Clustering of gout-related comorbidities and their relationship with gout flares: a data-driven cluster analysis of eight comorbidities.

OBJECTIVES: To study the aggregation of multiple comorbidities in people with gout and explore differences in prognosis of gout flares among different subgroups. METHODS: Hierarchical clustering was performed to identify homogeneous subgroups among 2639 people with gout using eight comorbidities. A one-year follow-up of acute gout flares in 463 of these people was conducted; the incidence and the timing of gout flares in each cluster were assessed to explore prognosis of gout flares. Binary logistic regression was applied to assess factors associated with gout flares. RESULTS: In baseline study, we identified five subgroups (C1-C5). C1 (n = 671, 25%) was characterized by isolated gout with few comorbidities. C2 (n = 258, 10%) were all obese. Almost all people in C3 (n = 335, 13%) had diabetes (99.7%). All people in C4 (n = 938, 36%) had dyslipidemia. C5 (n = 437, 17%) had the highest proportion of cardiovascular disease (CVD, 53%), chronic kidney disease (CKD, 56%), and cancer (7%). In follow-up study, C5 had the highest incidence (71.9%) and earliest onset (median 3 months) of gout flares. C2 had the lowest incidence (52.1%) and the latest onset (median 10 months) of gout flares. The highest relative risk for gout recurrent was seen for C5 (OR = 2.09). Other factors associated with the risk of gout flares were age at diagnosis of gout, duration of gout, presence of tophi, and smoking ≥ 20 cigarettes/day. CONCLUSIONS: We clustered people with gout into five groups with varying comorbidities. People with CVD, CKD, and cancer had the highest risk of gout flares and should receive comprehensive care.

Zhuyu Annao decoction promotes angiogenesis in mice with cerebral hemorrhage by inhibiting the activity of PHD3.

Some traditional Chinese decoctions, such as Zhuyu Annao, exert favorable therapeutic effects on acute cerebral hemorrhage, hemorrhagic stroke, and other neurological diseases, but the underlying mechanism remains unclear. This study aimed to determine whether Zhuyu Annao decoction (ZYAND) protects the injured brain by promoting angiogenesis following intracerebral hemorrhage (ICH) and elucidate its specific mechanism. The effect of ZYAND on the nervous system of mice after ICH was explored through behavioral experiments, such as the Morris water maze and Rotarod tests, and its effects on oxidative stress were explored by detecting several oxidative stress markers, including malondialdehyde, nitric oxide, glutathione peroxidase, and superoxide dismutase. Real-time quantitative RT-PCR and WB were used to detect the effects of ZYAND on the levels of prolyl hydroxylase domain 3 (PHD3), hypoxia-inducible factor-1α (HIF-1α), and vascular endothelial growth factor (VEGF) in the brain tissues of mice. The effect of ZYAND on the NF-κB signaling pathway was detected using a luciferase reporter gene. A human umbilical cord vascular endothelial cell angiogenesis experiment was performed to determine whether ZYAND promotes angiogenesis. The Morris water maze test and other behavioral experiments verified that ZYAND improved the neurobehavior of mice after ICH. ZYAND activated the PHD3/HIF-1α signaling pathway, inhibiting the oxidative damage caused by ICH. In angiogenesis experiments, it was found that ZYAND promoted VEGF-induced angiogenesis by upregulating the expression of HIF-1α, and NF-κB signaling regulated the expression of HIF-1α by inhibiting PHD3. ZYAND exerts a reparative effect on brain tissue damaged after ICH through the NF-κB/ PHD3/HIF-1α/VEGF signaling axis.

Cell death induced by ent-11alpha-hydroxy-15-oxo-kaur-16-en-19-oic-acid in anaplastic thyroid carcinoma cells is via a mitochondrial-mediated pathway.

The chemical compound ent-11alpha-hydroxy-15-oxo-kaur-16-en-19-oic-acid (5F), isolated from the Chinese herbal medicine plant Pteris semipinnata L, has been known to exert antitumor activity. However, the molecular mechanism of the action is not understood. In this study we demonstrated that apoptotic cell death induced by 5F in FRO cells was concentration- and time-dependent. The rapid increase in intracellular reactive oxygen species (ROS) levels was involved in the mechanism of cell death. c-Jun N-terminal kinase (JNK) activation and G2 block were related to cell death induced by 5F. Extracellular signal-related kinase (ERK) and p38 were also activated, but as survival signals in response to 5F treatment to counteract the induction of cell death. In the process of the induction of apoptotic cell death, Bax translocated into mitochondria, a reduction in Delta psi(m) was observed and a release of cytochrome c and apoptosis inducing factor (AIF) from mitochondria into the cytosol occurred, indicating that cell death induced by 5F was through a mitochondrial-mediated pathway.

Over-expression of Bcl-2 against Pteris semipinnata L-induced apoptosis of human colon cancer cells via a NF-kappa B-related pathway.

Ent-11 alpha-hydroxy-15-oxo-kaur-16-en-19-oic-acid (5F), an antitumor component, is a chemical compound isolated from Pteris semipinnata L (PsL), a Chinese traditional herb. We examined whether 5F could affect apoptosis in human colon cancer HT-29 cells, and test whether and how the over-expression of Bcl-2 and Bcl-xL could offset the effect of 5F on cell growth. The result demonstrated that 5F significantly induced apoptosis of HT-29, as shown by MTT assay and DNA fragmentation measurement. Treatment of HT-29 with 5F increased both p38 and iNOS levels, suggesting these two molecules may contribute to the apoptotic effect of 5F. Over-expression of Bcl-2 or Bcl-xL attenuated the increase of p38 and iNOS induced by 5F. The cells with Bcl-2 or Bcl-xL over-expression showed an elevation of nuclear factor kappa B (NF-kappa B) activity, accompanying a significant reduction of 5F-induced apoptosis. Furthermore, inhibition of NF-kappa B by I k B alpha SR, which is a powerful inhibitor of NF-kappa B, restored the ability of 5F to induce apoptosis in the cells transfected with Bcl-2. These data strongly indicated that the apoptotic effect of 5F on HT-29 was closely associated with the activity of NF-kappa B, which was up-regulated by Bcl-2 and Bcl-xL. In conclusion, 5F induced apoptosis in HT-29 cells and this apoptotic effect was associated with the high level of p38 and iNOS expression. The apoptotic effect of 5F could be significantly offset by over-expression of either Bcl-2 or Bcl-xL. Bcl-2, and to the less extent, Bcl-xL, were able to increase the activity of NF-kappa B, which was a known anti-apoptotic molecule in human colon cancer cells.

Preventive effects of ginsenosides on osteopenia of rats induced by ovariectomy.

AIM: To determine the effect of ginsenosides (GSL) on ovariectomized rats by analysis of cancellous bone histomorphometry. METHODS: Forty Sprague-Dawley female rats at age of 3 months were sham-operated (Sham, n = 8) and treated orally with vehicle, or ovariectomized (OVX, n = 32 which were divided into three group with n = 8 per group) and treated orally with either vehicle, 17alpha-ethynylestradiol (EE, 100 microg . kg-1 . d-1), or ginsenosides (GSL) at 100 or 300 mg . kg-1 . d-1 for 10 weeks. Double in vivo fluorochrome labeling was administrated. The undecalcified longitudinal proximal tibial metaphyseal sections were cut and stained with Goldner's Trichrome (4-micron thickness) or unstained (8-micron thickness) for the bone histomorphometric analysis. RESULTS: After 10 weeks post OVX the cancellous bone mass was lost markedly and showed high bone turnover indices (increased bone resorption and formation). EE decreased the resorptive surface and bone formation rate related to bone turnover and prevented bone loss. GSL at the two doses (100 and 300 mg . kg-1 . d-1) reduced the resorptive surfaces as did EE, but did not depress the mineral bone formation. High dose of GSL greatly increased bone mass and had a tendency to decrease bone turnover when compared with OVX group. CONCLUSION: GSL partially prevented OVX-induced cancellous bone loss by inhibiting osteoclast bone resorption and by a mild depression of bone turnover.

Effects of antitumor compounds isolated from Pteris semipinnata L on DNA topoisomerases and cell cycle of HL-60 cells.

AIM: To study the effect of the antitumor compounds 5F, 6F, and A from Pteris semipinnata L on the activities of DNA topoisomerases and cell cycle of HL-60 cells, and the synergism of compound 6F in combination with genistein in vitro. METHODS: DNA topoisomerases were isolated from HL-60 cell lines, and supercoiled pBR322 DNA was used as substrate to determine the activities of DNA topoisomerase I and II. Cell cycle was analyzed by flow cytometry (FCM). Cytotoxicity assay was tested by MTT method. RESULTS: Compounds 5F, 6F, and A inhibited the activities of DNA topoisomerase I and II. After exposure of the cells to compound 6F, an increase in cells in the S and G2/M phases and a decrease in cells in the G0/G1 phase of the cell cycle were observed. At low concentrations (57.8 and 115.6 nmol.L-1), compound 6F enhanced the cytotoxicity against HL-60 cell line in combination with genistein, q values were > 1.15. The enhancement times of 57.8 and 115.6 nmol.L-1 of 6F by genistein were 2.60 and 4.65, respectively. CONCLUSION: Compounds 5F, 6F, and A inhibited the activities of DNA topoisomerases of HL-60 cells. Compound 6F increased the number of cells in S and G2/M phases, decreased the population of G0/G1 phase cells, and enhanced the cytotoxicity of genistein, which had synergism with 6F in antitumor action.

[Comparison of the cytotoxicity of five constituents from Pteris semipinnata L. in vitro and the analysis of their structure-activity relationships].

The antitumor constituents were isolated from Pteris semipinnata L. (PsL), a Chinese traditional herb, and screened using tetrazolium salt (MTT) method. Their cytotoxic effects in vitro on several human tumor cell lines were studied. Compounds 5F, 6F, A and the ethanolic extract of PsL (PSE) were shown to have strong cytotoxicity against five cell lines: human liver adenocarcinoma cell line (HePG II), human lung adenocarcinom a cell line (SPC-A-1), human gastric adenocarcinoma cell line (MGC-803), human nasopharyngeal carcinoma cells in low differentiation (CNE-2Z) and human liver adenocarcinoma cell line (BEL-7402) in different degrees in a dose-dependent manner; compound 6F was the most active one, whose IC50 after 72 h treatment for the above five cell lines were 0.343 +/- 0.003, 0.115 +/- 0.022, 0.590 +/- 0.032, 0.328 +/- 0.066 and 0.221 +/- 0.058 microgram.ml-1, respectively. Compounds A and 5F were less active; no cytotoxicity of compounds 4F and B were detected on the five cell lines. Analysis of the relationship between structure and activity revealed that the antitumor activity portion in the structure is the alpha, beta-methylene cyclopentanone moiety, and the site and number of the hydroxy groups affect the cytotoxicity of these agents significantly.

Studies on the inhibitory effects of quercetin on the growth of HL-60 leukemia cells.

Quercetin, a naturally occurring flavonoid, has been shown to exert multiple pharmacological effects and to be an anticancer agent or a supplementary anticancer agent. In this report, the human HL-60 promyelocytic leukemia cell line was used to study the effects of quercetin on the growth, cell cycle, activities of cytosolic and membrane protein kinase C (PKC) and tyrosine protein kinase (TPK), and phosphoinositide production of the tumor cells. The results showed that quercetin inhibited the growth of HL-60 cells in a concentration-dependent manner, with an IC50 value of about 7.7 microM after 96 hr of treatment; when the concentration of quercetin was 10 microM, the percent inhibition on the growth of HL-60 cells was 17.1, 27.3, 40.1, and 52.7% after 24, 48, 72, and 96 hr of treatment, respectively. Flow cytometric analyses showed that quercetin caused an increase in cells in the G2/M phase and a decrease in cells in the G0/G1 phase of the cell cycle in a concentration-dependent manner; these effects were reversed when quercetin was removed from the culture medium. Quercetin strongly inhibited the activities of cytosolic PKC and membrane TPK from HL-60 cells in vitro, with IC50 values of about 30.9 and 20.1 microM, respectively, but did not affect membrane PKC or cytosolic TPK activity from HL-60 cells in vitro. Quercetin markedly inhibited in a concentration-dependent manner the production of phosphoinositides in intact HL-60 cells. The results provide evidence that the inhibitory effect of quercetin on the growth of HL-60 cells may be related to its inhibitory effects on PKC and/or TPK in vitro and/or on the production of phosphoinositides.

[Effects of fructus cnidii coumarins compared with nilestriol on osteoporosis in ovariectomized rats].

AIM: To compare the total coumarins from dried fruits of Cnidium monnieri (TCCM) and nilestriol (Nil) against osteoporosis. METHODS: SD rats (40, female, 3-month-old) were randomly divided into basal control, age control, ovariectomized (Ova), Ova + TCCM 67 mg.kg-1, Ova + TCCM 200 mg.kg-1, 6 times a week, and Ova + Nil 1 mg.kg-1, i.g. once a week. After 12 wk, sections (20 microns) of proximal tibiae were examined histologically. RESULTS: Ova reduced markedly the trabecular bone mass due to bone resorption excessed bone formation (% Tb. Ar -59%). Treatment with TCCM 67 mg.kg-1 partly suppressed bone turnover, but did not inhibit bone loss in Ova rats (% Tb.Ar -43%). Treatment with TCCM 200 mg.kg-1 and Nil 1 mg.kg-1 increased the trabecular area (% Tb. Ar +100% and +274%). CONCLUSION: Nil was more potent than TCCM in protecting against osteoporosis in Ova rats via supression of bone turnover.

Risedronate pretreatment does not hamper the anabolic effects of prostaglandin E2 in ovx rats.

Pretreatment of an anti-resorptive agent on the anabolic effects of prostaglandin E2 (PGE2) was studied on the proximal tibia and tibial shaft of ovariectomy (ovx) rats. Two days after ovx, rats were treated with either risedronate (Ris, 5 micrograms/kg twice weekly) or vehicle (V) for 60 days and then switched to 3 or 6 mg/kg/d PGE2 for 21 or 90 days. Bone area of both proximal tibial metaphysis (PTM) and tibial shaft (TX) were measured. Pretreatment with Ris increased the bone mass in PTM but not in TX of ovx rats. In the PTM, PGE2 produced the same percentage of new bone mass in both V- and Ris-pretreated ovx rats. The amount of new bone was almost the same after 3 weeks and 12 weeks of PGE2 treatment. There was no difference in the anabolic effects of 3 and 6 mg PGE2/kg/d in V-pretreated rats; however, the effects in Ris-pretreated groups were greater with 6 mg PGE2/kg/d than with 3 mg PGE2/kg/d. In TX, only the 6mg PGE2/kg/d administration added new bone on endocortical surfaces of both V- or Ris-pretreatment rats which leads to thickening the minimal cortical width, decreasing the marrow cavity and increasing total bone area. Both doses of PGE2 created new trabecular bone in the marrow cavity of tibial shaft in both vehicle- and Ris-pretreated ovx rats. These results suggest that Ris-pretreatment did not hamper the anabolic effects of PGE2 on either PTM or TX in ovx rats.

[Chemical components of Codonopsis pilosula (Franch.) Nannf. var. volubilis (Nannf.) L.T. Shen].

Ten compounds were isolated from Codono psis pilosula var. volubilis. Six of them were characterized as friedelin, taraxerol, alpha-spinasterol, alpha-spinasterol-beta-D-glucopyranoside, n-butyl-alpha-D-fructofuranoside and n-butyl-beta-D-fructopyranoside.

A new Plaque Glycolysis and Regrowth Method (PGRM) for the in vivo determination of antimicrobial dentifrice/rinse efficacy towards the inhibition of plaque growth and metabolism--method development, validation and initial activity screens.

A new method, the Plaque Glycolysis and Regrowth Method (PGRM), is described for the evaluation of antimicrobial effects on plaque metabolism in vivo. The method relies on the experimental observation that in vivo sampled dental plaques, collected from different quadrants of the dentition, produce equivalent rates of metabolic activity and regrowth when similarly dispersed and normalized into incubation media. In applications of the technique to antimicrobial evaluations, overnight fasted dental plaque is collected from a non-treated quadrant of the dentition along the gingival margin. Topical formulations are used in vivo. Following this, dental plaques are collected from other dentition quadrants at extended times, allowing for the back diffusion, clearance and natural intraoral deactivation of antimicrobials within the oral cavity. In vivo treated and non-treated plaque samples are subsequently tested for metabolic and regrowth activity under controlled and standardized conditions in vitro following normalization for biomass. The technique thus combines the necessary biological factors important to the legitimate evaluation of antimicrobial effects in vivo, while benefiting from the improved precision and control provided by in vitro assessment of plaque activity. In this paper evidence is presented validating the PGRM method, and initial activity screens of commercial antimicrobial mouthrinses and toothpastes, including a new stabilized stannous fluoride dentifrice, are described.

Effects of total coumarins of Fructus cnidii on skeleton of ovariectomized rats.

Total coumarins of Fructus Cnidii (TCFC), 5 g.kg-1 by intragastric gavage, 6 d/wk, x 7 wk, was effective for prevention of bone loss in ovariectomized (OVX) rats. In comparison to aging control rats, the proximal tibia of placebo-treated OVX rats were characterized by an increase in eroded perimeter (+298%), label perimeter (+77%), osteoid perimeter (+47%), mineral apposition rate (+32%) and bone formation rate (+130%). These changes indicated a high bone turnover in OVX rats leading to a rapid bone loss (-44%) in proximal tibial metaphysis. In contrast, the TCFC-treated OVX rats showed an increase of cancellous bone area (+41%) compared with placebo-treated OVX rats and decrease in all the above indices of bone turnover to near aging control levels except that of the osteoid area (+88%) which was higher than that in aging control, but mineralization lag time did not show significant changes. The results suggested that the TCFC inhibited the high bone turnover and reversed the bone loss at early menopausal stage.

Effect of ibopamine and the active metabolite epinine on the catecholamine content of rat hypothalamus and brainstem in vitro.

The effect of ibopamine (IBO) (SB 7505, SK&F 100168-A), a new drug for the treatment of congestive heart failure, and its active metabolite epinine (EPN) (N-methyldopamine), on the catecholamine content of hypothalamus and brainstem was studied in vitro after monoamine oxidase inhibition with pargyline. IBO and EPN increased levels of epinephrine (EPI) in a concentration- and time-dependent manner in both brain areas without significantly affecting the concentration of other catecholamines. Inhibition of either dopamine beta-hydroxylase, the neuronal EPI and norepinephrine uptake system, or esterase hydrolysis of IBO prevented the increase of EPI, whereas inhibition of phenylethanolamine N-methyltransferase, enzymatic dealkylation or the neuronal dopamine or serotonin uptake system had no influence on the increase of EPI levels. These results suggest that IBO after hydrolysis to EPN can be converted enzymatically to EPI by dopamine beta-hydroxylase in hypothalamus and brainstem. EPN seems to be accumulated into adrenergic and noradrenergic neurons by the high affinity uptake system. Changes in the EPI content of the central nervous system neurons might be responsible for some of the pharmacologic effects of IBO.

Resibufogenin-induced delayed afterdepolarization in rabbit heart in situ.

The pharmacological effects of resibufogenin (RGB), an effective cardiotonic obtained from a famous traditional Chinese medicine, on the electrical activity of the rabbit heart (n = 12) in situ were studied by using a new technique of recording monophasic action potential (MAP). RBG (0.3 mg/kg IV) induced DAD in 3/12 animals. DAD appeared within the first 0.5 to 1 min after RBG and lasted about 20-30 min. The position of DAD was in the end of phase 3 of MAP. The DADs of rabbit epicardium showed a transient, regular and phasic depolarization process. The triggered arrhythmias were not recorded in these experiments, as amplitudes of DAD were lower and did not reach the threshold. The results indicate that DADs are induced by RBG at first time in situ in rabbits. The characteristics of DAD were similar to those previously described in single cardiac fibers in vitro.

Release of endogenous brain epinephrine by the calcium ionophores X537A and A23187.

The Ca2+ ionophores X537A and A23187 produced dose-dependent release of endogenous epinephrine (Epi) as well as norepinephrine (NE) and dopamine (DA) from chopped rat hypothalamus and brainstem. The X537A-induced release of these catecholamines (CAs) was found to be Ca2+-independent, whereas the A23187-induced release was Ca2+-dependent. X537A and A23187 were approximately equipotent in causing the release of the hypothalamic Epi, NE and DA, but X537A was much more effective than A23187 in causing the maximal release. X537A, but not A23187, reduced the total endogenous CA content and increased the total 3,4-dihydroxyphenylacetic acid (DOPAC) content in the chopped hypothalamus. Ca2+-independent release of Epi, NE and DA in the chopped hypothalamus was also observed with indirectly acting sympathomimetic amines (e.g. tyramine and amphetamine). Tyramine and amphetamine did not affect the total endogenous CA contents nor the total DOPAC content. These results suggest that X537A caused release of endogenous brain Epi, NE and DA by transporting the biogenic amines across vesicular or intracellular storage sites. However, A23187 caused release of these CAs by exocytosis via transport of Ca2+ into the neurons.

Inhibitors of phenylethanolamine N-methyltransferase. 1. Effects of 2-cycloocytyl-2-hydroxyethylamine on rat brain and adrenal catecholamine content and blood pressure.

Acute administration of 2-cyclooctyl-2-hydroxyethylamine (CONH), a potent phenylethanolamine N-methyltransferase inhibitor, produced time- and dose-related reductions of blood pressure in spontaneously hypertensive rats (SHR). CONH did not affect the blood pressure of age-paired normotensive Wistar-Kyoto rats (WKY), or the heart rate of either animal group. CONH produced dose- and time-related reductions in epinephrine (Epi) levels in the hypothalamus and brainstem of both SHR and WKY. The drug also produced dose- and time-related reductions in hypothalamic and brainstem norepinephrine concentrations and elevations in dopamine and 3,4-dihydroxyphenylacetic acid concentrations in both SHR and WKY. CONH was about equally effective in reducing brain norepinephrine and Epi concentrations in both animal strains, but CONH was more effective in elevating brain dopamine and 3,4-dihydroxyphenylacetic acid concentrations in SHR than in WKY. Subchronic administration (3 days) of CONH produced a dose-related reduction in blood pressure in SHR, but had no affect on blood pressure in WKY. The changes in blood pressure in SHR were accompanied by a dose-related reduction in hypothalamic Epi concentrations, but the drug treatment produced no affect on the concentration of Epi in the brainstem. The results of this study suggest that brain Epi-containing neurons may be directly involved in the regulation of blood pressure or alternatively they may indirectly control cardiovascular function by modulating brain norepinephrine- and/or dopamine-neuronal activity.

Inhibitors of phenylethanolamine-N-methyltransferase. 2. Comparison of nonaromatic analogs of phenylethanolamines, 7,8-dichloro-1,2,3,4-tetrahydroisoquinoline (SKF-64139) and 2,3-dichloro-alpha-methylbenzylamine: effects on rat brain and adrenal catecholamine content and blood pressure.

Intraperitoneal injections of the phenylethanolamine N-methyltransferase inhibitors 2-cyclohexyl-2-hydroxyethylamine, 1-aminomethylcycloundecanol, 7,8-dichloro-1,2,3,4-tetrahydroisoquinoline and 2,3-dichloro-alpha-methylbenzylamine all lowered blood pressures in spontaneously hypertensive rats. The drugs did not affect the blood pressures of age-paired normotensive Wistar Kyoto rats or did the drugs alter the heart rates of either animal group. All of the phenylethanolamine N-methyltransferase inhibitors tested lowered hypothalamic epinephrine content which in spontaneously hypertensive rats could be correlated with the reduction in blood pressure. These data support the hypothesis that central nervous system epinephrine may play an important role in the regulation of blood pressure and in the genesis and/or maintenance of hypertension. The results also suggest the hypothesis that the blood pressure lowering effects of phenylethanolamine N-methyltransferase inhibitors may be mediated by their effects on hypothalamic epinephrine content.