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The mouse t-complex gene, Tcp-11, is under translational control.

The mouse t-complex is known to harbour genes which affect male fertility. Tcp-11 is a t-complex gene which is only expressed in male germ cells and from its position is a candidate for a distorter, one of the two types of genetic element involved in transmission ratio distortion. Antibodies raised to TCP-11 protein made in E. Coli were used on thin sections of testis and shown to recognise late spermatids. On Western blots the antibodies bound to a 68-kD protein present in protein extracts from testis. No specific signal could be detected using the antibody on protein extracts from other mouse tissues. Following gentle lysis of the germ cells and fractionation on sucrose gradients, all the material recognised by the anti-Tcp-11 antibody was found to be soluble and unassociated with any membrane fraction or organelle. A comparison of the time course of expression of the Tcp-11 mRNA and the TCP-11 protein revealed that expression of this gene is under translational control.

Animals↗

Mechanism of bone incorporation of beta-TCP bone substitute in open wedge tibial osteotomy in patients.

A histological study was performed of bone biopsies from 16 patients (17 biopsies) treated with open wedge high tibial osteotomies for medial knee osteoarthritis. The open wedge osteotomies were filled with a wedge of osteoconductive beta tricalcium phosphate (beta-TCP) ceramic bone replacement. At the time of removal of the fixation material, core biopsies of the area where the beta-TCP was located were taken at different follow-up periods (6-25 months). beta-TCP resorption, bone ingrowth and bone remodelling were studied. We hypothesized that the incorporation and remodelling process occurs similarly as in animals. Histology showed a good resorption of the beta-TCP with complete incorporation and remodelling into new bone. The different phases as described in animal studies were found. A correlation was found between histological findings and radiological assessment. In conclusion, beta-TCP appeared to be a bone replacement material with optimal biocompatibility, resorption characteristics and bone conduction properties for the clinical use.

Adult↗

Aerobic degradation of 2,4,6-TCP content in ECF bleached effluent.

Elemental chlorine-free (ECF) bleach effluents from kraft mill are characterised by: a chemical organic demand/biological organic demand (COD/BOD(5)) ratio of 4, chlorophenol content with low chlorine substitution, and toxicity. The effect of increasing the concentration of 2,4,6-trichlorophenol (2,4,6-TCP) content in ECF bleaching sequence effluent on the degradative activity of bacterial communities present in an aerobic system treatment was studied. An aerobic lagoon (AL) was used as a typical secondary treatment of kraft-mill effluent. AL displays a high performance of BOD(5) degradation (up to 90%); however, only 40% of the COD was removed. Simultaneously, the AL system shows a high ability to biodegrade 2,4,6-TCP up to 237 mg/l day. Kinetic parameters of the 2,4,6-TCP biodegradation by aerobic bacteria were determined. The K(s) and K(i) values were 34.3 and 50 mg/l 2,4,6-TCP, respectively. Moreover, the tolerance of aerobic bacteria was observed up to 1.3 g/l 2,4,6-TCP.

Bacteria, Aerobic↗

The low affinity PCP sites in the rat cerebellum not only bind TCP-like but also BTCP-like structures.

Congeners of the potent dopamine (DA) re-uptake inhibitor 1-[1-(2-benzo[b]thiophenyl)cyclohexyl]piperidine (BTCP) are unexpectedly able to bind in the rat cerebellum, although this structure is devoid of dopaminergic nerve endings. In line with previous studies the hypothesis that they bind to low affinity PCP sites labelled with [3H]TCP in the rat cerebellum, even though they do not bind to the high affinity PCP sites in the forebrain, was considered. Analogues of 1-[1-(2-thiophenyl)cyclohexyl]piperidine (TCP) and BTCP with a modified aromatic moiety and with O or S atoms substituted in the cyclohexyl ring were prepared and tested in competition experiments both in rat forebrain and cerebellum membranes labelled with [3H]TCP, and in rat striatum membranes labelled with [3H]BTCP. Results indicated that BTCP and congeners could bind to low affinity PCP sites labelled with [3H]TCP in the rat cerebellum with a decrease of the selectivity for the DA transporter. On the contrary, some TCP analogues displayed a very high selectivity for these low affinity sites; they might be important pharmacological tools to elucidate the nature and function at yet unknown of these sites.

Animals↗

Regulated expression of a novel TCP domain transcription factor indicates an involvement in the control of meristem activation processes in Solanum tuberosum.

In this study, the aim was to determine whether TCP transcription factors are implicated in meristem activation in potato (Solanum tuberosum). By searching a database of potato EST sequences, with a sequence characteristically conserved in TCP domains, a potato tcp gene was identified. A BAC clone containing the tcp sequence was isolated and the genomic sequence was determined. Using a CAPS marker assay, the potato tcp gene (sttcp1) was mapped to chromosome 8. In dormant buds, relatively high levels of sttcp1-specific transcript were detected by in situ hybridization. By contrast, in sprouting buds, no expression of the sttcp1 could be detected. Furthermore, an inverse relationship between axillary bud size and the steady-state level of the sstcp1 transcript was demonstrated. In non-growing buds exhibiting correlative inhibition, sttcpI-specific transcript levels were also relatively high, but rapidly decreased when apical dominance was removed by excision of the apical bud.

Amino Acid Sequence↗

Diversity and evolution of CYCLOIDEA-like TCP genes in relation to flower development in Papaveraceae.

Monosymmetry evolved several times independently during flower evolution. In snapdragon (Antirrhinum majus), a key gene for monosymmetry is CYCLOIDEA (CYC), which belongs to the class II TCP gene family encoding transcriptional activators. We address the questions of the evolutionary history of this gene family and of possible recruitment of genes homologous to CYC in floral development and symmetry in the Papaveraceae. Two to three members of the class II TCP family were found in each species analyzed, two of which were CYC-like genes, on the basis of the presence of both the TCP and R conserved domains. The duplication that gave rise to these two paralogous lineages (named PAPACYL1 and PAPACYL2) probably predates the divergence of the two main clades within the Papaveraceae. Phylogenetic relationships among angiosperm class II TCP genes indicated that (1) PAPACYL genes were closest to Arabidopsis (Arabidopsis thaliana) AtTCP18, and a duplication at the base of the core eudicot would have given rise to two supplementary CYC-like lineages; and (2) at least three class II TCP genes were present in the ancestor of monocots and eudicots. Semiquantitative reverse transcription-polymerase chain reaction and in situ hybridization approaches in three species with different floral symmetry indicated that both PAPACYL paralogs were expressed during floral development. A pattern common to all three species was observed at organ junctions in inflorescences and flowers. Expression in the outer petals was specifically observed in the two species with nonactinomorphic flowers. Hypotheses concerning the ancestral pattern of expression and function of CYC-like genes and their possible role in floral development of Papaveraceae species leading to bisymmetric buds are discussed.

DNA-Binding Proteins↗

A TCP-NTCP estimation module using DVHs and known radiobiological models and parameter sets.

Radiotherapy treatment plan evaluation relies on an implicit estimation of the tumor control probability (TCP) and normal tissue complication probability (NTCP) arising from a given dose distribution. A potential application of radiobiological modeling to radiotherapy is the ranking of treatment plans via a more explicit determination of TCP and NTCP values. Although the limited predictive capabilities of current radiobiological models prevent their use as a primary evaluative tool, radiobiological modeling predictions may still be a valuable complement to clinical experience. A convenient computational module has been developed for estimating the TCP and the NTCP arising from a dose distribution calculated by a treatment planning system, and characterized by differential (frequency) dose-volume histograms (DDVHs). The radiobiological models included in the module are sigmoidal dose response and Critical Volume NTCP models, a Poisson TCP model, and a TCP model incorporating radiobiological parameters describing linear-quadratic cell kill and repopulation. A number of sets of parameter values for the different models have been gathered in databases. The estimated parameters characterize the radiation response of several different normal tissues and tumor types. The system also allows input and storage of parameters by the user, which is particularly useful because of the rapidly increasing number of parameter estimates available in the literature. Potential applications of the system include the following: comparing radiobiological predictions of outcome for different treatment plans or types of treatment; comparing the number of observed outcomes for a cohort of patient DVHs to the predicted number of outcomes based on different models/parameter sets; and testing of the sensitivity of model predictions to uncertainties in the parameter values. The module thus helps to amalgamate and make more accessible current radiobiological modeling knowledge, and may serve as a useful aid in the prospective and retrospective analysis of radiotherapy treatment plans.

Body Burden↗

The physiological response associated with large particles of TCP, HA, and ALCAP implants using raw 264.7 cells.

Ceramic materials were introduced as implantable materials in the early 1960s. Currently, there are numerous types of ceramic materials under evaluation as ideal implant devices. The inflammatory responses generated by various calcium phosphate-based ceramics implants have not been clearly evaluated. The current investigation evaluates the response of RAW 264.7 macrophage cells to large particle size (> 38 microns) of hydroxyapatite (HA), tricalcium phosphate (TCP), and aluminum calcium phosphate (ALCAP). RAW 264.7 cells were cultured in a 24 well plate at a density of 1.5 x 10 cells per well. The plates were divided into seven equal groups (n = 6) (control, HA, TCP, ALCAP, ops HA, ops TCP, ops ALCAP). The total protein, maliondialdehyde (MDA), nitric oxide, and cell counts were measured using established lab protocols at 24, 48 and 72 hours of incubation. Cells grown on coverslips were used to evaluate morphological features. The data obtained from this investigation suggested that the nonopsonized particles of ALCAP and TCP materials exhibited an increase in cell number at both the 48 and 72 hour phases. However, a decrease was seen in cell number in all three opsonized groups at 48 and 72 hour phases when compared to the control. As for MDA levels in the opsonized group, all treatments showed an initial increase. Nitric oxide production increased in TCP and ALCAP at the 48-hour phase. Morphological evaluation revealed that upon the exposure of the three different type of ceramic, the cells appeared to be biocompatible.

Aluminum Compounds↗

[Manufacture and study of porous poly(l-lactic acid) (PLLA)/beta-tricalcium phosphate (beta-TCP) composite].

A promising alternative to supply bone substitutes is to develop living tissue substitutes based on biodegradable materials, which is called bone tissue engineering. One of the research high-lights of bone tissue engineering is to design and manufacture scaffolds for cell attaching, migrating, and proliferating. A process which consists of a solvent casting stage, a compression molding stage and a leaching stage has been used to fabricate macroporous composites of poly(l-lactic acid) (PLLA) and beta-tricalcium phosphate (beta-TCP). The effects of the weight fraction of porogen--NaCl, of the weight ratio of PLLA to beta-TCP and of the diameters of beta-TCP on the porosities, the average pore diameters and the compressive yield strength and compressive modulus have been studied. The results showed that the porosities and the average pore diameters increased and the compressive yield strength and modulus decreased when the weight fraction went from 50% to 90%. The compressive yield strength and compressive modulus could be improved by changing the weight ratio of PLLA to beta-TCP and the diameters of beta-TCP in low-porosity composites (lower than 70%). But high-porosity composites (90%) were not reinforced by changing the weight ratio.

Biocompatible Materials↗

Preparation and assessment of heterotopic osteoinduction of beta-TCP/rhBMP-2 composite.

OBJECTIVE: To search for ideal bone graft substitute. METHODS: The beta TCP/rhBMP-2 composite was constructed by combining beta-Tricalcium phosphat (beta-TCP) that was prepared by the authors with recombinant human morphogenetic protein-2 (rhBMP-2) and was implanted into the muscle pouches in the thigh of mice. beta-TCP alone was implanted on the opposite side as controls. At intervals of 1,3,7,14 and 28 days after the implantation, the specimens were obtained, and histologic study and alkaline phosphatase assay (7,14,28 days) were performed. RESULTS: There was a large amount of cartilage and bone formation within the composite, increasing with time; whereas there was no new bone formation where beta-TCP alone was implanted. Besides, the levels of alkaline phosphatase in the beta-TCP/rhBMP-2 implants also were increasing with time and were higher than those in controls. CONCLUSIONS: The results indicate that beta-TCP/rhBMP-2 composite possesses heterotopic osteoinductive potential.

Journal Article↗

Morphometric analysis of the hormonal effect on tissue-implant response associated with TCP bioceramic implants.

Recent studies in our laboratory have demonstrated the need for an experiment comparing the effect of steroid hormones on the cellular composition of the fibrous tissue capsules surrounding tricalcium phosphate (TCP) bioceramics. The effect of steroid hormones on inflammatory cells has been widely reported in the scientific literature. The purpose of this research is to contribute to the development of a long-term ceramic drug delivery system with human and veterinary applications. This study contributes information on the composition of fibrous tissue capsules surrounding hormone loaded TCP bioceramic implants at 90 days post-implantation. The data obtained in this study was gathered using protocols developed from ImagePro software. Sixteen animals in four experimental groups were implanted with one TCP bioceramic each. Group I animals were implanted with a sham TCP ceramic not containing a steroid hormone (control group). Group II animals received the testosterone loaded TCP ceramic. Group III animals were implanted with the dihydrotestosterone loaded ceramic. Group IV animals received the androstenedione ceramic. At 90 days post-implantation, the animals were euthanized. The implants and fibrous tissue capsules surrounding them were then extracted. The fibrous tissue capsules were evaluated microscopically using ImagePro software following routine II&E staining, modified Papanicolau, and Masson's trichrome. The research variables comparing the hormonal effects on fibrous tissue composition obtained by digital analysis were as follows: thickness of the fibrous tissue, quantification of macrophages, neutrophils, fibroblasts, vascularity, and area occupied by collagen. The results of this study support several conclusions. All three of the hormones in this study, particularly androstenedione, have significant affects on the thickness and cellular composition of the fibrous tissue capsules when compared to the control group. Digital analysis software enables more accurate and reproducible results to be obtained when specific procedures are followed.

Androgens↗

Immunohistochemical detection of cytokine expression in tissue-implant response associated with TCP bioceramic implants loaded with steroid hormones.

Recent studies in our laboratory have demonstrated the need for an experiment comparing cytokine expression in fibrous tissue capsules surrounding tricalcium phosphate (TCP) bioceramics loaded with androstenedione, dihydrotestosterone, and testosterone to that of a control to further elucidate the mechanisms involved in the tissue-implant response. The effect of steroid hormones on inflammatory cells has been widely reported in the scientific literature. The purpose of this research is to contribute to the development of a long-term ceramic drug delivery system with human and veterinary applications. This study contributes information on the presence of cytokines in fibrous tissue capsules surrounding hormone loaded TCP bioceramic implants at 90 days post-implantation. Sixteen animals in four experimental groups were implanted with one TCP bioceramic each. Group I animals were implanted with a sham TCP ceramic not containing a steroid hormone (control group). Group II animals received the testosterone loaded TCP ceramic. Group III animals were implanted with the dihydrotestosterone loaded ceramic. Group IV animals received the androstenedione ceramic. At 90 days post-implantation, the animals were euthanized. The implants and fibrous tissue capsules surrounding them were then extracted. The fibrous tissue capsules were evaluated microscopically following routine H&E staining and IHC staining of antibodies to IL-1 beta, Il-2, IL-6, TNF infinity, and cell specific markers for CD-4 and CD14 positive cells. The results of this study indicate that these hormones, particularly androstenedione, limit the expression of cytokines and greatly affect the cellular composition of the tissue-implant response.

Androgens↗

Beta-tricalcium phosphate (beta-TCP) graft combined with bone marrow stromal cells (MSCs) for posterolateral spine fusion.

Macaque lumber posterolateral spine fusion (PLF) was performed by using beta-TCP graft combined with bone marrow derived stromal cells (MSCs), to evaluate whether a beta-TCP/MSCs hybrid can be used for PLF instead of autogenous bone graft. Nine crab-eating macaque underwent bilateral PLF at L4-L5. The implants were divided into three groups: 1) beta-TCP/MSCs hybrid, 2) autogenous bone, and 3) beta-TCP. Six monkeys were sacrified at 12 weeks and three monkeys were sacrificed at 24 weeks after implantation. Manual palpation, radiography, micro computed tomography, peripheral quantitative computed tomography (pQCT), and histology were used to assess bone formation. Manual palpation and X-ray showed that 83.3% of hybrid groups and 66.7% of autogenous groups achieved solid spine fusion, whereas none of other groups fused. Histological analysis showed that all of the hybrid groups achieved massive bone formation. Bone mineral density (BMD) evaluated with pQCT in the hybrid groups increased by additional new bone. Beta-TCP/MSCs hybrid can be used for PLF instead of autogenous bone graft. Thus it can be hypothesized that the monkey PLF can simulate human PLF.

Animals↗

Gas chromatographic method for determination of chlorpyrifos and its metabolite 3,5,6-trichloro-2-pyridinol (TCP) in dates.

A method is described for the determination of the insecticide chlorpyrifos and its metabolite TCP in green, unprocessed, and processed dates with the seeds incorporated. After extraction, chloropyrifos is cleaned up using Florisil and analyzed using a gas chromatography (GC) equipped with a nitrogen/phosphorus detector. TCP is derivatized using bis-(trimethylsilyl)-acetamide (BSA) to form the TCP-derivative and analyzed by a gas chromatograph equipped with a Hall electrolytic conductivity detector. Recoveries of chlorpyrifos from all fortified dates (0.05 and 0.1 ppm) ranged from 86 to 110% with an average of 94.5%. Recoveries of TCP from all fortified dates (0.1 and 0.2 ppm) ranged from 79 to 99% with an average of 86%. Limits of detection for chlorpyrifos and TCP in green, unprocessed, and processed dates were 0.02 and 0.05 ppm, respectively.

Chlorpyrifos↗

Macrophages in degradation of collagen/hydroxylapatite(CHA), beta-tricalcium phosphate ceramics (TCP) artificial bone graft. An in vivo study.

The macrophages mediated biodegradation of two biomaterials, collagen/hydroxylapatite (CHA) and beta-tricalcium phosphate ceramics (TCP), was studied in 24 male Kunming mice and 20 male C57BL/6 mice with histopathologic, histochemical and ultrastructural observation. It was demonstrated that macrophages infiltrated after CHA, TCP were implanted. The macrophages could be differentiated from fibroblasts and the other infiltrated cells for special cellular profile and strong acid phosphatase activity. Morphologically, monocyte-macrophages and infused multinuclear giant cell degraded CHA and TCP by phagocytosis and extracellular resorption. The carbonic anhydrase activity of macrophages was demonstrated by histochemical technique. It suggested that macrophages secreted H+ and accomplished the decalcification of calcium phosphate compound of CHA and TCP. We conclude that macrophages are the main mediating cells which degraded CHA and TCP intracellularly and extracellularly.

Animals↗

Use of alpha-tricalcium phosphate (TCP) as powders and as an aqueous dispersion to modify processing, microstructure, and mechanical properties of polymethylmethacrylate (PMMA) bone cements and to produce bone-substitute compounds.

Addition of tricalcium phosphate (alpha-TCP) powders as an aqueous dispersion to a polymethylmethacrylate (PMMA) bone cement is shown to produce a class of composites that due to their microstructure and mechanical properties may be suitable for application as bone substitutes. The PMMA forms a solid cellular matrix with open cells about 100 micrometer in size and incorporating TCP clusters. The TCP aggregates inside the cells form a porous network, with average mesopore diameters of about 0.1 micrometer, that is accessible from the outer surface. If TCP is added to PMMA in the form of dried powders, the composites are not applicable as bone substitutes. The dynamic elastic modulus (DEM) and compressive and tensile strengths were measured and discussed for both classes of composites. The mechanical properties of the bone-substitute composites, although lower than the other class of composites, are still competitive with those properties of a porous ceramic matrix of hydroxyapatite and with those of natural bones.

Bone Cements↗

The expression of bone matrix protein mRNAs around beta-TCP particles implanted into bone.

Tissue response around beta-tricalcium phosphate (beta-TCP) particles (150-300 microm in diameter) implanted into rat tibiae was analyzed by in situ hybridization with digoxigenin-labeled procollagen alpha1(I) (COL), osteonectin, osteocalcin, and osteopontin (OPN) RNA probes. Specimens were collected at 3, 5, 7, and 10 days after the operation. Holes without implantation were used as control. In both the beta-TCP implanted and control groups, new bone was formed centripetally and all four kinds of mRNA were expressed in activated osteoblasts. A COL signal was expressed most strongly and widely, and was detected at the peripheral region of the hole at day 3. The other three mRNAs were also expressed in bone forming osteoblasts by day 7. However, in the earlier cell reaction stage, OPN expression in the beta-TCP implanted group was different than that in the control group: OPN mRNA was seen exclusively in the cells on the particles, and an OPN signal was detected not only in COL-positive cells, but also in COL-negative cells. The former cells may be osteoblasts and reflect the early process of bone formation on biomaterials. The latter cells may be macrophages and reflect foreign body reactions. Expression of these OPN mRNAs induced by implantation of beta-TCP may play a role in bone formation on the materials and in determining their biocompatibility.

Animals↗

Bone colonization of beta-TCP granules incorporated in brushite cements.

Injectable calcium phosphate hydraulic cements are known to have a high clinical potential in bone reconstruction for mini-invasive orthopaedic surgery, interventional radiology, and rheumatology. Previous in vivo experiments in rabbit have shown that the presence of beta-TCP granules in injectable bone cement help maintain the transient biomechanical function of the implanted bone and promote the formation of good-quality new bone. Histomorphometric analysis of two brushite hydraulic cement (BHC) mixtures selected from previous results (referred to in this work as BHC-A and BHC-B) was performed at three postoperative delays (0, 12, and 24 weeks): histomorphometric analysis of bone colonization within beta-TCP shows that, just before implantation, the beta-TCP granule area is significantly higher in BHC-B; the residual granule area decreases steadily over time in BHC-A, whereas it goes through a maximum of 30% at 12 weeks in BHC-B; the residual granule porosity increases steadily up to 35% in BHC-A, whereas it goes through a maximum of 35% at 12 weeks and decreases somewhat until 24 weeks in BHC-B. New bone formation within granules appears higher in BHC-A (58% Area) compared to BHC-B (38% area) at 12 weeks. At 24 weeks bone colonization levels off in both cements at about 50% area. Irrespective of the cement matrix composition, beta-TCP granules contribute actively to the conduction of new bone formation.

Animals↗