PDZD8-deficient mice accumulate cholesteryl esters in the brain as a result of impaired lipophagy.

Dyslipidemia including the accumulation of cholesteryl esters (CEs) in the brain is associated with neurological disorders, although the underlying mechanism has been unclear. PDZD8, a Rab7 effector protein, transfers lipids between endoplasmic reticulum (ER) and Rab7-positive organelles and thereby promotes endolysosome maturation and contributes to the maintenance of neuronal integrity. Here we show that CEs accumulate in the brain of PDZD8-deficient mice as a result of impaired lipophagy. This CE accumulation was not affected by diet, implicating a defect in intracellular lipid metabolism. Whereas cholesterol synthesis appeared normal, degradation of lipid droplets (LDs) was defective, in the brain of PDZD8-deficient mice. PDZD8 may mediate the exchange of cholesterol and phosphatidylserine between ER and Rab7-positive organelles to promote the fusion of CE-containing LDs with lysosomes for their degradation. Our results thus suggest that PDZD8 promotes clearance of CEs from the brain by lipophagy, with this role of PDZD8 likely contributing to brain function.

Protrudin and PDZD8 contribute to neuronal integrity by promoting lipid extraction required for endosome maturation.

Endosome maturation depends on membrane contact sites (MCSs) formed between endoplasmic reticulum (ER) and endolysosomes (LyLEs). The mechanism underlying lipid supply for this process and its pathophysiological relevance remains unclear, however. Here, we identify PDZD8-the mammalian ortholog of a yeast ERMES subunit-as a protein that interacts with protrudin, which is located at ER-LyLE MCSs. Protrudin and PDZD8 promote the formation of ER-LyLE MCSs, and PDZD8 shows the ability to extract various lipids from the ER. Overexpression of both protrudin and PDZD8 in HeLa cells, as well as their depletion in mouse primary neurons, impairs endosomal homeostasis by inducing the formation of abnormal large vacuoles reminiscent of those apparent in spastin- or REEP1-deficient neurons. The protrudin-PDZD8 system is also essential for the establishment of neuronal polarity. Our results suggest that protrudin and PDZD8 cooperatively promote endosome maturation by mediating ER-LyLE tethering and lipid extraction at MCSs, thereby maintaining neuronal polarity and integrity.

[Anesthetic management of a patient with congenital myopathy complicated with severely impaired pulmonary function].

A 17-year-old man, who had received a diagnosis of congenital myopathy, was scheduled for superficial temporal artery to middle cerebral artery anastomosis procedure. Preoperative respiratory examinations showed the breathing capacity of 450 ml because of a scoilosis, deformity of a thorax and severe muscular atrophy. Anesthesia was maintained with propofol, fentanyl, remifentanil and vecuronium to avoid malignant hyperthermia (MH). Endotracheal intubation was performed with a gum elastic bougie for difficult airway management (DAM). After intubation, we checked the positioning of the tracheal tube by a chest X ray and bronchofiber findings. During perioperative period, no symptom of MH and respiratory dysfunction was noticed. In the anesthesia for patients with congenital myopathy, deterioration of respiratoy function, prevention for MH, and possibility of DAM should be considered.

Pulse steroids as induction therapy for children with ulcerative colitis.

BACKGROUND: Corticosteroids therapy, classically the first-line treatment for ulcerative colitis (UC), often causes serious side-effects. Theoretically, pulse steroid therapy where high doses are given for a shorter period may have maximal beneficial effects and minimal side-effects as induction therapy for UC. We have therefore retrospectively compared induction therapy using pulse steroids with conventional steroid treatment for children and adolescents with moderate-to-severe UC. METHODS: We utilized conventional steroid treatment (prednisolone 1-1.5 mg/kg/day) as an induction treatment in 17 UC patients between 1985 and 2006. Alternatively we used a 3-day megadose pulse steroid therapy (methylprednisolone intravenously 20-30 mg/kg/day, max. 1000 mg/day) in 20 UC patients from 1993 to 2006. RESULTS: Pulse steroid therapy successfully induced rapid remission in UC patients with moderate-to-severe disease compared with conventional treatment (13.2 days vs 25.1 days; P < 0.05). The amelioration of Pediatric Ulcerative Colitis Activity Index score between before and 1 week after pulse steroid therapy was significantly more than that of conventional treatment (P < 0.01). No serious adverse effects were observed in the patients treated with pulse steroid therapy. However, the rate of the relapse episodes during the next 12 months after pulse steroid therapy was not significantly different from that after conventional treatment. CONCLUSION: These findings suggest that pulse steroid therapy is an option to be considered in children with moderate-to-severe UC.

Peroxisome proliferator-activated receptor γ 2 mutation may cause a subset of ulcerative colitis.

AIM: Previous studies suggest the homeostasis between acquisition of tolerance to the indigenous microflora and protective immune responses appears to be disrupted in inflammatory bowel disease (IBD). Some experimental studies indicate peroxisome proliferator-activated receptor γ (PPARγ) has been implicated as a regulator of intestinal inflammatory responses. In addition, the toll-like receptor (TLR)-4 can regulate expression of PPARγ in colonic epithelial cells. We attempted to demonstrate whether the functional imbalance between TLRs and PPARγ could lead to the onset and some polymorphisms of those genes could contribute to susceptibility to IBD. METHODS: RT-PCR analysis were performed to detect TLR4 and PPARγ mRNA associated with those of P65 of NFκB, TNFα, MyD88, NOD2/CARD15, TLR-2,5,9, in the diseased colonic mucosa in ulcerative colitis (UC; n = 13) and Crohn's disease (CD; n = 7) compared with normal controls (n = 18). Consequently, we genotyped UC (n = 29) and CD (n = 10) compared with normal controls (n = 134) for the prevalence of suspicious mutations. RESULTS: In a subset of UC patients who were revealed to carry PPARγ Pro12Ala mutation later, impaired expression of normal PPARγ mRNA was noted in the diseased mucosa accompanied with upregulations of MyD88 TLR-4, 5, 9, P65 and TNFα in mRNA levels. The prevalence of PPARγ Pro12Ala mutation was more frequently found in UC patients compared with CD patients and normal controls (P < 0.05). CONCLUSIONS: These findings suggested that imbalances between TLRs and PPARγ in response to luminal bacteria could lead to colonic inflammation in some UC patients. Alternative explanations will be needed for the onset of the rest of UC and CD.

Well-controlled proinflammatory cytokine responses of Peyer's patch cells to probiotic Lactobacillus casei.

SUMMARY: In order to clarify the probiotic features of immunomodulation, cytokine production by murine spleen and Peyer's patch (PP) cells was examined in response to probiotic and pathogenic bacteria. In spleen cells, probiotic Lactobacillus casei induced interleukin (IL)-12 production by CD11b(+) cells more strongly than pathogenic Gram-positive and Gram-negative bacteria and effectively promoted the development of T helper (Th) type 1 cells followed by high levels of secretion of interferon (IFN)-gamma. Although the levels of IL-12 secreted by PP cells in response to L. casei were lower in comparison with spleen cells, Th1 cells developed as a result of this low-level induction of IL-12. However, IFN-gamma secretion by the L. casei-induced Th1 cells stimulated with a specific antigen was down-regulated in PP cells. Development of IL-17-producing Th17 cells was efficiently induced in PP cells by antigen stimulation. Lactobacillus casei slightly, but significantly, inhibited the antigen-induced secretion of IL-17 without a decrease in the proportion of Th17 cells. No bacteria tested induced the development of IL-10-producing, transforming growth factor-beta-producing or Foxp3-expressing regulatory T cells, thus suggesting that certain probiotics might regulate proinflammatory responses through as yet unidentified mechanisms in PP cells. These data show probiotic L. casei to have considerable potential to induce IL-12 production and promote Th1 cell development, but the secretion of proinflammatory cytokines such as IL-12 and IL-17 may be well controlled in PP cells.

Anti-infectious activity of synbiotics in a novel mouse model of methicillin-resistant Staphylococcus aureus infection.

The anti-infectious activity of synbiotics against methicillin-resistant Staphylococcus aureus (MRSA) infection was evaluated using a novel lethal mouse model. Groups of 12 mice treated with multiple antibiotics were infected orally with a clinical isolate of MRSA at an inoculum of 10(8) CFU on day 7 after starting the antibiotics. A dose of 400 mg/kg 5-fluorouracil (5-FU) was injected intraperitoneally on day 7 after the infection. A dose of 10(8) CFU Bifidobacterium breve strain Yakult and 10 mg of galactooligosaccharides (GOS) were given orally to mice daily with the antibiotic treatment until day 28. The intestinal population levels of MRSA in the mice on multiple antibiotics were maintained stably at 10(8) CFU/g of intestinal contents after oral MRSA infection and the subsequent 5-FU treatment killed all the mice in the group within 14 days. B. breve administration saved most of the mice, but the synbiotic treatment saved all of the mice from lethal MRSA infection. The synbiotic treatment was effective for the treatment of intestinal infection caused by four MRSA strains with different toxin productions. There was a large difference among the six Bifidobacteria strains that were naturally resistant to the antibacterial drugs used. B. breve in combination with GOS is demonstrated to have valuable preventive and curative effects against even fatal MRSA infections.

Effects of the enteral administration of Bifidobacterium breve on patients undergoing chemotherapy for pediatric malignancies.

PURPOSE: Probiotics are expected to be effective in prophylaxis of infection in cancer patient, since infections in neutropenics are mainly caused by endogenous flora through the intestinal mucosa. However, the experience with the use of probiotics in immunocompromised patients is limited, and precise fecal bacteria analysis has not been reported. The aim of the study was to evaluate the effects of the enteral administration of the probiotic, Bifidobacterium breve strain Yakult, on its ability to prevent infection, fecal micro flora, and intestinal environments in cancer patients on chemotherapy. METHODS: A placebo-controlled trial was performed at Juntendo University Hospital. Patients with malignancies admitted for chemotherapy (n = 42) were randomized into two groups receiving probiotic or placebo. The effects on infectious complications, natural killer cells, fecal micro flora, fecal organic acid concentrations, and fecal pH were studied. RESULTS: The frequency of fever and the use of intravenous antibiotics were lower in the probiotic group than the placebo group. The probiotic administration enhanced the habitation of anaerobes. Disruption of the intestinal microbiota after chemotherapy such as the increase in the population levels of Enterobacteriaceae was observed at more pronounced manner in the placebo group in comparison to the probiotic group. The concentrations of total organic acids were maintained most of the time at the normal level, which constantly maintained the pH below 7.0 only in the probiotic group. CONCLUSION: These data, although based on a limited number of patients and samples, suggest that administration of B. breve strain Yakult could be an effective approach for achieving clinical benefits in immunocompromised hosts by improving their intestinal environments.

Bispectral index during epidural puncture predicts anterograde amnesia in patients given midazolam premedication.

PURPOSE: We hypothesized anterograde amnesia could be predicted by the bispectral index (BIS) during epidural puncture in patients premedicated with intramuscular midazolam. METHODS: We investigated 64 consecutive patients undergoing gynecological laparotomy under general anesthesia combined with epidural anesthesia. Midazolam (5 mg) was administered intramuscularly at 15 min before arrival at the operating room. The anesthesiologist informed the patient of the operating room number after evaluating her using the Observer's Assessment of Alertness and Sedation (OAA/S) scale. A BIS probe was then attached to the patient's forehead while she was in the lateral position for epidural puncture. Another anesthesiologist interviewed the patient on the day after surgery and asked her the operating room number and whether there was recall of pain. Group A comprised patients with no recall of the room number and no recall of pain during epidural puncture and group R comprised patients who remembered both the room number and the pain. Patients recalling only the room number or the pain were excluded. RESULTS: Forty patients were classified as group A and 20 as group R. Four patients remembered only the room number and they were excluded. There were significant differences in body weight, OAA/S scale on arrival at the operating room, and average BIS, and electromyogram (EMG) values during epidural puncture between the two groups. These four parameters were entered into a multiple logistic regression model for multivariate analysis. The analysis identified the BIS value as the only independent predictor of complete amnesia during epidural puncture. CONCLUSION: BIS assessment during epidural puncture is informative for the anesthesiologist to predict amnesia following midazolam premedication.

Breast cancer resistance protein/ABCG2 is differentially regulated downstream of extracellular signal-regulated kinase.

Breast cancer resistance protein (BCRP)/ABCG2 is a drug efflux pump responsible for multidrug resistance in cancer cells. We report that dephosphorylation of extracellular signal-regulated kinase (ERK) by treatment with mitogen-activated protein kinase/ERK kinase (MEK) inhibitors causes two opposing effects, transcriptional upregulation and prompted protein degradation of endogenous BCRP in breast cancer MCF-7 cells. Endogenous BCRP was eventually found to be upregulated. Conversely, treatment with epidermal growth factor was associated with its downregulation in the cells. MEK inhibitors also caused prompted degradation of exogenous BCRP in MCF-7 and gastric cancer NCI-N87 cells that express exogenous BCRP without affecting its transcriptional levels, and potentiated anticancer agents in the cells. A lysosomal inhibitor abolished this prompted degradation of exogenous BCRP, but a proteasome inhibitor did not. Inhibition of p90 ribosomal protein S6 kinase (RSK), one of the downstream effectors of ERK, resulted in transcriptional upregulation of endogenous BCRP but did not affect the protein degradation of exogenous BCRP. The data suggest that BCRP expression is differentially regulated downstream of the MEK-ERK pathway, transcriptionally upregulated through the inhibition of the MEK-ERK-RSK pathway, and posttranscriptionally downregulated through the inhibition of the MEK-ERK-non-RSK pathway. Although the immediate downstream effector of ERK remains to be elucidated, the data provide new insights into regulatory mechanisms of BCRP activity and may assist the development of BCRP-specific expression modulators.

Effect of suplatast tosilate on antileukotriene non-responders with mild-to-moderate persistent asthma.

BACKGROUND: Immunomodulatory therapy has been recently introduced for the management of asthma. Suplatast tosilate (ST), a new immune-modifying drug, is known to improve the airway function by inhibiting the release of Th-2 cytokines. However, its efficacy as a controller listed in the guideline, Global Initiative for Asthma 2005 has not been established. In this study we investigated the role of ST in leukotriene receptor antagonist (LTRA) non-responders with mild-to-moderate persistent asthma before initiating corticosteroids inhalation therapy. METHODS: This was a prospective open-level clinical trial. LTRAs was given to 41 patients with asthma for 4 weeks and clinical efficacy was assessed using daily symptom scores. The 10 patients, aged 2.5-8.5 years, who failed to show clinical improvement, were defined as LTRA non-responders. After a 1-week washout period, the efficacy of ST was investigated and compared with LTRA non-responders for the following 4 weeks. RESULTS: LTRA non-responders showed a significant improvement in the average symptom score, peak expiratory flow, use of rescue medication and the proportion of symptom-free days with ST therapy. CONCLUSIONS: ST is a good choice for patients who have failed to respond to LTRAs. ST should therefore be added to the list of treatment options for such patients.

Innate immune therapy with a Bacillus Calmette-Guérin cell wall skeleton after radical surgery for non-small cell lung cancer: a case-control study.

PURPOSE: We investigated whether adjuvant immunotherapy with Bacillus Calmette-Guérin (BCG) cell wall skeleton (CWS) and surgical resection was better than resection, with or without other adjuvant therapy, for patients with non-small cell lung cancer (NSCLC). METHODS: The case group comprised 71 patients who underwent radical surgery for NSCLC, followed by BCG-CWS immunotherapy, with follow-up data available. The case-control study was designed with one control selected for each case-group patient. Each control was matched by pathological stage and year of birth (+/-5 years). BCG-CWS 200 microg was inoculated intracutaneously in the upper arm four times per week (sensitization phase); then at 4-week intervals (therapeutic phase). RESULTS: The case-group patients received 45 +/- 22.6 (average +/- SD) cycles of BCG-CWS inoculation. Overall 5-year and 10-year survival rates were 71% and 61% for the case-group patients, and 63% and 43% for the control-group patients. The survival rate of the case group was better than that of the control group (not significant; P = 0.114). The same trend was seen in the patients with stage III or N+ NSCLC (not significant; P = 0.114, P = 0.168). There were no life-threatening adverse events. CONCLUSIONS: BCG-CWS immunotherapy seemed to improve survival after resection of NSCLC, especially locally advanced NSCLC. Moreover, this immunotherapy did not compromise quality of life during treatment.

Neonatal transient eosinophilic colitis causes lower gastrointestinal bleeding in early infancy.

BACKGROUND: Lower gastrointestinal bleeding (LGB), particularly in newborns, is of serious concern and requires urgent investigation and hospital care. Whereas allergic proctocolitis caused by food protein is a significant cause of LGB in infants with eosinophilia, there are several cases of diseases with symptoms similar to those of allergic proctocolitis but without an apparent allergic reaction influence. PATIENTS AND METHODS: We examined 2 neonates using rectosigmoidoscopy who showed eosinophilia and experienced fresh LGB soon after birth and before their first feedings. Serum eosinic cationic protein (ECP) and platelet activating factor (PAF) levels were also examined in the second case to confirm the involvement of eosinophils for its pathogenesis. RESULTS: Both patients were in a clinically stable condition, and their abdomens were soft. The results of their blood analyses, abdominal radiographs, and stool cultures were normal, but they had gross eosinophilia: the eosinophil counts were 9014/mm3 (patient 1) and 1955/mm3 (patient 2). Rectosigmoidoscopy with colonic mucosal biopsy revealed nodular lymphoid hyperplasia with a pale mucosal surface and massive oozing with diffuse eosinophilic infiltration in the lamina propria. In patient 2 the serum ECP and PAF levels were elevated to 123 microg/L (normal, <14.7) and 13.1 micromol/L/min (normal, <6). A few days after intravenous hydration therapy, LGB was no longer detected, and the serum ECP and PAF levels returned to normal. CONCLUSIONS: Inasmuch as these infants had LGB similar to allergic proctocolitis without any allergic reactions, we suggest that infiltrated eosinophils in the colonic mucosa could be involved in the pathogenesis of LGB in early infancy.

Brachial-ankle pulse wave velocity and metabolic syndrome in a Japanese population: the Minoh study.

To investigate the association between brachial-ankle pulse wave velocity (baPWV) and metabolic syndrome (MS), we examined 374 men and 622 women aged 40 to 69 years who did not have a past history of either coronary heart disease or stroke. We used a modified National Cholesterol Education Program definition of MS that utilizes body mass index instead of waist circumference. Age-adjusted mean values of baPWV were greater when obesity, high systolic and diastolic blood pressures, high triglyceride level, low high-density lipoprotein cholesterol, high fasting glucose level or MS itself were present. baPWV was also associated with fasting insulin levels and homeostasis model assessment of insulin resistance (HOMA-IR) values. Mean values of baPWV (adjusted for age, smoking status, and drinking status) in men with 0, 1, 2, and > or =3 features of MS were 1,409, 1,517, 1,640, and 1,665 cm/s, respectively (p for trend <0.001). The respective adjusted mean baPWV values for women were 1,368, 1,531, 1,547, and 1,661 cm/s (p for trend <0.001). As for insulin resistance, the adjusted mean baPWV values across quartiles of HOMA-IR (lowest to highest) were 1,488, 1,514, 1,566, and 1,624 cm/s (p for trend <0.001) for men. The respective adjusted mean baPWV values for women were 1,395, 1,441, 1,464, and 1,539 cm/s (p for trend <0.001). Our findings indicate that clustered features of MS and insulin resistance are strongly associated with the risk for increased baPWV in Japanese men and women.

Association between C-reactive protein and insulin resistance in a Japanese population: the Minoh Study.

OBJECTIVE: To investigate the association between C-reactive protein (CRP) and insulin resistance. MATERIALS AND METHODS: This study included 1,624 Japanese participants (652 men and 972 women) aged 40 to 69 years who were non-diabetics or did not have medication for hypertension or dyslipidemia, a history of cardiovascular disease or CRP levels >10 mg/l. Serum CRP level, fasting glucose level, and fasting insulin level were measured, and the degree of insulin resistance was assessed using the homeostasis model assessment of insulin resistance (HOMA-IR). Categories of CRP were defined by the following tertiles: <0.25 mg/l, 0.25-0.59 mg/l, and > or = 0.60 mg/l. RESULTS: Elevated CRP levels were associated with increased fasting insulin levels, fasting glucose levels, and HOMA-IR in both men and women. Although the adjustment for body mass index in addition to age, cigarette smoking, and alcohol consumption attenuated the associations between CRP and fasting insulin, fasting glucose, and HOMA-IR, elevated CRP levels were associated with increased insulin levels and HOMA-IR in both sexes. Stratified analyses by CRP level and obesity showed that obesity status was associated with increased fasting insulin levels, fasting glucose levels, and HOMA-IR in both sexes and that fasting insulin levels, fasting glucose levels, and HOMA-IR were higher among obese individuals than among non-obese individuals at the same level of CRP. CONCLUSION: These results suggest a possible role of subclinical inflammation in insulin resistance and glucose intolerance in Japanese, but it only partly explains the link between obesity and impaired glucose homeostasis.

Association between fasting glucose and C-reactive protein in a Japanese population: the Minoh study.

To investigate the association between fasting glucose and C-reactive protein (CRP), we examined 1715 Japanese individuals (723 men and 992 women) aged 40-69 years who did not have medication for hypertension, diabetes, or dyslipidemia, a history of cardiovascular disease or CRP levels>10mg/l. There was a statistically significant unadjusted correlation between CRP and each component of the metabolic syndrome, including fasting glucose, fasting insulin, body mass index, systolic blood pressure, diastolic blood pressure, high-density lipoprotein cholesterol (negative), and triglycerides in both men and women. With adjustment for age, cigarette smoking, alcohol intake, and other components of the metabolic syndrome, the CRP increments (as back-transformed) compared with the lowest tertile of normal fasting glucose were 0.99, 1.05, 1.21, and 1.34mg/l (P for trend=0.008) with the second lowest and highest tertiles of normal fasting glucose, impaired fasting glucose, and type-2 diabetes, respectively in men. The respective adjusted CRP increments were 1.12, 1.23, 1.33, and 1.93mg/l (P for trend<0.001) in women. In the stratified analyses of CRP levels by sex, obesity status, and fasting glucose category or the number of components of the metabolic syndrome, an increase in CRP levels was greater in women than men with obesity and higher fasting glucose category (gender interaction: P<0.001) or an increased number of components of the metabolic syndrome (gender interaction: P=0.003). These results indicate that CRP levels increase continuously across the spectrum of fasting glucose in both sexes. This association is more pronounced in women.

C-reactive protein concentration is more strongly related to metabolic syndrome in women than in men: the Minoh Study.

BACKGROUND: The gender differences in the association between C-reactive protein (CRP) and features of the metabolic syndrome (MS) need to be elucidated among Japanese. METHODS AND RESULTS: The study population included 715 men and 988 women aged 40-69 years who were not taking anti-hypertensive, lipid-lowering, hypoglycemic, anti-thrombotic, or non-steroidal anti-inflammation medications, and did not have a past history of cardiovascular disease or CRP concentration >10 mg/L. Except for high-density lipoprotein cholesterol, the unadjusted correlation between CRP and each MS component, including body mass index (BMI), systolic and diastolic blood pressures, triglycerides, fasting glucose, fasting insulin, and uric acid, was greater in women than in men. With adjustment for age, smoking status, and drinking status, the differences in CRP concentrations between those with the MS components of BMI, triglycerides, and uric acid and those without were greater in women than in men. Results of stratified analyses by the number of components of the MS of 0, 1, 2, 3, and > or = 4 revealed that an increase in CRP concentrations was greater in women than men with an increased number of components of the MS (gender interaction, p = 0.005). This tendency was observed in non-smokers, but not in current smokers (gender interaction, p = 0.013 and = 0.513, respectively). CONCLUSIONS: CRP concentrations are closely related to the MS-like state in both sexes, but an increase in CRP concentration associated with risk factor-clustering is more pronounced in women, particularly non-smokers.