pterygomaxillary suture in cellular density (P>0.05). Zygomaticotemporal suture showed no difference between 3-month experimental group and control group;however, 6-month experimental group was significantly higher than 3-month experimental group (P<0.01).For frontomaxillary suture, 3-month experimental group was significantly higher than the control group, and 6-month experimental group was significantly higher than later one (P<0.01).Zygomaticomaxillary suture in the experimental group was significantly higher than the control group (P<0.01);meanwhile, 3-month experimental group was significantly higher than 6-month experimental group (P<0.05). Furthermore, the width of sutures were different in these groups. Palatomaxillary suture became significant narrower in 3-month experimental group (P<0.01),but zygomaticotemporal suture became significant wider (P<0.01). The width of sutures restored at 6th month (P>0.05). The width of pterygomaxillary suture was not different between 3-month experimental group and the control group (P>0.05),but it became significant wider in 6-month experimental group (P<0.05). Frontomaxillary sutures of the experimental group were significantly narrower than the control group, and 6-month experimental group was significantly narrower than 3-month experimental group (P<0.01). For zygomaticomaxillary suture, the experimental groups were significantly wider than the control group;meanwhile, there was no difference between the experimental groups. CONCLUSION: Proper Class III orthopedic force can promote the physiological remodling of maxillary sutures.
OBJECTIVE: To describe the epidemiological characters of pregnancy-induced hypertension (PIH) in Jiaxing areas of Zhejiang province of China between 1995 and 2000. METHODS: We analyzed the perinatal health surveillance data that was collected as part of the Sino-American cooperative project on neural tube defects prevention established in 1992. The study population consisted of 136 070 pregnant women with at least 20 weeks of gestational age. National diagnostic criteria were used to identify the cases which were divided into three subgroups: mild, moderate and severe. RESULTS: 15 127 cases were identified and the overall incidence rate of PIH was 11.1% (95% CI : 11.0% - 11.3%). Among all the cases, mild, moderate and severe PIH were accounted for 71.4%, 22.3% and 6.3%, respectively. The proportions of PIH cases that occurred in the second trimester, third trimester and during delivery appeared to be 4.2%, 34.4% and 61.4%, respectively. There was a significant fall in the trend of PIH occurrence every year, which dropped from 10.7% in 1995 to 8.6% in 2000 by 19.6%. More risk of PIH seemed to be related to those mothers living in the urban areas with age under 20 or above 35, being peasants and having little educational, having had multiple gestations, conceiving in spring/summer or delivering in winter or spring etc. Compared with the results of national survey in 1988, the incidence rate of PIH was higher by 18.1%, while the proportion of severe PIH was much lower by 68.8%. Although the incidence rates of PIH in urban and rural areas were somehow similar, the proportion of severe PIH in rural areas was much higher than that in urban areas. CONCLUSION: Overall incidence rate and distribution of PIH were reported. Compared with the results in 1988, incidence rate of PIH was much higher, particularly for mild cases.
Certain human malignancies including prostate cancer overexpress sigma receptor, a membrane bound protein that binds haloperidol and various other neuroleptics with high affinity. An anisamide derivatized ligand possesses high affinity for sigma receptors and we hypothesized that its incorporation into the liposomes encapsulating doxorubicin (DOX) can specifically target and deliver the drug to prostate cancer cells that overexpress sigma receptors. A polyethylene glycol phospholipid was derivatized with an anisamide ligand, which was then incorporated into the DOX-loaded liposome. The resulting anisamide-conjugated liposomal DOX showed significantly higher toxicity to DU-145 cells than non-targeted liposomal DOX, the IC50 being 1.8 microM and 14 microM respectively. The cytotoxicity of the targeted liposomal DOX, however, was significantly blocked by haloperidol, suggesting that the enhanced cytotoxicity was specifically mediated by the sigma receptors. Fluorescence imaging studies after intravenous (i.v.) administration showed that incorporation of anisamide into liposomes significantly improved their accumulation into the tumor. A weekly injection of the targeted liposomal DOX for 4 weeks at a dose of 7.5 mg/kg led to a significant growth inhibition of established DU-145 tumor in nude mice with minimal toxicity. Free DOX was effective, but associated with significant toxicities. The present study is the first to demonstrate the use of small molecular weight ligand for mediating efficient targeting of liposomal drugs to sigma receptor expressing prostate cancer cells both in vitro and in vivo.
Bone marrow mesenchymal stem cells (MSCs) can differentiate into a variety of cell types, including vascular smooth muscle cells (SMCs), and have tremendous potential as a cell source for cardiovascular regeneration. We postulate that specific vascular environmental factors will promote MSC differentiation into SMCs. However, the effects of the vascular mechanical environment on MSCs have not been characterized. Here we show that mechanical strain regulated the expression of SMC markers in MSCs. Cyclic equiaxial strain downregulated SM alpha-actin and SM-22alpha in MSCs on collagen- or elastin-coated membranes after 1 day, and decreased alpha-actin in stress fibers. In contrast, cyclic uniaxial strain transiently increased the expression of SM alpha-actin and SM-22alpha after 1 day, which subsequently returned to basal levels after the cells aligned in the direction perpendicular to the strain direction. In addition, uniaxial but not equiaxial strain induced a transient increase of collagen I expression. DNA microarray experiments showed that uniaxial strain increased SMC markers and regulated the expression of matrix molecules without significantly changing the expression of the differentiation markers (e.g., alkaline phosphatase and collagen II) of other cell types. Our results suggest that uniaxial strain, which better mimics the type of mechanical strain experienced by SMCs, may promote MSC differentiation into SMCs if cell orientation can be controlled. This study demonstrates the differential effects of equiaxial and uniaxial strain, advances our understanding of the mechanical regulation of stem cells, and provides a rational basis for engineering MSCs for vascular tissue engineering and regeneration.
The present study was designed to select the effective dosage range of Z24 [3Z-3-[(1H-pyrrol-2-yl)-methylidene]-1-(1-piperidinylmethyl)-1,3-2H-indol-2-one], a novel synthetic indolin-2-ketone small-molecule compound, against tumorigenesis and angiogenesis in vitro and in vivo and to investigate the primary action mechanism of Z24 on the angiogenesis by comparing with SU5416 [3-[(2,4-dimethylpyrrol-5-yl)methyllidenyl]-indolin-2-one] in the selective effects on vascular endothelial growth factor (VEGF)/basic fibroblast growth factor (bFGF) signaling and Bcl-2-related cell vitality because Z24 is a potential inhibitor of the Bcl-2 that inhibits growth of multiple tumor types in vivo in our previous study. Per os Z24 inhibited dose-dependently the mouse S180 xenograft tumor growth and angiogenesis in mouse subcutaneous (s.c.) Matrigel plugs in vivo. The maximum growth inhibitory rate was 56.1% by 80 mg/kg/day on S180 mouse sarcoma cells; however, the maximum inhibitory potency on angiogenesis in C57BL/6 mouse subcutaneous Matrigel plug model was 50 mg/kg/day. Z24 inhibited angiogenesis in chicken chorioallantoic membrane (CAM) and invasion and inhibited tube formation of endothelial cells in a dose-dependent manner. Compared with SU5416, the IC50 (50% inhibition concentration) of Z24 on the proliferation of ECV-304 carcinoma cells induced by VEGF or bFGF was 24.4 and 17.99 microM, respectively, which is higher or lower, respectively, than that of SU5416 (14.2 microM for VEGF and 22.7 microM for bFGF). Furthermore, the IC50 of Z24 on the proliferation of Bcl-2 over-expressing HeLa cells and non-Bcl-2-expressing (wild-type) HeLa cells are 11.9 and 24.8 microM, respectively. SU5416 did not exert such a selective inhibiting effect on Bcl-2 over-expressing HeLa cells. These results suggest that Z24 per os has dose-dependent antitumor and antiangiogenesis pharmacological activity. The higher selectivity of Z24 on Bcl-2 protein and on bFGF other than VEGF signaling path may contribute to its efficiency against tumor and tumor-associated angiogenesis.
The interactions of chlorogenic acid and ferulic acid with human serum albumin (HSA) have been investigated by fluorescence and Fourier transformed infrared (FT-IR) spectrometry. Fluorescence results showed that one molecule of protein combined with one molecule of drugs at the molar ratio of drug to HSA ranging from 1 to 10, and their binding affinities (KA) are 4.37 x 10(4) M(-1) and 2.23 x 10(4) M(-1) for chlorogenic acid and ferulic acid, respectively. The primary binding site for chlorogenic acid is most likely located on IIA and that for ferulic acid in IIIA. The main mechanism of protein fluorescence quenching was static quenching process. Combining the curve-fitting results of infrared amide I and amide III bands, the alterations of protein secondary structure after drug complexation were estimated. With increasing the drug concentration, the protein alpha-helix structure decreased gradually and the reduction of protein alpha-helix structure reached about 7% and 5% for protein binding with chlorogenic acid and ferulic acid individually at the drug to protein molar ratio of 30. This indicated a partial unfolding of HSA in the presence of the two acids. From the fluorescence and FT-IR results, the binding mode was discussed.
Aberrant DNA methylation of CpG site is among the earliest and most frequent alterations in cancer. Detection of promoter hypermethylation of cancer-related gene may be useful for cancer diagnosis or the detection of recurrence. However, most of the studies have focused on a single gene only and gave little information about the concurrent methylation status of multiple genes. In this study, we attempted to develop a microarray method coupled with linker-PCR for detecting methylation status of multiple genes in the tumor tissue. A series of synthesized oligonucleotides were synthesised and purified to completely match with 16 investigated targets. Then they were immobilized on the aldehyde-coated glass slide to fabricate a DNA microarray for detecting methylation status of these genes. The results indicated that these genes were all methylated in the positive control. However, no methylated was found in these genes for the negative control. Only p16 and p15 genes were methylated in investigated genes for the gastric tumor tissue, whereas others were not methylated. The above results were validated by bisulfite DNA sequencing. Our experiments successfully demonstrated that the DNA microarray could be applied as a high-throughput tool to determine methylation status of the investigated genes.
Bone marrow mesenchymal stem cells (MSCs) can differentiate into different types of cells and have tremendous potential for cell therapy and tissue engineering. Transforming growth factor beta1 (TGF-beta) plays an important role in cell differentiation and vascular remodeling. We showed that TGF-beta induced cell morphology change and an increase in actin fibers in MSCs. To determine the global effects of TGF-beta on MSCs, we employed a proteomic strategy to analyze the effect of TGF-beta on the human MSC proteome. By using two-dimensional gel electrophoresis and electrospray ionization coupled to quadrupole/time-of-flight tandem mass spectrometers, we have generated a proteome reference map of MSCs, and we identified approximately 30 proteins with an increase or decrease in expression or phosphorylation in response to TGF-beta. The proteins regulated by TGF-beta included cytoskeletal proteins, matrix synthesis proteins, membrane proteins, metabolic enzymes, etc. TGF-beta increased the expression of smooth muscle alpha-actin and decreased the expression of gelsolin. Overexpression of gelsolin inhibited TGF-beta-induced assembly of smooth muscle alpha-actin; on the other hand, knocking down gelsolin expression enhanced the assembly of alpha-actin and actin filaments without significantly affecting alpha-actin expression. These results suggest that TGF-beta coordinates the increase of alpha-actin and the decrease of gelsolin to promote MSC differentiation. This study demonstrates that proteomic tools are valuable in studying stem cell differentiation and elucidating the underlying molecular mechanisms.
The potential for malignancy detection using dynamic infrared imaging (DIRI) has been investigated in an animal model of human malignancy. Malignancy was apparent in images formed at the vasomotor and cardiogenic frequencies of tumour bearing mice. The observation of malignancy was removed by the administration of an agent that blocks vasodilation caused by nitric oxide (NO). Image patterns similar to those that characterize malignancy could be mimicked in normal mice using an NO producing agent. Apparently DIRI allows for cancer detection in this model through vasodilation caused by malignancy generated NO. Dynamic infrared detection of vasomotor and cardiogenic surface perfusion was validated in human subjects by a comparison with laser Doppler flowmetry (LDF). Dynamic infrared imaging technology was then applied to breast cancer detection. It is shown that dynamic infrared images formed at the vasomotor and cardiogenic frequencies of the normal and malignant breast have image pattern differences, which may allow for breast cancer detection.
Although the CpG DNA immune response mediated by Toll-like receptor 9 (TLR9) has been extensively studied in a number of immune cells, the response to CpG DNA in endothelial cells (EC) is not well understood. In this study, we show that both mouse and rat lung EC display constitutive expression of TLR9 mRNA. Exposure to CpG DNA induced a potent proinflammatory response as manifested by an increased expression of IL-8 and ICAM-1 in mouse pulmonary EC. The proinflammatory response was sensitive to chloroquine, consistent with a role of endosomal contribution. A role for p38 MAPK and NF-kappaB pathway was apparent as the response was sensitive to inhibitors of p38 MAPK and NF-kappaB but was not affected by inhibitors of ERK1/2. A synergistic effect of CpG DNA and LPS on the inflammatory response is consistent with multiple TLR interaction in EC. This study suggests a possible role for CpG DNA-mediated EC immune response in the host defense system. It also has important implications in plasmid DNA-mediated pulmonary endothelium gene transfer.