Monday, September 5, 2011

Height Increase by inhibiting HSP90?

The effect of inhibition of heat-shock proteins on thiram-induced tibial dyschondroplasia.

"Tibial dyschondroplasia (TD)[growth plates that do not ossify] is a skeletal abnormality. Thiram-induced TD is characterized by enlarged, unvascularized growth plates, low levels of the vascular endothelial growth factor receptor Flk-1, abnormal chondrocyte differentiation, and lameness.  Heat-shock protein 90 (Hsp90) [is inolved] in chondrocyte differentiation and growth-plate vascularization. Inhibition of Hsp90 activity in thiram-induced TD resulted in increased Flk-1 levels, re-instated normal growth-plate angiogenesis and morphology, and abrogated lameness. We evaluated the efficacy of various concentrations of 17-(dimethylaminoethylamino)-17-demethoxygeldanamycin (17-DMAG), an inhibitor of Hsp90 activity, in preventing growth-plate histopathology and lameness in TD-affected chicks. Low doses of 17-DMAG (2 injections, each of 100 or 300 μg) did not prevent TD development even though Flk-1 levels were restored, which suggests that Flk-1 is not the only rate-limiting factor in growth-plate angiogenesis. High doses of 17-DMAG (2 injections, each of 600 or 900 μg) prevented BW loss, decreased the TD score, reduced lesion width, restored proper chondrocyte differentiation, increased blood vessel invasion, and eliminated lameness. To assess the specificity of Hsp90, we evaluated the efficacy of the flavonoid quercetin, an inhibitor of Hsp70 synthesis, in preventing TD development; it decreased Hsp70 levels but not those of Hsp90 in the control growth plates and prevented upregulation of Hsp70 in the TD-affected growth plates. Dietary quercetin (at 100 or 500 ppm) did not prevent the hypoxia that is characteristic of the TD-affected growth plate or development of thiram-induced TD and lameness. In contrast to the anti-angiogenic effect of 17-DMAG observed in mammals, inhibition of Hsp90 activity in the unvascularized TD-affected growth plates resulted in activation of the angiogenic switch and restored normal growth-plate morphology."

Couldn't get full study which is unfortunate as it mentions quercetin as an HSP70.

"Hsp90β is a member of the Hsp90 family of protein chaperones. This family plays essential roles in the folding, maturation and activity of many proteins that are involved in signal transduction and transcriptional regulation.
Human OA chondrocytes were isolated from cartilage obtained from patients undergoing joint replacement surgery, and primary cultured. Cells were stimulated with proinflammatory cytokines (IL-1β or TNF-α) and nitric oxide donors (NOC-12 or SNP). For Hsp90β inhibition, two different chemical inhibitors (Geldanamycin and Novobiocin) were employed, or siRNA transfection procedures were carried out.
Hsp90β was found to be increased by proinflammatory cytokines. Inhibition of Hsp90β by the chemicals Geldanamycin (GA) and Novobiocin (NB) caused a dose-dependent decrease of the NO production induced by IL-1β in chondrocytes, up to basal levels. Immunofluorescence analyses demonstrate that the NO donors NOC-12 and SNP also increased Hsp90β. Chemical inhibition or specific gene silencing of this chaperone reduced the DNA condensation and fragmentation, typical of death by apoptosis, that is induced by NO donors in chondrocytes."

"the proinflammatory cytokine IL-1β acts as a positive modulator of Hsp90β abundance"

"Hsp90 [interacts with] glucocorticoid receptors, Akt/Protein kinase B and Raf-1, the tumor suppressor protein p53, and NOS family members"

"silencing Hsp90β significantly increased MMP-13"

"Hsp90 inhibition (of both α and β forms) blocked IL-1β-induced up-regulation of MMP-13 in equine articular chondrocytes"

"Thiram-induced tibial dyschondroplasia (TD) and vitamin-D deficiency rickets are avian bone disorders of different etiologies characterized by abnormal chondrocyte differentiation, enlarged and unvascularized growth plates, and lameness. Heat-shock protein 90 (Hsp90) is a proangiogenic factor in mammalian tissues and in tumors; therefore, Hsp90 inhibitors were developed as antiangiogenic factors. Administration of the Hsp90 inhibitor to TD- and rickets-afflicted chicks at the time of induction resulted in reduction in growth-plate size and, contrary to its antiangiogenic effect in tumors, a major invasion of blood vessels occurred in the growth plates. This was the result of upregulation of the VEGF receptor Flk-1, the major rate-limiting factor of vascularization in TD and rickets. In addition, the abnormal chondrocyte differentiation, as characterized by collagen type II expression and alkaline phosphatase activity, and the changes in hypoxia-inducible factor-1α (HIF-1α) in both disorders were restored. All these changes resulted in prevention of lameness. Inhibition of Hsp90 activity reduced growth-plate size, increased vascularization, and mitigated lameness also in TD chicks with established lesions.  In contrast to the antiangiogenic effect of Hsp90 inhibitors observed in mammals, inhibition of Hsp90 activity in the unvascularized TD- and rickets-afflicted chicks resulted in activation of the angiogenic switch and reinstated normal growth-plate morphology."

HSP90 stimulation of NO pathway may be what increases growth plate size.

"HIF-1α is one of the major client proteins of heat-shock protein 90 (Hsp90), and it is required for the functioning and the rapid hypoxic stabilization of HIF-1α, which otherwise is degraded by the ubiquitin-proteasome protein system. Hsp90 is implicated in angiogenesis by affecting the VEGF/VEGF-receptor system at various levels"

HSP 90 is induced by high hydrostatic pressure.

Specific induction of heat shock protein 90beta by high hydrostatic pressure.

"In chondrocytes, a low-amplitude intermittent hydrostatic pressure induces production of extracellular matrix molecules, while high hydrostatic pressure inhibits it. High pressure increases cellular heat shock protein 70 level in a number of cell types on account of increased stabilisation of the heat shock protein 70 mRNA. In our experiments, only bovine primary chondrocytes, but not an immortalized chondrocytic cell line, could resist the induction of the stress response in the presence of continuous 30 MPa hydrostatic pressure. We have recently shown that protein synthesis is required for the stabilization. According to two-dimensional gel electrophoresis the synthesis of heat shock protein 90 was also increased in a chondrocytic cell line and in HeLa cells, and mass spectrometric analysis suggested that the induction was rather due to increase in heat shock protein 90beta than in heat shock protein 90alpha. The stress response was rather intense in HeLa cells, therefore, we investigated the effect of continuous 30 MPa hydrostatic pressure on the expression of the two heat shock protein 90 genes in HeLa cells using Northern and Western blot analyses. Heat shock protein 90beta mRNA level increased within 6 hours of exposure to 30 MPa hydrostatic pressure, while hsp90alpha level remained stable. At protein level there was a clear increase in the heat shock protein 90beta/heat shock protein 90alpha ratio, too."

The intracellular Ca(2+)-pump inhibitors thapsigargin and cyclopiazonic acid induce stress proteins in mammalian chondrocytes., states HSP90 can also be induced by inhitibion of the calcium pump.

Identification of differentially expressed genes in the growth plate of broiler chickens with thiram-induced tibial dyschondroplasia.

"Tibial dyschondroplasia (TD) is characterized by expansion of the proximal growth plates of the tibiotarsus that fail to form bone, lack blood vessels, and contain non-viable cells. Thiram (a carbamate pesticide), when fed to young broiler chicks, induces TD with high regularity and precision. We used this experimental model to understand the cause of the defects associated with TD by selecting and identifying the genes differentially expressed in the TD growth plate of broiler chickens. Broiler chicks at 7 days of age were randomly divided into two groups. After fasting overnight, they were fed with regular diet (control) or the same diet containing 100 mg/kg thiram for 96 h to induce TD (thiram-fed). mRNA was purified from the growth plates of control and thiram-fed broilers. Forward and reverse-subtracted cDNA libraries were generated by suppression subtractive hybridization technology. Ten selected genes from cDNA libraries were identified by real-time quantitative polymerase chain reaction. All were differentially expressed in TD growth plates. The levels of collagen type X (Col X){up}, pro-alpha-1 collagen type I (Col I alpha1){up}, collagen type IX (Col IX), NADH dehydrogenase (NADH DH), cytochrome C oxidase subunit III (COX III), enolase 1, alpha (ENO1){down}, carbonic anhydrase II (CA2) and heat shock protein 90 (Hsp90) mRNA transcripts were up-regulated, while the expression levels of Matrilin 3 (MATN3){up} and chondromodulin-I (ChM-I) were down-regulated. Col I and Hsp90 were detected by immunohistochemistry at different stages. Given that these genes are involved in matrix formation, endochondral ossification, developmental regulation, electron transport in the mitochondrial respiratory chain and vascularization."

"increased Col I was detected in the thiram-fed chicken growth plate on days 4, 9, 16 and 23 or around the TD lesion at the 16th and 23rd days."

"NADH DH, cytochrome C oxidase (COX) and ENO1 are important enzymes for transmitting electrons in the mitochondrial respiratory chain and in glucose metabolism. Thiram may oxidize cellular thiol-group-containing compounds, resulting in cellular injury and death by interfering with the –SH group(s) of numerous critical proteins and enzymes"

According to Genistein effects on growth and cell cycle of Candida albicans., Genistein increases HSP90 levels in plants.
I think the best thing is to inhibit HSP90 and to stimulate the CNP pathway over the NO pathway.

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