AQP4 protein expression in each astrocyte group.

<p>(A) The Western blot image of AQP4 protein. (B) Semi-quantification showed that rhVEGF165 increased AQP4 protein expression in astrocytes, which was inhibited by SP600125 and U0126, but not SB239063 or Ly294002 (n = 6, *<i>p</i><0.05 vs. control, #<i>p</i><0.05 vs. VEGF). (C) Negative control gave negative result without detectable AQP4 labeling. (D) Group of control. (E) Group of VEGF. (F) Group of VEGF plus SP600125. (G) Group of VEGF plus U0126. (H) Group of VEGF plus SB239063. (I) Group of VEGF plus Ly294002. Nuclei are highlighted by DAPI staining. Scale bar: 50 µm.</p

Western blot analysis of AQP4 protein expression in each normal AQP4^+/+ group.

<p>(A, B) Western blot analysis showed rhVEGF165 injected intracerebroventricularly up-regulated AQP4 protein expression at striatum in normal AQP4^+/+ mice at 1 d, 3 d, and 7 d (n = 6, *<i>p</i><0.05 vs. control). (C, D) Western blot analysis showed rhVEGF165 up-regulated perihemotomal AQP4 protein expression at 1 d, 3 d, and 7 d after intracerebroventricular injection (n = 6, *<i>p</i><0.05 vs. sham operation, #<i>p</i><0.05 vs. ICH).</p

Immunofluorescence of AQP4 protein expression in each normal AQP4^+/+ group.

<p>(A) Negative control without primary antibody gave negative result with no detectable AQP4 labeling. (B, C) High magnification for groups of control and VEGF 3 d after injection showed AQP4 labeling was concentrated in glial end-feet surrounding intracerebral capillaries (arrows). (D–G) Groups of control, VEGF 1 d, 3 d, and 7 d after injection. Vermiform AQP4 with green fluorescence was abundantly expressed after rhVEGF165 injection. Immunofluorescence revealed similar results as Western blot analysis. Scale bar: A, D–G: 100 µm; B, C: 50 µm.</p

Immunofluorescence of AQP4 protein expression in each ICH AQP4^+/+ group.

<p>(A) Group of sham operation. (B–D) Groups of ICH 1 d, 3 d, and 7 d. (E–G) Groups of ICH plus VEGF 1 d, 3 d, and 7 d after injection. (H–J) High magnification for groups of sham operation, ICH and ICH plus VEGF 3 d after injection. Arrows show AQP4 labeling is concentrated in glial end-feet surrounding intracerebral capillaries. Scale bar: A–G: 100 µm; H–J: 50 µm.</p

Nissl’s staining of AQP4^+/+ and AQP4^−/− mice.

<p>Injection of rhVEGF165 intracerebroventricularly had no effect on Nissl’s staining in normal mice. At the three time points, marked neuron loss perihemotoma were found in AQP4^−/− mice when compared to AQP4^+/+ mice. Besides, rhVEGF165 increased perihemotomal Nissl’s staining cells in both AQP4^+/+ and AQP4^−/− mice. (A) Groups of control, control plus VEGF and sham operation at 3 d. (B) Groups of ICH and ICH plus VEGF at 1 d, 3 d and 7 d. Astericks represent the hemotoma zones. Scale bar: 50 µm.</p

Brain water content, brain specific gravity and EB extravasation amount of AQP4^+/+ and AQP4^−/− mice.

<p>(A) Brain tissue water content surrounding the hemotoma in AQP4^+/+ mice was markedly reduced by rhVEGF165 at 1 d, 3 d, and 7 d after intracerebroventricular injection (n = 6, *<i>p</i><0.05 vs. sham operation; & <i>p</i><0.05 vs. ICH), but no effect was observed in AQP4^−/− mice. AQP4^−/− mice had more perihemotomal brain tissue water content than AQP4^+/+ mice in both ICH and ICH plus VEGF groups (n = 6, #<i>p</i><0.05). (B) The specific gravity of brain tissue surrounding hematoma from AQP4^+/+ mice was significantly decreased by rhVEGF165 at 1 d, 3 d, and 7 d after injected intracerebroventricularly (n = 6, *<i>p</i><0.05 vs. sham operation; & <i>p</i><0.05 vs. ICH), while no influence was detected in AQP4^−/− mice. In addition, AQP4^+/+ mice had more specific gravity of perihemotomal brain tissues than AQP4^−/− mice in both ICH and ICH plus VEGF groups (n = 6, #<i>p</i><0.05). Put together, results of the two sections were consistent with each other. (C) In normal mice, the amount of EB extravasation was increased 1 d after intracerebroventricular rhVEGF165 injection in both types of mice, but this effect was only detected in AQP4^−/− mice at 3 d (n = 6, *<i>p</i><0.05 vs. control). No change was found at 7 d. RhVEGF165 had no effect on EB extravasation resulting from ICH whether 1 d, 3 d, or 7 d after intracerebroventricular injection in AQP4^+/+ mice, while an increasing effect in AQP4^−/− mice was found at all time points observed (n = 6, $ <i>p</i><0.05 vs. sham operation; & <i>p</i><0.05 vs. ICH). Meanwhile, this section also showed that AQP4^−/− mice had more EB extravasation than AQP4^+/+ mice in VEGF 1 d, 3 d, all time points of ICH and ICH plus VEGF groups (n = 6, #<i>p</i><0.05).</p

Western blot of phosphorylation of MAPKs and Akt influenced by rhVEGF165.

<p>RhVEGF165 up-regulated p-JNK and p-ERK at 12 h after administration and expression of the two proteins reached maximum at 48 h (n = 6, *<i>p</i><0.05 vs. 0 h). While no up-regulation of p-p38 or p-Akt was observed within 72 h after rhVEGF165 administration. Besides, expression of AQP4 protein was increased by rhVEGF165 after 24 h and peaked at 48 h (n = 6, *<i>p</i><0.05 vs. 0 h). (A) Western blot image. (B) Semi-quantification analysis.</p

Metabolomic Study of Biochemical Changes in the Plasma and Urine of Primary Dysmenorrhea Patients Using UPLC–MS Coupled with a Pattern Recognition Approach

Primary dysmenorrhea (PD) is characterized by painful menstrual cramps without any organic pathology and has a prevalence of up to 90% in adolescents. Recent advances in its etiology and pathogenesis are providing more speculative hypotheses focused on integral systems. Using a targeted tandem mass spectrometry (MS/MS)-based metabolomic platform, we explored the changes of metabolic profiling in plasma/urine simultaneously between PD patients and healthy controls before and after a 3-month herbal medicine (namely Shaofu Zhuyu formula concentrated-granule, SFZYFG) therapy. To detect and identify potential biomarkers associated with PD and SFZYFG treatment, we also performed a combined UPLC–QTOF-MS/MS-based metabolomic profiling of the plasma/urine samples, indicating a further deviation of the patients’ global metabolic profile from that of controls. The total thirty-five metabolites (nineteen in plasma and sixteen in urine), up-regulated or down-regulated (<i>p</i> < 0.05 or 0.01), were identified and contributed to PD progress. These promising identified biomarkers underpinning the metabolic pathway including sphingolipid metabolism, steroid hormone biosynthesis, and glycerophospholipid metabolism are disturbed in PD patients, which were identified by using pathway analysis with MetPA. Twenty-four altered metabolites and fourteen biochemical indicators were restored back to the control-like level after the treatment of SFZYFG and could be potential biomarkers for monitoring therapeutic efficacy. These findings may be promising to yield a valuable insight into the pathophysiology of PD and to advance the approaches of treatment, diagnosis, and prevention of PD and related syndromes

Cultural policy of Prague

This thesis follows mainly approach of the city of Prague to culture and cultural policy. Part one contains mainly the definition of culture, specifics of Czech environment a culture as public service. It follows approach since 2000 to today, from the view of strategic documents and its effects on culture, culture funding and advocacy

Altered Cytokine Gene Expression in Peripheral Blood Monocytes across the Menstrual Cycle in Primary Dysmenorrhea: A Case-Control Study

<div><p>Primary dysmenorrhea is one of the most common gynecological complaints in young women, but potential peripheral immunologic features underlying this condition remain undefined. In this paper, we compared 84 common cytokine gene expression profiles of peripheral blood mononuclear cells (PBMCs) from six primary dysmenorrheic young women and three unaffected controls on the seventh day before (secretory phase), and the first (menstrual phase) and the fifth (regenerative phase) days of menstruation, using a real-time PCR array assay combined with pattern recognition and gene function annotation methods. Comparisons between dysmenorrhea and normal control groups identified 11 (nine increased and two decreased), 14 (five increased and nine decreased), and 15 (seven increased and eight decreased) genes with ≥2-fold difference in expression (<em>P</em><0.05) in the three phases of menstruation, respectively. In the menstrual phase, genes encoding pro-inflammatory cytokines (IL1B, TNF, IL6, and IL8) were up-regulated, and genes encoding TGF-β superfamily members (BMP4, BMP6, GDF5, GDF11, LEFTY2, NODAL, and MSTN) were down-regulated. Functional annotation revealed an excessive inflammatory response and insufficient TGF-β superfamily member signals with anti-inflammatory consequences, which may directly contribute to menstrual pain. In the secretory and regenerative phases, increased expression of pro-inflammatory cytokines and decreased expression of growth factors were also observed. These factors may be involved in the regulation of decidualization, endometrium breakdown and repair, and indirectly exacerbate primary dysmenorrhea. This first study of cytokine gene expression profiles in PBMCs from young primary dysmenorrheic women demonstrates a shift in the balance between expression patterns of pro-inflammatory cytokines and TGF-β superfamily members across the whole menstrual cycle, underlying the peripheral immunologic features of primary dysmenorrhea.</p> </div