Tetrodotoxin Framework Construction from Linear Substrates Utilizing a Hg(OTf)2-Catalyzed Cycloisomerization Reaction: Synthesis of the Unnatural Analogue 11-nor-6,7,8-Trideoxytetrodotoxin.

In this contribution, we propose a new synthetic approach to tetrodotoxin (TTX), one of the most famous marine toxins that, after first preparing a functionalized linear substrate, forms a cyclohexane core from the substrate utilizing our mercuric triflate (Hg(OTf)2)-catalyzed cycloisomerization reaction. The concept was applied to the synthesis of 11-nor-6,7,8-trideoxyTTX and 11-nor-4,9-anhydro-6,7,8-trideoxyTTX, which are unnatural TTX analogues, demonstrating the validity of our new approach.

A novel anti-histone H1 monoclonal antibody, SSV monoclonal antibody, improves lung injury and survival in a mouse model of lipopolysaccharide-induced sepsis-like syndrome.

BACKGROUND: Histones play important roles in both host defenses and inflammation related to microbial infection. A peptide mimotope (SSV) was identified from a novel histone H1 monoclonal antibody (16G9 mAb) that was shown to inhibit the mixed lymphocyte reaction. In the present study, an anti-SSV producing hybridoma was established. We investigated the effects of SSV mAb in a mouse acute inflammation model induced by intraperitoneal injection of lipopolysaccharide (LPS). METHODS: SSV mAb was generated and characterized. Mice were treated with SSV mAb or a control IgG antibody prior to LPS injection. Evaluation of survival rate and lung tissue on histological score was performed. The levels of inflammatory cytokines and histones H1, H3, and H4 in plasma and lung tissue were measured by ELISA. RESULTS: Competitive ELISA revealed that SSV mAb binds to histone H1. SSV mAb improved lung injury and prolonged the survival of LPS-injected mice. Increased levels of histones H1, H3, and H4 and inflammatory cytokines (TNF-α, IL-1ß, and IL-6) in plasma and lung tissue after LPS injection were ameliorated by SSV mAb. CONCLUSION: SSV mAb is shown to have anti-inflammatory activity and organ-protective effects, highlighting the importance of controlling histone H1 as well as H3 and H4 levels during inflammation.

Phototherapy with artificial light suppresses dextran sulfate sodium-induced colitis in a mouse model.

BACKGROUND AND AIM: Medical treatment for inflammatory bowel disease (IBD) requires chronic administration and causes side effects. Recently, anti-inflammatory effects of phototherapy were reported in animal models. The present study evaluated whether phototherapy improves dextran sulfate sodium (DSS)-induced colitis in a mouse model of IBD. METHODS: Mice were divided into four equal groups: Control, DSS, DSS + light low (LL), and DSS + light high (LH) groups. Normal fluorescent light intensity in the Control and DSS groups was 200 lux. Artificial light intensities were as follows: DSS + LL group, 1000 lux; DSS + LH group, 2500 lux. After administering phototherapy for 7 days, we evaluated disease activity index (DAI), histological score, colon length/weight, serum 1,25-dihydroxyvitamin D(3) level, and serum and colonic cytokines in the mice. RESULTS: DAI and histological scores were significantly lower in the DSS + LL group than in the DSS group (both, P < 0.05). Colon length and weight were significantly higher in the DSS + LL group than in the DSS group (both, P < 0.05). Serum interleukin (IL)-6, TNF-α, and IL-17 in the DSS + LL group were significantly lower, and serum and colonic IL-10 were significantly higher in the DSS + LL group than in the DSS group (all, P < 0.05). Serum 1,25-dihydroxyvitamin D(3) levels in the DSS + LH group were significantly increased compared with those in the DSS + LL and DSS groups. CONCLUSION: Artificial light phototherapy suppressed DSS-induced colitis in mice by suppression of pro-inflammatory cytokines and promotion of anti-inflammatory cytokines.

New evaluation of fetal oxidative stress: measurement of the umbilical cord blood dimethyl sulfate-induced ascorbyl-free radical by an electron spin resonance method.

OBJECTIVE: The aim is to evaluate intrapartum fetal oxidative stress in real-time by umbilical cord blood dimethyl sulfate (DMSO)-induced ascorbyl-free radical (AFR) measured by an electron spin resonance (ESR) method. METHODS: Seventy-five mothers delivering at gestational age after 37 weeks were recruited. They were divided into three groups: spontaneous vaginal birth (n = 27), elective cesarean section (CS) (n = 34), and emergency CS due to non-reassuring fetal status (n = 14). Umbilical artery (UA) and venous (UV) cord blood gas analysis was performed. Serum levels of DMSO-induced AFR (AFR/DMSO) that reflect vitamin C concentrations were measured by ESR spectroscopy. RESULTS: Blood gas analysis showed no significant differences among the groups. UA-AFR/DMSO level of elective CS group was significantly lower compared with spontaneous delivery group (0.32 ± 0.12 versus 0.46 ± 0.14, p < 0.005). Emergency CS group showed significantly lower levels of UA-AFR/DMSO compared with elective CS group (0.25 ± 0.11 versus 0.32 ± 0.12, p < 0.005). UV-AFR/DMSO levels had no significant difference among the groups. CONCLUSIONS: It is suggested that fetal cord blood AFR/DMSO is a sensitive marker to assess fetal oxidative stress during delivery.

The Japanese guidelines for the management of sepsis.

This is a guideline for the management of sepsis, developed by the Sepsis Registry Committee of The Japanese Society of Intensive Care Medicine (JSICM) launched in March 2007. This guideline was developed on the basis of evidence-based medicine and focuses on unique treatments in Japan that have not been included in the Surviving Sepsis Campaign guidelines (SSCG), as well as treatments that are viewed differently in Japan and in Western countries. Although the methods in this guideline conform to the 2008 SSCG, the Japanese literature and the results of the Sepsis Registry Survey, which was performed twice by the Sepsis Registry Committee in intensive care units (ICUs) registered with JSICM, are also referred. This is the first and original guideline for sepsis in Japan and is expected to be properly used in daily clinical practice. This article is translated from Japanese, originally published as "The Japanese Guidelines for the Management of Sepsis" in the Journal of the Japanese Society of Intensive Care Medicine (J Jpn Soc Intensive Care Med), 2013; 20:124-73. The original work is at http://dx.doi.org/10.3918/jsicm.20.124.

Uncoupling of peripheral and master clock gene rhythms by reversed feeding leads to an exacerbated inflammatory response after polymicrobial sepsis in mice.

Reversed feeding uncouples peripheral and master clock gene rhythms and leads to an increased risk of disease development. The aim of this study was to determine the effects of clock gene uncoupling on sepsis-induced inflammation using a mouse cecal ligation and puncture (CLP) model. C57BL/6N mice were entrained to a 12-h light-dark cycle (lights on at 7 AM). Mice were permitted ad libitum feeding either during the night (7 PM-7 AM) or the nonphysiological light phase (7 AM-7 PM) for a week before CLP. In daytime-fed mice, phase inversion of clock gene expression was observed in the liver, but not in the suprachiasmatic nucleus. Daytime-fed mice also had decreased body weight and food intake. Survival rate was significantly lower in daytime-fed mice (29%) compared with nighttime-fed mice (54%) 72 h after CLP (P = 0.03). Serum levels of interleukin 6 (IL-6), tumor necrosis factor α, high mobility group box 1, IL-1α, IL-9, eotaxin, and granulocyte colony-stimulating factor increased in daytime-fed mice compared with nighttime-fed mice after CLP. Baseline expression levels of sirtuin peroxisome 1 and proliferator-activated receptor γ coactivator 1α in the liver decreased in daytime-fed mice compared with nighttime-fed mice. Thus, daytime feeding induces clock gene uncoupling, which leads to decreased expression of longevity-related and energy metabolism proteins. Daytime feeding may also increase the levels of inflammatory cytokines, thereby increasing mortality in a mouse sepsis model. Our findings suggest that uncoupling of peripheral and master clock gene rhythms by reversed feeding exacerbates inflammatory responses.

[Relationships between damage-associated molecular patterns and organ injury].

Recently studies have demonstrated that cell components called damage-associated molecular patterns (DAMPs) are secreted from cells in response to inflammation and organ injury. While DAMPs are maintained within the cell under normal conditions, they are secreted in response to systemic or chronic inflammation. DAMPs are recognized by pattern recognition receptors (PRRs) such as Toll-like receptor (TLR) and receptor for advanced glycation end products (RAGE). DAMPs also induce the phosphorylation of various intracellular proteins and activate NF-kappaB signaling. This induces an inflammatory response via cytokine production and activation of macrophages and dendritic cells. In essence, DAMPs alert the immune system to danger. Some DAMPs are considered therapeutic targets for acute systemic inflammation (e.g., sepsis). Indeed, anti-HMGB1 and anti-histone antibodies attenuated the inflammatory response and organ injury in a systemic inflammation model. Anti-RAGE antibodies were also shown to have beneficial effects in an animal inflammation model. These findings suggest that DAMPs may serve as novel therapeutic targets against severe systemic inflammation as well. We anticipate that in the near future, anti-DAMP therapy may become more widespread in the clinical setting.

Window current through the T-type Ca2+ channel triggers the mechanism for cellular apoptosis via mitochondrial pathways.

We hypothesized that Ca(2+) entry through the window T-type Ca(2+) current causes apoptosis. To test this hypothesis, we transfected human embryonic kidney (HEK) 293 cells to express recombinant Cav3.2 T-type Ca(2+) channels (hereafter called HEK-Cav3.2 cells). After incubation in media containing a high concentration (7.2 mM) of Ca(2+), intracellular Ca(2+) levels increased in HEK-Cav3.2 cells without electrical stimulation but not in untransfected HEK293 cells. In quiescent HEK-Cav3.2 cells exposed to high Ca(2+) media, apoptosis, as indicated by the appearance of hypodiploid cells, loss of mitochondrial transmembrane potential, and activation of caspases-3 and -9 was observed, while caspase-8 was not activated. These apoptosis-associated changes were blunted by pretreatment with the R(-)-isomer of efonidipine, a selective blocker of T-type Ca(2+) channels. High Ca(2+) did not induce apoptosis in untransfected HEK293 cells. Our findings show that Ca(2+) entry through the steady-state window current of T-type Ca(2+) channels causes apoptosis via mitochondrial pathways, and suggests that T-type Ca(2+) channels may be novel therapeutic targets for several diseases associated with abnormal apoptosis.

Bolus injection of newly synthesized vitamin E derivative ETS-GS for the treatment of acute severe ulcerative colitis in a mouse model. New vitamin E derivative for acute severe UC.

PURPOSE: Vitamin E with its antioxidant action has therapeutic effects on ulcerative colitis (UC), but use of vitamin E is limited because of its insolubility in water. We developed ETS-GS (γ-L-glutamyl-S-[2-[[[3,4-dihydro-2,5,7,8-tetramethyl-2-(4,8,12-trimethyltri-decyl)-2 H-1-benzopyran-6-yl]oxy]carbonyl]-3-oxo-3-[(2-sulfoethyl)amino]propyl]-L-cysteinylglycine sodium salt), a newly synthesized soluble vitamin E derivative with strong antioxidant action. We evaluated the therapeutic effects of bolus injection of ETS-GS on acute severe UC in a mouse model. METHODS: An animal model of acute severe UC was induced by feeding mice 5 % dextran sulfate sodium (DSS) for 5 days, followed by 1 % DSS on days 5-8, the experimental period. ETS-GS or saline was administered by subcutaneous bolus injection during the experimental period. We examined disease activity index (DAI) score, histological score, colon length, colon weight, and serum cytokines in the mice. RESULTS: The following results at day 8 in the DSS + ETS-GS group were significantly lower than those in the DSS + Saline group: DAI score, 2.6 ± 0.6 vs. 3.1 ± 0.5; histological score, 2.1 ± 1.0 vs. 3.1 ± 0.8; serum interleukin (IL)-6, 15 ± 9.4 vs. 39 ± 23 pg/ml; and keratinocyte-derived chemokine (KC), 122 ± 61 vs. 228 ± 66 pg/ml (P < 0.05). Colon length, colon weight, and serum IL-10 in the DSS + ETS-GS group were significantly higher than those in the DSS + Saline group (88 ± 12 vs. 75 ± 5.7 mm, 0.48 ± 0.09 vs. 0.38 ± 0.05 g, and 55 ± 18 vs. 31 ± 10 pg/ml, respectively; P < 0.05). CONCLUSIONS: Bolus injection of ETS-GS may be one therapeutic modality for acute severe UC. Its effects are associated with suppression of serum IL-6 and serum KC and promotion of serum IL-10.

Should nicorandil infusion be adapted in dialysis-dependent patients undergoing continuous renal replacement therapy after cardiac surgery?

In this report, we studied whether plasma concentration of nicorandil is maintained effectively and safely in dialysis-dependent patients with stage 5 chronic kidney disease (CKD5D) undergoing continuous renal replacement therapy (CRRT). Participants consisted of 10 patients undergoing CRRT after cardiac surgery. CRRT was performed with an effluent flow rate of either 600 mL/h (low-flow group; n = 5) or 1800 mL/h (high-flow group; n = 5). Nicorandil was infused intravenously at 0.1 mg/kg/h for more than 15 h starting 8 h before and 7 h after the start of CRRT. Plasma nicorandil concentrations were measured from arterial blood lines 1 h before and 7 h after CRRT initiation. Nicorandil clearance by CRRT was also calculated 1 h after CRRT initiation. Nicorandil plasma concentrations before and 7 h after CRRT initiation were 68.0 ng/mL and 74.6 ng/mL, respectively. Nicorandil clearance 1 h after CRRT initiation was 20.2 mL/min. Increasing the effluent flow rate from 600 mL/h to 1800 mL/h tended to increase nicorandil clearance. When nicorandil was infused intravenously during CRRT at 0.1 mg/kg/h in patients with CKD5D, plasma nicorandil concentrations were maintained within an effective concentration range.

[Severe hypotension and ventricular fibrillation during combined general and epidural anesthesia in a patient on major tranquilizers].

A 54-year-old man (height 155 cm, weight 49 kg) was scheduled for retroperitoneoscopic nephrectomy. He had a history of schizophrenia that had been controlled with propericiazine 10 mg and bromperidol 3 mg daily for 34 years. After induction of anesthesia, 1% mepivacaine 5 ml was administered via an epidural catheter. Blood pressure decreased 15 minutes later to 47/25 mmHg and heart rate dropped to 50 beats x min(-1). Ventricular fibrillation occurred despite titrated injection of ephedrine (40 mg total), phenylephrine (1 mg total), atropine (0.5 mg total), and rapid infusion of crystalloid and colloid solutions. Chest compression and defibrillation were required to restore spontaneous circulation. Surgery was cancelled and he was extubated 45 minutes later without any complications. These findings suggest that caution must be exercised when combining general and epidural anesthesia for patients on long-term major tranquilizers. In the event of refractory hypotension, the use of direct-acting vasoconstrictors such as noradrenaline or vasopressin should be considered.

Ultrasound visibility of regional anesthesia catheters: an in vitro study.

BACKGROUND: Ultrasound subjective visibility of in-plane needles is correlated with the intensity difference between the needle surface and the background. Regional anesthesia catheters are difficult to visualize by an ultrasound. In the present study, we investigated the ultrasound visibility of the catheters. METHODS: Six catheters were placed at 0° and 30° relative to and at a depth of 1 cm below the pork phantom surface. Ultrasound images of in-plane catheters were evaluated, subjectively and objectively. Outer and inner objective visibilities were defined as the difference in the mean pixel intensity between the catheter surface and adjacent background, and between the surface and the center of the catheter, respectively. Evaluations were made based on the portion of the catheters. A P value < 0.05 was considered significant. RESULTS: Subjective visibility was more strongly correlated with the inner objective visibility than with the outer objective visibility at both angles. Metallic 19-gauge catheters were more subjectively visible than the non-metallic 20-gauge catheters at 30° degrees (P < 0.01). Subjective, and outer and inner objective visibility were significantly lower at 30° than at 0° (P < 0.01, P < 0.01, P = 0.02). Perifix ONE at 0° and Perifix FX at 30° were the most visible catheters (P < 0.01 for both). CONCLUSIONS: Subjective visibility of catheters can not be evaluated in the same manner as that of the needles. For the best possible visualization, we recommend selecting a catheter with a structure that enhances the dark at the center of catheter, rather than basing the catheter selection on the bore size.

Use of perioperative ureteral stent in abdominal aortic aneurysm with retroperitoneal fibrosis - A report of two cases -.

Retroperitoneal fibrosis is associated with fibroblast proliferation due to inflammatory changes in adipose/fibrous tissue. Given that aortic dilation in abdominal aortic aneurysm can cause compression of the ureter, abdominal aortic aneurysm complicated by retroperitoneal fibrosis is likely to result in urinary tract obstruction. Accordingly, close attention to changes in perioperative urine volume is warranted when operating on patients with abdominal aortic aneurysm complicated by retroperitoneal fibrosis. We have recently performed laparotomies on two cases of abdominal aortic aneurysm complicated by retroperitoneal fibrosis. In the first case, surgery was performed without the placement of a ureteral stent. The patient developed postrenal acute renal failure caused by postoperative urinary retention. In the second case, ureteral stent placement in advance enabled perioperative management without complications. The clinical course of these cases suggests that, in laparotomy with concomitant retroperitoneal fibrosis, preoperative ureteral stent placement can prevent postoperative complications in the renal and urinary systems.

Establishment of a model of atrial fibrillation associated with chronic kidney disease in rats and the role of oxidative stress.

BACKGROUND: An animal model of atrial fibrillation (AF) associated with chronic kidney disease (CKD) has not been available. OBJECTIVE: The purpose of this study was to test the validity of 5/6 nephrectomy (5.6Nx) as an appropriate model of AF associated with CKD and to investigate the role of oxidative stress. METHODS: Male Sprague-Dawley rats were subjected to 5.6Nx. A novel derivative of lipoic acid, sodium zinc dihydrolipoylhistidinate (DHLHZn), was subcutaneously infused. Four weeks later, hearts were isolated. RESULTS: We observed 5 main findings. (1) 5.6Nx induced renal dysfunction with elevation of systolic blood pressure and impaired glucose tolerance. (2) In the left atrium (LA), expressions of α-smooth muscle action and collagen type I, the compositional proteins of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, and malondialdehyde were increased by 5.6Nx, which was reversed by DHLHZn treatment. (3) In the LA, the tissue content of angiotensin II was elevated by 5.6Nx, which was also reversed by DHLHZn. (4) Masson trichrome staining revealed that heterogeneous LA interstitial fibrosis was induced by 5.6Nx, which was attenuated by DHLHZn. (5) In isolated perfused heart experiments, 5.6Nx caused slowing of interatrial conduction. In the hearts of rats of the 5.6Nxgroup, right atrial extrastimuli invariably induced AF (8/8 [100%]), which were suppressed by DHLHZn (3/8 [38%], P <.05). CONCLUSION: We successfully established an appropriate model of AF associated with CKD in rats. Because the amount of NADPH oxidase was increased and the potent antioxidant agent DHLHZn was effective, oxidative stress may be involved in the pathogenesis of LA fibrosis and enhanced AF vulnerability in our model.

[Reappraisal of current practice of preoperative fasting in Japan].

BACKGROUND: The American Society of Anesthesiologists (ASA) published a clinical practice guideline of preoperative fasting in 1999. A nationwide survey conducted in Japan in 2003 reveals that many hospitals have a much longer fasting period. We conducted a similar survey in three limited areas in Japan to assess the changes in fasting practice. METHODS: A written questionnaire for preoperative fasting was sent to 50 hospital in 3 prefectures. RESULTS: The duration of fasting for liquids tends to be shorter than those in the 2003 survey. The rates of application of the ASA guideline, however, are still low specifically in adults (4.2%), which is significantly lower than those in children (17.7%), or in infants (39.0%). The reasons for noncompliance are mainly due to organizational problems associated with scheduling of operation. Most hospitals aspire to have Japanese guideline about preoperative fasting periods. CONCLUSIONS: Longer preoperative fasting periods are still common practice in Japanese hospitals.

New evaluation of fetal oxidative stress: measurement of the umbilical cord blood dimethyl sulfate-induced ascorbyl free radical by an electron spin resonance method.

OBJECTIVE: The aim is to evaluate intrapartum fetal oxidative stress in real-time by umbilical cord blood dimethyl sulfate (DMSO)-induced ascorbyl free radical (AFR) measured by an electron spin resonance (ESR) method. METHODS: 75 mothers delivering at gestational age after 37 weeks were recruited. They were divided into three groups: spontaneous vaginal birth (n = 27), elective cesarean section (CS) (n = 34), and emergency CS due to non-reassuring fetal status (n = 14). Umbilical artery (UA) and venous (UV) cord blood gas analysis was performed. Serum levels of DMSO-induced AFR (AFR/DMSO) that reflect vitamin C concentrations, was measured by ESR spectroscopy. RESULTS: Blood gas analysis showed no significant differences among the groups. UA-AFR/DMSO level of elective CS group was significantly lower compared with spontaneous delivery group (0.32 ± 0.12 vs. 0.46 ± 0.14, p < 0.005). Emergency CS group showed significantly lower levels of UA-AFR/DMSO compared with elective CS group (0.25 ± 0.11 vs. 0.32 ± 0.12, p < 0.005). UV-AFR/DMSO levels had no significant difference among the groups. CONCLUSIONS: It is suggested that fetal cord blood AFR/DMSO is a sensitive marker to assess fetal oxidative stress during delivery.

The vitamin E derivative, EPC-K1, suppresses inflammation during hepatic ischemia-reperfusion injury and exerts hepatoprotective effects in rats.

BACKGROUND: An important component of postoperative management includes alleviation of hepatic ischemia-reperfusion (I/R) injury, which commonly results from liver surgery. EPC-K1 is a hydroxyl radical scavenger reported to have mitigating effects on I/R injury in many organs. This study evaluates the effects of EPC-K1 on hepatic I/R injury. MATERIALS AND METHODS: Rats were injected subcutaneously with either EPC-K1 (100 mg/kg) or saline. The hepatic artery and left branch of the portal vein were clamped for 45 min under general anesthesia. Indicators of liver function, including aspartate aminotransferase (AST), alanine aminotransferase (ALT), and lactate dehydrogenase (LDH), and of liver tissue damage were evaluated after 6h and 24h of reperfusion. Serum levels of tumor necrosis factor (TNF)-α, interleukin (IL)-6, and high-mobility group box 1 (HMGB1) protein were measured, and apoptosis was quantified via caspase 3/7 activity and TUNEL assay. RESULTS: AST, ALT, and LDH levels increased significantly as a result of hepatic I/R injury, but were attenuated by EPC-K1 administration. Histologic findings revealed that normal structure of the hepatic parenchyma was maintained in rats pretreated with EPC-K1. TNF-α, IL-6, and HMGB1 levels rose significantly after reperfusion, together with activation of the inflammatory response. However, EPC-K1 administration suppressed levels of inflammatory markers and attenuated the inflammatory response. Moreover, EPC-K1 administration prevented apoptosis as determined by inhibition of caspase 3/7 activity and a decrease in apoptotic cells. CONCLUSIONS: Results demonstrate that EPC-K1 inhibits the inflammatory response and suppresses apoptosis during hepatic I/R injury. This suggests that EPC-K1 has hepatoprotective effects, and may be a valuable and novel therapeutic agent in the clinical setting.

New anthranilic acid derivative, EAntS-GS, attenuates Freund's complete adjuvant-induced acute pain in rats.

BACKGROUND: Acute and chronic pain commonly accompany various clinical conditions such as contusion, fracture, osteoarthritis, peripheral neuropathy, and postherpetic neuralgia. Recent studies have found that antioxidative drugs can have analgesic effects. The present study tested the hypothesis that a new anthranilic acid derivative, EAntS-GS, exerts antinociceptive effects on inflammatory pain in a rat model. METHODS: We induced subacute pain with a plantar injection of Freund's complete adjuvant (FCA) in Sprague-Dawley rats. EAntS-GS (1 mg/kg subcutaneous injection or 1% application) was administered every 12 h beginning 24 h after FCA administration, and the plantar test was used to determine its effect on pain. Levels of myeloperoxidase, inducible nitric oxide synthase (iNOS), and protease activated receptor 2 (PAR2) were measured to elucidate the mechanism of action of EAntS-GS. RESULTS: EAntS-GS significantly reduced FCA-induced pain and myeloperoxidase, iNOS, and PAR2 levels. Our findings suggest that the new anthranilic acid derivative, EAntS-GS, exerts antinociceptive effects, and that the mechanism involves iNOS and PAR2. CONCLUSION: We conclude that EAntS-GS should be considered a new therapeutic tool to treat acute and chronic pain.