Limb ischemia reperfusion (LIRI) injury is associated with serious local and systemic effects. Reperfusion may augment tissue injury in excess of that produced by ischemia alone. Calcium overloading and inflammation are considered to be two of the pathological mechanisms of limb ischemia/ reperfusion (I/R) injury. Tao-Hong-Si-Wu decoction (THSWD) is a traditional Chinese herbal medicine with a powerful anti-inflammatory properties. We studied the probable restorative effect of THSWD on limb I/R-induced calcium overloading and inflammation in myoblast obtained from gastrocnemius muscle tissues of Sprague-Dawley rats (Frizzled Z5,a wnt5a blocker; KN-93, a calmodulin-dependent protein kinase II (CamkII) blocker; XeC, a IP3R blocker as positive controls). The simulated ischemia and reperfusion(I/R) solutions were used to imitate LIRI environment. The results showed that after I/R treatment, the secretion of proinflammatory factors (TNF-α and IL-1β) andWnt5a/Ca 2+ signal molecules (wnt5a, camkII, and IP3R) upregulated significantly, the Ca 2+ concentration enhanced too in myoblast cells. THSWD pretreatment decreased the secretion of TNF-α and IL-1β, Ca 2+ concentration; and abated the Wnt5a/Ca 2+ signal molecules of wnt5a, camkII and IP3R expression activated by I/R injury; but could not abated the Wnt11 and protein kinase C (PKC) expression significantly, the results was similar with Frizzled Z5 treatment cells. Our research illustrated that THSWD may have a mitigating effect on LIRI targeting Wnt/IP3R/CAMKII but not Wnt/IP3R/PKC signaling pathway for the first time. This study may encourage the use of THSWD in the critical clinical settings with LIRI.

Background Liuwei Dihuang Pill (LP) was verified to alleviate postmenopausal osteoporosis (PMOP) development. Nevertheless, the major constituent of LP and the related network pharmacology study remain unexplored. Methods Protein–protein interaction was established to identify the downstream target of LP in PMOP, and the related signaling pathway was investigated by bioinformatics analysis. MC3T3-E1 cells were added to ferric ammonium citrate (FAC) to mimic osteoporosis in vitro. The osteoblasts were identified by Alizarin red staining. Western blot was applied to evaluate protein levels. In addition, Cell Counting Kit-8 (CCK8) assay was applied to assess cell viability, and cell apoptosis was assessed by flow cytometry. Results Quercetin was the major constituent of LP. In addition, quercetin significantly reversed FAC-induced inhibition of osteogenic differentiation in MC3T3-E1 cells. In addition, quercetin notably abolished the FAC-induced upregulation of Bax, Caspase-3, FOS, JUN, TGFB1 and PPARD. In contrast, Bcl-2, p-mTOR/mTOR, p-AKT/AKT and p-PI3K/PI3K levels in MC3T3-E1 cells were reduced by FAC, which was restored by quercetin. Meanwhile, FAC notably inhibited the viability of MC3T3-E1 cells via inducing apoptosis, but this impact was abolished by quercetin. Furthermore, quercetin could reverse pcDNA3.1-FOS-mediated growth of FAC-treated osteoblasts by mediating PI3K/AKT/mTOR signaling. Conclusion Quercetin alleviated the progression of PMOP via activation of PI3K/AKT/mTOR signaling. Hence, this study would shed novel insights into discovering new methods against PMOP.