FKBP25, a novel regulator of the p53 pathway, induces the degradation of MDM2 and activation of p53

The p53 tumour suppressor protein is tightly controlled by the E3 ubiquitin ligase, mouse double minute 2 (MDM2), but maintains MDM2 expression as part of a negative feedback loop. We have identified the immunophilin, 25 kDa FK506-binding protein (FKBP25), previously shown to be regulated by p53-mediated repression, as an MDM2-interacting partner. We show that FKBP25 stimulates auto-ubiquitylation and proteasomal degradation of MDM2, leading to the induction of p53. Depletion of FKBP25 by siRNA leads to increased levels of MDM2 and a corresponding reduction in p53 and p21 levels. These data are consistent with the idea that FKBP25 contributes to regulation of the p53-MDM2 negative feedback loop.

Structured summary

MINT-6823686:MDM2 (uniprotkb:Q00987) physically interacts (MI:0218) with FKBP25 (uniprotkb:Q00688) by anti bait coimmunoprecipitation (MI:0006)MINT-6823707, MINT-6823722:MDM2 (uniprotkb:Q00987) physically interacts (MI:0218) with FKBP25 (uniprotkb:Q62446) by pull down (MI:0096)MINT-6823775:P53 (uniprotkb:Q04637) physically interacts (MI:0218) with MDM2 (uniprotkb:Q00987) by anti bait coimmunoprecipitation (MI:0006)MINT-6823735, MINT-6823749:FKBP25 (uniprotkb:Q62446) binds (MI:0407) to MDM2 (uniprotkb:Q00987) by pull down (MI:0096)MINT-6823761:Ubiquitin (UNIPROTKB:62988)P physically interacts (MI:0218) with MDM2 (uniprotkb:Q00987) by pull down (MI:0096)MINT-6823669:MDM2 (uniprotkb:Q00987) physically interacts (MI:0218) with FKBP25 (uniprotkb:Q00688) by two hybrid (MI:0018)     

Regulation of phospholipase D (PLD) in growth plate chondrocytes by 24R,25-(OH)2D3 is dependent on cell maturation state (resting zone cells) and is specific to the PLD2 isoform

Many of the effects of 1α,25-(OH)₂D₃ and 24R,25-(OH)₂D₃ on costochondral chondrocytes are mediated by the protein kinase C (PKC) signal transduction pathway. 1α,25-(OH)₂D₃ activates PKC in costochondral growth zone chondrocytes through a specific membrane receptor (1α,25-mVDR), involving rapid increases in diacylglycerol via a phospholipase C (PLC)-dependent mechanism. 24R,25-(OH)₂D₃ activates PKC in resting zone chondrocytes. Although diacylglycerol is increased by 24R,25-(OH)₂D₃, PLC is not involved, suggesting a phospholipase D (PLD)-dependent mechanism. Here, we show that resting zone and growth zone cells express mRNAs for PLD1a, PLD1b, and PLD2. Both cell types have PLD activity, but levels are higher in resting zone cells. 24R,25-(OH)₂D₃, but not 24S,25-(OH)₂D₃ or 1α,25-(OH)₂D₃, stimulates PLD activity in resting zone cells within 3 min via nongenomic mechanisms. Neither 1α,25-(OH)₂D₃ nor 24R,25-(OH)₂D₃ affected PLD in growth zone cells. Basal and 24R,25-(OH)₂D₃-stimulated PLD were inhibited by the PLD inhibitors wortmannin and EDS. Inhibition of phosphatidylinositol 3-kinase (PI 3-kinase), PKC, phosphatidylinositol-specific PLC (PI-PLC), and phosphatidylcholine-specific PLC (PC-PLC) had no effect on PLD activity. Thus, 24R,25-(OH)₂D₃ stimulates PLD, and PI 3-kinase, PI-PLC and PKC are not involved, whereas PLD is required for stimulation of PKC by 24R,25-(OH)₂D₃. Pertussis toxin, GDPβS, and GTPγS had no effect on 24R,25-(OH)₂D₃-dependent PLD when added to cell cultures, indicating that G-proteins are not involved. These data show that PKC activation in resting zone cells is mediated by PLD and suggest that a functional 24R,25-(OH)₂D₃-mVDR is required. The results also support the conclusion that the 24R,25-(OH)₂D₃-responsive PLD is PLD2, since this PLD isoform is G-protein-independent.     

Feedback regulation of Ras2 guanine nucleotide exchange factor (Ras2-GEF) activity of Cdc25p by Cdc25p phosphorylation in the yeast Saccharomyces cerevisiae

Ras-GEF Cdc25p has been found to be hyperphosphorylated upon glucose addition. This work provides evidence indicating that PKA activity positively regulates the degree of Cdc25p phosphorylation, and that the intracellular association of Cdc25p and Ras2p is independent of PKA activity. In vitro experiments revealed that the Ras2-GEF activity of Cdc25p is inhibited by Cdc25p phosphorylation. These data suggest a negative feedback mechanism by which intracellular cAMP synthesis is inhibited by PKA through Cdc25p phosphorylation.

Structured summary

MINT-8053016: CDC25p (uniprotkb:P04821) physically interacts (MI:0915) with ras2p (uniprotkb:P01120) by anti tag co-immunoprecipitation (MI:0007)MINT-8053030: ras2p (uniprotkb:P01120) physically interacts (MI:0915) with CDC25p (uniprotkb:P04821) by anti bait co-immunoprecipitation (MI:0006)     

(25R)-Isonuatigenin,an unusual steroidal sapogenin from Vestia lycioides

(25R)-Isonuatigenin, a ⁵-spirosten-3β,25-diol, was isolated from aerial parts of Vestia lycioides. Its structure was elucidated mainly by ¹H NMR and ¹³C NMR spectroscopy. A mixture of (25R)-nuatigenin and (25S)-isonuatigenin was also characterized. This is the first report on the natural occurrence of the two (25R)-isomers.     

Vitamin D-Binding Protein Levels in Female Patients with Primary Hyperparathyroidism

ObjectiveTo determine whether low levels of vitamin D-binding protein (DBP) are related to 25-hydroxyvitamin D (25[OH]D) deficiency in female patients with primary hyperparathyroidism (PHPT).MethodsTwenty-five female patients with PHPT (serum calcium level >10.2 mg/dL and intact parathyroid hormone (iPTH) level >66 pg/mL) and 25 healthy age- and body mass index-matched female control subjects were xaminod. Serum calcium and iPTH levels were determined by commercial laboratories. Levels of 25(OH)D and 1,25-dihydroxyvitamin D (1,25[OH]₂D) were determined by radioimmunoassay, and DBP level was determined by enzyme-linked immunosorbent assay.ResultsSerum iPTH and calcium levels were higher in PHPT patients than control subjects (P<.001). Levels of 25(OH)D, albumin, and DBP were lower in the serum of PHPT patients than control subjects (P<.01). There were no significant differences in 1,25(OH)₂D and free 25(OH) D levels between PHPT patients and control subjects. DBP level was inversely correlated with calcium (r = -0.47; P<.01) and iPTH (r = −0.31; P<.05) levels. The 25(OH)D level correlated positively with both DBP (r = 0.28; P <.05) and albumin (r = 0.44; P<.05) levels.ConclusionsBoth serum 25(OH)D and DBP levels were lower in female patients with PHPT compared with control subjects. We suggest that a low DBP level contributes to the low 25(OH)D level observed in female PHPT patients. The etiology of the decrease in DBP and its relationship to calcium, 25(OH)D, and PTH levels require further investigation. (Endocr Pract. 2013;19:609-613)     

Steroid sapogenins from Tristagma uniflorum

From bulbs of Tristagma uniflorum the known sapogenins tigogenin, neotigogenin and (20S,22R,25S)-5α-spirostan-3β,25-diol, as well as the new (20S,22R,25R)-5α-spirostan-3β,25-diol, (20S,22S,25S)-5α-furostan-22,25-epoxy-3β,26-diol and (20S,22S,25R) -5α-furostan-22,25-epoxy-3β,26-diol, were isolated and characterized by spectroscopic (IR, ¹H NMR, ¹³C NMR, MS) methods.     

T-Cell Regulation in Lepromatous Leprosy

Regulatory T (T_reg) cells are known for their role in maintaining self-tolerance and balancing immune reactions in autoimmune diseases and chronic infections. However, regulatory mechanisms can also lead to prolonged survival of pathogens in chronic infections like leprosy and tuberculosis (TB). Despite high humoral responses against Mycobacterium leprae (M. leprae), lepromatous leprosy (LL) patients have the characteristic inability to generate T helper 1 (Th1) responses against the bacterium. In this study, we investigated the unresponsiveness to M. leprae in peripheral blood mononuclear cells (PBMC) of LL patients by analysis of IFN-γ responses to M. leprae before and after depletion of CD25⁺ cells, by cell subsets analysis of PBMC and by immunohistochemistry of patients'' skin lesions. Depletion of CD25⁺ cells from total PBMC identified two groups of LL patients: 7/18 (38.8%) gained in vitro responsiveness towards M. leprae after depletion of CD25⁺ cells, which was reversed to M. leprae-specific T-cell unresponsiveness by addition of autologous CD25⁺ cells. In contrast, 11/18 (61.1%) remained anergic in the absence of CD25⁺ T-cells. For both groups mitogen-induced IFN-γ was, however, not affected by depletion of CD25⁺ cells. In M. leprae responding healthy controls, treated lepromatous leprosy (LL) and borderline tuberculoid leprosy (BT) patients, depletion of CD25⁺ cells only slightly increased the IFN-γ response. Furthermore, cell subset analysis showed significantly higher (p = 0.02) numbers of FoxP3⁺ CD8⁺CD25⁺ T-cells in LL compared to BT patients, whereas confocal microscopy of skin biopsies revealed increased numbers of CD68⁺CD163⁺ as well as FoxP3⁺ cells in lesions of LL compared to tuberculoid and borderline tuberculoid leprosy (TT/BT) lesions. Thus, these data show that CD25⁺ T_reg cells play a role in M. leprae-Th1 unresponsiveness in LL.     

Hashimoto Thyroiditis not Associated with Vitamin D Deficiency

Objective: Vitamin D deficiency is associated with several autoimmune diseases. This study assessed whether vitamin D deficiency is associated with Hashimoto thyroiditis (HT).Methods: Two groups of patients were selected for which serum 25-hydroxyvitamin D (25(OH)D) levels had been measured: (1) a study group of patients diagnosed with HT as indicated by thyroid antibodies, and (2) a healthy control group. Each group was separated by sex and then controlled for age and body mass index (BMI). Groups' mean 25(OH)D levels were compared by analysis of variance (ANOVA), and percent frequencies of vitamin D sufficiency, insufficiency, and deficiency were compared with a Z-test. The correlations between 25(OH)D levels and thyroid antibodies and thyroid-stimulating hormone (TSH) levels were also tested.Results: The mean 25(OH)D levels for the HT and control groups were significantly different in females (30.75 vs. 27.56 ng/mL, respectively) but not in males (14.24 vs. 13.26 ng/mL). HT females had a higher rate of vitamin D sufficiency (51.7% vs. 31.1%) and a lower rate of insufficiency (48.3% vs. 68.9%) relative to control females. No such differences were found in the male groups. None of the females were vitamin D deficient, but almost all males were. A significant (P = .016) positive correlation (r_s = 0.436) between 25(OH)D and TPOAb was observed in males.Conclusion: HT is not associated with higher rates of vitamin D deficiency relative to a control group.Abbreviations:BMI = body mass indexHT = Hashimoto thyroiditis25(OH)D = 25-hydroxyvitamin DTgAb = thyroglobulin antibodyTSH = thyroid-stimulating hormoneTPOAb = thyroid-peroxidase antibodyVDR = Vitamin D receptor     

Regulation of Adipogenesis and Key Adipogenic Gene Expression by 1, 25-Dihydroxyvitamin D in 3T3-L1 Cells

The functions of 1, 25-dihydroxyvitamin D (1, 25-(OH)₂D₃) in regulating adipogenesis, adipocyte differentiation and key adipogenic gene expression were studied in 3T3-L1 preadipocytes. Five concentrations (0.01, 0.1, 1, 10, 100nM) of 1, 25-(OH)₂D₃ were studied and lipid accumulation measured by Oil Red O staining and expression of adipogenic genes quantified using quantitative real-time PCR. Adipogenic responses to 1, 25-(OH)₂D₃ were determined on 6, and 12 h, and days 1-10 after induction of adipogenesis by a hormonal cocktail with or without 1, 25-(OH)₂D₃. In response to 1, 25-(OH)₂D₃ (1, 10, and 100 nM), lipid accumulation and the expression of PPARγ, C/EBPα, FABP4 and SCD-1 were inhibited through day 10, and vitamin D receptor expression was inhibited in the early time points. The greatest inhibitory effect was upon expression of FABP4. Expression of SREBP-1c was only affected on day 2. The lowest concentrations of 1, 25-(OH)₂D₃ tested did not affect adipocyte differentiation or adipogenic gene expression. The C/EBPα promoter activity response to 1, 25-(OH)₂D₃ was also tested, with no effect detected. These results indicate that 1, 25-(OH)₂D₃ inhibited adipogenesis via suppressing adipogenic-specific genes, and is invoked either during PPARγ activation or immediately up-stream thereof. Gene expression down-stream of PPARγ especially FABP4 was strongly inhibited, and we suggest that the role of 1, 25-(OH)₂D₃ in regulating adipogenesis will be informed by further studies of adipogenic-specific gene promoter activity.     

Relationship Between Glycated Hemoglobin and Circulating 25-Hydroxyvitamin D Concentration In African American And Caucasian American Men

ObjectiveTo examine whether (1) serum 25-hydroxy- vitamin D level (25[OH]D) is a risk factor for hyperglycemia, as assessed by glycated hemoglobin (HbA1c), in African American men (AAM) and (2) 25(OH)D is a predictor of HbA1c in AAM and Caucasian American men (CAM).MethodsWe prospectively assessed 25(OH)D and HbA1c in 1,074 men, outpatients with and without diabetes, at an urban Veteran Administration Medical Center (66.8% AAM, 26.4% CAM, 6% Hispanic, 0.4% Asian, and 0.4% Native American men). Multivariate regression analyzed the determinants of HbA1c after accounting for potential confounders.ResultsWe found high prevalence of low (< 30 ng/mL) 25(OH)D (81%) and elevated (≥5.7%) HbAlc (53.5%). The 25(OH)D was inversely associated with HbA1c in all men (r = −0.12, P<.001), in AAM (r = −0.11, P = .003), and in CAM (r = −0.15, P = .01). In the entire group the independent determinants of HbA1c included body mass index (BMI), age, 25(OH)D levels, systolic blood pressure (BP), triglycerides, high-density lipoprotein (HDL), and current alcohol use (P<.0001, .013, .009, .01, .008, .034, and .048, respectively) while glomerular filtration rate (GFR) and marital status showed borderline significance (P = .08 and .09, respectively). In AAM these determinants included BMI, 25(OH)D levels, systolic BP, and current alcohol use (P<.0001, .01, .02, and .03, respectively), while age had borderline significance (P = .06). In CAM, these included BMI, age, and triglycerides (P = .01, .03, and .004, respectively) but not 25(OH)D levels (P = .50).ConclusionCirculating low 25(OH)D is a risk factor for hyperglycemia, as assessed by HbA1c, in AAM. The 25(OH)D level is an independent determinant of HbA1c in AAM, but not in CAM, including men with and without diabetes. (Endocr Pract. 2013;19:73-80)     

Saponins from Astragalus hareftae (NAB.) SIRJ.

Four cycloartane- (hareftosides A–D) and oleanane-type triterpenoids (hareftoside E) were isolated from Astragalus hareftae along with fifteen known compounds. Structures of the compounds were established as 3,6-di-O-β-d-xylopyranosyl-3β,6α,16β,24(S),25-pentahydroxycycloartane (1), 3,6,24-tri-O-β-d-xylopyranosyl-3β,6α,16β,24(S),25-pentahydroxycycloartane (2), 3-O-β-d-xylopyranosyl-3β,6α,16β,25-tetrahydroxy-20(R),25(S)-epoxycycloartane (3), 16-O-β-d-glucopyranosyl-3β,6α,16β,25-tetrahydroxy-20(R),24(S)-epoxycycloartane (4), 3-O-[β-d-xylopyranosyl-(1→2)-O-β-d-glucopyranosyl-(1→2)-O-β-d-glucuronopyranosyl]-soyasapogenol B (5) by the extensive use of 1D- and 2D-NMR experiments along with ESI-MS and HR-MS analyses.     

A novel chemically modified analogue of xenin-25 exhibits improved glucose-lowering and insulin-releasing properties

Background: Xenin-25 is a K-cell derived gut peptide with insulin-releasing activity which is rapidly degraded following release into the circulation. We hypothesized that substitution of all naturally-occurring Lys and Arg residues with Gln would lead to prolonged enzyme resistance and enhanced biological efficacy.Methods: Peptide stability was assessed using murine plasma, in vitro insulin-releasing actions evaluated in BRIN-BD11 cells and acute glucose-lowering and insulin-releasing actions examined in high fat fed mice. For sub-chronic studies, a range of metabolic parameters and pancreatic histology were assessed in high fat fed mice which had received saline vehicle or xenin-25(gln) twice-daily for 21 days.Results: In contrast to native xenin-25, xenin-25(gln) was resistant to plasma-mediated degradation and significantly stimulated insulin secretion in BRIN-BD11 cells. Acute administration of xenin-25(gln) in high fat fed mice significantly reduced blood glucose and increased plasma insulin concentrations. Twice-daily administration of xenin-25(gln) in high fat fed mice did not affect food intake, body weight or circulating insulin concentrations but significantly decreased blood glucose from day 9 onwards. Furthermore, glucose tolerance, glucose-mediated insulin secretion, insulin sensitivity and GIP-stimulated insulin-release were significantly enhanced in xenin-25(gln)-treated mice. Pancreatic immunohistochemistry revealed decreased alpha cell area with increased beta cell area and beta-to-alpha cell ratio in xenin-25(gln)-treated mice. In addition, xenin-25(gln) exerted similar beneficial actions in ob/ob mice as demonstrated by reduced blood glucose, superior glycaemic response and glucose-mediated insulin release.Conclusions: Xenin-25(gln) is resistant to plasma-mediated degradation and exerts sustained and beneficial metabolic actions in high fat fed and ob/ob mice.General significance: Glutamine (gln)-modified analogues of xenin may represent an attractive therapeutic approach for type 2 diabetes.     

Trabecular Bone Score Change Differs with Regard to 25(OH)D Levels in Patients Treated for Adult-Onset Growth Hormone Deficiency

Objective: Vitamin D is important in bone health. However, potential relationships of concomitant vitamin D deficiency with growth hormone deficiency (GHD) and the possibility that vitamin D inadequacy may alter the skeletal effects of growth hormone (GH) replacement therapy have not been adequately evaluated.Methods: A prospective study was conducted in adult-onset GHD patients treated with recombinant human GH (rhGH) for 2 years. Trabecular bone score (TBS), lumbar spine (LS) bone mineral density (BMD), total hip (TH) BMD, and 25-hydroxyvitamin D (25(OH)D) levels were assessed at baseline and 24 months. The study cohort was divided based on 25(OH)D levels into 2 groups with the cutoff defined as the 50^th percentile at each follow-up time point.Results: Fifty-seven patients (29 males/28 females, mean age 34.4 years) were studied. After 24 months of GH replacement, LS BMD increased by 7.6% and TH BMD increased by 4.5% (both P<.05), with no difference according to 25(OH)D levels. TBS increased (+1.39 ± 3.6%) in those whose 25(OH)D was above the 50^th percentile but decreased (-1.36 ± 5.6%, P<.05) in the cohort below the 50^th percentile of 25(OH)D. Positive correlations were observed between baseline levels of IGF-1 and 25(OH)D (R = 0.37, P<.001) and between 24-month 25(OH)D and TBS (R = 0.25, P<.05).Conclusion: A differential effect of GH on TBS change was observed; TBS increased only in the cohort with 25(OH)D above the 50^th percentile. Vitamin D sufficiency may be required to obtain optimal effects of GH treatment on bone quality, as assessed by TBS, in GHD adults.Abbreviations:AO-GHD = adult-onset GHDBMD = bone mineral densityBMI = body mass indexCa = calciumCTx = carboxyterminal collagen crosslinksCV = coefficient of variationDXA = dual energy X-ray absorptiometryECLIA = enzyme-labeled chemiluminescent immunometric assayGH = growth hormoneGHD = growth hormone deficiencyIGF-1 = insulin-like growth factor 1LS BMD = lumbar spine BMDOC = osteocalcin25(OH)D = 25-hydroxyvitamin DP = phosphorusPTH = parathyroid hormonerhGH = recombinant human GHTBS = trabecular bone scoreTH BMD = total hip BMD     

Human Immunodeficiency Virus Type 1 Vpr Induces Cell Cycle G2 Arrest through Srk1/MK2-Mediated Phosphorylation of Cdc25

Human immunodeficiency virus type 1 (HIV-1) Vpr induces cell cycle G₂ arrest in fission yeast (Schizosaccharomyces pombe) and mammalian cells, suggesting the cellular pathway(s) targeted by Vpr is conserved among eukaryotes. Our previous studies in fission yeast demonstrated that Vpr induces G₂ arrest in part through inhibition of Cdc25, a Cdc2-specific phosphatase that promotes G₂/M transition. The goal of this study was to further elucidate molecular mechanism underlying the inhibitory effect of Vpr on Cdc25. We show here that, similar to the DNA checkpoint controls, expression of vpr promotes subcellular relocalization of Cdc25 from nuclear to cytoplasm and thereby prevents activation of Cdc2 by Cdc25. Vpr-induced nuclear exclusion of Cdc25 appears to depend on the serine/threonine phosphorylation of Cdc25 and the presence of Rad24/14-3-3 protein, since amino acid substitutions of the nine possible phosphorylation sites of Cdc25 with Ala (9A) or deletion of the rad24 gene abolished nuclear exclusion induced by Vpr. Interestingly, Vpr is still able to promote Cdc25 nuclear export in mutants defective in the checkpoints (rad3 and chk1/cds1), the kinases that are normally required for Cdc25 phosphorylation and nuclear exclusion of Cdc25, suggesting that others kinase(s) might modulate phosphorylation of Cdc25 for the Vpr-induced G₂ arrest. We report here that this kinase is Srk1. Deletion of the srk1 gene blocks the nuclear exclusion of Cdc25 caused by Vpr. Overexpression of srk1 induces cell elongation, an indication of cell cycle G₂ delay, in a similar fashion to Vpr; however, no additive effect of cell elongation was observed when srk1 and vpr were coexpressed, indicating Srk1 and Vpr are likely affecting the cell cycle G₂/M transition through the same cellular pathway. Immunoprecipitation further shows that Vpr and Srk1 are part of the same protein complex. Consistent with our findings in fission yeast, depletion of the MK2 gene, a human homologue of Srk1, either by small interfering RNA or an MK2 inhibitor suppresses Vpr-induced cell cycle G₂ arrest in mammalian cells. Collectively, our data suggest that Vpr induces cell cycle G₂ arrest at least in part through a Srk1/MK2-mediated mechanism.     

The synthesis of cholestane and furostan saponin analogues and the determination of sapogenin's absolute configuration at C-22

A facile and efficient way for the synthesis of cholestane and furostan saponin analogues was established and adopted for the first time. Following this strategy, starting from diosgenin, three novel cholestane saponin analogues: (22S,25R)-3β,22,26-trihydroxy-cholest-5-ene-16-one 22-O-[O-α-l-rhamnopyranosyl-(1 → 2)-β-d-glucopyranoside] 11, (25R)-3β,16β,26-trihydroxy-cholest-5-ene-22-one 16-O-[O-α-l-rhamnopyranosyl-(1 → 2)-α-d-glucopyranoside] 14 and (25R)-3β,16β,26-trihydroxy-cholest-5-ene-22-one 16-O-[O-α-l-rhamnopyranosyl-(1 → 2)-β-d-glucopyranoside] 17, three novel furostan saponin analogues: (22S,25R)-furost-5-ene-3β,22,26-triol 22-O-(α-d-glucopyranoside) 23, (22R,25R)-furost-5-ene-3β,22,26-triol 22-O-(α-d-glucopyranoside) 24 and (22S,25R)-furost-5-ene-3β,22,26-triol 22-O-[O-α-l-rhamnopyranosyl-(1 → 2)-α-d-glucopyranoside] 26, were synthesized ultimately. The structures of all the synthesized analogues were confirmed by spectroscopic methods. The S-chirality at C-22 of cholestane was confirmed by Mosher's method. The absolute configuration at C-22 of furostan saponin analogues was distinguished by conformational analysis combined with the NMR spectroscopy. The cytotoxicities of the synthetic analogues toward four types of tumor cells were shown also.     

Inverse Association of Serum Vitamin D in Relation to Carotid Intima-Media Thickness in Chinese Postmenopausal Women

Objective

This study aimed to investigate the relationship between serum vitamin D level and carotid intima-media thickness (C-IMT) in Chinese postmenopausal women.

Methods

Nine hundred and twenty six Chinese postmenopausal women without carotid artery plaque or history of cardiovascular disease were selected for analysis. Measurements of serum 25 hydroxyvitamin D₃ (25(OH)D₃) concentration and C-IMT were made by electrochemiluminescence immunoassay and B-mode ultrasound, respectively. Trend analysis was conducted according to tertiles of C-IMT.

Results

The median serum 25(OH)D₃ level was 11.03 ng/mL, with an interquartile range of 8.22–14.70. A decreasing trend of serum 25(OH)D₃ level was accompanied by increased C-IMT tertiles (P for trend = 0.001). Correlation analysis found an inverse relationship between serum 25(OH)D₃ level and C-IMT (r = –0.113, P = 0.001). After adjustment for confounding factors, multiple regression analysis showed that serum 25(OH)D₃ level independently and negatively associated with C-IMT (Standard β = –0.112, P < 0.001). Moreover, the inverse correlation of serum 25(OH)D₃ with C-IMT was also found in a subgroup of women with normal glucose tolerance, blood pressure and body mass index, and without undergoing lipid-lowering therapy (standard β = –0.140, P = 0.018).

Conclusions

Serum 25(OH)D₃ level was inversely correlated with C-IMT in Chinese postmenopausal women.     

Microbiological transformation of diosgenin by resting cells of filamentous fungus,Cunninghamella echinulata CGMCC 3.2716

Microbial transformation of the steroidal sapogenin diosgenin (1) by resting cells of the filamentous fungus, Cunninghamella echinulata CGMCC 3.2716 was studied. Four metabolites were isolated and unambiguously characterized as (25R)-spirost-5-ene-3β,7β-diol-11-one (2), (25R)-spirost-5-ene-3β,7β-diol (3), (25R)-spirost-5-ene-3β,7β,11α-triol (4), and (25R)-spirost-5-ene-3β,7β,12β-triol (5), by various spectroscopic methods (¹H, ¹³C NMR, DEPT, ¹H–¹H COSY, HMBC, HSQC and NOESY). Compound 2 is a new metabolite. The NMR data and full assignment for the known metabolites (25R)-spirost-5-ene-3β,7β-diol (3) and (25R)-spirost-5-ene-3β,7β,11α-triol (4) are described here for the first time. The biotransformation characteristics observed included were C-7β, C-11α and C-12β hydroxylations. Compounds 1–5 exhibited no significant cytotoxic activity to human glioma cell line U87.     

In Vitro Metabolism of 20(R)-25-Methoxyl-Dammarane-3, 12, 20-Triol from Panax notoginseng in Human,Monkey, Dog,Rat, and Mouse Liver Microsomes

The present study characterized in vitro metabolites of 20(R)-25-methoxyl-dammarane-3β, 12β, 20-triol (20(R)-25-OCH₃-PPD) in mouse, rat, dog, monkey and human liver microsomes. 20(R)-25-OCH₃-PPD was incubated with liver microsomes in the presence of NADPH. The reaction mixtures and the metabolites were identified on the basis of their mass profiles using LC-Q/TOF and were quantified using triple quadrupole instrument by multiple reaction monitoring. A total of 7 metabolites (M1–M7) of the phase I metabolites were detected in all species. 25(R)-OCH₃-PPD was metabolized by hydroxylation, dehydrogenation, and O-demethylation. Enzyme kinetic of 20(R)-25-OCH₃-PPD metabolism was evaluated in rat and human hepatic microsomes. Incubations studies with selective chemical inhibitors demonstrated that the metabolism of 20(R)-25-OCH₃-PPD was primarily mediated by CYP3A4. We conclude that 20(R)-25-OCH₃-PPD was metabolized extensively in mammalian species of mouse, rat, dog, monkey, and human. CYP3A4-catalyzed oxygenation metabolism played an important role in the disposition of 25(R)-OCH₃-PPD, especially at the C-20 hydroxyl group.