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Purpose: To investigate the presence and change of nerve fibers and neuropeptide during early development of articular cartilage in neonatal rats. Methods: Articular cartilage in distal-femoral epiphyses was collected from neonatal Sprague Dawley rats, which were 1-day, 5-day, and 10-day postnatal (P1, P5 and P10). Microscopy, immunofluorescence, transmission and scanning electron microscopy (TEM and SEM) were performed for detection of nerve fibers. Quantitative analysis for substance P (SP) and neuropeptide Y (NPY) was conducted using immunofluorescence and enzyme-linked immunosorbent assay (ELISA). Results: TEM showed the existence of myelinated nerve fibers in the extracellular matrix of articular cartilage in both P1, P5 and P10 rats, and they formed synaptic contacts with chondrocytes. During this time, chondrocytes proceeded with their development, and the nerve fibers gradually degraded. The ELISA results showed significant increase of the sensory neuropeptide SP and the sympathetic neuropeptide NPY in the cartilage tissue. Immunofluorescence results showed the distribution of SP and NPY in the perichondrium, the cartilage canals, the plasma of chondrocytes, and extracellular matrix in the cartilage tissue. Conclusions: Nerve fibers exist in the matrix of articular cartilage during early development of knee joints in neonatal rats. Nerve fibers form synaptic contacts with chondrocytes at the early stage and then degrade gradually in the course of chondrocyte development. SP and NPY significantly increase in articular cartilage during this very period. These results indicate that the nerve fibers and the neuropeptide they secrete may exert important effect on the development of articular cartilage. [ABSTRACT FROM AUTHOR]

Background: Mechanical loading has been widely considered to be essential for growth plate to maintain metabolism and development. Cyclic mechanical strain has been demonstrated to induce autophagy, whereas the relationship between cyclic tensile strain (CTS) and autophagy in growth plate chondrocytes (GPCs) is not clear. The objective of this study was to investigate whether CTS can regulate autophagy in GPCs in vitro and explore the potential mechanisms of this regulation. Methods: The 2-week-old Sprague–Dawley rat GPCs were subjected to CTS of varying magnitude and duration at a frequency of 2.0 Hz. The mRNA levels of autophagy-related genes were measured by RT-qPCR. The autophagy in GPCs was verified by transmission electron microscopy (TME), immunofluorescence and Western blotting. The fluorescence-activated cell sorting (FACS) was employed to detect the percentage of apoptotic and necrotic cells. Results: In GPCs, CTS significantly increased the mRNA and protein levels of autophagy-related genes, such as LC3, ULK1, ATG5 and BECN1 in a magnitude- and time-dependent manner. There was no significant difference in the proportion of apoptotic and necrotic cells between control group and CTS group. The autophagy inhibitors, 3-methyladenine (3MA) and chloroquine (CQ) reversed the CTS-induced autophagy via promoting the formation of autophagosomes. Cytochalasin D (cytoD), an inhibitor of G-actin polymerization into F-actin, could effectively block the CTS-induced autophagy in GPCs. Conclusion: Cyclic mechanical strain with high-tensile triggers autophagy in GPCs, which can be suppressed by 3MA and CQ, and cytoskeletal F-actin microfilaments organization plays a key role in chondrocytes' response to mechanical loading. [ABSTRACT FROM AUTHOR]

The study is aimed at investigating the changes in expressions of heat shock protein 27 (HSP27), HSP70, and soluble glycoprotein (SGP) in heart failure (HF) rats complicated with pulmonary edema and exploring their potential correlations with cardiopulmonary functions. The rat model of HF was established, and the rats were divided into HF model group (model group, n = 15) and normal group (n = 15). After successful modeling, MRI and ECG were applied to detect the cardiac function indexes of the rats. The myocardial function indexes were determined, the injury of myocardial tissues was observed via hematoxylin and eosin (HE) staining, and the content of myeloperoxidase (MPO), matrix metalloproteinase-9 (MMP-9), and tumor necrosis factor-alpha (TNF-α) in the blood was measured. The partial pressure of oxygen (PaO2) and oxygenation index (OI) were observed, and the airway resistance and lung compliance were examined. Moreover, quantitative polymerase chain reaction (qPCR) and Western blotting assay were performed to detect the gene and protein expression levels of HSP27, HSP70, and SGP130. The levels of serum creatine kinase (CK), creatine (Cr), and blood urea nitrogen (BUN) were increased markedly in model group (p < 0.05). Model group had notably decreased fractional shortening (FS) and ejection fraction (EF) compared with normal group (p < 0.05), while the opposite results of left ventricular end-diastolic diameter (LVEDD) and left ventricular end-systolic diameter (LVESD) were detected. In model group, the content of serum MPO, MMP-9, and TNF-α was raised remarkably (p < 0.05), OI and PaO2 were reduced notably (p < 0.05), the airway resistance was increased (p < 0.05), and the lung compliance was decreased (p < 0.05). Obviously elevated gene and protein expression levels of HSP27, HSP70, and SGP130 were detected in model group (p < 0.05). The expressions of HSP27, HSP70, and SGP130 are increased in HF rats complicated with pulmonary edema, seriously affecting the cardiopulmonary functions of the rats. [ABSTRACT FROM AUTHOR]

Objectives. To study the protective effects and mechanisms of Huoxue Jiedu Huayu formula on cell pyroptosis through the NLRP3/caspase-1/IL-1β pathway in contralateral kidneys in 6-month-old unilateral ureteral obstruction (UUO) rats. Methods. Wistar rats were randomly assigned to 5 groups: a Sham group, a unilateral nephrectomy group (UNX group), a UUO group, a UUO treated with spironolactone group (Spi group), and a UUO treated with Huoxue Jiedu Huayu formula group (HJHF group). After 6 months of oral drug intervention, blood and contralateral kidneys were collected for research. Results. The morphology and function of the contralateral kidneys were essentially normal after unilateral nephrectomy. HJHF obviously decreased serum creatinine, urea, and inflammatory lesions and depressed cell pyroptosis based on the NLRP3/caspase-1/IL-1β pathway. Moreover, spironolactone, a mineralocorticoid receptor (MR) blocker, suppressed cell pyroptosis through SGK-1 and NF-кB. Conclusion. HJHF and spirolactone inhibited excessive activation of MR and then reduced cell pyroptosis, which was dependent on the NLRP3/caspase-1/IL-1β pathway, to protect the contralateral kidneys of 6-month-old UUO rats. [ABSTRACT FROM AUTHOR]

Ethnopharmacological relevance Euonymus alatus, Radix trichosanthis, Panax notoginseng and Coptis chinensis are popular plants used in traditional Chinese medicine to treat diabetes. Aim of the study The aim of the study is to investigate the therapeutic effect of the active components of Euonymus alatus, Radix trichosanthis, Panax notoginseng and Coptis chinensis (cERPC) on diabetic peripheral neuropathy in the rats and explore the underlying mechanism involved. Methods After diabetes was induced in rats for 20 weeks, cERPC or water was administered for 12 weeks. After a hot plate test, motor nerve conduction velocity and sciatic nerve blood flow were determined; the sciatic nerves were isolated for toluidine blue staining; and the fibre area, fibre diameter, axon area, axon diameter and myelin thickness were evaluated. The levels of the myelin basic protein, myelin protein zero, Oct6 and Krox20 were measured by western blot or immunofluorescence. Results cERPC was efficient in reducing the response latency, increasing motor nerve conduction velocity, enhancing sciatic nerve blood flow and ameliorating the pathological changes in diabetic rats. cERPC also had a role in increasing the levels of myelin basic protein and myelin protein zero and improving the expression of Oct6 and Krox20 in sciatic nerves of diabetic rats. Conclusions cERPC ameliorates diabetic peripheral neuropathy by attenuating electrophysiological, circulatory and morphological alterations, which is mediated by the Oct6-Krox20 pathway. [ABSTRACT FROM AUTHOR]

Celastrus orbiculatus Thunb., an important folk medicine, has long been used for the treatment of rheumatoid arthritis and its ethyl acetate extract (COE) has been reported to possess anticancer, antiinflammation and antiarthritic effects. However, the therapeutic effect and mechanism of COE treatment in rheumatoid arthritis has been rarely studied especially from the perspective of metabolomics. To reveal the therapeutic effects of COE on adjuvant–induced arthritis (AIA) rats through histopathological analysis, non-targeted metabolomics, and molecular docking study. Forty-three Wistar rats were randomly divided into normal group, AIA model group, methotrexate group, and COE groups (80 mg/kg, 160 mg/kg and 320 mg/kg of ethyl acetate extract). Paw swelling and arthritis score were monitored through the experiment. Serum levels of tumor necrosis factor α (TNF-α) and nitric oxide were determined and histopathological evaluation was performed. Furthermore, Ultra-high performance liquid chromatography-linear trap quadrupole-Orbitrap-based metabolomics was employed to characterize metabolic changes of AIA rats after COE treatment and molecular docking was performed to predict the potential phytochemicals of COE against TNF-α. COE at three dosages could significantly relieve paw swelling and reduce arthritis scores of AIA rat. Histopathological analysis revealed remarkable decrease in synovial inflammation and bone erosion after COE treatment, especially at middle and high dosage. Additionally, COE down-regulated serum levels of TNF-α and nitric oxide. Serum metabolomics showed that 22 potential biomarkers for the COE treatment of AIA rats were identified, which were closely related to fatty acid metabolism, glycerophospholipid catabolism, and tryptophan metabolism. The molecular docking models predicted that olean-type triterpenes in COE may contribute most to therapeutic effects of rheumatoid arthritis through targeting TNF-α. COE could significantly relieve the arthritic symptoms in AIA rats and the ultra-high performance liquid chromatography-mass spectrometry based metabolomics proved to be an efficient method to characterize subtle metabolic changes of AIA rats after COE treatment. [Display omitted] [ABSTRACT FROM AUTHOR]

The Qing' E Formula (QEF) is a compound preparation that was originally recorded in the 'Prescriptions of the Bureau of Taiping People's Welfare Pharmacy' during the Song Dynasty (10th century CE). It consists of four Chinese medicinal herbs, Eucommiae Cortex (Eucommia ulmoides), Psoraleae Fructus (Psoralea corylifolium), Juglandis Semen (Juglans regia), and Garlic Rhizoma. According to traditional Chinese medicine (TCM), QEF has the ability to tonify the kidney and strengthen muscle and bone. According to the 'kidney governing bone' theory in TCM, QEF is also used to treat the symptoms of climacteric syndrome, especially osteoporosis caused by reduced production of estrogen during the perimenopausal period; however, the therapeutic roles of the individual components of the QEF and their compatibility within the formula has not been investigated. In this study, the compatibility mechanism and estrogen-like action properties of the four herbal components in the QEF was elucidated according to the organizing principle of Chinese medicine formulas using both in vitro and in vivo models. The estrogen-like effects of QEF and its herbal components were investigated in MCF7 and HEK293 cells as well as ovariectomized (OVX) rats. The estrogen-like effects of the QEF and its components were analyzed in vitro using Cell Counting Kit-8 and Luciferase reporter gene assays. In the in vivo studies, the blood plasma levels of hormones, lipids, neurotransmitters, aromatase, superoxide dismutase (SOD), and malondialdehyde (MDA) were measured through enzyme-linked immunosorbent assays (ELISAs). The histological morphologies of the target organs after exposure to QEF were investigated by HE staining and immunohistochemical methods. The expression levels of estrogen pathway-related proteins and genes in the OVX rats were measured by Western blotting and real time quantitative PCR (RT-qPCR), respectively. The in vitro results showed that the QEF, Eucommia (EC) and Psoralea (PF) promoted the proliferation of MCF-7 cells and upregulated the expression of ERα, ERβ and pS2 genes in the MCF-7 cells. Notably, the QEF demonstrated the most active estrogen-like effects compared to the individual ingredients. The in vivo results showed that the QEF, EC, and PF increased the uterine coefficient, upregulated the expression of both ERs (ERα and ERβ) in the uterus, and increased blood serum hormone levels. QEF and its individual components ameliorated menopausal-derived lipid metabolism dysfunction, increased neurotransmitter production by stimulating the adrenal glands, enhanced the antioxidant activity in the serum by increasing the concentration of SOD, reversed ovariectomy-derived atrophy in the uterus, and reduced the weight gain associated with estrogen reduction in the OVX rats. The QEF also antagonize the loss of appetite of OVX animals caused by feeding Psoralea alone, which could explain the compatibility mechanism of Qing' E Formula with reducing toxicity and increasing efficiency. The estrogen-like effects of Eucommia and Psoralea were mainly mediated through activation of ERα and ERβ. The phytoestrogen components regulated hormone production and the expression of related proteins and genes, which indicated that these components exhibited estrogen-like therapeutic effects. However, the QEF showed the greatest estrogen-like effects compared to the individual components. Overall, this corroborated the therapeutic prowess of the QEF and clarified the pharmacodynamic interactions between the different components extracts in the QEF. [Display omitted] • The compatibility of the herbs in QEF was investigated and QEF is more effective than a single drug. • QEF, Psoraleae and Eucommia have significant estrogenic activities in vivo and vitro. • The function may be effected by classical ER pathway. [ABSTRACT FROM AUTHOR]

BabaoDan (BBD) is a famous traditional Chinese formula frequently used in TCM clinics to eliminate jaundice and treat infectious viral hepatitis. This paper assesses BBD's preventive and therapeutic effects on hepatic encephalopathy after liver cirrhosis (CHE) and acute liver failure (AHE) in rats and explains its possible mechanism of action. CHE rat model was established by injection of carbon tetrachloride (CCl4) twice a week for a total of 9 weeks and then by injection of thioacetamide (TAA) to induce hepatic encephalopathy. AHE rat model was established by injection of TAA once a day for a total of 3 days. In CHE rat model, BBD was gavaged once a day at the end of the 6th week until the experiment ended. In AHE rat model,BBD was gavaged once a day 3 days before TAA injection until the experiment ended. The preventive and therapeutic effects of BBD on brain dysfunction, as well as liver injury, pathology and fibrosis were evaluated in vivo. The role of BBD in the regulation of inflammatory factors and myeloid differentiation factor 88/Toll-like receptor 4/nuclear factor kappa-B (TLR4/MyD88/NK-κ B) pathway was detected in both liver and brain in vivo. The rat bone marrow derived macrophages (BMDMs) were activated by Lipopolysaccharide (LPS), and the role of BBD in the regulation of inflammatory factors and NK-κ B pathway were detected in vitro. In CHE rat model: BBD significantly improved the total distance as well as the activity rate of rats. BBD also improved the learning and memory abilities of rats compared with the control group. In addition, BBD effectively decreased ammonia levels and significantly decreased the levels of alanine aminotransferase (ALT), aspartate transaminase (AST), total bilirubin (TBil) and total bile acid (TBA), as well as improved the levels of total protein (TP) and albumin (Alb). In the liver, BBD not only inhibited the gene expressions of tumor necrosis factor alpha (TNF-α), interleukini-6 (IL-6), TLR4, MyD88, and NF-κ B but also inhibited the protein expressions of TLR4, MyD88, NK-κ B and TNF-α. In the brain, BBD inhibited the gene expressions of iNOS, IL-6, TNF-α, TLR-4, MyD88, and NF-κ B, as well as inhibited the protein expressions of TLR4, MyD88, P65 TNF-α and ionized calcium binding adapter molecule 1 (Iba-1). BBD also decreased NO and TNF-α in the blood. BBD improved neurological scores, blood ammonia levels and the brain inflammatory gene expressions of iNOS, TNF-α and IL-1β. BBD also improved liver function biomarkers such as ALT, TBil, TBA, TP, ALB and inflammatory and apoptotic gene expressions of TNF-α, IL-1β, IL-6, Bax, Bcl-2, caspase-9, caspase-3 and NF-κ B. In LPS-activated rat BMDMs, BBD decreased NO and TNF-α production in BMDM culture supernatant. In addition, BBD inhibited the gene expressions of TNF-α, IL-1 β and IL-6 as well as the phosphorylation of P65. BBD can prevent and cure hepatic encephalopathy (HE) derived from both chronic and acute liver diseases. BBD can reduce hyperammonemia as well as the systematic and neurological inflammation. Inflammation is likely an important target of BBD to treat HE. The anti-inflammatory role of BBD may lie in its regulation of the TLR4/MyD88/NF-κ B pathways. Image 1 [ABSTRACT FROM AUTHOR]