Expression of a Truncated, Kinase-Defective TGF-β Type II Receptor in Mouse Skeletal Tissue Promotes Terminal Chondrocyte Differentiation and Osteoarthritis

Members of the TGF-β superfamily are important regulators of skeletal development. TGF-βs signal through heteromeric type I and type II receptor serine/threonine kinases. When over-expressed, a cytoplasmically truncated type II receptor can compete with the endogenous receptors for complex formation, thereby acting as a dominant-negative mutant (DNIIR). To determine the role of TGF-βs in the development and maintenance of the skeleton, we have generated transgenic mice (MT-DNIIR-4 and -27) that express the DNIIR in skeletal tissue. DNIIR mRNA expression was localized to the periosteum/perichondrium, syno-vium, and articular cartilage. Lower levels of DNIIR mRNA were detected in growth plate cartilage. Transgenic mice frequently showed bifurcation of the xiphoid process and sternum. They also developed progressive skeletal degeneration, resulting by 4 to 8 mo of age in kyphoscoliosis and stiff and torqued joints. The histology of affected joints strongly resembled human osteo-arthritis. The articular surface was replaced by bone or hypertrophic cartilage as judged by the expression of type X collagen, a marker of hypertrophic cartilage normally absent from articular cartilage. The synovium was hyperplastic, and cartilaginous metaplasia was observed in the joint space.

We then tested the hypothesis that TGF-β is required for normal differentiation of cartilage in vivo. By 4 and 8 wk of age, the level of type X collagen was increased in growth plate cartilage of transgenic mice relative to wild-type controls. Less proteoglycan staining was detected in the growth plate and articular cartilage matrix of transgenic mice. Mice that express DNIIR in skeletal tissue also demonstrated increased Indian hedgehog (IHH) expression. IHH is a secreted protein that is expressed in chondrocytes that are committed to becoming hypertrophic. It is thought to be involved in a feedback loop that signals through the periosteum/ perichondrium to inhibit cartilage differentiation. The data suggest that TGF-β may be critical for multifaceted maintenance of synovial joints. Loss of responsiveness to TGF-β promotes chondrocyte terminal differentiation and results in development of degenerative joint disease resembling osteoarthritis in humans.

     

β-Amino acid residues: Conformational characterization of an N- and C-protected homo-β-(S)-leucine

Summary An N- and C-protected derivative ofhomo-β-leucine, Fmoc-homo-β-(S)-leucine methyl ester, synthesized from the corresponding proteinogenic parent α-amino acid in enantiopure form has been fully characterized in the solid state by X-ray diffraction analysis. The crystal conformation of this new residue indicates and extended conformation for thishomo-β-residue, with the ϕ torsion angle being more constrained than the μ and ψ angles.     

The Ecological Footprint Intensity of National Economies

At least three perspectives—industrial ecology (IE), ecological modernization theory (EMT), and the “environmental Kuznets curve” (EKC)—emphasize the potential for sustainability via refinements in production systems that dramatically reduce the environmental impacts of economic development. Can improvements in efficiency counterbalance environmental impacts stemming from the scale of production? To address this question we analyze cross‐national variation in the ecological footprint (EF) per unit of gross domestic product (GDP). The EF is a widely recognized indicator of human pressure on the environment. The EF of a nation is the amount of land area that would be required to produce the resources it consumes and to absorb the wastes it generates. The most striking finding of our analyses is that there is limited variation across nations in EF per unit of GDP. This indicates limited plasticity in the levels of EF intensity or eco‐efficiency among nations, particularly among affluent nations. EF intensity is lowest (ecoefficiency is highest) in affluent nations, but the level of efficiency in these nations does not appear to be of sufficient magnitude to compensate for their large productive capacities. These results suggest that modernization and economic development will be insufficient, in themselves, to bring about the ecological sustainability of societies.     

β-Elimination in aldonolactones. Synthesis of 3,6-dideoxy L-arabino-hexose (ascarylose)

Benzoylation of L-rhamnono-1,5-lactone (1) with an excess of benzoyl chloride and pyridine for 16 h afforded 2,4-O-benzoyl-3,6-dideoxy-L-erythro-hex-2-enono-1,5-lactone (2). Catalytic hydrogenation of 2 was stereoselective and gave crystalline 2,4-di-O-benzoyl-3,6-dideoxy-L-arabino-hexono-1,5-lactone (3). Reduction of the lactone 3 with disiamylborane afforded 2,4-di-O-benzoyl-3,6-dideoxy-L-arabino-hexopyranose (4) which, on debenzoylation, gave 3,6-dideoxy-L-arabino-hexose (ascarylose) (7) in good overall yield. The sugar was identified as the corresponding alditol (ascarylitol) and by convertion into methyl 3,6-dideoxy-α-L-arabino-hexopyranoside (methyl ascaryloside, 6).     

Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry for the Discrimination of Food-Borne Microorganisms

A methodology based on matrix-assisted laser desorption ionization-time of flight mass spectrometry of intact bacterial cells was used for rapid discrimination of 24 bacterial species, and detailed analyses to identify Escherichia coli O157:H7 were carried out. Highly specific mass spectrometric profiles of pathogenic and nonpathogenic bacteria that are well-known major food contaminants were obtained, uploaded in a specific database, and made available on the Web. In order to standardize the analytical protocol, several experimental, sample preparation, and mass spectrometry parameters that can affect the reproducibility and accuracy of data were evaluated. Our results confirm the conclusion that this strategy is a powerful tool for rapid and accurate identification of bacterial species and that mass spectrometric methodologies could play an essential role in polyphasic approaches to the identification of pathogenic bacteria.     

NUCLEAR FEATURES AND EFFECT OF NUTRIENTS ON GYMNODINIUM CATENATUM (DINOPHYCEAE) SEXUAL STAGES1

Gymnodinium catenatum Graham is an unarmored, cyst‐forming dinoflagellate species responsible for outbreaks of paralytic shellfish poisoning. The nuclear development of the cells during the sexual cycle and the effect of different nitrate and phosphate external levels on sexual stages were studied. Nuclear fusion of gametes occurred before or at the same time as cytoplasmic fusion. During this process, either both nuclei migrated to a central area in the sulcal region, or only one of them migrated to the other nucleus. The motile and longitudinally biflagellated zygote presented a large, pear‐shaped nucleus, and either divided or encysted. Planozygotes and germlings underwent similar division processes, which suggested an uncoordinated meiosis in both encysting and non‐encysting zygotes. Encystment in culture was greater under low nitrate and phosphate limitation (L/15) than when only one or neither of these nutrients were added (L‐N, L‐P, and ‐N‐P). However, planozygotes individually monitored achieved the maximum encystment (40%) in a medium with no phosphate or nitrate added (‐N‐P), while most of them divided (70%–90%) in replete (L1) or half‐replete (L‐N and L‐P) media. Low levels of nitrate in the medium of cyst formation promoted a deficient development of the cyst wall. On the other hand, low phosphate levels in the medium of germination prevented both planozygote and germling division and lowered the final germination frequencies of cysts. The minimum dormancy, with an average value of 13.7±5.5 days, was not affected by any of the nutritional conditions studied.     

MULTIPLE ROUTES OF SEXUALITY IN ALEXANDRIUM TAYLORI (DINOPHYCEAE) IN CULTURE1

Alexandrium taylori Balech is a cyst‐forming dinoflagellate species responsible for recurrent blooms in Mediterranean coastal waters. The nuclear development of the cells during the sexual cycle and the effect of different external nitrate and phosphate levels were studied. Nuclear fusion of gametes occurred 6–12 h after the complete cytoplasmic fusion. The U‐shaped nuclei fused through the end of one nucleus and the mid‐area of the other. The mobile and biflagellated zygote had a large, U‐shaped nucleus and may follow three different fates: direct division, short‐term encystment (ecdysal), and long‐term encystment (resting). Ecdysal cysts may divide in >24–96 h into two, four, six, or eight cells before germinating. Meiosis presumably occurred in three locations: in the planozygote, within the ecdysal cyst, and in the planomeiocyte (germling) liberated either from ecdysal or resting cysts. The effects of nutrients on these routes were studied in individually isolated sexual stages. (1) Direct divisions occurred mainly under replete conditions (L1), whereas no direct planozygote divisions were recorded in media with no phosphate added (L‐P). (2) Short‐term encystment was larger in media lacking phosphate (L‐P and L/30) than in medium with no nitrate added (L‐N) or under replete conditions (L1). (3) Long‐term encystment was only observed in medium with no nitrate added (L‐N). The long‐lived resting cyst, not previously described for this species, had a clear double wall, an irregular shape, a flat morphology, and a middle orange spot. No cysts germinated in 1–2 months, whereas 86% of the cysts germinated 2–3 months after being formed. A flow cytometry analysis showed that sexual induction and zygote formation were very fast and highly common processes, zygotes being nearly half of the population at days 3 and 5 after the induction of sexuality in the cultures.