Zishen Yutai pills restore fertility in premature ovarian failure through regulating arachidonic acid metabolism and the ATK pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: The Zishen Yutai pills (ZYP), a Chinese medicinal formulation derived from the Qing Dynasty prescription "Shou Tai pills", have been documented to exhibit beneficial effects in clinical observations treating premature ovarian failure (POF). However, the anti-POF effects and its comprehensive systemic mechanism have not yet been clarified. AIM OF THE REVIEW: Therapeutic effects and systemic mechanism of ZYP in POF were evaluated. MATERIALS AND METHODS: After pulverization, sieving, and stirring, ZYP was administered intragastrically to cisplatin-induced POF mice at a dose of 1.95 mg/kg/d for 14 days. The anti-POF effects of ZYP were investigated by assessing the number of ovarian follicles at different developmental stages, as well as measuring serum estradiol (E2) levels and ovarian-expressed anti-Müllerian hormone (AMH). Reproductive performance and offspring health were evaluated to predict fertility restoration. Furthermore, a combination of proteomic and metabolomic profiling was employed to elucidate the underlying molecular mechanism of ZYP in treating POF. Western blot (WB) analyses and real-time quantitative polymerase chain reaction (RT-qPCR) were conducted to explore the mechanisms through which ZYP exerted its anti-POF effects. RESULTS: We have demonstrated that oral administration of ZYP reversed the reduction in follicles at different developmental stages and stimulated the expressions of serum E2 and ovarian-expressed AMH in a cisplatin-induced POF model. Additionally, ZYP ameliorated follicle apoptosis in ovaries affected by cisplatin-induced POF. Furthermore, treatment with ZYP restored the quantity and quality of oocytes, as well as enhanced fertility. Our results revealed 62 differentially expressed proteins (DEPs) through proteomic analyses and identified 26 differentially expressed metabolites (DEMs) through metabolomic analyses. Both DEPs and DEMs were highly enriched in the arachidonic acid (AA) metabolism pathway. ZYP treatment effectively upregulated the protein and mRNA expression of critical targets in AA metabolism and the AKT pathway, including CYP17α1, HSD3ß1, LHR, STAR, and AKT, in cisplatin-induced POF mice. CONCLUSIONS: These results indicated that ZYP exerted protective effects against POF and restored fertility from cisplatin-induced apoptosis. ZYP could be a satisfying alternative treating POF.

Protective effect of ZYMT, a traditional Chinese patent medicine in a mouse model of retinitis pigmentosa.

Retinitis pigmentosa (RP) is the most common genetic disorder that causes blindness. At present, there exists no remedy for the disease. The aim of the current research was to investigate the protective effect of Zhangyanming Tablets (ZYMT) in a mouse model of RP, and explore the underlying mechanism. Eighty RP mice were randomly divided into two groups. The mice in ZYMT group were administered with ZYMT suspension(0.0378 g/mL), while the mice in model group were given the same volume of distilled water. At day 7 and day 14 after intervention, electroretinogram (ERG), fundus photography, and histological examination were used to assess the retinal function and structure. TUNEL, immunofluorescence and qPCR were used to evaluate cell apoptosis and expressions of Sirt1, Iba1, Bcl-2, Bax and Caspase-3. A significantly shortened latency of ERG waves was observed in ZYMT-treated mice, in comparison to those in the model group (P < 0.05). Histologically, ultrastructure of the retina was better preserved, and the outer nuclear layer (ONL) exhibited marked increase in thickness and cell count in ZYMP group (P < 0.05). The apoptosis rate was decreased markedly in ZYMT group. Immunofluorescence analysis showed that the expressions of Iba1 and Bcl-2 in the retina were increased, Bax and Caspase-3 were decreased after ZYMT intervention, while the qPCR revealed that the expressions of Iba1 and Sirt1 were significantly increased (P < 0.05). This study indicated that ZYMT has protective effect on retinal function and morphology of inherited RP mice in the early stage, possibly mediated via the regulation of antioxidant and anti-/pro-apoptotic factors expressions.

Mechanisms of action of Zishen Yutai pills in treating premature ovarian failure determined by integrating UHPLC-Q-TOF-MS and network pharmacology analysis.

BACKGROUND: Zishen Yutai (ZSYT) pill, a patent Chinese medicine, has been widely used in the treatment of infertility, abortion, and adjunctive treatment of in vitro fertilization (IVF) for decades. Recently, the results of clinical observations showed that premature ovarian failure (POF) patients exhibited improved expression of steroids and clinical symptoms associated with hormone disorders after treatment with Zishen Yutai pills. However, the pharmacological mechanism of action of these pills remains unclear. METHODS: The compounds of Zishen Yutai pills found in blood circulation were identified via ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS) technique in the serum of POF mice after oral administration of Zishen Yutai pills. The potential targets of compounds were screened using Traditional Chinese Medicine Systems Pharmacology Database, Traditional Chinese Medicine Database@Taiwan, Drugbank Database, PubChem, HIT, Pharmapper, and Swiss Target Prediction. The target genes associated with POF were collected from Online Mendelian Inheritance in Man Database, PharmGkb, Genecards, Therapeutic Target Database, and Genetic Association Database. The overlapping genes between the potential targets of Zishen Yutai pills' compounds and the target genes associated with POF were clarified via protein-protein interaction (PPI), pathway, and network analysis. RESULTS: Nineteen compounds in Zishen Yutai pills were detected in the serum of POF mice after oral administration. A total of 695 Zishen Yutai (ZSYT) pill-related targets were screened, and 344 POF-related targets were collected. From the results of Zishen Yutai (ZSYT) pill-POF PPI analysis, CYP19A1, AKR1C3, ESR1, AR, and SRD5A2 were identified as key targets via network analysis, indicating their core role in the treatment of POF with Zishen Yutai pills. Moreover, the pathway enrichment results suggested that Zishen Yutai pills treated POF primarily by regulating neuroactive ligand-receptor interaction, steroid hormone biosynthesis, and ovarian steroidogenesis. CONCLUSIONS: Via virtual screening, we found that regulation of neuroactive ligand-receptor interaction, steroid hormone biosynthesis, and ovarian steroidogenesis was the potential therapeutic mechanism of Zishen Yutai pills in treating POF. Our study suggested that combining the analysis of Zishen Yutai pills' compounds in blood in vivo in the POF model and network pharmacology prediction might offer a tool to characterize the mechanism of Zishen Yutai pills in the POF.

Bone-targeted lncRNA OGRU alleviates unloading-induced bone loss via miR-320-3p/Hoxa10 axis.

Unloading-induced bone loss is a threat to human health and can eventually result in osteoporotic fractures. Although the underlying molecular mechanism of unloading-induced bone loss has been broadly elucidated, the pathophysiological role of long noncoding RNAs (lncRNAs) in this process is unknown. Here, we identified a novel lncRNA, OGRU, a 1816-nucleotide transcript with significantly decreased levels in bone specimens from hindlimb-unloaded mice and in MC3T3-E1 cells under clinorotation-unloading conditions. OGRU overexpression promoted osteoblast activity and matrix mineralization under normal loading conditions, and attenuated the suppression of MC3T3-E1 cell differentiation induced by clinorotation unloading. Furthermore, this study found that supplementation of pcDNA3.1(+)-OGRU via (DSS)6-liposome delivery to the bone-formation surfaces of hindlimb-unloaded (HLU) mice partially alleviated unloading-induced bone loss. Mechanistic investigations demonstrated that OGRU functions as a competing endogenous RNA (ceRNA) to facilitate the protein expression of Hoxa10 by competitively binding miR-320-3p and subsequently promote osteoblast differentiation and bone formation. Taken together, the results of our study provide the first clarification of the role of lncRNA OGRU in unloading-induced bone loss through the miR-320-3p/Hoxa10 axis, suggesting an efficient anabolic strategy for osteoporosis treatment.

Targeted overexpression of the long noncoding RNA ODSM can regulate osteoblast function in vitro and in vivo.

Ameliorating bone loss caused by mechanical unloading is a substantial clinical challenge, and the role of noncoding RNAs in this process has attracted increasing attention. In this study, we found that the long noncoding RNA osteoblast differentiation-related lncRNA under simulated microgravity (lncRNA ODSM) could inhibit osteoblast apoptosis and promote osteoblast mineralization in vitro. The increased expression level of the lncRNA ODSM partially reduced apoptosis and promoted differentiation in MC3T3-E1 cells under microgravity unloading conditions, and the effect was partially dependent on miR-139-3p. LncRNA ODSM supplementation in hindlimb-unloaded mice caused a decrease in the number of apoptotic cells in bone tissue and an increase in osteoblast activity. Furthermore, targeted overexpression of the lncRNA ODSM in osteoblasts partially reversed bone loss induced by mechanical unloading at the microstructural and biomechanical levels. These findings are the first to suggest the potential value of the lncRNA ODSM in osteoporosis therapy and the treatment of pathological osteopenia.

[Preparation instructions for Experts consensus statement on Cheezheng Xiaotong Tiegao in clinical practice].

As a topical plaster developed by modern pharmaceutical technology based on traditional Tibetan medicine,Cheezheng Xiaotong Tiegao has functions of promoting blood circulation,relieving swelling and relieving pain. Since its introduction in 1993,it has been widely used in the treatment of various types of acute and chronic musculoskeletal pain and various types of spinal,joint and soft tissue diseases. In order to better standardize the clinical application and improve the clinical efficacy of Cheezheng Xiaotong Tiegao,the research and development work of the Experts consensus statement on Cheezheng Xiaotong Tiegao in clinical practice was officially launched on October 19,2017,upon approval from China Association of Chinese Medicine. In this paper,main R&D process and related technical links for the experts consensus on Cheezheng Xiaotong Tiegao would be summarized,which will help the various medical workers understand,master and apply more accurately,and also provide reference for the development of experts consensus on clinical application of other topical Chinese medicines.

Bushen Yijing Fang Reduces Fall Risk in Late Postmenopausal Women with Osteopenia: A Randomized Double-blind and Placebo-controlled Trial.

Falls in late postmenopausal women with osteopenia usually cause fractures with severe consequences. This 36-month randomized, double-blind and placebo-controlled trial with a 10-year observational follow-up study aimed to investigate the long-term effect of herbal formula Bushen Yijing Fang (BSYJF) on fall risk in the late postmenopausal women with osteopenia. 140 late postmenopausal women (Femoral neck T-score, -2.5~-2 SD) were recruited and randomized to orally receive calcium carbonate 300 mg daily with either BSYJF or placebo for 36 months. The effect was further investigated for another 10-year follow-up. During the 36-month administration, there were 12 falls in BSYJF group and 28 falls in placebo group, respectively, indicating 64% lower risk of falls (RR 0.36 [95% CI, 0.18 to 0.71]; P = 0.004) in BSYJF group. During the 10-year follow-up, 36% lower fall risk (RR 0.64 [95% CI, 0.46 to 0.89]; P = 0.009) was observed in BSYJF group. No significant difference was found in safety profile between two groups. Thirty-six-month administration of BSYJF reduced fall risk with an increase in bone mass, and its latent effect on fall risk was continually observed in the 10-year follow-up in late postmenopausal women with osteopenia. This clinical trial was registered at Chinese clinical trial registry (ChiCTR-IOR-16008942).

Osteoblast-targeted delivery of miR-33-5p attenuates osteopenia development induced by mechanical unloading in mice.

A growing body of evidence has revealed that microRNAs (miRNAs) play crucial roles in regulating osteoblasts and bone metabolism. However, the effects of miRNAs in osteoblast mechanotransduction remain to be defined. In this study, we investigated the regulatory effect of miR-33-5p in osteoblasts and tested its anti-osteopenia effect when delivered by an osteoblast-targeting delivery system in vivo. First, we demonstrated that miR-33-5p could promote the activity and mineralization of osteoblasts without influencing their proliferation in vitro. Then our data showed that supplementing miR-33-5p in osteoblasts by a targeted delivery system partially recovered the osteopenia induced by mechanical unloading at the biochemical, microstructural, and biomechanical levels. In summary, our findings demonstrate that miR-33-5p is a key factor in the occurrence and development of the osteopenia induced by mechanical unloading. In addition, targeted delivery of the mimics of miR-33-5p is a promising new strategy for the treatment of pathological osteopenia.

Targeted Delivery Systems for Molecular Therapy in Skeletal Disorders.

Abnormalities in the integral components of bone, including bone matrix, bone mineral and bone cells, give rise to complex disturbances of skeletal development, growth and homeostasis. Non-specific drug delivery using high-dose systemic administration may decrease therapeutic efficacy of drugs and increase the risk of toxic effects in non-skeletal tissues, which remain clinical challenges in the treatment of skeletal disorders. Thus, targeted delivery systems are urgently needed to achieve higher drug delivery efficiency, improve therapeutic efficacy in the targeted cells/tissues, and minimize toxicities in non-targeted cells/tissues. In this review, we summarize recent progress in the application of different targeting moieties and nanoparticles for targeted drug delivery in skeletal disorders, and also discuss the advantages, challenges and perspectives in their clinical translation.

Effects of hypocrellin A on expression of vascular endothelial growth factor and endothelin-1 in human umbilical endothelial cells.

Increased endothelin-1 (ET-1), vascular endothelial growth factor (VEGF) and activation of protein kinase C (PKC) are co-contributors to endothelial hyperpermeability in diabetes. Several lines of evidence have suggested a hypothesis that activation of specific PKC isoforms are the causative factor in ET-1 and VEGF mediated endothelial dysfunction. In the present study, we tested this hypothesis with hypocrellin A, a naturally occurring PKC inhibitor from a Chinese plant. Human umbilical vein endothelial cells (HUVECs) were incubated with 20 mM glucose in both the presence and absence of hypocrellin A, after which, the protein expression and release of VEGF and mRNA expression and release of ET-1 were measured. VEGF and ET-1 were released into the medium and expressions of VEGF protein and ET-1 mRNA were significantly increased in HUVECs incubated with 20 mM glucose. Hypocrellin A (150 nM) significantly decreased VEGF release (117 +/- 3 vs. 180 +/- 11 pg/mg, p < 0.05) and VEGF protein expression (from 130 +/- 14% to 88 +/- 18.5%, p < 0.05). ET-1 release was also reduced in hypocrellin A treated HUVECs (63.3 +/- 9.9 vs. 75.2 +/- 12.6 ng/mg). Hypocrellin A significantly reversed the effect of high glucose on ET-1 mRNA expression (p < 0.05). The results revealed that PKC activation plays a pivotal role in VEGF and ET-1 mediated endothelial permeability. The naturally occurring compound hypocrellin A may be a potentially novel treatment for endothelial dysfunction in diabetes.

Inhibitory effect of hypocrellin A on protein kinase C in liver and skeletal muscle of obese Zucker rats.

In this ex vivo study, the inhibitory activity of hypocrellin A (HA), a perylene quinonoid pigment isolated from the Chinese medicinal fungus Hypocrella bambuase, on protein kinase C (PKC) enzyme activity in insulin target tissues of obese Zucker rats was assessed. Pre-incubation with HA for 30 minutes significantly inhibited the activity of partially purified PKC enzyme from liver and soleus skeletal muscle in a dose-dependent manner (IC50 = 0.07 and 0.26 microg/ml, respectively). HA produced a greater inhibitory effect in enzyme prepared from the liver than enzyme prepared from soleus muscle. Since total PKC activity in these two insulin target tissues is the net result of several different isoforms of PKC, and PKC-theta is a major isoform expressed in the soleus skeletal muscle, the present data suggest that the naturally occurring compound, HA, may selectively inhibit certain PKC isoforms other than PKC-theta. Further investigations are required to determine which PKC isoforms are most susceptible to HA and whether changes in PKC signaling during treatment with HA can reverse abnormalities of glucose and lipid metabolism in insulin resistant and diabetic states.