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Quantitative Polymerase Chain Reaction Analysis (quantitative + polymerase_chain_reaction_analysis)
Selected AbstractsHow many symbionts are provided by mothers, acquired by offspring, and needed for successful vertical transmission in an obligate insect,bacterium mutualism?MOLECULAR ECOLOGY, Issue 24 2007TAKAHIRO HOSOKAWA Abstract Vertical symbiont transmission is among the most pivotal processes for maintenance of symbiotic associations. However, it is poorly understood whether and how the levels of resource allocation and investment upon vertical transmission are regulated. The stinkbug Megacopta punctatissima is obligatorily associated with the gut symbiotic bacterium ,Candidatus Ishikawaella capsulata', whose transmission is mediated by a unique mechanism called ,symbiont capsule'. We investigated the population dynamics of the symbiont during vertical transmission in the host,symbiont mutualism. The stinkbug mothers produced one capsule for around 3.6 eggs irrespective of clutch size, suggesting a strict maternal control over symbiont supply for the offspring. However, experimental manipulation of egg/capsule ratios revealed that one capsule is sufficient for symbiont transmission to six nymphs. Quantitative polymerase chain reaction analyses demonstrated that a capsule contains 1.2 × 108 symbionts, a newborn nymph possesses 2 × 107 symbionts from a capsule, and thus one capsule certainly contains a sufficient amount of symbiont cells for six nymphs. These results indicated that the stinkbug mothers produce 1.7 times more symbiont capsules than needed. The newborn nymphs consistently harboured around 2 × 107 symbionts, also suggesting a nymphal control over symbiont transmission. The threshold symbiont titre minimally needed for successful vertical transmission was estimated to be 1.9 × 106 symbionts, which is only 1/10 of the actual symbiont titre detected in a newborn nymph. These results illuminate several ecological factors that may be relevant to parental and offspring controls over symbiotic resource allocation through host insect generations. [source] Thrombopoietin, flt3-ligand and c-kit-ligand modulate HOX gene expression in expanding cord blood CD133+ cellsCELL PROLIFERATION, Issue 4 2004C. P. McGuckin Extrinsic modulators include growth factors and cell adhesion molecules, whereas intrinsic regulation is achieved with many transcription factor families, of which the HOX gene products are known to be important in haemopoiesis. Umbilical cord blood CD133+ HSPC proliferation potential was tested in liquid culture with ,TPOFLK' (thrombopoietin, flt-3 ligand and c-kit ligand, promoting HSPC survival and self-renewal), in comparison to ,K36EG' (c-kit-ligand, interleukins-3 and -6, erythropoietin and granulocyte colony-stimulating factor, inducing haemopoietic differentiation). TPOFLK induced a higher CD133+ HSPC proliferation (up to 60-fold more, at week 8) and maintained a higher frequency of the primitive colony-forming cells than K36EG. Quantitative polymerase chain reaction analysis revealed opposite expression patterns for specific HOX genes in expanding cord blood CD133+ HSPC. After 8 weeks in liquid culture, TPOFLK increased the expression of HOX B3, B4 and A9 (associated with uncommitted HSPC) and reduced the expression of HOX B8 and A10 (expressed in committed myeloid cells) when compared to K36EG. These results suggest that TPOFLK induces CD133+ HSPC proliferation, self-renewal and maintenance, up-regulation of HOX B3, B4 and A9 and down-regulation of HOX B8 and A10 gene expression. [source] Design and assessment of a tissue-engineered model of human phalanges and a small jointORTHODONTICS & CRANIOFACIAL RESEARCH, Issue 4 2005WJ Landis Structured Abstract Authors ,, Landis WJ, Jacquet R, Hillyer J, Lowder E, Yanke A, Siperko L, Asamura S, Kusuhara H, Enjo M, Chubinskaya S, Potter K, Isogai N. Objectives ,, To develop models of human phalanges and small joints by suturing different cell-polymer constructs that are then implanted in athymic (nude) mice. Design ,, Models consisted of bovine periosteum, cartilage, and/or tendon cells seeded onto biodegradable polymer scaffolds of either polyglycolic acid (PGA) or copolymers of PGA and poly-L-lactic acid (PLLA) or poly- , -caprolactone (PCL) and PLLA. Constructs were fabricated to produce a distal phalanx, middle phalanx, or distal interphalangeal joint. Setting and Sample Population ,, Studies of more than 250 harvested implants were conducted at the Northeastern Ohio Universities College of Medicine. Experimental Variable ,, Polymer scaffold, cell type, and implantation time were examined. Outcome Measure ,, Tissue-engineered specimens were characterized by histology, transmission electron microscopy, in situ hybridization, laser capture microdissection and qualitative and quantitative polymerase chain reaction analysis, magnetic resonance microscopy, and X-ray microtomography. Results ,, Over periods to 60 weeks of implantation, constructs developed through vascularity from host mice; formed new cartilage, bone, and/or tendon; expressed characteristic genes of bovine origin, including type I, II and X collagen, osteopontin, aggrecan, biglycan, and bone sialoprotein; secreted corresponding proteins; responded to applied mechanical stimuli; and maintained shapes of human phalanges with small joints. Conclusion ,, Results give insight into construct processes of tissue regeneration and development and suggest more complete tissue-engineered cartilage, bone, and tendon models. These should have significant future scientific and clinical applications in medicine, including their use in plastic surgery, orthopaedics, craniofacial reconstruction, and teratology. [source] Proinflammatory role of the Th17 cytokine interleukin-22 in collagen-induced arthritis in C57BL/6 miceARTHRITIS & RHEUMATISM, Issue 2 2009Lies Geboes Objective To investigate the role of interleukin-22 (IL-22) in collagen-induced arthritis (CIA), an animal model of rheumatoid arthritis. Methods C57BL/6 mice were immunized with type II collagen (CII) in Freund's incomplete adjuvant with added Mycobacterium tuberculosis, and levels of IL-22 and its specific receptor, IL-22 receptor type I (IL-22RI), were measured in sera and tissue by enzyme-linked immunosorbent assay and real-time quantitative polymerase chain reaction analysis. Clinical and histologic signs of arthritis were recorded and compared with those in C57BL/6 mice deficient in the IL-22 gene (IL-22,/,). Humoral and cellular immune responses against CII were analyzed. In vitro osteoclastogenesis assays were performed on splenocytes. Results Upon immunization with CII in Freund's incomplete adjuvant plus heat-killed Mycobacterium tuberculosis, sera from C57BL/6 mice were found to contain high levels of IL-22, and the specific IL-22RI was expressed in lymphoid tissue, including splenocytes. IL-22,/, mice were less susceptible to CIA than were wild-type mice, as evidenced by their decreased incidence of arthritis and decreased pannus formation. Remarkably, the less severe form of arthritis in IL-22,/, mice was associated with increased production of CII-specific and total IgG antibodies, whereas cellular CII responses were unchanged. In vitro, IL-22 was found to promote osteoclastogenesis, a process that might contribute to its proinflammatory activity in CIA. Conclusion Endogenous IL-22 plays a proinflammatory role in CIA in C57BL/6 mice. Our data also indicate that IL-22 promotes osteoclastogenesis and regulates antibody production. [source] | |||||||||||||