[Acupuncture combined with auricular point sticking for girls aged 3-8 years with incomplete precocious puberty: a randomized controlled trial].

OBJECTIVE: To observe the efficacy and safety of acupuncture combined with auricular point sticking for girls aged 3-8 years with incomplete precocious puberty (IPP). METHODS: Sixty girls with IPP were randomly divided into an observation group (30 cases, 2 cases dropped off) and a control group (30 cases, 2 cases were eliminated). The girls in the control group were treated with healthy diet and proper exercise for 12 weeks. On the basis of the treatment in the control group, the girls in the observation group were treated with acupuncture combined with auricular point sticking. The acupuncture was applied at Sanyinjiao (SP 6), Guanyuan (CV 4), Guilai (ST 29), etc., the needles were retained for 20 min, acupuncture was given twice a week (once every 3 days). The auricular point sticking was applied at Luanchao (TF2), Neishengzhiqi (TF2), Neifenmi (CO18), Yuanzhong (AT2,3,4i), etc., twice a week. The treatment was given for 12 weeks. Before treatment, after treatment and in follow-up after 12 weeks of treatment completion, the Tanner stage of breast, serum contents of sex hormone (luteinizing hormone [LH], follicle-stimulating hormone [FSH], estradiol [E2]) were observed. The ovarian volume, the number of follicles with diameter>4 mm, and the uterine volume were measured by abdominal color Doppler ultrasound. In addition, the safety of the observation group was evaluated. RESULTS: Compared with before treatment, the Tanner stage of breast in the observation group was improved after treatment and in follow-up (P<0.05); after treatment and in follow-up, the Tanner stage of breast in the observation group was better than that in the control group (P<0.05). Compared with before treatment, the serum levels of LH and E2 in the observation group were increased (P<0.05), and the volume of bilateral ovaries was larger (P<0.05) in follow-up. Compared with before treatment, the serum contents of LH, FSH and E2 in the control group were increased (P<0.05), the volume of bilateral ovaries was larger (P<0.05), and the number of follicles was increased (P<0.05) after treatment and in follow-up. The serum levels of LH, FSH and E2 in the observation group were lower than those in the control group (P<0.05), the volume of bilateral ovaries was smaller than that in the control group (P<0.05), and the number of follicles was lower than that in the control group (P<0.05). Compared with before treatment, the uterine volume in the two groups was larger in follow-up (P<0.05). There was no statistically significant difference between the two groups after treatment and in follow-up (P>0.05). During the treatment, 3 cases in the observation group had slight abdominal pain and subcutaneous blood stasis, without serious adverse reactions. CONCLUSION: Acupuncture combined with auricular point sticking could improve the Tanner stage of breast, reduce the level of sex hormone, slow down the development and maturation of ovary and follicle, and control the degree and speed of sexual development in girls aged 3-8 years with IPP.

[Effect of transcutaneous electrical acupoint stimulation combined with auricular acupressure on sexual hormone level and gonadal development in girls with precocious puberty].

OBJECTIVE: To explore the effect of transcutaneous electrical acupoint stimulation (TEAS) combined with auricular acupressure on serum sexual hormone level, and the ovarian, follicular and uterine development in the girls with precocious puberty. METHODS: Sixty girls with precocious puberty were randomly divided into a control group (30 cases, 2 cases dropped off) and a treatment group (30 cases, 3 cases dropped off). In both the control group and the treatment group, the healthy life-style intervention was provided for 12 weeks. Besides, in the treatment group, TEAS was delivered for 20 minutes each time, twice a week; and bilateral auricular acupressure was combined. The duration of treatment was 12 weeks. The levels of serum luteinizing hormone (LH), follicle stimulating hormone (FSH) and estradiol (E2), as well as ovarian volume, maximum follicular diameter and uterine volume before and after treatment were detected, and the safety was evaluated separately. RESULTS: Compared with before treatment, the contents of serum LH and FSH were increased (P<0.05), and the ovarian volume and the maximum follicle diameter were increased (P<0.05) in the control group after treatment; the contents of serum FSH and E2 were decresed (P<0.05), and the maximum follicular diameter was reduced significantly (P<0.05) in the treatment group after treatment. Compared with the control group, the contents of serum LH, FSH and E2 were decreased (P<0.05), while the ovarian volume and the maximum follicle diameter were decreased (P<0.05) in the treatment group after treatment. CONCLUSION: TEAS combined with auricular acupressure can effectively decrease the level of sex hormone, improve the ovarian and uterine development and retard the gonadal development. Such combined therapy is of high safety and conductive to regulating the development for the girls with precocious puberty.

Hu'po Anshen Decoction Accelerated Fracture-Healing in a Rat Model of Traumatic Brain Injury Through Activation of PI3K/AKT Pathway.

Hu'po Anshen decoction (HPASD) is a traditional Chinese medicine formula comprising five herbal medicines for the treatment of concussion and fracture healing, but its pharmacological mechanism is still unclear. Ultra-performance liquid chromatography coupled with quadrupole time of flight mass spectrometry (UPLC/Q-TOF MS) was used to analyze the main active components of HPASD. Rats were randomly assigned to fracture group, fracture combined with traumatic brain injury (TBI) group (FBI) and FBI combined with HPASD treatment group (FBIH). Rats in the FBIH group were given oral doses of HPASD (2.4 g/kg, 4.8 g/kg and 9.6 g/kg) for 14 or 21 consecutive days. The fracture callus formation and fracture sites were determined by radiographic analysis and micron-scale computed tomography (micro-CT) analysis. Hematoxylin and eosin (H&E) staining and a three-point bending test were applied to assess histological lesions and biomechanical properties, respectively. The levels of cytokines-/protein-related to bone formation and differentiation as well as PI3K/AKT pathway-related proteins were determined by Enzyme-linked immunosorbent assay (ELISA), quantitative reverse transcription-polymerase chain reaction (qRT-PCR), or western blot assays, respectively. UPLC-Q/TOF-MS-based serum metabolomic analysis was also performed to investigate the therapeutic effects of HPASD in the treatment of FBI. UPLC/Q-TOF MS analysis showed the chemical components in HPASD, including flavonoids, amino acids, saponins, and phenylpropanoid constituents, etc. HPASD dose-dependently promoted callus formation, increased bone density, improved mechanical parameters and morphological scores, and facilitated the expressions of VEGF, PDGF, bFGF, VEGFA, CoL1A1, RUNX2, BMP2, and Aggrecan, inhibited the expression of MMP13, and activated PI3K/AKT pathway. Metabolomics analysis revealed abnormalities of malate-aspartate shuttle and glucose-alanine. HPASD accelerates fracture healing by promoting bone formation and regulating the malate-aspartate shuttle and glucose-alanine cycle, which might be associated with the activation of the PI3K/AKT pathway.

The water extracts of Euonymus alatus (Thunb.) Siebold attenuate diabetic retinopathy by mediating angiogenesis.

ETHNOPHARMACOLOGICAL RELEVANCE: Euonymus alatus (Thunb.) Siebold (family Celastraceae) is a deciduous woody shrub that is recorded in ShenNong BenCaoJing. It has been widely used for diabetes in traditional Chinese medicine. AIM OF THE STUDY: This study aimed to identify the most effective extract of Euonymus alatus (EA) against high glucose-induced endothelial cells in vitro, evaluate its pharmacological effect on retinopathy in diabetic mice and explore its underlying mechanism by RNA sequencing. METHODS: Retinal vascular endothelial cells (RF/6A) were treated with normal glucose (5.5 mmol/L glucose), high glucose (25 mmol/L glucose) or high glucose plus methanol extracts of EA (MEA), ethyl acetate extracts of EA (EEA) or water extracts of EA (WEA). The cytotoxicity and cell viability were determined by Cell Counting Kit-8 (CCK-8) assay. Cell migration was examined using the Transwell assay, and tube formation ability was measured using the Matrigel assay. Then, the KK-Ay mice were administered WEA or water for 12 weeks. The velocities of ocular blood flow were determined by Doppler ultrasound. RNA sequencing and reverse transcription quantitative PCR (RT-qPCR) were performed on WEA-stimulated RF/6A cells to reveal the underlying mechanism. RESULTS: The cytotoxicity assay found that 30 µg/mL MEA, 20 µg/mL EEA and 30 µg/mL WEA had no toxic effect on RF/6A cells. The cell viability results showed that MEA, EEA and WEA all decreased cell viability. Compared with the high-glucose group, both MEA and WEA decreased the number of migrated cells, while the inhibition rate of WEA was higher. The Matrigel results showed that 30 µg/mL WEA effectively reduced the total tube length. Moreover, WEA improved the haemodynamics of the central retinal artery. RNA sequencing coupled with RT-qPCR verified that WEA regulated angiogenesis-related factors in high glucose-stimulated RF/6A cells. CONCLUSIONS: WEA inhibits the migration and tube formation of RF/6A cells and improves diabetic retinopathy (DR) by mediating angiogenesis.

Fufang Xueshuantong alleviates diabetic retinopathy by activating the PPAR signalling pathway and complement and coagulation cascades.

ETHNOPHARMACOLOGICAL RELEVANCE: Fufang Xueshuantong (FXST) is a traditional Chinese patent medicine composed of Panax notoginseng (Burkill) F.H.Chen (Araliaceae), Salvia miltiorrhiza Bunge (Lamiaceae), Astragalus propinquus Schischkin (Leguminosae), and Scrophularia ningpoensis Hemsl. (Scrophulariaceae). It has been widely used for the treatment of diabetic retinopathy (DR) and exerts a positive clinical therapeutic effect. AIM OF THE STUDY: The aim of this study was to observe the effect of FXST on diabetic rat retinas and investigate its pharmacological mechanism for improving DR. METHODS: The diabetic rat model was established by intraperitoneal injection of streptozotocin. The rats were divided into a normal group, diabetic group, and FXST group. The rats in the FXST group were treated with FXST by intragastric administration for 12 weeks while other rats were given the same volume of normal saline. The haemodynamic parameters of the central retinal artery in the rats were measured by ultrasound. Haematoxylin-eosin staining was utilised to observe the pathological structural changes in the retina. The apoptosis of retinal nerve cells was detected by terminal deoxynucleotidyl transferase dUTP nick end labelling. RNA sequencing was used to screen the differentially expressed genes (DEGs), and enrichment analyses were performed. The DEGs were validated through real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR). RESULTS: The peak systolic velocity, end diastolic velocity, and mean velocity decreased while the resistance index and pulsatility index increased in the diabetic rat retinas. FXST also improved haemodynamics. In contrast with the diabetic group, FXST allayed the disorder and oedema of the retinal structure in addition to reversing the reductions in retinal thickness and retinal ganglion cell number. It also decreased the apoptosis index of retinal cells. A total of 1134 DEGs were identified by RNA sequencing in the FXST group compared to the diabetic group, including 814 upregulated genes and 320 downregulated genes. These genes were enriched in the complement and coagulation cascades as well as the peroxisome proliferator-activated receptor (PPAR) signalling pathway. Several DEGs, including PPAR gamma, perilipin 4, acyl-CoA dehydrogenase long chain, CD55 molecule, and plasminogen activator urokinase, were identified by qRT-PCR, and the results were consistent with the RNA sequencing data. CONCLUSIONS: FXST alleviates DR by improving the haemodynamics and morphological alterations of diabetic rat retinas, which are mediated by complement and coagulation cascades and the PPAR signalling pathway.

New cytotoxic biflavones from Selaginella doederleinii.

Three new biflavones, apigenin-(3',8″)-chrysin (1), (2S)-2,3-Dihydroametoflavone 5,4'-dimethyl ether (2), and (2S)-5″,7″-Dihydroxy-2″-phenoxychromonyl-(4'″,3')-naringenin (3), together with seven known biflavones (4-10) were isolated from the 75% EtOH extract of Selaginella doederleinii. The structures of new compounds were established by application of spectroscopic methods, including 1D and 2D NMR, HRMS, and CD measurements. In addition, all new compounds were evaluated for their cytotoxic potential against three human cancer cell lines A549, MCF-7, and SMMC-7721 in vitro. Compound 2 exhibited potent cytotoxic activity with IC50 values ranging from 6.35 to 10.18 µM.

HuoXue JieDu formula improves diabetic retinopathy in rats by regulating microRNAs.

ETHNOPHARMACOLOGICAL RELEVANCE: HuoXue JieDu Formula (HXJDF) originates from classical formulas and was formed based on clinical experience. It is composed of Euonymus alatus (Thunb.) Siebold, Panax notoginseng (Burkill) F.H. Chen, the roots of Anguina kirilowii (Maxim.) Kuntze, and Coptis omeiensis (C. Chen) C.Y.Cheng. HXJDF prevents the deterioration of diabetic retinopathy. AIM OF THE STUDY: To evaluate the effects of HXJDF on diabetic retinopathy in rats and investigate the roles of miRNAs in the effects of HXJDF. MATERIALS AND METHODS: A single intraperitoneal injection of streptozotocin (STZ) (65 mg/kg) was used to induce diabetes in rats. Rats were divided into three groups: normal, diabetic, and diabetic + HXJDF. Rats were treated with HXJDF (15.4 g/kg) or water by oral gavage for twelve weeks. At the end of the treatment, rats were anaesthetized, and retinal haemodynamic changes were measured. Then, the retinas were removed and examined by haematoxylin and eosin (HE) staining and TUNEL assays. In addition, miRNA expression profiling was performed using miRNA microarrays and further validated by quantitative real-time PCR (qRT-PCR). RESULTS: Diabetes reduced peak systolic velocity (PSV), end-diastolic velocity (EDV), mean velocity (MV) and central retinal vein velocity (CRV) but increased the resistance index (RI) and pulsatility index (PI). In addition, in the diabetic group, retinal cell arrangement was disordered and loosely arranged, the retinal thickness and retinal ganglion cell (RGC) number decreased, and retinal cell apoptosis increased. In addition, 11 miRNAs were upregulated and 4 miRNAs were downregulated. After treatment, HXJDF improved retinal haemodynamics and morphologic changes, restored retinal thickness and RGC number and decreased retinal cell apoptosis. Furthermore, the changes in miRNA expression were significantly abolished by HXJDF. CONCLUSION: HXJDF may prevent DR by regulating the expression of miRNAs.

New prenylflavonol glycosides with xanthine oxidase inhibitory activity from the leaves of Cyclocarya paliurus.

Eight new prenylflavonol glycosides (1-8), along with five known analogues (9-13) were isolated from the n-butanol extract of the dried leaves of Cyclocarya paliurus (family Juglandaceae) for the first time. The structures of these compounds were characterized by comprehensive analysis of 1D, 2D NMR, HRESIMS, UV data and acid hydrolysis. In bioassay, all these thirteen prenylflavonol glycosides exhibited inhibitory effects on xanthine oxidase (XOD) activity. Especially compounds 2 and 7, showed outstanding IC50 values of 31.81 ± 2.20 and 29.71 ± 3.69 µM, respectively.

Brachyanins A-C, pinene-derived meroterpenoids and phloroglucinol derivative from Leptospermum brachyandrum.

A pair of epimer brachyanins A (1) and B (2), along with a new phloroglucinol brachyanin C (3), were isolated from the leaves of Leptospermum brachyandrum. Brachyanins A (1) and B (2) were the first example of novel meroterpenoid with a unique skeleton that combined a synacrpic acid and a pinene units via a benzyl moiety. Their structures were elucidated through the application of extensive spectroscopic measurements and single-crystal X-ray diffraction analysis and with the absolute configurations of 1 and 2 were confirmed by the quantum chemical CD calculation. The hetero Diels-Alder as the key biotransformation was proposed to account for the biosynthesis of brachyanins A and B sheding light by the potential procursor brachyanin C.

Protective effects of quercetin against chronic mixed reflux esophagitis in rats by inhibiting the nuclear factor-κB p65 and interleukin-8 signaling pathways.

OBJECTIVE: To observe the effects of quercetin on chronic mixed reflux esophagitis (RE) in rats by inhibiting the nuclear factor-κB p65 (NF-κBp65) and interleukin-8 (IL-8) signaling pathways. METHODS: Forty-eight healthy male Sprague-Dawley rats were randomly divided into six groups, with 8 rats in each group: the normal intact group, the sham operation group, the RE control group, the RE group treated with omeprazole or 100 mg/kg and 200 mg/kg quercetin. The animals were sacrificed after 6 weeks of different interventions. The pathological characteristics of esophageal mucosa were observed according to the diagnostic criteria and the expressions of NF-κBp65 and IL-8 were assessed by immunohistochemistry and real-time polymerase chain reaction. RESULTS: Compared with the RE control group, esophageal mucosal injury was improved and the expressions of NF-κBp65 and IL-8 were significantly decreased in the RE group treated with omeprazole or quercetin (P < 0.05). Compared with the omeprazole group, the gross and microscopic scores of esophageal mucosal injury and the expressions of NF-κBp65 and IL-8 in the 100 mg/kg and 200 mg/kg quercetin groups were not increased (P > 0.05). There was no statistically significant difference between the RE groups treated with 100 mg/kg quercetin and 200 mg/kg quercetin. CONCLUSION: Quercetin can prevent esophageal mucosal injury in RE rats by suppressing the NF-κBp65 and IL- 8 signaling pathways.