calpain and 3-(5--hydroxymethyl-2--furyl)-1-benzylindazole

Osteoporosis is a major health problem in postmenopausal women and the elderly that leads to fractures associated with substantial morbidity and mortality. Current osteoporosis therapies have significant drawbacks, and the risk of fragility fractures has not yet been eliminated. There remains an unmet need for a broader range of therapeutics. Previous studies have shown that YC-1 has important regulatory functions in the cardiovascular and nervous systems. Many of the YC-1 effector molecules in platelets, smooth muscle cells and neurons, such as cGMP and μ-calpain, also have important functions in osteoclasts. In this study, we explored the effects of YC-1 on bone remodeling and determined the potential of YC-1 as a treatment for postmenopausal osteoporosis. Micro-computed tomography of lumbar vertebrae showed that YC-1 significantly improved trabecular bone microarchitecture in ovariectomized rats compared with sham-operated rats. YC-1 also significantly reversed the increases in serum bone resorption and formation in these rats, as measured by enzyme immunoassays for serum CTX-1 and P1NP, respectively. Actin ring and pit formation assays and TRAP staining analysis showed that YC-1 inhibited osteoclast activity and survival. YC-1 induced extrinsic apoptosis in osteoclasts by activating caspase-3 and caspase-8. In osteoclasts, YC-1 stimulated μ-calpain activity and inhibited Src activity. Our findings provide proof-of-concept for YC-1 as a novel antiresorptive treatment strategy for postmenopausal osteoporosis, confirming an important role of nitric oxide/cGMP/protein kinase G signaling in bone.

Topics: Actins; Animals; Apoptosis; Bone Density Conservation Agents; Bone Resorption; Calpain; Cell Survival; Female; Humans; Indazoles; Lumbar Vertebrae; Osteoblasts; Osteoclasts; Osteoporosis; Ovariectomy; Rats, Sprague-Dawley; Signal Transduction; src-Family Kinases; X-Ray Microtomography

Neurodegenerative brain disorders such as Alzheimer's disease (AD) have been well investigated. However, significant methods for the treatment of the progression of AD are unavailable currently. Heat shock protein 70 (Hsp70) plays important roles in neural protection from stress by assisting cellular protein folding. In this study, we investigated the effect and the molecular mechanism of YC-1, an activator of guanylyl cyclase (GC), on Aβ25-35-induced cytotoxicity in differentiated PC12 cells. The results of this study showed that Aβ25-35 (10 µM) significantly increased p25 protein production in a pattern that was consistent with the increase in μ-calpain expression. Moreover, Aβ25-35 significantly increased tau hyperphosphorylation and induced differentiated PC12 cell death. YC-1 (0.5-10 µM) prevented the cell death induced by Aβ25-35. In addition, YC-1 (1, 10 µM) significantly blocked Aβ25-35-induced μ-calpain expression and decreased the formation of p25 and tau hyperphosphorylation. Moreover, YC-1 (5-20 µM) alone or combined with Aβ25-35 (10 µM) significantly increased the expression of Hsp70 in differentiated PC12 cells. The neuroprotective effect of YC-1 was significantly attenuated by an Hsp70 inhibitor (quercetin, 50 µM) or in PC12 cells transfected with an Hsp70 small interfering RNA. However, pretreatment of cells with the GC inhibitor ODQ (10 µM) did not affect the neuroprotective effect of YC-1 against Aβ25-35 in differentiated PC12 cells. These results suggest that the neuroprotective effect of YC-1 against Aβ25-35-induced toxicity is mainly mediated by the induction of Hsp70. Thus, YC-1 is a potential agent against AD.

Topics: Amyloid beta-Peptides; Animals; Calpain; Cell Differentiation; Guanylate Cyclase; HSP70 Heat-Shock Proteins; Indazoles; Neuroprotective Agents; Neurotoxins; PC12 Cells; Peptide Fragments; Phosphorylation; Rats; Signal Transduction; tau Proteins