Intravenously engrafted human multilineage-differentiating stress-enduring (Muse) cells rescue erectile function after rat cavernous nerve injury.

OBJECTIVE: To evaluate the effect of intravenous administration of human multilineage-differentiating stress-enduring (Muse) cells on rat postoperative erectile dysfunction (ED) with cavernous nerve (CN) injury without an immunosuppressant. MATERIALS AND METHODS: Male Sprague-Dawley rats were randomised into three groups after CN crush injury. Either human-Muse cells, non-Muse mesenchymal stem cells (MSCs) (both 1.0 × 105 cells), or vehicle was infused intravenously at 3 h after CN injury without immunosuppressant. Erectile function was assessed by measuring intracavernous pressure (ICP) and arterial pressure (AP) during pelvic nerve electrostimulation 28 days after surgery. At 48 h and 28 days after intravenous infusion of Muse cells, the homing of Muse cells and non-Muse MSCs was evaluated in the major pelvic ganglion (MPG) after CN injury. In addition, expressions of C-X-C motif chemokine ligand (Cxcl12) and glial cell line-derived neurotrophic factor (Gdnf) in the MPG were examined by real-time polymerase chain reaction. Statistical analyses and comparisons among groups were performed using one-way analysis of variance followed by the Tukey test for parametric data and Kruskal-Wallis test followed by the Dunn-Bonferroni test for non-parametric data. RESULTS: The mean (SEM) ICP/AP values at 28 days were 0.51 (0.02) in the Muse cell group, 0.37 (0.03) in the non-Muse MSC group, and 0.36 (0.04) in the vehicle group, showing a significant positive response in the Muse cell group compared with the non-Muse and vehicle groups (P = 0.013 and P = 0.010, respectively). In the MPG, Muse cells were observed to be engrafted at 48 h and expressed Schwann cell markers S100 (~46%) and glial fibrillary acidic protein (~24%) at 28 days, while non-Muse MSCs were basically not engrafted at 48 h. Higher gene expression of Cxcl12 (P = 0.048) and Gdnf (P = 0.040) was found in the MPG of the Muse group than in the vehicle group 48 h after infusion. CONCLUSION: Intravenously engrafted human Muse cells recovered rat erectile function after CN injury in a rat model possibly by upregulating Cxcl12 and Gdnf.

17ß-neriifolin from unripe fruits of Cerbera manghas suppressed cell proliferation via the inhibition of HOXA9-dependent transcription and the induction of apoptosis in the human AML cell line THP-1.

Homeobox A9 (HOXA9) is a transcription factor that is overexpressed in acute myeloid leukemia (AML). It is associated with the pathogenesis and progression of AML, and is a factor responsible for a poor prognosis. Therefore, the development of HOXA9-targeting molecules may contribute to not only better understanding of the mechanism of HOXA9 regulation, but also the development of therapeutic applications. We constructed a reporter assay system using the promoter region of the KBTBD10 gene, to which HOXA9 directly binds and regulates transcription, in the human acute monocytic leukemia cell line THP-1. Using this luciferase gene assay, we screened 1120 plant extracts and a methanol extract of the unripe fruits of Cerbera manghas was found to suppress the reporter gene expression mediated by the KBTBD10 promoter. From the extract, five steroid-type compounds were identified as the active constituents: 7α-neriifolin (1), 17ß-neriifolin (2), 17α-digitoxigenin ß-D-glucosyl-(1 → 4)-α-L-thevetoside (3), 17ß-digitoxigenin ß-D-glucosyl-(1 → 4)-α-L-thevetoside (4), and acetylthevetin B (5). Among the five compounds, 17ß-neriifolin most potently inhibited HOXA9-dependent gene expression without affecting the HOXA9 mRNA levels, and suppressed cell proliferation by inducing apoptosis. The findings on the structure-activity relationships of the compounds from C. manghas may contribute to the development of small molecule inhibitors of HOXA9.

Identification of a Selective RelA Inhibitor Based on DSE-FRET Screening Methods.

Nuclear factor-κB (NF-κB) is an important transcription factor involved in various biological functions, including tumorigenesis. Hence, NF-κB has attracted attention as a target factor for cancer treatment, leading to the development of several inhibitors. However, existing NF-κB inhibitors do not discriminate between its subunits, namely, RelA, RelB, cRel, p50, and p52. Conventional methods used to evaluate interactions between transcription factors and DNA, such as electrophoretic mobility shift assay and luciferase assays, are unsuitable for high-throughput screening (HTS) and cannot distinguish NF-κB subunits. We developed a HTS method named DNA strand exchange fluorescence resonance energy transfer (DSE-FRET). This assay is suitable for HTS and can discriminate a NF-κB subunit. Using DSE-FRET, we searched for RelA-specific inhibitors and verified RelA inhibition for 32,955 compounds. The compound A55 (2-(3-carbamoyl-6-hydroxy-4-methyl-2-oxopyridin-1(2H)-yl) acetic acid) selectively inhibited RelA-DNA binding. We propose that A55 is a seed compound for RelA-specific inhibition and could be used in clinical applications.

Effects of Combined Treatment with Branched-Chain Amino Acids, Citric Acid, L-Carnitine, Coenzyme Q10, Zinc, and Various Vitamins in Tumor-Bearing Mice.

A functional dietary supplement (FDS) containing Coenzyme Q10, branched-chain amino acids and L-carnitine was administered to tumor-bearing mice, investigating its effects on tumor and muscle tissues. Experiment (A): B16 melanoma cells were implanted subcutaneously into the right side of the abdomen of 8- to 9-week-old C57BL/6J mice. The mice were divided into two groups: a FDS group that received oral administration of FDS (n=10), and a control group that received oral administration of glucose (n=10). The moribund condition was used as the endpoint, and median survival time was determined. Experiment (B): On day 21 after tumor implantation, tumors, soleus muscle, gastrocnemius muscle, and suprahyoid muscles were collected. Tumor and muscle weight and other aspects were evaluated in each group: FDS group (n=15) and control group (n=15). The median survival time was comparable (21 d in the FDS group vs. 18 d in the control group, p=0.30). However, cumulative food intake was significantly higher in the FDS group than the control group (p=0.011). Metastasis of melanoma to the lung was observed in the control group but not in the FDS group (p=0.043). The weight of the suprahyoid muscles was significantly higher in the FDS group than in the control group (p=0.0045). The weight of the tumor was significantly lower in the FDS group than in the control group (p=0.013). The results possibly suggest oral administration of FDS in tumor-bearing mice enhances the maintenance of suprahyoid muscles, resulting in an extended feeding period and suppression of tumor growth and metastasis.

Impact of Tissue Sealing Sheet on Erectile Dysfunction in a Rat Model of Nerve-Sparing Radical Prostatectomy.

INTRODUCTION: The tissue sealing sheet has recently been used to prevent intraoperative bleeding from the neurovascular bundles in radical prostatectomy. Surgical stress or inflammatory changes likely play a role in erectile dysfunction after cavernous nerve injury. However, the efficacy of a tissue sealing sheet for preventing erectile function after nerve-sparing radical prostatectomy remains unclear. AIM: To evaluate the effect of a tissue sealing sheet on erectile dysfunction after cavernous nerve dissection. METHODS: Male Sprague-Dawley rats were randomly divided into three groups and subjected to sham operation or bilateral cavernous nerve dissection with (sheet group) or without (non-sheet group) a tissue sealing sheet. In the sheet group, cavernous nerves were sealed with a tissue sealing sheet immediately after cavernous nerve dissection. MAIN OUTCOME MEASURES: Erectile function was assessed by measuring intracavernous pressure and arterial pressure during pelvic nerve electrostimulation at 4 weeks after surgery. Expressions of interleukin-6, tumor growth factor-ß1, and heme-oxygenase-1 in the major pelvic ganglion were examined by real-time polymerase chain reaction. RESULTS: Mean intracavernous pressure along with mean arterial pressure in the sheet group were similar to those in the sham group and showed a significant positive response compared with the non-sheet group (P < .05). Furthermore, expressions of interleukin-6, tumor growth factor-ß1, and heme-oxygenase-1 were significantly lower in the sheet group than in the non-sheet group (P < .05). CONCLUSION: Use of a tissue sealing sheet attenuated postoperative inflammatory changes and oxidative stress and improved erectile function after cavernous nerve injury in rats. The tissue sealing sheet might become a useful therapeutic approach to preserve erectile function after nerve-sparing radical prostatectomy.

The effect of the kampo medicine yokukansan on preoperative anxiety and sedation levels.

Background. Preoperative anxiety can lead to unfavorable physiological response such as tachycardia and hypertension. Prevention of preoperative anxiety improves surgical outcome and decreases inpatient stay. Yokukansan is one of prescriptions in Kampo, traditional Japanese herbal medicine, and is known to exert anxiolytic effects. The aim of the present study was to compare the effects of diazepam and Yokukansan on preoperative anxiety, salivary amylase activity, and sedation levels. Methods. Seventy American Society of Anesthesiologists physical status I or II patients presenting for hemicolectomy under general anesthesia combined with epidural anesthesia were enrolled. The Diazepam group received diazepam 5 mg orally and the Yokukansan group received Yokukansan 2.5 g orally. Results. Although levels of anxiety and salivary amylase activity were not different between the two groups, the modified Observer's Assessment of Alertness/Sedation Scale of the Yokukansan group was significantly higher compared to that of the Diazepam group. Conclusion. Yokukansan alleviated preoperative anxiety without undesirable sedation, when compared with diazepam.

Auricular acupuncture at the "shenmen" and "point zero" points induced parasympathetic activation.

Purpose. Since auricular acupuncture is a diagnostic and treatment system based on normalizing the body's dysfunction, auricular acupuncture has been applied for pain relief, relaxation, and so on. These techniques would modulate the autonomic nerve system, thereby inducing the above-mentioned effects. The aim was to see the effect of auricular acupuncture applied to the "Shenmen" and "Point Zero" points on the postoperative heart rate variability (HRV). Methods. Twenty-six patients who underwent hemicolectomy under general anesthesia were randomized into the control or the acupuncture group. After the operation and before emergence, the acupuncture group received auricular acupuncture. An electrocardiographic unit was placed for recording the autonomic nervous activities. Results. The low frequency (LF)/high frequency (HF) ratio of HRV increased (P = 0.0007) in the control, but the ratio in the acupuncture did not change. There were significant differences between the ratios of the two groups at 3 : 00, 4 : 00, and 5 : 00. HF of the acupuncture group tended to be higher. HFs of the acupuncture group were significantly higher than those of the control group at 3 : 00, 4 : 00, and 5 : 00. Conclusion. Auricular acupuncture kept the LF/HF ratio at lower levels and HF at higher levels during postoperative period in the patients who had undergone hemicolectomy.

Inhibition of ENNG-induced pyloric stomach and small intestinal carcinogenesis in mice by high temperature- and pressure-treated garlic.

High temperature- and pressure-treated garlic (HTPG) has been shown to have enhanced antioxidative activity and polyphenol contents. Previously, we reported that HTPG inhibited 1,2-dimethylhydrazine-induced mucin depleted foci (premalignant lesions) and O6-methylguanine DNA adduct formation in the rat colorectum. In the present study, we investigated the modifying effects of HTPG on N-ethyl-N'-nitro-N-nitrosoguanidine (ENNG)- induced pyloric stomach and small intestinal carcinogenesis in mice. Male C57BL/6 mice were given ENNG (100 mg/l) in drinking water for the first 4 weeks, then a basal diet or diet containing 2% or 5% HTPG for 30 weeks. The incidence and multiplicity of pyloric stomach and small intestinal (duodenal and jejunal) tumors in the 2% HTPG group (but not in the 5% HTPG group) were significantly lower than those in the control group. Cell proliferation of normal-appearing duodenal mucosa was assessed by MIB-5 immunohistochemistry and shown to be significantly lower with 2% HTPG (but again not 5% HTPG) than in controls. These results in dicate that HTPG, at 2% in the diet, inhibited ENNG-induced pyloric stomach and small intestinal (especially duodenal) tumorigenesis in mice, associated with suppression of cell proliferation.

Transcutaneous Electrical Nerve Stimulation on the PC-5 and PC-6 Points Alleviated Hypotension after Epidural Anaesthesia, Depending on the Stimulus Frequency.

Neuraxial blockade causes arterial hypotension. Transcutaneous electrical nerve stimulation (TENS) at the Neiguan (PC-6) and Jianshi (PC-5) reduces the severity of hypotension after spinal anaesthesia, but did not clarify the optimal stimulus frequency. We hypothesized that the stimulus frequency of TENS at the PC-6 and PC-5 points would influence the severity of hypotension after epidural anaesthesia. 65 ASA I or II male patients presenting for inguinal hernia repair were randomized to five groups: the control group received no treatment; the 2 Hz, 10 Hz, 20 Hz, and 40 Hz groups received TENS at a frequency of 2 Hz, 10 Hz, 20 Hz, and 40 Hz, respectively. The lowest SBP was significantly higher in the 40 Hz group [the control, 84 (74-110) mmHg; the 2 Hz, 96 (62-116) mmHg; the 10 Hz, 100 (68-110) mmHg; the 20 Hz, 96 (64-115) mmHg; the 40 Hz, 104 (75-140) mmHg: P = 0.004]. Significantly less patients experienced hypotension in the 40 Hz group [the control, 78%; the 2 Hz, 43%; the 10 Hz, 38%; the 20 Hz, 38%; the 40 Hz, 8%: P = 0.008]. TENS on the PC-6 and PC-5 points reduced the severity and incidence of hypotension after epidural anaesthesia, depending on the stimulus frequency.

Development of live-cell imaging probes for monitoring histone modifications.

The combination of histone posttranslational modifications occurring in nucleosomal histones determines the epigenetic code. Histone modifications such as acetylation are dynamically controlled in response to a variety of signals during the cell cycle and differentiation, but they are paradoxically maintained through cell division to impart tissue specific gene expression patterns to progeny. The dynamics of histone modifications in living cells are poorly understood, because of the lack of experimental tools to monitor them in a real-time fashion. Recently, FRET-based imaging probes for histone H4 acetylation have been developed, which enabled monitoring of changes in histone acetylation during the cell cycle and drug treatment. Further development of this type of fluorescent probes for other modifications will make it possible to visualize complicated epigenetic regulation in living cells.

Auricular Acupunctures are Effective for the Prevention of Postoperative Agitation in Old Patients.

Postoperative cognitive problems and delirium are not uncommon in the elderly. We reported four cases in which auricular acupunctures on the 'Shenmen' and 'Point Zero' points successfully managed postoperative problematic behaviors of the three patients with dementia and the one patient postoperatively demonstrating an agitated behavior.

Radical prostatectomy for high-risk prostate cancer: biochemical outcome.

OBJECTIVES: To determine the biochemical outcome following radical prostatectomy alone in patients with high-risk prostate cancer. METHODS: Between January 2002 and August 2007, 252 patients underwent radical retropubic prostatectomy. Those who received neoadjuvant hormone therapy were excluded from this analysis. Based on pre-operative data, we stratified the patients into low, intermediate, and high-risk groups according to the risk criteria of the National Comprehensive Cancer Network in 2003, respectively. Prostate-specific antigen (PSA) failure was defined as any detectable PSA level higher than 0.2 ng/mL. RESULTS: The PSA failure-free survival rate for the high-risk group (n = 46) was 64.5% after a median follow-up period of 39 months. Among patients with high-risk disease, none with pathologically organ-confined cancer (n = 19) and a negative surgical margin had PSA failure. The PSA failure-free rate in patients with non organ-confined cancer (n = 27) was 39.5%. Among the pretreatment variables, a positive biopsy core percentage (the number of positive biopsy cores/total biopsy core) >or=30 was a significant independent predictor of extra prostatic extension. CONCLUSIONS: Radical prostatectomy is feasible in high-risk prostate cancer patients, only if they have a pathologically organ-confined disease.

Ginkgolic acid inhibits protein SUMOylation by blocking formation of the E1-SUMO intermediate.

Protein modification by small ubiquitin-related modifier proteins (SUMOs) controls diverse cellular functions. Dysregulation of SUMOylation or deSUMOylation processes has been implicated in the development of cancer and neurodegenerative diseases. However, no small-molecule inhibiting protein SUMOylation has been reported so far. Here, we report inhibition of SUMOylation by ginkgolic acid and its analog, anacardic acid. Ginkgolic acid and anacardic acid inhibit protein SUMOylation both in vitro and in vivo without affecting in vivo ubiquitination. Binding assays with a fluorescently labeled probe showed that ginkgolic acid directly binds E1 and inhibits the formation of the E1-SUMO intermediate. These studies will provide not only a useful tool for investigating the roles of SUMO conjugations in a variety of pathways in cells, but also a basis for the development of drugs targeted against diseases involving aberrant SUMOylation.

Nerves at the ventral prostatic capsule contribute to erectile function: initial electrophysiological assessment in humans.

BACKGROUND: Recent microanatomical studies have identified a significant amount of nerve fibers along the ventral circumference of the prostate capsule and confirmed the concept of a periprostatic nerve network. However, functional investigations have not yet proved whether nerve fibers distributed on the prostate capsule, particularly those outside the neurovascular bundle (NVB), contribute to erection. OBJECTIVE: To confirm whether nerve fibers distributed on the prostate capsule contribute to erectile function, the present study was performed using electrophysiologic testing. DESIGN, SETTING, AND PARTICIPANTS: The circumference of the prostate capsule was electrically stimulated during radical retropubic prostatectomy (RRP) in 12 patients with clinically localized prostate cancer (PCa). We defined the ventromedian circumference of the prostate capsule as the 12 o'clock position and the posterolateral position of the typical NVB as the 5 o'clock position. Periprostatic nerve fibers at the 12, 1, 2, 3, 4, and 5 o'clock positions of the midprostate were stimulated using bipolar electrodes. MEASUREMENTS: Changes in pressure at the middle of the urethra were measured using an inserted balloon catheter to detect increases in cavernosal pressure. RESULTS AND LIMITATIONS: Although the study included only 12 patients, every stimulation at all positions between 1 and 5 o'clock evoked urethral pressure responses in all patients. Mean pressure response was most powerful for 5 o'clock stimulation and decreased with stimulated points further from the 5 o'clock position. CONCLUSIONS: We demonstrated that electrostimulation at not only the posterolateral but also the anterior and lateral circumference of the prostate capsule can increase cavernosal pressure. These findings indicate that the periprostatic nerve network contributes to erections.

Ipsilateral seminal emission generated by electrostimulation of the lumbar sympathetic nerve during nerve sparing laparoscopic retroperitoneal lymph node dissection for testicular cancer.

PURPOSE: To confirm nerve preservation during laparoscopic retroperitoneal lymph node dissection we perform electrostimulation of the lumbar sympathetic nerves relevant to ejaculation. In recent cases we monitored seminal emission using endoscopy of the posterior urethra to observe the response to stimulation. MATERIALS AND METHODS: Six patients with testicular tumors underwent intraoperative stimulation of the lumbar sympathetic nerves during laparoscopic retroperitoneal lymph node dissection with a unilateral modified template. A long handled pair of bipolar electrodes was inserted through a laparoscopic port, and ipsilateral and contralateral lumbar sympathetic nerves were individually electrostimulated. Ipsilateral stimulation was performed at the preserved lumbar splanchnic nerves and contralateral stimulation was performed at the union of nerve fibers derived from the lumbar splanchnic nerve just above the superior hypogastric plexus. The side of generated seminal emission was monitored simultaneously by endoscopy of the posterior urethra. RESULTS: In all patients each electrostimulation generated unilateral seminal emission. Each stimulation of a lumbar splanchnic nerve, whether ipsilateral or contralateral, caused seminal emission only from the ejaculatory orifice of the stimulated side. CONCLUSIONS: Intraoperative electrostimulation of the lumbar sympathetic nerves generated only ipsilateral seminal emission. This suggests that some efferent sympathetic signals for emission might run ipsilaterally in humans.

Electrostimulation of sympathetic nerve fibers during nerve-sparing laparoscopic retroperitoneal lymph node dissection in testicular tumor.

A long-handled pair of electrodes with sufficient length to allow stimulation during laparoscopic retroperitoneal lymph node dissection (RPLND) was designed at our institute. We clinically utilized this electrode in the treatment of a 37-year-old patient with testicular tumor who underwent right orchidectomy and nerve-sparing laparoscopic RPLND. During laparoscopic RPLND, sympathetic nerve fibers relevant to ejaculation were electrically stimulated and changes in pressure at the bladder neck were observed. Nerve preservation was confirmed by increased pressure at the bladder neck and ejaculation immediately after the electrostimulation. The application of laparoscopic electrostimulation may become widespread, particularly since it meets the increasing demand for minimally invasive surgery.