Towards an integrative, eco-evolutionary understanding of ecological novelty: studying and communicating interlinked effects of global change

Global change has complex eco-evolutionary consequences for organisms and ecosystems, but related concepts (e.g., novel ecosystems) do not cover their full range. Here we propose an umbrella concept of “ecological novelty” comprising (1) a site-specific and (2) an organism-centered, eco-evolutionary perspective. Under this umbrella, complementary options for studying and communicating effects of global change on organisms, ecosystems, and landscapes can be included in a toolbox. This allows researchers to address ecological novelty from different perspectives, e.g., by defining it based on (a) categorical or continuous measures, (b) reference conditions related to sites or organisms, and (c) types of human activities. We suggest striving for a descriptive, non-normative usage of the term “ecological novelty” in science. Normative evaluations and decisions about conservation policies or management are important, but require additional societal processes and engagement with multiple stakeholders

Deficiency of Thrombospondin-4 in Mice Does Not Affect Skeletal Growth or Bone Mass Acquisition, but Causes a Transient Reduction of Articular Cartilage Thickness

<div><p>Although articular cartilage degeneration represents a major public health problem, the underlying molecular mechanisms are still poorly characterized. We have previously utilized genome-wide expression analysis to identify specific markers of porcine articular cartilage, one of them being Thrombospondin-4 (Thbs4). In the present study we analyzed <i>Thbs4</i> expression in mice, thereby confirming its predominant expression in articular cartilage, but also identifying expression in other tissues, including bone. To study the role of Thbs4 in skeletal development and integrity we took advantage of a <i>Thbs4</i>-deficient mouse model that was analyzed by undecalcified bone histology. We found that <i>Thbs4</i>-deficient mice do not display phenotypic differences towards wildtype littermates in terms of skeletal growth or bone mass acquisition. Since <i>Thbs4</i> has previously been found over-expressed in bones of Phex-deficient <i>Hyp</i> mice, we additionally generated <i>Thbs4</i>-deficient <i>Hyp</i> mice, but failed to detect phenotypic differences towards <i>Hyp</i> littermates. With respect to articular cartilage we found that <i>Thbs4</i>-deficient mice display transient thinning of articular cartilage, suggesting a protective role of Thbs4 for joint integrity. Gene expression analysis using porcine primary cells revealed that Thbs4 is not expressed by synovial fibroblasts and that it represents the only member of the Thbs gene family with specific expression in articular, but not in growth plate chondrocytes. In an attempt to identify specific molecular effects of Thbs4 we treated porcine articular chondrocytes with human THBS4 in the absence or presence of conditioned medium from porcine synovial fibroblasts. Here we did not observe a significant influence of THBS4 on proliferation, metabolic activity, apoptosis or gene expression, suggesting that it does not act as a signaling molecule. Taken together, our data demonstrate that Thbs4 is highly expressed in articular chondrocytes, where its presence in the extracellular matrix is required for articular cartilage integrity.</p></div

Intact skeletal growth in <i>Thbs4</i>-deficient mice.

<p>(A) Xray analysis demonstrates absence of gross skeletal abnormalities in 6 weeks old <i>Thbs4</i>-deficient mice. (B) Length of lumbar spine and femur in 6 and 26 weeks old wildtype (WT) and <i>Thbs4</i>-deficient (KO) mice. (C) Toluidine blue staining of the tibia growth plates from 6 weeks old wildtype (+/+) and <i>Thbs4</i>-deficient (-/-) mice. (D) Quantification of the tibial growth plate width (GPW) in wildtype (WT) and <i>Thbs4</i>-deficient (KO) mice at 6 and 26 weeks of age. (E) Quantification of the lengths of the proliferative zone (PZ) and hypertrophic zone (HZ) in the same sections. All bars represent mean ± SD (n = 6 per group).</p

Chondrogenesis is unaffected in articular cartilage in <i>Thbs4</i>-deficient mice.

<p>(A) Quantification of the chondrocyte number per cartilage area in femur and tibia sections from 6, 26 and 52 weeks wildtype and <i>Thbs4</i>-deficient mice. Bars represent mean ± SD (n = 6 per group). (B) Representative images showing TUNEL-positive cells in articular cartilage from the femora of 26 weeks old wildtype and <i>Thbs4</i>-deficient mice. The percentage of TUNEL-positive cells is givenon the right. Values represent mean ± SD (n = 6 per group). (C) qRT-PCR monitioring expression of the indicated genes in articular cartilage of 26 weeks wildtype <i>Thbs4</i>-deficient mice. Bars represent mean ± SD (n = 4). Asterisks indicate statistically significant differences between WT and KO (p<0.05).</p

Porcine <i>THBS4</i> is specifically expressed in articular chondrocytes.

<p>Shown are the results of qRT-PCR expression analyses for all members of the <i>THBS</i> family, as well as markers for synovial fibroblasts (<i>FN1</i>), chondrocytes (<i>ACAN</i>, <i>COL2A1</i>) and hypertrophic chondrocytes (<i>COL10A1</i>). Primary cells (AC, articular chondrocytes; GP, growth plate chondrocytes; SF, synovial fibroblasts) were derived from 6 weeks old minipigs. Shown is the relative expression (after normalization to <i>GAPDH</i>) towards the cell type displaying the highest expression level. All bars represent mean ± SD (n = 3).</p

No impact of <i>Thbs4</i>-deficiency on bone mass or matrix mineralization on a wildtype or <i>Hyp</i> genetic background.

<p>(A) Von Kossa/van Gieson staining of spine sections from 6 and 26 weeks old wildtype (+/+) and <i>Thbs4</i>-deficient (-/-) mice. (B) Quantification of the trabecular bone volume per tissue volume (BV/TV) and trabecular thickness (Tb.Th.) in wildtype (WT) and <i>Thbs4</i>-deficient (KO) mice at both ages. Bars represent mean ± SD (n = 6 per group). (C) Xray analysis (top panels) and von Kossa/van Gieson staining of tibia (middle panels) or spine sections (bottom panels) from 6 weeks old Hyp mice with (+/+) or without (-/-) <i>Thbs4</i>. (D) Quantification of the growth plate width (GPW) in the tibia (top) or the osteoid volume per bone volume (OV/BV) from mice of the indicated genotypes. Bars represent mean ± SD (n = 5 per group). Asterisks indicate statistically significant differences towards WT controls (p<0.05).</p

THBS4 does not affect gene expression in porcine articular chondrocytes.

<p>(A) qRT-PCR monitoring OAS gene expression in porcine articular chondrocytes treated with human THBS4 and/or SF-CM for 6 hours. (B) qRT-PCR monitoring expression of <i>COL2A1</i>, <i>ACAN</i>, <i>THBS4 and SDC4</i> expression in the same samples. Bars represent mean ± SD (n = 4). Asterisks indicate statistically significant differences towards untreated cells (p<0.05).</p

THBS4 does not affect the molecular behavior of porcine articular chondrocytes.

<p>(A) BrdU incorporation monitoring cellular proliferation of porcine articular chondrocytes in the presence of increasing concentrations of human THBS4 and/or conditioned medium from porcine synovial fibroblasts (SF-CM), as indicated. Bars represent mean ± SD (n = 5). (B) MTT conversion monitoring metabolic activity of porcine articular chondrocytes under the same conditions. Bars represent mean ± SD (n = 5). (C) Caspase-3/7 activity monitoring apoptosis of porcine articular chondrocytes under the same conditions. Bars represent mean ± SD (n = 5). (D) Cell attachment to non-coated plates or plates coated with THBS4 or THBS5/COMP in the absence or presence of a ß1-integrin antibody. Bars represent mean ± SD (n = 6). The asterisk indicates a statistically significant of the antibody (p<0.05).</p

Murine <i>Thbs4</i> is highly expressed in skeletal tissues.

<p>(A) qRT-PCR monitioring <i>Thbs4</i> expression in various tissues of 15 weeks old widltype (WT) and <i>Thbs4</i>-deficient mice (KO). Li, liver; Lu, lung; He, heart; Ao, aorta; Br, brain; St, stomach; Te, tendon; Sp, spleen; CB, cortical bone; AC, articular cartilage. Bars represent mean ± SD (n = 4). <i>Thbs4</i> expression was undetectable in all KO tissues. (B) qRT-PCR monitoring <i>Thbs4</i> expression in primary osteoclasts (Ocl) and osteoblasts (Obl) from wildtype mice at different stages of differentiation. Bars represent mean ± SD (n = 3). (C) qRT-PCR monitioring expression of the indicated genes in articular cartilage of 15 weeks <i>Thbs4</i>-deficient mice. The dotted red line indicates the expression in wildtype littermates. Bars represent mean ± SD (n = 4).</p