Challenges in the diagnosis of the enigmatic primary adrenal leiomyosarcoma: two case reports and review of the literature.

BACKGROUND: Primary adrenal leiomyosarcoma is a rare and aggressive mesenchymal tumor derived from the smooth muscle wall of a central adrenal vein or its tributaries; therefore, tumors tend to invade the inferior vena cava and cause thrombosis. The great majority of tumors grow rapidly, which makes the disease difficult to diagnose in its early clinical stages and needs differentiation from adrenocortical carcinomas for the selection of chemotherapy including mitotane which causes adrenal insufficiency. CASE PRESENTATION: We presented two patients with adrenal leiomyosarcoma who were referred to our hospital with abdominal pain and harboring large adrenal tumors and inferior vena cava thrombosis. The endocrine findings, including serum catecholamine levels, were unremarkable. These two patients were considered clinically inoperable, and CT-guided core needle biopsy was performed to obtain the definitive histopathological diagnosis and determine the modes of therapy. The masses were subsequently diagnosed as primary adrenal leiomyosarcoma based on the histological features and positive immunoreactivity for SMA (smooth muscle actin), desmin, and vimentin. CONCLUSIONS: Adrenal leiomyosarcoma derived from the smooth muscle wall of a central adrenal vein or its tributaries is rare but should be considered a differential diagnosis in the case of nonfunctioning adrenal tumors extending directly to the inferior vena cava. CT-guided biopsy is considered useful for histopathological diagnosis and clinical management of patients with inoperable advanced adrenal tumors without any hormone excess.

A greater effect on clarithromycin resistance of mef(A)-associated msr(D) than mef(E)-associated msr(D) in Streptococcus pyogenes.

The mef(A)- and its subclass mef(E) systems had long been considered to constitute one of the primary macrolide-resistant mechanisms in Streptococcus pyogenes. However, we have previously demonstrated that the msr(D) gene located immediately downstream of the mef(A)/mef(E) genes plays a predominant role in these systems. In previous studies, furthermore, mef(A)-associated msr(D)10-85 of an S. pyogenes strain (10-85) exhibited a greater increase in clarithromycin minimum inhibitory concentration (MIC) than mef(E)-associated msr(D)13-O-10 of another strain (13-O-10). Both msr(D) genes encode 487 amino acid residues, 13 amino acid residues of which are different from each other. In this study, we performed mutational analysis of the msr(D) genes and showed that a single-nucleotide polymorphism to cause a substitution of Asp238 with Gly is mainly associated with the greater increase in clarithromycin MIC by the msr(D)10-85 than by the msr(D)13-O-10 allele. In addition, another substitution of Ser with Arg at codon 194 is partially associated with the greater increase by the msr(D)10-85 than by the msr(D)13-O-10 allele.

Streptococcus pyogenes TrxSR Two-Component System Regulates Biofilm Production in Acidic Environments.

Bacteria form biofilms for their protection against environmental stress and produce virulence factors within the biofilm. Biofilm formation in acidified environments is regulated by a two-component system, as shown by studies on isogenic mutants of the sensor protein of the two-component regulatory system in Streptococcus pyogenes. In this study, we found that the LiaS histidine kinase sensor mediates biofilm production and pilus expression in an acidified environment through glucose fermentation. The liaS isogenic mutant produced biofilms in a culture acidified by hydrochloric acid but not glucose, suggesting that the acidified environment is sensed by another protein. In addition, the trxS isogenic mutant could not produce biofilms or activate the mga promoter in an acidified environment. Mass spectrometry analysis showed that TrxS regulates M protein, consistent with the transcriptional regulation of emm, which encodes M protein. Our results demonstrate that biofilm production during environmental acidification is directly under the control of TrxS.

Association of urinary free cortisol with bone formation in patients with mild autonomous cortisol secretion.

CONTEXT: Mild autonomous cortisol secretion (ACS) is associated with an increased risk of vertebral fractures (VFx). However, the influence of this condition on bone turnover or its association with mild ACS is still controversial. OBJECTIVE: This study aimed to evaluate the impact of mild ACS on bone quality among patients living with the disease. DESIGN AND SETTING: A retrospective study was conducted using data from 55 mild ACS and 12 nonfunctioning adrenal tumour (NFT) patients who visited Chiba University Hospital, Japan, from 2006 to 2018. PATIENTS AND MAIN OUTCOME MEASURES: We analysed clinical features and bone-related factors, including bone mineral density (BMD) and VFx, performed blood tests to assess bone metabolism markers in patients with mild ACS and NFT, and assessed the associations between bone-related markers and endocrinological parameters in patients with mild ACS. RESULTS: No significant differences between mild ACS and NFT patients were observed with respect to the presence or absence of VFx and BMD. Urinary free cortisol (UFC) was higher in mild ACS patients with VFx than those without (p = .037). The T-score and young adult mean (YAM) of the BMD of the femoral neck in mild ACS patients with a body mass index <25 were positively correlated with dehydroepiandrosterone sulphate levels (ρ: 0.42, p = .017; ρ: 0.40, p = .024, respectively). Pearson's correlation analysis showed that bone-specific alkaline phosphatase was negatively correlated with UFC in the patients with mild ACS (ρ: -0.37, p = .026). CONCLUSIONS: These results suggest that urinary free cortisol may be useful for predicting bone formation in mild ACS patients.

p53-inducible DPYSL4 associates with mitochondrial supercomplexes and regulates energy metabolism in adipocytes and cancer cells.

The tumor suppressor p53 regulates multiple cellular functions, including energy metabolism. Metabolic deregulation is implicated in the pathogenesis of some cancers and in metabolic disorders and may result from the inactivation of p53 functions. Using RNA sequencing and ChIP sequencing of cancer cells and preadipocytes, we demonstrate that p53 modulates several metabolic processes via the transactivation of energy metabolism genes including dihydropyrimidinase-like 4 (DPYSL4). DPYSL4 is a member of the collapsin response mediator protein family, which is involved in cancer invasion and progression. Intriguingly, DPYSL4 overexpression in cancer cells and preadipocytes up-regulated ATP production and oxygen consumption, while DPYSL4 knockdown using siRNA or CRISPR/Cas9 down-regulated energy production. Furthermore, DPYSL4 was associated with mitochondrial supercomplexes, and deletion of its dihydropyrimidinase-like domain abolished its association and its ability to stimulate ATP production and suppress the cancer cell invasion. Mouse-xenograft and lung-metastasis models indicated that DPYSL4 expression compromised tumor growth and metastasis in vivo. Consistently, database analyses demonstrated that low DPYSL4 expression was significantly associated with poor survival of breast and ovarian cancers in accordance with its reduced expression in certain types of cancer tissues. Moreover, immunohistochemical analysis using the adipose tissue of obese patients revealed that DPYSL4 expression was positively correlated with INFg and body mass index in accordance with p53 activation. Together, these results suggest that DPYSL4 plays a key role in the tumor-suppressor function of p53 by regulating oxidative phosphorylation and the cellular energy supply via its association with mitochondrial supercomplexes, possibly linking to the pathophysiology of both cancer and obesity.

Steroid metabolites for diagnosing and predicting clinicopathological features in cortisol-producing adrenocortical carcinoma.

BACKGROUND: Approximately 60% of adrenocortical carcinomas (ACC) are functional, and Cushing's syndrome is the most frequent diagnosis that has been revealed to have a particularly poor prognosis. Since 30% of ACC present steroid hormone-producing disorganization, measurement of steroid metabolites in suspected ACC is recommended. Previous reports demonstrated that steroid hormone precursors or their urine metabolites, which can be assessed using liquid chromatography tandem mass spectrometry (LC-MS/MS) or gas chromatography mass spectrometry (GC-MS) respectively, are useful for distinguishing ACC from cortisol-producing adenomas (CPA); however, despite high precision, LC-MS/MS and GC-MS require a highly trained team, are expensive and have limited capacity. METHODS: Here, we examined 12 serum steroid metabolites using an immunoassay, which is a more rapid and less costly method than LC-MS/MS, in cortisol-producing ACC and CPA. Further, the correlation of each steroid metabolite to the classification stage and pathological status in ACC was analyzed. RESULTS: Reflecting disorganized steroidogenesis, the immunoassay revealed that all basal levels of steroid precursors were significantly increased in cortisol-producing ACC compared to CPA; in particular, 17-hydroxypregnenolone (glucocorticoid and androgen precursor) and 11-deoxycorticosterone (mineralocorticoid precursor) showed a large area under the ROC curve with high sensitivity and specificity when setting the cut-off at 1.78 ng/ml and 0.4 mg/ml, respectively. Additionally, a combination of androstenedione and DHEAS also showed high specificity with high accuracy. In cortisol-producing ACC, 11-deoxycortisol (glucocorticoid precursor) showed significant positive correlations with predictive prognostic factors used in ENSAT classification, while testosterone showed significant positive correlations to the Ki67-index in both men and women. CONCLUSION: Less expensive and more widely available RIA and ECLIA may also biochemically distinguish ACC from CPA and may predict the clinicopathological features of ACC.

New index of abdominal obesity, a body shape index, is BMI-independently associated with systemic arterial stiffness in real-world Japanese population.

OBJECTIVE: To clarify that the new index of abdominal obesity, a body shape index (ABSI), is associated with obesity-related metabolic disorders and arterial stiffness. MATERIALS: We analyzed the cross-sectional data from 62,514 Japanese subjects (mean age 44.4 years, mean body mass index (BMI) 22.2 kg/m2) without a past history of cardiovascular disease, stroke, or treatment for obesity-related metabolic disorders. METHODS: Various body adiposity indices including BMI, waist circumference (WC), and ABSI were evaluated for abilities to indicate metabolic disorders and arterial stiffness assessed by cardio-ankle vascular index (CAVI). RESULTS: WC, WC/height ratio, and WC/BMI ratio correlated with BMI regardless of gender or obesity, whereas ABSI hardly correlated with BMI. ROC analyses demonstrated that ABSI had the highest discriminatory power in predicting high CAVI (≥ 90th percentile) compared to other body adiposity indices, and the cut-off value was 0.080. Increases in ABSI as well as BMI reflected severity of metabolic disorders. After adjusting for confounders identified by multiple regression analysis, adjusted CAVI correlated positively with ABSI, whereas an inverted relationship was observed between adjusted CAVI and BMI. Additionally, the contribution of high ABSI (≥ 0.080) for high CAVI was independent of gender, age, obesity, and obesity-related metabolic disorders in the multivariate logistic regression model. CONCLUSION: ABSI is an easily calculated index of abdominal obesity which reflects metabolic disorders and systemic arterial stiffening, and may be useful in primary health screening even without any medical equipment for visceral fat quantification.

Prevalence of emm1 Streptococcus pyogenes having a novel type of genomic composition.

Streptococcus pyogenes is a causative agent of streptococcal toxic shock syndrome (STSS). The complete genome sequence of a S. pyogenes strain 10-85 isolated from a STSS patient was recently announced. In this study, the genome sequence was dissected and it was found that the genomic region around 200 kbp (region A) and the genomic region around 1600 kbp (region B) were replaced by each other in strain 10-85, when compared with those in reference strains SF370 and A20. In order to address whether this replacement is unique to 10-85, we further analyzed 163 emm1-type strains. The results indicated that none of the strains isolated before 1990 had the replacement. In contrast, most of the strains isolated at least after 2000 appeared to have the 10-85-type replacement.

7-Ketocholesterol induces ROS-mediated mRNA expression of 12-lipoxygenase, cyclooxygenase-2 and pro-inflammatory cytokines in human mesangial cells: Potential role in diabetic nephropathy.

7-Ketocholesterol (7-KCHO) is a highly proinflammatory oxysterol and plays an important role in the pathophysiology of diabetic nephropathy (DN). Lipoxygenases (LOXs) and cyclooxygenases (COXs) are also involved in the development of DN. The aim of this study was to clarify the effects of 7-KCHO on mRNA expression of LOXs and COXs as well as pro-inflammatory cytokines in human mesangial cells (HMC). We evaluated cell viability by WST-8 assay and measured mRNA expression by reverse transcription-polymerase chain reaction. Intracellular reactive oxygen species (ROS) production was evaluated by flow cytometry. Although 7-KCHO did not affect cell viability of HMC, 7-KCHO stimulated significant increases in mRNA expression of 12-LOX, COX-2 and pro-inflammatory cytokines. 7-KCHO also induced an increase in ROS production, while N-acetylcysteine partially suppressed the increase. The 12-LOX and COX-2 inhibitors also suppressed mRNA expression of cytokines. These findings may contribute to the elucidation of the molecular mechanism of the pathophysiology of DN.

Fracture risk assessment tool (FRAX) and for the diagnosis of osteoporosis in Japanese middle-aged and elderly women: Chiba bone survey.

Osteoporosis not only increases bone fracture risk but also affects survival in postmenopausal women. Although osteoporosis is diagnosed based on low bone mineral density (BMD) determined by dual energy X-ray absorptiometry (DXA), BMD measurement is sometimes difficult because DXA is not widely available in the community. The Fracture Risk Assessment tool (FRAX) can predict 10-year major osteoporotic fracture risk and hip fracture risk with or without femoral neck BMD. The FRAX has not been investigated adequately in community-dwelling Japanese women. We administered the FRAX tool in 13,421 Japanese women who underwent DXA-based forearm BMD measurement in Chiba Bone Survey, a population-based, multicenter, cross-sectional study of postmenopausal osteoporosis conducted in Chiba, Japan. Mean age was 57.77 ± 9.24 years. Mean forearm BMD was 87.94 ± 17.00% of young adult mean (YAM). Mean FRAX major osteoporotic fracture risk without femoral neck BMD was 7.06 ± 5.22%. BMD decreased and percentage of osteoporosis increased from age 55 onward. Age distribution of percentage of subjects with FRAX major osteoporotic fracture risk >15% was similar to that of percentage of osteoporosis subjects. We identified the cutoff value of FRAX major osteoporotic fracture risk for diagnosis of osteoporosis as 7.2%. With this cutoff, the positive likelihood ratio was over 1.0 at age 55 and above but accuracy was low. In conclusion, FRAX without femoral neck BMD reflects bone status, and may be useful to diagnose osteoporosis in Japanese women aged 55 and above, although the sensitivity was low for osteoporosis screening, especially in middle-aged women.

[Body weight and bone/calcium metabolism. Response of bone metabolism after bariatric surgery.]

While bariatric surgery is an effective treatment for severe obesity, these procedures may have unfavorable effects on bone and mineral metabolism. The most pronounced abnormalities are disorders of calcium/vitamin D metabolism, bone loss, as well as increased risk of bone fracture. These changes that occur after surgery vary by procedure type. Proposed mechanisms include skeletal unloading, malabsorption, secondary hyperparathyroidism and changes in gut hormones. This review explores bone response to bariatric surgery, potential mechanisms for these changes.

Resveratrol Ameliorates Arterial Stiffness Assessed by Cardio-Ankle Vascular Index in Patients With Type 2 Diabetes Mellitus.

Resveratrol has been reported to have potent anti-atherosclerotic effects in animal studies. However, there are few interventional studies in human patients with atherosclerogenic diseases. The cardio-ankle vascular index (CAVI) reflects arterial stiffness and is a clinical surrogate marker of atherosclerosis. The aim of the present study was to investigate the effect of resveratrol on arterial stiffness assessed by CAVI in patients with type 2 diabetes mellitus (T2DM).In this double-blind, randomized, placebo-controlled study, 50 patients with T2DM received supplement of a 100mg resveratrol tablet (total resveratrol: oligo-stilbene 27.97 mg/100 mg/day) or placebo daily for 12 weeks. CAVI was assessed at baseline and the end of study. Body weight (BW), blood pressure (BP), glucose and lipid metabolic parameters, and diacron-reactive oxygen metabolites (d-ROMs; an oxidative stress marker) were also measured.Resveratrol supplementation decreased systolic BP (-5.5 ± 13.0 mmHg), d-ROMs (-25.6 ± 41.8 U.CARR), and CAVI (-0.4 ± 0.7) significantly (P < 0.05) and decreased BW (-0.8 ± 2.1 kg, P = 0.083) and body mass index (-0.5 ± 0.8 kg/m2, P = 0.092) slightly compared to baseline, while there were no significant changes in the placebo group. Decreases in CAVI and d-ROMs were significantly greater in the resveratrol group than in the placebo group. Multivariate logistic regression analysis identified resveratrol supplementation as an independent predictor for a CAVI decrease of more than 0.5.In conclusion, 12-week resveratrol supplementation may improve arterial stiffness and reduce oxidative stress in patients with T2DM. Resveratrol may be beneficial in preventing the development of atherosclerosis induced by diabetes. However, a large-scale cohort study is required to validate the present findings.

Predominant role of msr(D) over mef(A) in macrolide resistance in Streptococcus pyogenes.

In Japan, the number of patients with streptococcal toxic shock syndrome is reported to be increasing. mef(A) gene-positive macrolide-resistant emm1 strains are thought to possibly contribute to the rise in the frequency of STSS. Although analyses of macrolide-resistant mechanisms, including mef(A) resistance, have been performed mainly in Streptococcus pneumoniae, the role of this gene in Streptococcus pyogenes has not been completely investigated. Therefore, to the best of our knowledge, we established the first mef(A)-knockout strain using an emm1-type S. pyogenes strain, and tested its susceptibility to erythromycin, clarithromycin and azithromycin. We found that the antimicrobial susceptibilities were almost identical to those of the parental strain. Hence, we established a knockout strain for another gene, msr(D), that is located immediately downstream of mef(A). The macrolide resistances of the resulting strain significantly decreased, and were further altered when both mef(A) and msr(D) were knocked out. The introduction of the msr(D) gene into a macrolide-sensitive strain conferred more resistance than the introduction of the mef(A) gene. The erythromycin susceptibilities of knockout strains were further dissected using two additional emm4- and emm75-type S. pyogenes strains. We found almost identical results for both strains except for the mef(A) knockout emm4 type, whose susceptibility was altered, although the change was less than that for the msr(D) knockout. These results suggest that both mef(A) and msr(D) are involved in macrolide resistance in S. pyogenes, and that the msr(D) gene plays a more predominant role in macrolide resistance than mef(A).

Two-Component Systems Involved in Susceptibility to Nisin A in Streptococcus pyogenes.

UNLABELLED: Two-component systems (TCSs) are regulatory systems in bacteria that play important roles in sensing and adapting to the environment. In this study, we systematically evaluated the roles of TCSs in the susceptibility of the group A Streptococcus (GAS; Streptococcus pyogenes) SF370 strain to several types of lantibiotics. Using individual TCS deletion mutants, we found that the deletion of srtRK (spy_1081-spy_1082) in SF370 increased the susceptibility to nisin A, which is produced by Lactococcus lactis ATCC 11454, but susceptibility to other types of lantibiotics (nukacin ISK-1, produced by Staphylococcus warneri, and staphylococcin C55, produced by Staphylococcus aureus) was not altered in the TCS mutants tested. The expression of srtFEG (spy_1085 to spy_1087), which is located downstream of srtRK and is homologous to ABC transporters, was increased in response to nisin A. However, srtEFG expression was not induced by nisin A in the srtRK mutant. The inactivation of srtFEG increased the susceptibility to nisin A. These results suggest that SrtRK controls SrtFEG expression to alter the susceptibility to nisin A. Further experiments showed that SrtRK is required for coexistence with L. lactis ATCC 11454, which produces nisin A. Our results elucidate the important roles of S. pyogenes TCSs in the interactions between different bacterial species, including bacteriocin-producing bacteria. IMPORTANCE: In this study, we focused on the association of TCSs with susceptibility to bacteriocins in S. pyogenes SF370, which has no ability to produce bacteriocins, and reported two major new findings. We demonstrated that the SrtRK TCS is related to susceptibility to nisin A by controlling the ABC transporter SrtFEG. We also showed that S. pyogenes SrtRK is important for survival when the bacteria are cocultured with nisin A-producing Lactococcus lactis This report highlights the roles of TCSs in the colocalization of bacteriocin-producing bacteria and non-bacteriocin-producing bacteria. Our findings provide new insights into the function of TCSs in S. pyogenes.

GERMLINE DELETION OF ARMC5 IN FAMILIAL PRIMARY MACRONODULAR ADRENAL HYPERPLASIA.

OBJECTIVE: Primary macronodular adrenal hyperplasia (PMAH) is considered a predominantly sporadic disease, but familial forms are well recognized. Genetic studies revealed germline mutations in the armadillo repeat containing 5 gene (ARMC5) in the majority of PMAH cases. Furthermore, somatic ARMC5 mutations, as different types of second-hit mutations and loss of heterozygosity have been reported in each adrenal nodule in PMAH. Here, we describe the involvement of ARMC5 alteration in a familial case of PMAH. METHODS: In our study, we performed clinical and genetic evaluations in a mother and her son with familial PMAH. To search for mutations and deletion of ARMC5, we used Sanger sequencing and droplet digital polymerase chain reaction (ddPCR), respectively. RESULTS: Both patients showed the same phenotype of subclinical Cushing syndrome, with mild excess of mineralocorticoids and vasopressin-responsive cortisol secretion. The ddPCR analysis demonstrated that both mother and son had germline deletions in exons 1 to 5 of the ARMC5 gene locus. Furthermore, Sanger sequencing of DNA from the right and left adrenal nodules as well as peripheral blood of the son revealed the presence of another germline, missense mutation in ARMC5 exon 3 (p.P347S). CONCLUSION: This is the first report demonstrating germline deletion of ARMC5 in familial PMAH. In addition to investigating mutations, germline and somatic deletions of ARMC5 could be examined by ddPCR, which permits rapid and accurate evaluation of the ARMC5 allelic status.

Current smoking is an independent risk factor for new-onset diabetes mellitus during highdose glucocorticoid treatment.

AIMS/INTRODUCTION: Although high-dose glucocorticoids have been reported to cause new-onset diabetes mellitus (glucocorticoid-induced diabetes mellitus), its risk factors have remained to be determined. We investigated the risk factors related to glucocorticoid-induced diabetes mellitus diagnosed within 2 months after the high-dose treatment (newly treated with an initial high dose of > 20 mg prednisolone (PSL) equivalent per day for at least more than 6 months) in collagen vascular diseases. METHODS: A total of 2,631 patients with collagen vascular diseases was registered between 1986 and 2006 in the Chiba-Shimoshizu Rheumatic Cohort. We analyzed 681 patients newly treated with high-dose glucocorticoid who did not have diabetes mellitus and/or its previous diagnosis (age: 46.3 ± 16.7 years, PSL dose: 40.0 ± 14.1 mg/day). Glucocorticoid-induced diabetes mellitus was diagnosed by two or more glucose measurements in patients with fasting glycaemia ≥ 7 mmol/L and 120 minutes post-load glycaemia ≥ 11.1 mmol/L. RESULTS: Glucocorticoid-induced diabetes mellitus was observed in 26.3% of patients, and the glucocorticoid-induced diabetes mellitus group had higher age, higher BMI, lower rates of females and systemic lupus erythematosus, higher rates of smoking, alcohol use, and microscopic polyangiitis. Multivariate logistic regression analysis demonstrated that the risk of glucocorticoid-induced diabetes mellitus was independently higher in every 10-year increment of initial age with adjusted odds ratio (OR) 1.556 (95% confidence interval: 1.359 - 1.783), in every 1 kg/m2 increment of BMI with OR 1.062 (1.002 - 1.124), in current smoking with OR 1.664 (1.057 - 2.622), and in every 10 mg increment of initial dose of prednisolone with OR 1.250 (1.074 - 1.454). CONCLUSIONS: High-dose glucocorticoids caused diabetes mellitus with high prevalence within a short period, and current smokers should be considered at higher risk of glucocorticoidinduced diabetes mellitus in addition to age, BMI, and initial dose.

Effects of serotonin on expression of the LDL receptor family member LR11 and 7-ketocholesterol-induced apoptosis in human vascular smooth muscle cells.

Serotonin (5-HT) is a known mitogen for vascular smooth muscle cells (VSMCs). The dedifferentiation and proliferation/apoptosis of VSMCs in the arterial intima represent one of the atherosclerotic changes. LR11, a member of low-density lipoprotein receptor family, may contribute to the proliferation of VSMCs in neointimal hyperplasia. We conducted an in vitro study to investigate whether 5-HT is involved in LR11 expression in human VSMCs and apoptosis of VSMCs induced by 7-ketocholesterol (7KCHO), an oxysterol that destabilizes plaque. 5-HT enhanced the proliferation of VSMCs, and this effect was abolished by sarpogrelate, a selective 5-HT2A receptor antagonist. Sarpogrelate also inhibited the 5-HT-enhanced LR11 mRNA expression in VSMCs. Furthermore, 5-HT suppressed the 7KCHO-induced apoptosis of VSMCs via caspase-3/7-dependent pathway. These findings provide new insights on the changes in the differentiation stage of VSMCs mediated by 5-HT.

Adhesion of enterohemorrhagic Escherichia coli O157:H7 to the intestinal epithelia is essential for inducing secretory IgA antibody production in the intestine of mice.

We examined whether adherence of enterohemorrhagic Escherichia coli (EHEC) O157:H7 to intestinal epithelial cells contributed to the induction of secretory immunoglobulin A (IgA) antibody production in mice. Wild-type EHEC O157:H7 and its mutants deficient in the espA, sepL, tir and eae genes, encoding adherent factors on the locus of enterocyte effacement (LEE), were inoculated intragastrically into mice. Inoculation of wild-type EHEC induced fecal IgA antibodies specific to EHEC at 4 weeks after the inoculation, but that of espA- and sepL-deletion mutants did not. Furthermore, even 4 inoculations at weekly intervals with espA-deletion mutant, heat-killed wild-type EHEC and nonpathogenic E. coli did not induce fecal IgA antibodies, although these bacterial inoculations induced serum antibodies. Kanamycin (Km)-treated mice showed prolonged and similar fecal shedding of Km-resistant mutants of EHEC O157:H7 including A2-F6 having intact LEE, A6-E7 (sepL-insertion mutant), G1-E11 (tir-insertion mutant) and Δeae (eae-deletion mutant). In this case, A2-F6 induced fecal IgA antibodies, but the other mutants with defective LEE did not. In contrast to the fecal IgA antibodies, serum IgM and IgG antibodies were induced in mice inoculated with any of the LEE defective mutants as well as A2-F6. Thus, adhesion of EHEC to epithelial cells is essential for inducing the mucosal immune response in the intestine but not for inducing the systemic immune response.

Metformin reduces circulating malondialdehyde-modified low-density lipoprotein in type 2 diabetes mellitus.

PURPOSE: Type 2 diabetes is known to be associated with increasing cardiovascular mortality. Malondialdehyde-modified LDL (MDA-LDL) is an oxidized LDL and is increased in patients with diabetes or hypertriglyceridemia. Elevated MDA-LDL has been reported to be a risk factor of atherosclerosis or cardiovascular disease. Sitagliptin is a dipeptidyl peptidase-4 inhibitor and a new class of hypoglycemic agents. In this study, the effects of increasing the dose of metformin and add-on sitagliptin on MDA-LDL were examined in type 2 diabetes patients. METHODS: Seventy patients with type 2 diabetes, inadequately controlled despite on-going treatment with metformin 500 mg/day, were enrolled in this randomized controlled trial. The patients received additional metformin (500 mg/day) or sitagliptin (50 mg/day) for 6 months, and changes in metabolic parameters including MDA-LDL were evaluated. RESULTS: After 6 months of treatment, add-on sitagliptin (n=35) improved fasting blood glucose (FBG) and hemoglobin A1c (HbA1c) to significantly greater extent than increasing the dose of metformin (n=35). There were no differences in total cholesterol and low-density lipoprotein cholesterol levels between two groups. MDA-LDL levels (mean ± S.E.) decreased significantly with increasing the dose of metformin (from 94.40 ± 6.35 to 77.83 ± 4.74 U/L, P < 0.005), but remained unchanged with add-on sitagliptin treatment (from 89.94 ± 5.59 to 98.46 ± 6.78 U/L, p > 0.05). Multiple linear regression analysis identified increasing the dose of metformin treatment as the only independent factor associated with decreased MDA-LDL (ß coefficient 0.367, P < 0.0119), and no significant correlation between change in MDA-LDL and fasting blood glucose or HbA1c. CONCLUSION: These results suggest that increasing the dose of metformin improves serum MDA-LDL levels in type 2 diabetes mellitus.

C5 variant is associated with decreased risk of coronary artery disease in outpatients with severe hypercholinesterasemia.

OBJECTIVE: Although the C5 variant of cholinesterase is known to be a cause of hypercholinesterasemia, the pathophysiological significance of the C5 variant and the C5 variant-related hypercholinesterasemia in cardiovascular diseases remain unclear. The present study aimed to clarify the pathophysiological significance of the C5 variant as a risk or protective factor for coronary artery disease (CAD) in patients with severe hypercholinesterasemia. METHODS: Severe hypercholinesterasemia was defined as serum cholinesterase (ChE) activity >= 450 IU/L (>= 2.0 SD). We screened 11,648 consecutive outpatients between 2005 and 2011 at Toho University, Sakura Medical Center. In patients with severe hypercholinesterasemia, phenotyping of the C5 variant was conducted using polyacrylamide gel electrophoresis and alpha-naphthyl butyrate staining. RESULTS: 157 subjects (1.4% of 11,648 outpatients screened) were diagnosed with severe hypercholinesterasemia (mean serum ChE activity 574 ± 109 IU/L), and the frequency of the C5 variant was 45.2%. Subjects with the C5 variant had higher age, lower body mass index, milder dyslipidemia and liver dysfunction, and lower rates of hypertension and CAD compared with subjects without the C5 variant. Multivariate logistic regression model demonstrated that the presence of C5 variant independently lowered the risk of CAD, with odds ratio 0.071 (95% confidence interval (CI) 0.007 - 0.763, p = 0.029). CONCLUSION: The prevalence of the C5 variant was relatively high, and the C5 variant is associated with decreased risk of CAD in outpatients with severe hypercholinesterasemia.