Dexamethasone stimulates expression of C-type Natriuretic Peptide in chondrocytes-3

<p><b>Copyright information:</b></p><p>Taken from "Dexamethasone stimulates expression of C-type Natriuretic Peptide in chondrocytes"</p><p>BMC Musculoskeletal Disorders 2006;7():87-87.</p><p>Published online 20 Nov 2006</p><p>PMCID:PMC1660540.</p><p></p>encoding cGMP-dependent protein kinase I, A) and (encoding cGMP-dependent protein kinase II, B) mRNA levels were determined by real-time PCR relative to . While expression remained relatively unchanged over time and in response to DEX treatment, levels were significantly increased during the time course and in response to DEX. Data represent mean ± SEM from three or four independent trials (*P < 0.05)

Dexamethasone stimulates expression of C-type Natriuretic Peptide in chondrocytes-2

<p><b>Copyright information:</b></p><p>Taken from "Dexamethasone stimulates expression of C-type Natriuretic Peptide in chondrocytes"</p><p>BMC Musculoskeletal Disorders 2006;7():87-87.</p><p>Published online 20 Nov 2006</p><p>PMCID:PMC1660540.</p><p></p>encoding CNP, A), (encoding the CNP signaling receptor, B) and (encoding the CNP decoy receptor, C) mRNA levels were determined by real-time PCR relative to . expression is significantly increased upon DEX treatment, while DEX down-regulated slightly and up-regulated at the 48 and 72 hr time points. Data represent mean ± SEM from three or four independent trials (*P < 0.05)

The PI3K pathway regulates endochondral bone growth through control of hypertrophic chondrocyte differentiation-6

S), LY294002 or CNP + LY294002. Bone growth measurements showed increased bone growth in the CNP treatment, and no significant difference between the LY294002 and CNP+LY294002. (B) E 15.5 tibiae treated for 6 days with the treatments mentioned above, were fixed, embedded, sectioned and stained with H&E. The length of hypertrophic zone was increased in the CNP treatment compared to control, decreased in the LY294002 treatment and similar between LY294002 and CNP+LY294002<p><b>Copyright information:</b></p><p>Taken from "The PI3K pathway regulates endochondral bone growth through control of hypertrophic chondrocyte differentiation"</p><p>http://www.biomedcentral.com/1471-213X/8/40</p><p>BMC Developmental Biology 2008;8():40-40.</p><p>Published online 11 Apr 2008</p><p>PMCID:PMC2329617.</p><p></p

The PI3K pathway regulates endochondral bone growth through control of hypertrophic chondrocyte differentiation-2

In & eosin (H&E) staining shows decreased length of hypertrophic (HZ) and proliferative zones (PZ) and increased resting zone (RZ) length in the LY294002 treated bones. (B) Measurements of three zones of the growth plate confirmed the reduction in length for the HZ and PZ in the LY294002 treated tibiae, but the RZ length zone was not found to be significantly increased. (C) The HZ length expressed as ratio from the entire growth plate length was found to be significantly decreases and the RZ significantly increased in the case of LY294002 treatment. The PZ ratio was found to be similar between the treatments.<p><b>Copyright information:</b></p><p>Taken from "The PI3K pathway regulates endochondral bone growth through control of hypertrophic chondrocyte differentiation"</p><p>http://www.biomedcentral.com/1471-213X/8/40</p><p>BMC Developmental Biology 2008;8():40-40.</p><p>Published online 11 Apr 2008</p><p>PMCID:PMC2329617.</p><p></p

The PI3K pathway regulates endochondral bone growth through control of hypertrophic chondrocyte differentiation-4

cell area (marked by dashed blue circles), which showed positive collagen X stain, was smaller in the LY294002 treated bones. (B) Tibiae were also analyzed by immunohistochemistry for cyclin-dependent kinase inhibitor 1C (p57), a marker of cell cycle arrest in G1 and chondrocyte differentiation. There was a narrower area of p57 positive cells (showed by brackets) in the LY294002 treated bones. (C) E 15.5 tibiae treated for 6 days with LY294002 shoed an increased number of apoptotic cells in the hypertrophic and mineralized areas (black arrows) as shown by colorimetric TUNEL assay.<p><b>Copyright information:</b></p><p>Taken from "The PI3K pathway regulates endochondral bone growth through control of hypertrophic chondrocyte differentiation"</p><p>http://www.biomedcentral.com/1471-213X/8/40</p><p>BMC Developmental Biology 2008;8():40-40.</p><p>Published online 11 Apr 2008</p><p>PMCID:PMC2329617.</p><p></p

The PI3K pathway regulates endochondral bone growth through control of hypertrophic chondrocyte differentiation-3

Oglycan content. The chondrocyte morphology was analyzed by comparing the chondrocyte size in the three zones of the growth plate. We noticed a reduction of hypertrophic cell size in the case of hypertrophic chondrocytes. (B) E18.5 tibiae treated with LY294002 for 6 days showed the same reduction in hypertrophic zone length and cell size as the E15.5 treated tibiae.<p><b>Copyright information:</b></p><p>Taken from "The PI3K pathway regulates endochondral bone growth through control of hypertrophic chondrocyte differentiation"</p><p>http://www.biomedcentral.com/1471-213X/8/40</p><p>BMC Developmental Biology 2008;8():40-40.</p><p>Published online 11 Apr 2008</p><p>PMCID:PMC2329617.</p><p></p

ATRX-null growth plates are indistinguishable from controls.

<p>(A) Growth plate sections from control and <i>Atrx^Col2Cre</i> mice at P0.5 and P21 were stained with safranin-O/Fast green, demonstrating unaltered growth plate architecture and morphology in mutant mice. Scale bar: 100 µm. (B) No significant difference in the length of the resting, proliferating or hypertrophic zones could be detected between genotypes at birth or weaning (N = 3 littermate pairs; two-tailed T-test). Error bars depict standard error of the mean (SEM). (C) Expression of Sox9, an early chondrocyte differentiation marker expressed in resting and proliferating chondrocytes, is not altered in the absence of ATRX at P0.5 or P21. Scale bar: 50 µm. (D) Immunohistochemistry for the differentiation marker p57 reveals that ATRX loss has no effect on the proportion of chondrocytes reaching terminal differentiation. Scale bar: 100 µm.</p

The PI3K pathway regulates endochondral bone growth through control of hypertrophic chondrocyte differentiation-0

2 or DMSO as vehicle control. They were stained after 6, 9 and 12 days, respectively for different chondrocyte differentiation markers: Alcian blue for glycosaminoglycans, Alkaline phosphatase activity and Alizarin red for the calcium content. The intensity of these markers is reduced in the LY294002 treated cultures. (B) After 9 and 12 days respectively the Alcian blue content of the micromasses was spectrophotometrically measured at 620 nm, after extraction with 6 M Guanidine hydrochloride. We noticed decreased absorbance levels for the LY294002 treated cultures. (C) Measurements of Hoechst fluorescence intensity (excitation/emission: 350/450 nm) showed no significant difference in the DNA content between the LY294002 and DMSO treated micromass cultures. (D, E) RNA was isolated from the micromass cultures after 6 and 9 days of culture and real time analysis was performed. The relative transcript levels for and were decreased in the LY294002 treated micromasses compared to the vehicle control.<p><b>Copyright information:</b></p><p>Taken from "The PI3K pathway regulates endochondral bone growth through control of hypertrophic chondrocyte differentiation"</p><p>http://www.biomedcentral.com/1471-213X/8/40</p><p>BMC Developmental Biology 2008;8():40-40.</p><p>Published online 11 Apr 2008</p><p>PMCID:PMC2329617.</p><p></p

The PI3K pathway regulates endochondral bone growth through control of hypertrophic chondrocyte differentiation-1

Emistry, at the end of the time course. Phosphorylation of Akt at Ser473 was found to be reduced under PI3K inhibition with LY294002 (10 μM). Black arrows show cells expressing phosphorylated Akt. (B) The difference between the length of the tibiae in the beginning and at the end of the time course represents the bone growth and it was found to be significantly reduced in bones treated with LY294002 (45% reduction) or PI3-K α inhibitor IV (500 nM) (35% reduction) compared to DMSO. (C) At the end of culture period the tibiae were also stained with Alcian blue/Alizarin red. We notice decreased length of both proximal (gp1) and distal (gp2) growth plates of tibiae treated with LY294002 or PI3-K α inhibitor IV. (D) Measurements of the growth plates and mineralized area (min) length confirmed the reduction of these under PI3K inhibition with LY294002. (E) When the gp1, gp2 and min were expressed as ratio of the entire bone length there is an increased ratio of the mineralized area in the LY294002 treated tibiae.<p><b>Copyright information:</b></p><p>Taken from "The PI3K pathway regulates endochondral bone growth through control of hypertrophic chondrocyte differentiation"</p><p>http://www.biomedcentral.com/1471-213X/8/40</p><p>BMC Developmental Biology 2008;8():40-40.</p><p>Published online 11 Apr 2008</p><p>PMCID:PMC2329617.</p><p></p

The PI3K pathway regulates endochondral bone growth through control of hypertrophic chondrocyte differentiation-7

2 or DMSO as vehicle control. They were stained after 6, 9 and 12 days, respectively for different chondrocyte differentiation markers: Alcian blue for glycosaminoglycans, Alkaline phosphatase activity and Alizarin red for the calcium content. The intensity of these markers is reduced in the LY294002 treated cultures. (B) After 9 and 12 days respectively the Alcian blue content of the micromasses was spectrophotometrically measured at 620 nm, after extraction with 6 M Guanidine hydrochloride. We noticed decreased absorbance levels for the LY294002 treated cultures. (C) Measurements of Hoechst fluorescence intensity (excitation/emission: 350/450 nm) showed no significant difference in the DNA content between the LY294002 and DMSO treated micromass cultures. (D, E) RNA was isolated from the micromass cultures after 6 and 9 days of culture and real time analysis was performed. The relative transcript levels for and were decreased in the LY294002 treated micromasses compared to the vehicle control.<p><b>Copyright information:</b></p><p>Taken from "The PI3K pathway regulates endochondral bone growth through control of hypertrophic chondrocyte differentiation"</p><p>http://www.biomedcentral.com/1471-213X/8/40</p><p>BMC Developmental Biology 2008;8():40-40.</p><p>Published online 11 Apr 2008</p><p>PMCID:PMC2329617.</p><p></p