Bone morphogenetic protein-7 (osteogenic protein-1, OP-1) and tooth development

The photodynamic effects of temoporfin (meso-tetrahydroxyphenylchlorin, mTHPC) and merocyanine 540 (MC540) in murine myeloid leukemia M1 and WEHI 3B (JCS) cells were compared. The mTHPC was found to be more potent and selective. At a lethal dosage of 90% killing (LD90), only 1.3 microM of mTHPC and 4.2 kJ/m2 of light irradiation was required, which was a 20-fold lower drug concentration and 11-fold smaller light dose than that required when using MC540. Meanwhile, three times less, or 15%, of the coincubated erythrocytes were destroyed by mTHPC than by MC540. Confocal micrographs showed that both drugs accumulated diffusely inside the cytoplasm in a very similar fashion, but mTHPC induced a more extensive apoptosis in photosensitized JCS cells. For example, at LD90, mTHPC practically killed all JCS cells via apoptosis and cleaved the DNA to extremely small 150 base-pair fragments. In contrast, among the JCS cells killed by MC540, about 88% died via apoptosis and large DNA fragments were abundant. Relative to MC540, the ability of mTHPC to trigger large-scale and thorough apoptosis in leukemia cells may help explain its potency and selectivity.     

Intracellular signaling of osteogenic protein-1 through Smad5 activation

This research examines job satisfaction and need for autonomy of 190 registered nurses in Hong Kong using a cross-sectional survey design. The level of job satisfaction towards six job components (autonomy, professional status, pay, interaction, task requirements and organizational policies) was measured using the Index of Work Satisfaction. Results showed that the sample was dissatisfied more than satisfied, they valued the job components of autonomy, professional status and pay more than interaction, task requirements and organizational policies. In addition, comparisons were made between nurses working in different hospitals and also different nursing units within a hospital. The level of need for autonomy was assessed using the autonomy subscale of the Edwards Personal Preference Schedule. Results showed that the level of need for autonomy of this group of nurses was below the mid-score of the sub-scale and there was no significant relationship between their satisfaction with job autonomy and their individual need for autonomy.     

Identification of genes differentially expressed in Mycobacterium tuberculosis by differential display PCR

RNA arbitrarily-primed differential display PCR (RAP-PCR) was used to identify and isolate genes differentially expressed between attenuated (H37Ra) and virulent (H37Rv, Erdman) laboratory strains of Mycobacterium tuberculosis (Mtb). Using this method, cDNA fragments showing homology to three known mycobacterial genes and six putative novel genes in mycobacterial cosmid vectors were identified. Among the putative novel Mtb genes identified, we found: (1) gene MTV041.29, containing multiple tandem repetitive sequences and encoding a putative Gly-, Ala, Asn-rich protein (PPE family); (2) gene MTV004.03, containing the AT10S repetitive gene sequence; (3) gene MTV028.09, encoding a hypothetical protein of unknown function; (4) genes MTCY78.20,21, possibly encoding two hypothetical proteins of unknown function; (5) gene MTCY01A6.09, encoding a putative novel ferrodoxin dependent glutamate synthase; and (6) gene MTCY31.20, encoding a putative cyclohexanone monooxygenase. Using gene specific primers in a second differential display PCR and by RT-PCR amplification, novel genes 1, 2, 3 and 4 were shown to be differentially up-regulated in the attenuated Mtb strain H37Ra compared to H37Rv and Erdman strain. Overall, we demonstrated that RAP-PCR, as a first step, is a quick and sensitive method for the identification and isolation of novel genes expressed in Mtb. Because of limitations inherent to the lack of specificity of arbitrary primers in the RAP-PCR method, a second differential display PCR and RT-PCR amplification with gene-specific primers was necessary in order to confirm differential expression of the identified genes.     

Induction of reparative dentine formation in monkeys by recombinant human osteogenic protein-1

Osteogenic protein-1 (OP-1, BMP-7), a member of the transforming growth factor-beta supergene family, induces cartilage and bone formation when implanted in intra- and extraskeletal sites in vivo. The human OP-1 gene has been cloned and biologically active recombinant OP-1 homodimers (hOP-1) produced. The amount of bone induced by hOP-1 in vivo is related to the amount of protein implanted. Dentine possesses bone morphogenetic protein (BMP) activity. Impure material from allogenic bone with BMP activity induced reparative dentine formation in dogs. The objective of this study was to determine if the amount of reparative dentine stimulated by hOP-1 is related to the amount of protein utilized in direct pulp-capping experiments. Freshly exposed molar and premolar pulps were treated with varying amounts of a complex comprising hOP-1 and a carrier matrix of purified bovine type-1 collagen powder (CM) moistened with sterile saline. Reparative dentine was present in all hOP-1/CM treated teeth (12 of 15) that remained sealed for the 6 weeks' healing. Substantially more new dentine was present in teeth treated with hOP-1/CM than in those treated with Ca(OH)2 paste and the amount of reparative dentine formed was proportional to the amount of hOP-1/CM (P < 0.05). No reparative dentine formed in collagen carrier or untreated teeth. The appearances of the new tissue suggested that much of the mass of the hOP-1/CM was replaced first by a pulp-like connective tissue, which mineralized to form reparative dentine.(ABSTRACT TRUNCATED AT 250 WORDS)     

Isolation of developmentally regulated genes by differential display screening of cDNA libraries

mRNA differential display RT-PCR has been extensively used for the isolation of genes differentially expressed between RNA populations. We have assessed its utility for the identification of developmentally regulated genes in plasmid cDNA libraries derived from individual tissues dissected from early mouse embryos. Using plasmid Southern blot hybridisation as a secondary screen, we are able to identify such genes and show by whole-mount in situ hybridisation that their expression pattern is that expected from the differential display profile.     

Inhibition of ultraviolet B-induced activator protein-1 (AP-1) activity by aspirin in AP-1-luciferase transgenic mice

Aspirin is under consideration as a promising chemopreventative agent for human cancers. To study the usefulness of aspirin as a chemopreventative agent for UV-induced human skin cancer, we investigated the effect of aspirin on UVB-induced activator protein-1 (AP-1) activity. In the JB6 cell culture system, aspirin or sodium salicylate (SA) inhibited UVB-induced AP-1 activity in a dose-dependent manner; this inhibitory effect occurred only in cells pretreated with aspirin or SA before UVB irradiation but not cells treated with aspirin or SA after UVB irradiation. Furthermore, these inhibitory effects on UVB-induced AP-1 activity appeared to be mediated through blocking of activation of MAP kinase family members, including extracellular signal-regulated protein kinases, c-Jun N-terminal kinases, and p38. It was not due to absorption of UVB light by aspirin. In the skin of AP-1-luciferase transgenic mice, UVB irradiation induced a rapid increase in AP-1 activity, which reached the peak at 48 h post-UVB irradiation. The topical pretreatment of mouse skin with aspirin markedly blocked the UVB-induced AP-1 transactivation in vivo. These data provide the first evidence that aspirin and SA are inhibitors of UV-induced signal transduction and thus could be used as a chemopreventative agent for skin cancer.     

Acute and relapsing experimental autoimmune encephalomyelitis are regulated by differential expression of the CC chemokines macrophage inflammatory protein-1alpha and monocyte chemotactic protein-1

Myasthenia gravis (MG) patients develop autoantibodies primarily against the acetylcholine receptor in the motor endplate, but also against intracellular striated muscle proteins, notably titin, the giant elastic protein of the myofibrillar cytoskeleton. Titin antibodies have previously been shown to be directed against a single epitope on the molecule, located at the A-band/I-band junction and referred to as the main immunogenic region (MIR) of titin. By using immunofluorescence microscopy on stretched single myofibrils, we now report that approximately 40% of the sera from 18 MG/thymoma patients and 8 late-onset MG patients with thymus atrophy contain antibodies that bind to a more central I-band titin region. This region consists of homologous immunoglobulin domains and is known to be differentially spliced dependent on muscle type. All patients with I-band titin antibodies also had antibodies against the MIR. Although a statistically significant correlation between the occurrence of I-band titin antibodies and MG severity was not apparent, the results could hint at an initial immunoreactivity to titin's MIR, followed by reactivity along the titin molecule in the course of the disease.     

Applicability of nonsyngeneic cell models for screening of genes in monogenetic diseases via differential display technique

Conventional subtraction library techniques or DNA-transfection studies are standard techniques applied for identification and isolation of genes relevant in monogenetic diseases like Fanconi anemia (FA). The differential display technique (DDT) was developed to compare mRNA expression between a mutant cell line and its syngeneic control and allows comparison of almost all mRNA species within a short time. However, for identification of genes relevant in monogenetic diseases, no syngeneic cell model is available. In this report, we show that the use of nonsyngeneic diploid human fibroblasts does not increase the number of differentially displayed bands due to diversity of untranslated regions. cDNA bands with a length of up to 1000 bp were obtained and applied to DDT. After screening of about 13000 cDNA bands, only 0.5% were found to be differentially expressed between FA and control cells. Finally, three mRNAs were cloned and verified in Northern blot experiments to be differentially expressed in FA fibroblasts. The low number of differentially displayed cDNA bands in DDT indicates the usefulness of this statistical, molecular approach for identification of multiple genes dysregulated in gene regulation cascades potentially relevant for cell cycle disturbances.     

Isolation and characterization of immune-related genes from the fall webworm, Hyphantria cunea, using PCR-based differential display and subtractive cloning

Following injection of bacteria into the hemocoel of the fall webworm, Hyphantria cunea, several inducible genes were identified and characterized using PCR-based differential display (DD-PCR) and subtractive cloning. Ten immune-related cDNA clones (Hdd1, Hdd2, Hdd3, Hdd11, Hdd13, Hdd15, Hdd17, Hdd23, Hs106, Hs302) were isolated and characterized. The deduced amino acid sequence of Hdd2 was shown to be a member of the copper, zinc superoxide dismutase (Cu-Zn SOD) family. The H. cunea Cu-Zn SOD is novel in that it is up-regulated following a bacterial challenge and has a putative signal peptide suggesting its secretion and involvement in the insect immune response. Hdd3 was found to encode a new member of the serpin (serine protease inhibitor) family. The putative lectin corresponding to Hdd15 is of a different kind in that it has two lectin C domains in a single molecule. These two lectin C domains show significant homology to the lectin C domain of Periplaneta lipopolysaccharide binding protein (LPS-BP). Three cloned genes, Hdd17, Hs106 and Hs302, encode a homologue to Bombyx mori Gram negative binding protein, a hemolin-like protein and a attacin-like protein, respectively. The deduced amino acid sequences from Hdd11 showed weak homology with a Locusta migratoria hemolymph protein. On the contrary, Hdd1, Hdd13 and Hdd23 did not reveal any significant homology with known proteins. All of the 10 genes were clearly inducible by E. coli and M. luteus injection. Injection of distilled water only slightly induced mRNA levels. Comparison of temporal mRNA expression following E. coli injection showed three types of expression patterns.     

Identification of new early auxin markers in tobacco by mRNA differential display

Cerebrovascular disease (CVD) is associated with high fibrinogen levels and lipid fractions leading to an increase of both plasma and whole blood viscosity as well as raised aggregability of blood cells. One important goal in the treatment of cerebral multiinfarct dementia (MID) therefore should be to reduce fibrinogen and lipoproteins and thereby to improve the haemorheological state. The effect of heparin-induced extracorporeal LDL precipitation (H.E.L.P.), a method for safe and immediate reduction of parameters relevant to haemorheology, such as plasma fibrinogen and the lipoproteins, was investigated in 98 patients with MID. All the patients underwent two H.E.L.P. applications within 8 days. The impact of H.E.L.P. on CVD was studied by changes of laboratory data and by evaluation of clinical symptoms before and after treatment. Each H.E.L.P. session caused an immediate, safe and significant reduction of important rheological parameters such as fibrinogen (P < 0.001), whole blood viscosity at high and low shear rate, plasma viscosity and red cell transit time (P < 0.01 each). Also total cholesterol and low density lipoprotein (P < 0.0001 each), lipoprotein (a) (P < 0.003) and the triglycerides (P < 0.0001) had been reduced. The results in laboratory measurement were followed by a statistically significant improved neurologic recovery, represented in the values of the Mathew Scale, the Mini Mental State Examination and the Activities-of-Daily-Living-Test. These results can indicate the importance and influence of haemorheology on clinical symptoms in CVD.     

Effects of resorbable membrane placement and human osteogenic protein-1 on hard tissue healing after periradicular surgery in cats

Periradicular surgeries were performed on the maxillary cuspid teeth of twelve cats. Before reapproximation of the surgical flaps, eight of the osteotomies were covered with a resorbable membrane and eight were filled with human osteogenic protein-1 (hOP-1) on a collagen carrier. The remaining eight sites received no further treatment and served as controls. The animals were euthanized after 12 wk, and the specimens were examined histomorphometrically for the presence or absence of osseous regeneration, inflammation, and cementum formation on the root ends. The results showed that the sites treated with the membrane exhibited significantly more inflammation adjacent to the resected root ends (p < 0.05), and that the use of the membrane had no statistically significant effect on osseous healing or new cementum formation. The use of hOP-1 was associated with a significant decrease in the thickness of new cementum formed on the resected root ends (p < 0.05), but had no statistically significant effect on osseous healing or degree of inflammation. Based on these results, it seems that neither the use of hOP-1 nor resorbable membranes have a positive effect on periradicular tissue healing in endodontic surgery.     

Lipopolysaccharide and its analog antagonists display differential serum factor dependencies for induction of cytokine genes in murine macrophages

Monocytes/macrophages play a central role in mediating the effects of lipopolysaccharide (LPS) derived from gram-negative bacteria by the production of proinflammatory mediators. Recently, it was shown that the expression of cytokine genes for tumor necrosis factor alpha (TNF-alpha), interleukin-1beta (IL-1beta), and interferon-inducible protein-10 (IP-10) by murine macrophages in response to low concentrations of LPS is entirely CD14 dependent. In this report, we show that murine macrophages respond to low concentrations of LPS (相似文献   

Analysis of gene expression in lung and thymus of TCDD treated C57BL/6 mice using differential display RT-PCR

The differential display RT-PCR technique offers a means to identify genes which are regulated by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Treatment of female C57BL/6 mice with 10 micrograms TCDD/kg body weight followed by DDTR-PCR analysis with a set of different primers revealed differential expression for a number of mRNAs. RNA slot blot analysis of six of these clones confirmed differential expression. Further characterization of one of these clones obtained from lung tissue demonstrated that the corresponding mRNA is about 1 kb and shows significant TCDD induced expression in Northern blot analysis. A Genbank search of this clone showed no similarity to known sequences.     

Analysis of urinary proteins by disc electrophoresis: a method for the differential diagnosis of kidney diseases (author's transl)

After introductory observations on the role of the kidney in protein metabolism, the methodology and results of disc electrophoresis on polyacrylamide, using detergents, are discussed. Separation of urinary proteins according to molecular weight permits the differentiation of kidney diseases into those of the glomerulus and of the tubules. Proteinurias of prerenal and postrenal origin are also differentiated. Owing to the considerable time involved, this methodology cannot at present be introduced for routine purposes.     

Identification of elicitor-induced cytochrome P450s of soybean (Glycine max L.) using differential display of mRNA

Magnesium (Mg) absorption across the intestinal epithelium is crucial for Mg homeostasis in all vertebrates. Besides paracellular transport, it involves a cell-mediated component, and this predicts the presence of specific Mg carriers at both the apical and basolateral pole of the epithelial cells. Although the mechanism of transmembrane Mg transport in enterocytes, as in most cell types, has remained an elusive topic for many years, recent studies have provided promising new insights. We here recapitulate the progress that has been made in this field, and advance evidence for membrane carriers that are involved in transcellular Mg transport in the intestine.