Maintenance of increased bone mass after PTH withdrawal by sequential medicarpin treatment via augmentation of cAMP‐PKA pathway

Abstract

Osteoporosis is a metabolic bone disorder associated with impaired bone microarchitecture leading to fragility fractures. Long‐term usage of parathyroid hormone (PTH) enhances bone resorption and leads to osteosarcoma in rats which limits its exposure to maximum 2 years in human. Notably, the anabolic effects of PTH do not endure in the absence of sustained administration. Studies in our lab identified osteogenic and antiresorptive activity in medicarpin, a phytoestrogen belonging to the pterocarpan class. Considering dual‐acting property of medicarpin and limitations of PTH therapy, we envisaged that medicarpin sequential treatment after PTH withdrawal could serve as promising therapeutic approach for osteoporosis treatment. As PTH exerts its bone anabolic effect by increasing osteoblast survival, our study aims to determine whether medicarpin amplifies this effect of PTH. Our results show that PTH withdrawal led to reduced bone mineral density and bone parameters, while sequential treatment of medicarpin after PTH withdrawal significantly enhanced these parameters. Remarkably, these effects were more pronounced than 8‐week PTH treatment. Sequential therapy also significantly increased P1NP levels and decreased CTX levels and TRAP positive cells compared to PTH 8W group where CTX levels were quite high due to bone resorptive action of PTH. Protein expression studies revealed that medicarpin along with PTH betters the antiapoptotic potential compared to PTH alone, through augmentation of cyclic adenosine monophosphate‐PKA‐CREB pathway. These results proclaim that medicarpin sequential treatment prevented the reduction in bone accrual and strength accompanying PTH withdrawal and also aided in antiapoptotic role of PTH. The study points toward the potential use of medicarpin as a replacement therapeutic option postdiscontinuation of PTH.

MMP14 cleaves PTH1R in the chondrocyte-derived osteoblast lineage, curbing signaling intensity for proper bone anabolism

“…Questions for future study are whether the response is as robust in aged mice (2 years or older) and whether, compared to other sources of osteoblasts, the history of PTHRP/PTH1R signaling in pre-hypertrophic chondrocytes ( Mizuhashi et al, 2018 ) produces a molecular memory that is advantageous for the ability of HC descendants to mount a response to PTH. Intermittent treatment with PTH/teraparatide also induces bone resorption leading to an ‘anabolic window’ of bone mass gain that restricts long-term use of this treatment strategy ( Hattersley et al, 2016 ; Sharma et al, 2022 ). It would be important in future to determine the long-term impact of increased PTH signaling on bone anabolism and physiology in Mmp14ΔHC mice as they age.…”

Design, synthesis and biological evaluation of novel pyrimidine derivatives as bone anabolic agents promoting osteogenesis via the BMP2/SMAD1 signaling pathway

“…The primarily osteogenic potential of synthetic compounds was assessed by ALP activity on calvarial osteoblast cells according to a previously published protocol. 26 ALP is a crucial primary osteoblast differentiation marker located at the outer surface of osteoblast cells and plays an essential role in skeletal mineralization. 27 Briefly, osteoblast cells were treated with different concentrations of compounds ranging from 1 pM to 1 μM for 48 h. Medicarpin (Med), a pterocarpan that stimulated osteoblast differentiation at 100 pM, was taken as a positive control.…”

“…Cells were used for further experiments after attaining 80% to 90% confluency. 26 Mouse bone marrow cell isolation and culture. For the mineralization assay, the long bones of 6 to 8 weeks old BALB/c mice were surgically removed, adherent flesh around the legs was removed to clean the bones and bone marrow cells were flushed out in 10% α-MEM complete media and centrifuged at 1500 rpm.…”

“…This journal is © The Royal Society of Chemistry 2024 substrate was added to each well and ALP activity was measured by spectrophotometry at 405 nm. 26 Cell viability assay. To check the effect of test compounds on cell viability, osteoblast cells at a density of 2 × 10 3 were seeded in 96-well plates.…”

“…Then, 100 μL p -nitrophenyl phosphate (PNPP) substrate was added to each well and ALP activity was measured by spectrophotometry at 405 nm. 26…”