Effects of schisandrin B on hypoxia-related cognitive function and protein expression in vascular dementia rats.

Vascular dementia (VD) a heterogenous group of brain disorders in which cognitive impairment is attributable to vascular risk factors and cerebrovascular disease. A common phenomenon in VD is a dysfunctional cerebral regulatory mechanism associated with insufficient cerebral blood flow, ischemia and hypoxia. Under hypoxic conditions oxygen supply to the brain results in neuronal death leading to neurodegenerative diseases including Alzheimer's (AD) and VD. In conditions of hypoxia and low oxygen perfusion, expression of hypoxia-inducible factor 1 alpha (HIF-1α) increases under conditions of low oxygen and low perfusion associated with upregulation of expression of hypoxia-upregulated mitochondrial movement regulator (HUMMR), which promotes anterograde mitochondrial transport by binding with trafficking protein kinesin 2 (TRAK2). Schisandrin B (Sch B) an active component derived from Chinese herb Wuweizi prevented ß-amyloid protein induced morphological alterations and cell death using a SH-SY5Y neuronal cells considered an AD model. It was thus of interest to determine whether Sch B might also alleviate VD using a rat bilateral common carotid artery occlusion (BCAO) dementia model. The aim of this study was to examine the effects of Sch B in BCAO on cognitive functions such as Morris water maze test and underlying mechanisms involving expression of HIF-1α, TRAK2, and HUMMR levels. The results showed that Sch B improved learning and memory function of rats with VD and exerted a protective effect on the hippocampus by inhibition of protein expression of HIF-1α, TRAK2, and HUMMR factors. Evidence indicates that Sch B may be considered as an alternative in VD treatment.

Long-time qingyan formula extract treatment exerts estrogenic activities on reproductive tissues without side effects in ovariectomized rats and via active ER to ERE-independent gene regulation.

The reproductive tissues are negatively influenced by estrogens in hormone therapy. Qingyan formula ethanol extract (QYFE)'s estrogenic effects and safety on reproductive tissues after long-term administration and its mechanism via estrogen receptor (ER) pathway haven't been studied. Here, we characterized its estrogenic effects using ovariectomized rats together with in vitro studies for further molecular characterization. Ovariectomized rats were treated with QYFE at doses of 0.7, 1.4, and 2.8g/kg for 12 weeks. The results showed QYFE has a potent estrogenic activity, as indicated by restoring the disappeared estrous cycle, antagonizing the atrophy of uterus, vagina and mammary gland, and the estrogen decline in circulation caused by ovariectomy. In addition, QYFE upregulated ERα and ERß expressions and had a less stimulatory effect on PCNA and ki-67 antigen in reproductive tissues compared with estradiol valerate. QYFE components can bind to ERα and ERß, significantly increased ERα/ß-ERE luciferase reporter gene expression, upregulated the expressions of ERs, PR and pS2 in MCF-7 cells at protein and gene level. All these activities were significantly inhibited by the ER antagonist ICI182,780. QYFE's estrogenic activity maybe mediated by stimulating biosynthesis of estrogen and increasing the quantity of ERs in target tissue and via active ER to ERE-independent gene regulation.

Estrogenic Effect of the Extract of QingYan Formula on Reproductive Tissues in Immature Mice.

A Chinese herbal preparation, QingYan formula (QYF), has been used clinically for kidney-invigorating. However, no evidence base links QYF to estrogen replacement therapy. In this study, the estrogenic effects of QingYan formula 70% ethanol extract (QYFE) were investigated in immature mice. Immature mice were treated with QYFE at doses of 1, 2, and 4g/kg for 7 days. QYFE treatments promoted vaginal cornification and prolonged the estrus status of the immature mice, promoted the growth and development of uterus and vagina, upregulated ERα and ERß expression at protein level in uterus and vagina, increased the level of estradiol (E2), and decreased concentration of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in serum. This study demonstrated that QYFE exerts estrogenic effects by stimulating biosynthesis of estrogen and increasing estrogen receptors (ERs) in target tissues and provided an evidence base for QYFE treatment instead of estrogen replacement therapy.

Incompatibility Mechanism Between Radix Paeoniae Alba and Veratrum nigrum Focusing on Estrogen-Estrogen Receptor Pathway in Immature/Ovariectomized Mice.

Radix paeoniae alba (RPA) and Veratrum nigrum (VN) L. belong to the 18 incompatible medicaments and have been prohibited for thousands of years in China. Previous studies focused on the chemical constituents that induced the toxicological response of the two agents, but this study offers preliminary insight into the pharmacodynamics and mechanism on estrogenic activity, which is responsible for their incompatibility. We undertook a characterization of the interaction on estrogenic activity of RPA and VN using in vivo models of immature and ovariectomized (OVX) mice and in vitro studies focused on estrogen receptor (ER) pathway for further mechanism. VN disturbed the estrogenic efficacy of RPA in promoting development of uterus and vagina in immature mice, and reversing the atrophy of reproductive tissues in OVX mice by decreasing the increase of serum estrogen level and upregulation of ER expression in reproductive tissues by treatment with RPA. Besides, VN antagonized the estrogenic efficacy of RPA in stimulating the binding with ERα and ERß, increasing ERα/ß-estrogen response element (ERE) luciferase reporter gene expression and promoting MCF-7 cell viability. This study provided evidence that VN antagonized the estrogenic efficacy of RPA by decreasing the up-regulations of estrogen biosynthesis in circulation and ERs in target tissues caused by RPA, and through ER-ERE-dependent pathway.

Radix Paeoniae Alba increases serum estrogen level and up-regulates estrogen receptor expression in uterus and vagina of immature/ovariectomized mice.

Radix Paeoniae Alba (RPA) is widely used in clinical treatment for gynecological diseases, particularly abnormal menstruation, menstrual pain, and breast tenderness; however, no scientific evidence base links RPA to estrogen replacement therapy. In this study, we characterize estrogenic activity of RPA using immature and ovariectomized (OVX) mice together with in vitro studies focus on estrogen receptor (ER) pathway for molecular mechanism. RPA treatments demonstrated significant estrogenic activity, as indicated by promoting the development of uterus and vagina in immature mice, reversing the atrophy of uterus and vagina in OVX mice, up-regulating the expressions of ERα and ERß at protein and mRNA level in reproductive tissues. Meanwhile, RPA significantly increased serum estradiol and clearly decreased serum luteinizing hormone and follicle-stimulating hormone of immature/OVX mice. Moreover, RPA could induce ER positive MCF-7 cell from S-phase to G2 stage and induce proliferation and no influence on ER negative MDA-MB-231 cell. RPA could bind with ERα and ERß and significantly stimulate ERα/ß-estrogen response element (ERE) luciferase reporter gene expression. All activities were inhibited by the ER antagonist ICI 182,780. This study illustrates RPA exerts estrogenic effects by stimulating biosynthesis of estrogen in circulation, up-regulating ERs in target tissues, and mimicking the estrogen through ER-ERE-dependent pathway.

Veratrum nigrum inhibits the estrogenic activity of salvia miltiorrhiza bunge in vivo and in vitro.

BACKGROUND: As recorded in the 18 incompatible medicaments of Traditional Chinese Medicine theory, the combined use of Salvia miltiorrhiza bunge (SM) and Veratrum nigrum (VN) could induce toxicity and has been prohibited for thousands of years in China. However, the theory has been validated due to lack of evidence. Previous studies have focused on the chemical constituents that are responsible for the toxicity of the two agents. PURPOSE: This study offers preliminary insight into the pharmacodynamics and mechanism of estrogenic activity responsible for their incompatibility. STUDY DESIGN: We undertook a characterization of the interaction between estrogenic activities of SM and VN using in vivo models of immature and ovariectomized (OVX) mice, and in vitro studies focused on the estrogen receptor (ER) pathway for further mechanism. METHODS: Immature and OVX mice were treated intragastrically with SM at doses of 1.6, 3.2 g/kg, or combine with 0.045 g/kg VN and 0.005 g/kg the ER antagonist ICI182, 780 for elucidating the effects on estrogenic activity in reproductive tissues, E2 secretion, and the ER mechanism. ERα/ß binding experiments and ERα/ß transcriptional activity were performed in order to evaluate the biological action exerted through ERs. RESULTS: VN decreased the estrogenic efficacy of SM in promoting the development of the uterus and vagina in immature mice, and reversing the atrophy of reproductive tissues in OVX mice. VN interfered with the estrogenic efficacy of SM by decreasing the serum estradiol and the upregulation of ERα and ERß expressions in reproductive tissues by treatment with SM. VN antagonized the estrogenic efficacy of SM in promoting the viability of MCF-7 cells and stimulating the binding ability with ERα and ERß, and increasing ERα/ß-estrogen response element (ERE) luciferase activity. CONCLUSIONS: This study provided evidence that the combined use of SM and VN could induce unfavorable effects. VN decreased the estrogenic activity of SM, which might be related to the regulation of estrogen secretion and ERs through the ER-ERE pathway.

[Research progress in phytoestrogens of traditional Chinese medicine].

Phytoestrogens are plant-derived compounds, which have a similarity in structure with human endogenous estrogen 17-ß-estradiol. Structural likeness enables phytoestrogens to interact with estrogen receptors, not simply mimicking the effects of human steroidal estrogen but also exhibiting similar and divergent actions. The global literature relating to phytoestrogen in recent years was systematically summarized in this paper. Chemical compositions of phytoestrogens were mainly flavonoids, coumarins, lignans, terpenoids, steroids, etc., with a character of prevention and treatment of perimenopausal syndrome, osteoporosis, cardiovascular disease, metabolic diseases, cancer, regulation of brain function and other pharmacological effects. The mechanisms of action mainly included classical estrogen receptor pathway, epigenetic effect, activation of 5'-adenosyl-phospho-activated protein kinase, inhibition of kinase, activation of peroxisome proliferator-activated receptor, regulation of apoptosis-related proteins, inhibition of nuclear factor κB signaling pathway and so on. According to their efficacy classification, phytoestrogens were mainly distributed in the tonifying medicines, blood-activating and stasis-resolving medicines and heat-clearing medicines. The classical prescriptions with estrogen activity included tonifying formula, Qi-regulating formula and harmonizing formula, etc. This review was aimed at providing a certain reference for the further study of phytoestrogens by researchers and clinicians.

The influence of Schisandrin B on a model of Alzheimer's disease using ß-amyloid protein Aß1-42-mediated damage in SH-SY5Y neuronal cell line and underlying mechanisms.

Schisandrin B, an active substance, is derived from Chinese herb fruit Wuweizi, which exerts various pharmacological activities and has displayed significant beneficial effects in ameliorating Alzheimer's disease (AD). The aim of this study was to further extend our examination for the use of schisandrin B extract in the potential treatment of AD effects by investigating DNA methylation (DNMT), known to be modified in this disease using SH-SY5Y neuronal cell line exposed to ß-amyloid protein (Aß1-42). In particular, the purpose of this investigation was to examine alterations in mRNA and protein expression of DNMT. Data demonstrated that schisandrin B blocked Aß1-42-mediated injury in SH-SY5Y neuronal cell line as evidenced by a restoration of cellular morphology and cell viability to approximate control levels at the highest 10 µg/ml Schisandrin B. Incubation with Aß1-42 significantly decreased mRNA and protein expression of DNMT3A and DNMT1 in SH-SY5Y neuronal cell line. Incubation with Aß1-42 followed by 24 treatment with schisandrin B significantly inhibited the Aß1-42 -induced changes in mRNA and protein expression of DNMT3A and DNMT3B in a concentration-dependent manner. It is of interest that the mRNA expression of DNMT3A and DNMT1 were significantly higher than control. Data thus indicate schisandrin B was effective in inhibiting the actions of Aß1-42 on cell survival and morphology and that DNA methylation may be associated with the beneficial findings.

Salvia miltiorrhiza bunge increases estrogen level without side effects on reproductive tissues in immature/ovariectomized mice.

Salvia miltiorrhiza bunge(SM) is a popular herb for alleviating menopausal symptoms, although the scientific evidence of applying SM to estrogen replacement therapy is limited. In this study, we characterized the estrogenic activity of SM using in vivo models of immature and ovariectomized (OVX) mice and performed in vitro studies focusing on the estrogen receptor (ER) pathway for further molecular characterizations. SM treatments demonstrated significant estrogenic activity by promoting the development of uterus and vagina in immature mice, restoring the estrus cycle and reversing the atrophy of reproductive tissues in OVX mice, as well as increasing the expressions of ERα and ERß at protein and mRNA level in the reproductive tissues. Meanwhile, SM significantly increased estradiol in serum, and decreased follicle-stimulating hormone (FSH) and luteinizing hormone (LH) in the circulation of immature and OVX mice. SM could stimulate the binding effect of ERα and ERß, and significantly induce ERα/ß-estrogen response element (ERE) luciferase reporter gene expression. All these activities were inhibited by the ER antagonist ICI182, 780. This study demonstrates SM exerts estrogenic effects by stimulating biosynthesis of estrogen and increasing ERs in target tissues without side effects on reproductive tissues and through ER-ERE-dependent pathway.

[Effect of Coixenolide on Foxp3+ CD4+ CD25+ Regulatory T Cells in Collagen-induced Arthritis Mice].

OBJECTIVE: To study the effect of coixenolide on Foxp3+ CD4+ CD25+ regulatory T cells (Treg) in collagen induced arthritis (CIA) mice, and to explore its possible mechanism for treating rheumatiol arthritis. METHODS: Five mice were recruited as a normal control group from 25 mice, and the rest 20 were used in CIA modeling. After successful modeling they were randomly divided in the model control group and the coixenolide group, 10 in each group. Coixenolide injection at 25 mL/kg was intraperitoneally injected to mice in the coixenolide group, while normal saline at 25 mL/kg was intraperitoneally injected to mice in the normal control group and the model control group. The injection lasted for 21 days. Scoring for CIA was performed after injection and arthritis index was calculated. The peripheral blood Foxp3+ CD4+ CD25+ Treg ratio was determined by flow cytometry (FCM). RESULTS: Compared with the normal control group, the arthritis index obviously increased in the model control group (P < 0.01). The arthritis index obviously decreased more in the coixenolide group than in the model control group (P < 0.01). Foxp3+ CD4+ CD25+ Treg levels obviously decreased more in the model control group than in the normal control group (P < 0.01 ). Foxp3+ CD4+ CD25+ Treg levels obviously increased more in the coixenolide control group than in the model control group (P < 0.01). CONCLUSION: Coixenolide could up-regulate Foxp3+ CD4+ CD25+ Treg ratios in CIA mice, which might play certain immunoregulation roles in the incidence of CIA.

Identification and characterization of a novel gene, c1orf109, encoding a CK2 substrate that is involved in cancer cell proliferation.

BACKGROUND: In the present study we identified a novel gene, Homo Sapiens Chromosome 1 ORF109 (c1orf109, GenBank ID: NM_017850.1), which encodes a substrate of CK2. We analyzed the regulation mode of the gene, the expression pattern and subcellular localization of the predicted protein in the cell, and its role involving in cell proliferation and cell cycle control. METHODS: Dual-luciferase reporter assay, chromatin immunoprecipitation and EMSA were used to analysis the basal transcriptional requirements of the predicted promoter regions. C1ORF109 expression was assessed by western blot analysis. The subcellular localization of C1ORF109 was detected by immunofluorescence and immune colloidal gold technique. Cell proliferation was evaluated using MTT assay and colony-forming assay. RESULTS: We found that two cis-acting elements within the crucial region of the c1orf109 promoter, one TATA box and one CAAT box, are required for maximal transcription of the c1orf109 gene. The 5' flanking region of the c1orf109 gene could bind specific transcription factors and Sp1 may be one of them. Employing western blot analysis, we detected upregulated expression of c1orf109 in multiple cancer cell lines. The protein C1ORF109 was mainly located in the nucleus and cytoplasm. Moreover, we also found that C1ORF109 was a phosphoprotein in vivo and could be phosphorylated by the protein kinase CK2 in vitro. Exogenous expression of C1ORF109 in breast cancer Hs578T cells induced an increase in colony number and cell proliferation. A concomitant rise in levels of PCNA (proliferating cell nuclear antigen) and cyclinD1 expression was observed. Meanwhile, knockdown of c1orf109 by siRNA in breast cancer MDA-MB-231 cells confirmed the role of c1orf109 in proliferation. CONCLUSIONS: Taken together, our findings suggest that C1ORF109 may be the downstream target of protein kinase CK2 and involved in the regulation of cancer cell proliferation.

Knockdown of Rab5a expression decreases cancer cell motility and invasion through integrin-mediated signaling pathway.

BACKGROUND: Rab GTPases function as modulators in intracellular transport. Rab5a, a member of the Rab subfamily of small GTPases, is an important regulator of vesicle traffic from the plasma membrane to early endosomes. Recent findings have reported that Rab5a gene was involved in the progression of cancer. In the present study, we investigated the effect of Rab5a on cervical cancer invasion and metastasis and the molecular mechanism underlying the involvement of Rab5a. METHODS: Rab5a expression was assessed by immunohistochemical analysis on a cervical cancer tissue microarray. RNA interference (RNAi) was performed to knock down the endogenous expression of Rab5a gene in HeLa and SiHa cells. Cell motility was evaluated using invasion assay and wound migration assay in vitro. The expression levels of integrin-associated molecules were detected by Western blot and immunofluorescence. RESULTS: We found that Rab5a was expressed at a high level in cervical cancer tissues. Silencing of Rab5a expression significantly decreased cancer cell motility and invasiveness. The down-regulation of integrin-associated focal adhesion signaling molecules was further detected in Rab5a knockdown cells. Meanwhile, active GTP-bound Rac1, Cdc42, and RhoA were also down-regulated, accompanied with the reduction in the number and size of filopodia and lamellipodia. CONCLUSIONS: Taken together, these data suggest that Rab5a functions in regulating the invasion phenotype, and we propose that this regulation may be via integrin-mediated signaling pathway in cervical cancer cells.

Regulatory effect of Langchuang serial recipes on T-lymphocyte subsets Th and Tc in patients with systemic lupus erythematosus.

OBJECTIVE: To study the principle of clearing Fei (), cooling blood, and detoxification as well as nourishing yin and moisening Fei (abbr. as CCD-NM) in regulating the levels of peripheral T-lymphocyte subsets Th and Tc cells to explore its mechanism for lowering the incidence of infection in patients with systemic lupus erythematosus (SLE). METHODS: Sixty SLE patients without complicated infection were assigned to the treatment group and the control group, 30 in each group. The control group was treated with Western medicine alone, while the treatment group was treated with the same program of Western medicine, but additionally administered with either Langchuang No.1 (I) or 2 (II), serial concerted Chinese recipes, applied respectively in patients in the active stage or in the resting stage. The total time of treatment for both groups was 1 year. Further, a healthy control group was set up with 20 healthy subjects. The expressions of Th1, Th2, and Tc1 and Tc2 cells in peripheral blood were detected and compared with those in the healthy control group. RESULTS: (1) As compared with the healthy control group, ratios of Th1/Th2 and Tc1/Tc2 in SLE patients, whether complicated with infection or not, were significantly lower (P<0.05 or P<0.01). (2) Comparison between patients with complications and those uncomplicated with infection showed that the two ratios and Th1 expression were lower and Tc2 was higher in the former than those in the latter (all P<0.05). (3) Ratios of Th1/Th2 and Tc1/Tc2 increased after treatment in patients of both the treatment group and the control group (P<0.05 and P<0.01), but the changes in the treatment group were more significant (P<0.05). CONCLUSION: The principle of CCD-NM could regulate the Th and Tc subsets toward equilibrium in SLE patients, which might be one of the mechanisms of action for alleviating complicated infection.

[Clinical research on effect of TCM treatment beginning with Fei in treating systemic lupus erythematosus for reducing incidence of complicated infection].

OBJECTIVE: To observe the effect of TCM treatment beginning with Fei in reducing the incidence of complicated infection and the antibiotic utilization rate in patients with systemic lupus erythematosus (SLE). METHODS: One hundred and ten SLE patients were randomly assigned to 2 groups equally, the control group treated with the conventional Western medicinal treatment and the treated group treated with the same conventional treatment and SLE I formula (in active stage) or SLE II formula (in silent period) additionally. RESULTS: After 3-month and 6-month treatment, the total effective rate was 83.64% , 87.27% in the treated group, and 78.18%, 81.82% in the control group respectively, showing insignificant difference between the two groups. It lowered in both groups after 1-year treatment, however, which in the treated group (78.18%) was higher than that in the control group (60.00%, P < 0.05). But the difference became insignificant again after 2-year treatment, it being 87.27% in the treated group and 72.73% in the control group. The incidence of complicated infection and antibiotic utilization rate in the 2-year treatment was 23.6%, 55.0% respectively in the treated group, markedly lower than those (50.9% and 100%) in the control group respectively (P < 0.01). CONCLUSION: TCM treatment beginning with Fei could decrease the incidence of complicated infection and the antibiotic utilization rate in SLE patients.

[Observation on therapeutic effect of acupuncture and moxibustion in 60 cases of Sjogren syndrome].

OBJECTIVE: To observe clinical therapeutic effect of acupuncture on Sjogren syndrome. METHODS: Treatment group (n=60) were treated with acupuncture of clearing away dryness and toxic substance and removing obstruction in collaterals at Quze (PC 3), Taichong (LR 3), Xuehai (SP 10), Sanyinjiao (SP 6) and Taixi (KI 3), and the control group (n=60) with prednisone. Their therapeutic effects were compared. RESULTS: The total effective rate was 73.3% in the treatment group and 56.7% in the control group with a significant difference between the two groups (P < 0.05); there was no reverse effect in the treatment group. CONCLUSION: The needling method for clearing dryness and removing obstruction in collaterals is effective and safe for treatment of Sjogren syndrome.

Interleukin-10 production genotype protects against acute persistent rejection after lung transplantation.

BACKGROUND: Our previous studies demonstrated that cytokine gene polymorphisms are related to acute rejection in pediatric heart transplantation; a decreased tumor necrosis factor (TNF)-alpha production genotype combined with an increased or intermediate interleukin (IL)-10 production genotype was associated with the smallest incidence of acute rejection. The objective of this study was to determine whether cytokine genotypes TNF-alpha, IL-10, IL-6, interferon-gamma, and transforming growth factor beta were associated with acute persistent rejection after lung transplantation. METHODS: Cytokine genotyping was performed in 119 adult lung transplantation recipients who underwent surveillance transbronchial biopsies during their first year after transplantation. We categorized recipients with acute persistent rejection if they had 2 consecutive biopsy specimens at >/=Grade A2 despite anti-rejection treatment. We performed cytokine genotyping using the polymerase chain reaction-sequence specific primers technique, with a commercially available kit. RESULTS: We analyzed the IL-10 genotype in 116 patients. For the increased IL-10 production genotype, 7 of 20 patients (35%) were persistent rejecters. In comparison, 57 of 96 patients (59%) with intermediate or decreased IL-10 production genotype had acute persistent rejection (p = 0.046). For IL-10 haplotypes associated with intermediate IL-10 production, 30 of 45 patients with GCC/ACC haplotype (67%) had acute persistent rejection compared with 10 of 22 patients with GCC/ATA (45%). In the patients with intermediate IL-10 production, 17 of 22 (77%) with IL-10 GCC/ACC and IL-6 G/C had acute persistent rejection, whereas only 2 of 7 patients (29%) with IL-10 GCC/ATA and IL-6 G/G had acute persistent rejection (p = 0.018). CONCLUSIONS: In lung transplant recipients, the increased IL-10 production genotype protects against acute persistent rejection when compared with the intermediate or decreased IL-10 production genotypes. The intermediate IL-10 production genotype in lung transplant recipients can be differentiated into 2 haplotype responses, with the GCC/ACC haplotype associated more with acute persistent rejection. In lung transplant recipients, the immunomodulatory effects of IL-6 are differentiated in the G/C and G/G alleles in conjunction with IL-10 haplotypes, with G/C being associated with more acute persistent rejection in conjunction with the IL-10 GCC/ACC haplotype. Future pharmacogenomic models may incorporate these associations with acute persistent rejection in lung transplant recipients to formulate individualized therapeutic regimens.