DMN Group Research Articles

PLGA Nanoparticle-Based Dissolving Microneedle Vaccine of Clostridium perfringens ε Toxin

Epsilon toxin (ETX) is an exotoxin produced by type B and D Clostridium perfringens that causes enterotoxemia or necrotic enteritis in animals such as goats, sheep, and cattle. Vaccination is a key method in preventing such diseases. In this study, we developed a new type of dissolving microneedle patch (dMN) with a nanoparticle adjuvant for enhanced immune response to deliver the rETXY196E-C protein vaccine. We chose FDA-approved poly(lactic-co-glycolic acid) (PLGA) to prepare nanospheres as the vaccine adjuvant and introduced dimethyldioctadecylammonium bromide (DDAB) to make the surface of PLGA nanoparticles (PLGA NPs) positively charged for antigen adsorption. PLGA NPs with a diameter of 100~200 nm, a surface ZETA potential of approximately +40 mV, and good safety were successfully prepared and could effectively adsorb rETXY196E-C protein. Using non-toxic and antibacterial fish gelatin as the microneedle (MN) matrix, we prepared a PLGA-DDAB dMN vaccine with good mechanical properties that successfully penetrated the skin. After immunization of subcutaneous (SC) and dMN, antibody titers of the PLGA and Al adjuvant groups were similar in both two immune ways. However, in vivo neutralization experiments showed that the dMN vaccines had a better protective effect. When challenged with 100 × LD50 GST-ETX, the survival rate of the MN group was 100%, while that of the SC Al group was 80%. However, a 100% protective effect was achieved in both immunization methods using PLGA NPs. In vitro neutralization experiments showed that the serum antibodies from the dMN and SC PLGA NPs groups both protect naive mice from 10 × LD50 GST-ETX attack after being diluted 20 times and could also protect MDCK cells from 20 × CT50 GST-ETX attack. In conclusion, the PLGA-DDAB dMN vaccine we prepared has good mechanical properties, immunogenicity, and protection, and can effectively prevent ETX poisoning. This provides a better way of delivering protein vaccines.

Machine learning models for diabetic neuropathy diagnosis using microcirculatory parameters in type 2 diabetes patients.

Diabetic peripheral neuropathy (DPN) is a primary cause of diabetic foot, early detection of DPN is essential. This study aimed to construct a machine learning model for DPN diagnosis based on microcirculatory parameters, and identify the most predictive parameters for DPN. Our study involved 261 subjects, including 102 diabetics with neuropathy (DMN), 73 diabetics without neuropathy (DM), and 86 healthy controls (HC). DPN was confirmed by nerve conduction velocity and clinical sensory tests. Microvascular function was measured by postocclusion reactive hyperemia (PORH), local thermal hyperemia (LTH), and transcutaneous oxygen pressure (TcPO<inf>2</inf>). Other physiological information was also investigated. Logistic regression (LR) and other machine learning (ML) algorithms were used to develop the model for DPN diagnosis. Kruskal-Wallis Test (non-parametric) were performed for multiple comparisons. Several performance measures, such as accuracy, sensitivity and specificity, were used to access the efficacy of the developed model. All the features were ranked based on the importance score to find features with higher DPN predictions. There was an overall decrease in microcirculatory parameters in response to PORH and LTH, as well as TcPO<inf>2</inf>, in DMN group compared to DM group and HC group. Random forest (RF) was found to be the best model, and achieved 84.6% accuracy along with 90.2% sensitivity and 76.7% specificity. RF_PF% of PORH was the main predictor of DPN. In addition, diabetic duration was also an important risk factor. PORH Test is a reliable screening tool for DPN, which can accurately distinguish DPN from diabetics using RF.

Granulin Expression in Hamsters during Opisthorchis viverrini Infection-Induced Cholangiocarcinogenesis

The secreted growth factor granulin (GRN) is upregulated during diverse epithelial cancers. GRN stimulates cell growth and development while inhibiting apoptosis. Orthologues of vertebrate granulins evolved in other animals including the liver fluke Opisthorchis viverrini. Curiously, liver fluke granulin, termed Ov-GRN-1 promotes cholangiocarcinogenesis during chronic opisthorchiasis but, by contrast, limited information is available concerning mammalian GRN during liver fluke infection-induced cholangiocarcinoma (CCA). Here we investigated the expression of mammalian granulin in the O. viverrini-associated a hamster model of opisthorchiasis and liver fluke infection-induced CCA. Male Syrian golden hamsters were assigned to one of four treatment groups, each group included 30 hamsters: 1) normal (control), 2) infected with O. viverrini (OV); 3) exposed to N-dimethylnitrosamine in drinking water (DMN); and 4) infected with O. viverrini and exposed to DMN (OVDMN). Immunohistochemistry using an anti-granulin specific probe for mammalian granulin was undertaken to monitor expression and location in hepatobiliary tissues of the hamsters. In parallel, cognate studies of transcription of mRNA and protein. Histopathological examination revealed development of proliferative lesions from the onset and eruption of CCA onwards, an outcome that was most prominent in the OVDMN hamsters. Proliferating cell nuclear antigen (PCNA) index rose continuously from initiation of infection and increased with lesion progression in OV, DMN and markedly in OVDMN hamsters. Expression of GRN in biliary was elevated in biliary epithelial cells in CCA lesions in hamsters in the DMN and OVDMN groups. Expression of GRN as assayed by western blot and RT-PCR reflected the same trend as seen with PCNA. Together the histopathogical and molecular assay based findings revealed marked expression of granulin during cholangiocarcinoma in these hamsters, and highlighted the prospect that granulin represents a potential prognostic marker for cholangiocarcinoma.

Decreased risk of cholangiocarcinogenesis following repeated cycles of Opisthorchis viverrini infection-praziquantel treatment: Magnetic Resonance Imaging (MRI) and histopathological study in a hamster model

It has been suggested that repeated infection of Opisthorchis viverrini followed by repeated treatment with praziquantel (PZQ) increases risk of development of cholangiocarcinoma (CCA). Evidence for the prediction has accumulated based on findings of indirect approaches involving molecular changes and epidemiological trends. By contrast, here we directly monitored the impact of repeated liver fluke infection and treatment with PZQ on cholangiocarcinogenesis in a rodent model of human opisthorchiasis, using magnetic resonance imaging (MRI) and histopathology. Twenty five Syrian golden hamsters were assigned to five treatment groups: 1) infection with O. viverrini (OV group), 2) treatment with the carcinogen N-nitrosodimethylamine (NDMA) at 12.5ppm (DMN), 3) O. viverrini infection in tandem with NDMA (OD), 4) O. viverrini infection, NDMA, and treatment with PZQ (ODP), and 5) uninfected, untreated control. The repeated infections were established by intragastric inoculation of 50 metacercariae of O. viverrini to the OV, OD and ODP hamsters at weeks 0, 5 and 10. PZQ at 300mg/kg body weight was given to each hamster of the ODP group on weeks 4, 9 and 13 (four weeks after each infection). Imaging by MRI was undertaken on weeks 5, 10 and 14 (i.e. one week after each PZQ treatment). MRI revealed that the ODP hamsters did not develop CCA, whereas necropsy at week 40 revealed CCA in hamsters of the OD and DMN groups. Findings for histopathology and for proliferating cell nuclear antigen index conformed to the MRI findings. In overview, and notwithstanding that the immune response of individual hosts may play roles in cholangiocarcinogenesis, three cycles of the infection with O. viverrini followed treatment of the infection with PZQ did not increase the risk of bile duct cancer in this hamster model of liver fluke infection-induced CCA.

Implant based differences in adverse local tissue reaction in failed total hip arthroplasties: a morphological and immunohistochemical study.

BackgroundAdverse local tissue reaction (ALTR) is characterized by periprosthetic soft tissue inflammation composed of a mixed inflammatory cell infiltrate, extensive soft tissue necrosis, and vascular changes. Multiple hip implant classes have been reported to result in ALTR, and clinical differences may represent variation in the soft tissue response at the cellular and tissue levels. The purpose of this study was to describe similarities and differences in periprosthetic tissue structure, organization, and cellular composition by conventional histology and immunohistochemistry in ALTR resulting from two common total hip arthroplasty (THA) implant classes.MethodsConsecutive patients presenting with ALTR from two major hip implant classes (N = 54 patients with Dual-Modular Neck implant; N = 14 patients with Metal-on-Metal implant) were identified from our prospective Osteolysis Tissue Database and Repository. Clinical characteristics including age, sex, BMI, length of implantation, and serum metal ion levels were recorded. Retrieved synovial tissue morphology was graded using light microscopy and cellular composition was assessed using immunohistochemistry.ResultsLength of implantation was shorter in the DMN group versus MoM THA group (21.3 [8.4] months versus 43.6 [13.8] months respectively; p < 0.005) suggesting differences in implant performance. Morphologic examination revealed a common spectrum of neo-synovial proliferation and necrosis in both groups. Macrophages were more commonly present in diffuse sheets (Grade 3) in the MoM relative to DMN group (p = 0.016). Perivascular lymphocytes with germinal centers (Grade 4) were more common in the DMN group, which trended towards significance (p = 0.066). Qualitative differences in corrosion product morphology were seen between the two groups. Immunohistochemistry showed features of a CD4 and GATA-3 rich lymphocyte reaction in both implants, with increased ratios of perivascular T-cell relative to B-cell markers in the DMN relative to the MoM group (p = 0.032).ConclusionOur results demonstrate that both implant classes display common features of neo-synovial proliferation and necrosis with a CD4 and GATA-3 rich inflammatory infiltrate. Qualitative differences in corrosion product appearance, macrophage morphology, and lymphocyte distributions were seen between the two implant types. Our data suggests that ALTR represents a histological spectrum with implant-based features.

RE-EVALUATION OF TWO-DIMENSIONAL STRAIN IN THE PATIENTS WITH TYPE 2 DIABETES MELLITUS

ObjectivesBased on the ventricular-arterial coupling, the longitudinal strain (LS) of myocardium in the patients with type 2 diabetes mellitus (DM) were discussed in detail.MethodsEighty patients with DM were divided into...

Study on ventricular-arterial coupling in patients with type 2 diabetes mellitus by ultrasound

Objective To study the ventricular-arterial coupling in the patients with type 2 diabetes mellitus (DM) by ultrasound.Methods Eighty patients with DM were divided into 2 groups:normal left ventricular ejection fraction (EF) group (DMN,EF≥50 ％,n =40) and abnormal EF group (DMA,EF＜50％,n =40).At the same time,42 healthy volunteers were selected as a control group.The left ventricular (LV) relative wall thickness (RWT),mass index (LVMI),stroke volume (SV),stroke works (SW),ratepressure product (RPP),systemic vascular resistance index (SVRI),LV end-systolic elastance (Ees),arterial elastance index (Ea) and ventricular vascular index (VVI) were measured and calculated.Longitudinal strain (LS) of myocardial segment including base(LSBA),papillary muscle (LSPM) and apex (LSAP) were analyzed by two-dimensional speckle tracking imaging.Results (1) Compared with the control,the RWT and LVMI were all increased (P ＜0.01),but the LSBA,LSPM and LSAP were decreased (P ＜ 0.01) in the DMN and DMA groups (P ＜0.01).In DMA group,the RWT,LSBA,LSPM and LSAP were decreased (P ＜0.01).Compared with the DMN group,the LSBA,LSPM and LSAP were decreased in DMN group (P ＜0.01).(2) Compared with the control,the Ea,VVI,RPP and SVRI were increased,but the Ees was decreased (P ＜0.05)in DMA group (P ＜0.01).In the DMN,RPP was increased compared with the control group (P ＜0.01).Compared with the DMN group,the Ea,VVI,RPP and SVRI were increased,but Ees was decreased in the DMA (P ＜0.05).(3)The LSBA,LSPM and LSAP were all positively correlated with VVI(P ＜0.01).Multiple regression analysis showed that the LSAP,RWT,LVMI and SW were independent predictor of VVI (b' =-0.613,-0.290,0.290 and-0.205,P ＜ 0.05).Conclusions The ventricular-arterial coupling is normal in the DMN group,but is abnormal in DNA group.LSAP is independent predictors for ventricular-arterial coupling. Key words: Echocardiography; Diabetes mellitus,type 2; Ventricular function,left; Ventriculararterial coupling

Hyperfiltration and effect of nitric oxide inhibition on renal and endothelial function in humans with uncomplicated type 1 diabetes mellitus

Studies of experimental diabetes mellitus (DM) suggest that increased nitric oxide (NO) bioactivity contributes to renal hyperfiltration. However, the role of NO in mediating hyperfiltration has not been fully elucidated in humans. Our aim was to examine the effect of NO synthase inhibition on renal and peripheral vascular function in normotensive subjects with uncomplicated type 1 DM. Renal function and brachial artery flow-mediated vasodilatation (FMD) were measured before and after an intravenous infusion of the NO synthase inhibitor N(G)-nitro-l-arginine methyl ester (l-NMMA) in 21 healthy control and 37 type 1 DM patients. Measurements in DM participants were made under clamped euglycemic conditions. The effect of l-NMMA on circulating and urinary NO metabolites (NO(x)) and cGMP and on urinary prostanoids was also determined. Baseline characteristics were similar in the two groups. For analysis, the DM patients were divided into those with hyperfiltration (DM-H, n = 18) and normal glomerular filtration rate (GFR) levels (DM-N, n = 19). Baseline urine NO(x) and cGMP were highest in DM-H. l-NMMA led to a decline in GFR in DM-H (152 ± 16 to 140 ± 11 ml·min(-1)·1.73 m(-2)) but not DM-N or healthy control participants. The decline in effective renal plasma flow in response to l-NMMA (806 ± 112 to 539 ± 80 ml·min(-1)·1.73 m(-2)) in DM-H was also exaggerated compared with the other groups (repeated measures ANOVA, P < 0.05), along with declines in urinary NO(x) metabolites and cGMP. Baseline FMD was lowest in DM-H compared with the other groups and did not change in response to l-NMMA. l-NMMA reduced FMD and plasma markers of NO bioactivity in the healthy control and DM-N groups. In patients with uncomplicated type 1 DM, renal hyperfiltration is associated with increased NO bioactivity in the kidney and reduced NO bioactivity in the systemic circulation, suggesting a paradoxical state of high renal and low systemic vascular NO bioactivity.

The protective effect of thalidomide on left ventricular function in a rat model of diabetic cardiomyopathy

To evaluate the protective effect of thalidomide, a potent anti-inflammatory drug, on the development of diabetic cardiomyopathy (DMCMP). We induced type 1 diabetes using streptozocin in 8-week-old Sprague-Dawley rats, divided them into two groups-a thalidomide treatment group (DM-T, n = 15) and a non-treatment group (DM-N, n = 15)-and compared them with a normal control (n = 10). Ten weeks after diabetes induction, heart and lung mass indices were higher in the DM-N group compared with the control group. In the DM-T group, increases in heart and lung mass indices were attenuated compared with the DM-N group. On echocardiographic examination, systolic and diastolic mitral annulus velocities were impaired in the DM-N group, but they remained normal in the DM-T group. On haemodynamic analyses, left ventricular (LV) systolic function, represented by end-systolic elastance (0.35 ± 0.14 vs. 0.18 ± 0.07 mmHg/μl, P < 0.001) and preload-recruitable stroke work (90.5 ± 24.3 vs. 51.8 ± 22.0 mmHg, P < 0.001), was preserved in the DM-T group compared with the DM-N group. Likewise, deterioration of LV diastolic function was attenuated in the DM-T group. Increases in serum levels of TNF-α were attenuated in the DM-T group compared with the DM-N group. On histological analysis, thalidomide treatment lowered total myocardial collagen content and the expression of TNF-α, IL-1β, ICAM-1, and VCAM-1. In an animal model of DMCMP, deterioration of LV systolic and diastolic function was partially prevented by thalidomide treatment.

Antifibrotic effects of CGX, a traditional herbal formula, and its mechanisms in rats

Aim CGX is a modification of a traditional herbal medicine for “liver cleaning,” which is used to treat various chronic liver disorders in oriental clinics. This study investigated the antifibrotic effects and associated mechanisms of CGX. Materials and methods Liver fibrosis was induced in rats by dimethylnitrosamine (DMN; 10 mg kg −1, ip) injection on 3 consecutive days per week for 4 weeks. CGX (100 or 200 mg kg −1, po) was administrated once a day for 4 weeks. Three cell lines (HepG2, RAW 264.7, and HSC-T6) were used to examine its mechanisms. Results CGX treatment dramatically ameliorated the change in liver and spleen weight and serum albumin ( p < 0.01), aspartate transaminase ( p < 0.01), alanine transaminase ( p < 0.01), alkaline phosphatase ( p < 0.01), and total bilirubin ( p < 0.01) levels. Histopathologically, CGX administration decreased necrosis, inflammatory cell infiltration, and collagen accumulation. The antifibrotic effects of CGX were confirmed from hydroxyproline determination and the reduction in the numbers of activated hepatic stellate cells. In addition, antioxidant proteins, glutathione content, and glutathione peroxidase, catalase, and superoxide dismutase activities were maintained in the CGX-treated groups compared with the DMN group. CGX downregulated fibrosis-related genes (inducible nitric oxide synthase, tumor necrosis factor-alpha, transforming growth factor-beta, connective tissue growth factor, and platelet-derived growth factor-beta) and decreased the protein levels of profibrotic cytokines (transforming growth factor-beta and platelet-derived growth factor-beta) in liver tissues. In the cell line-based studies, CGX showed supportive effects, such as the protection of hepatocytes from CCl 4-toxicity, inhibition of NO production in RAW 264.7 cells, and inactivation of hepatic stellate cells. Conclusion These results demonstrated the antifibrotic effects of CGX and the corresponding mechanisms associated with sustaining the antioxidative system and inhibiting hepatic stellate cell activation via the downregulation of fibrogenic cytokines.

Inhibition of hepatic stellate cell activation following Yinchenhao decoction administration to dimethylnitrosamine‐treated rats

In an effort to investigate the mechanism by which Yinchenhao decoction (YCHD) acts on liver injury, we investigated the potential antifibrogenic effects of YCHD in an experimental liver fibrosis rat model, with special focus on the mechanisms inhibiting the activation and promoting apoptosis of hepatic stellate cells (HSC). The rats were initially randomized into two groups: the control (n = 10) and dimethylnitrosamine-treated (DMN; n = 30) groups. DMN (10 mg/kg body weight) was administered intraperitoneally to the DMN-treated rats for three consecutive days each week. At the end of the second week, three rats from the control and six rats from the DMN-treated groups were killed for the fibrosis development assessment. The remaining DMN rats were further randomized into two groups: the DMN-water group (n = 12) and the DMN-YCHD group (n = 12). Both groups continued to receive weekly DMN treatment for another 2 weeks in addition to daily administration of either water or YCHD, which were given intragastrically at a dose of 0.418 g/100 g body weight. Hepatic hydroxyproline content decreased and had improved histopathology in the DMN-YCHD rats. Compared to the DMN group, alpha-smooth muscle actin (SMA) and CD68 expression in the DMN-YCHD group was reduced significantly; however, alpha-SMA-positive HSC apoptosis was not observed by confocal microscopy; Fibrogenic proteins (tissue inhibitor matrix proteinases-1 and 2 and matrix metalloproteinase [MMP]-2/14) and cytokines (tumor necrosis factor-alpha and transforming growth factor-beta(1)) were decreased; MMP-9 was significantly upregulated. Yinchenhao administration attenuates liver fibrosis at least in part by inhibiting HSC activation directly, rather than promoting cell apoptosis of activated HSC, and the suppressive activation of Kupffer cells.

Acquired Loss of Erythrocyte Complement Receptor Type 1 in Patients with Diabetic Nephropathy Undergoing Hemodialysis

Background: Complement receptor type 1 on erythrocytes (E-CR1) plays important roles not only in the regulation of complement activation, but also the clearance of immune complexes. Reduced E-CR1 was previously found in patients undergoing hemodialysis (HD). We investigated whether the E-CR1level in HD patients with diabetic nephropathy (DMN) is decreased. The levels of decay accelerating factor (DAF) and CD59 on erythrocytes (E) were also determined to ascertain whether the loss of CR1 is a specific phenomenon or other complement regulatory proteins are also affected. Methods: The levels of CR1, DAF, and CD59 on E were analyzed in 176 HD patients with DMN, 101 HD patients with non-diabetes mellitus renal diseases (non-DMN), and 108 healthy individuals. Hind III restriction fragment length polymorphism of intron 27 of the CR1 gene was analyzed. The serum-soluble CR1 levelwas measured by ELISA. Results: The E-CR1 level was significantly lower in the DMN group than the non-DMN group (p < 0.0001) and healthy individuals (p < 0.05). The E-CR1 level was significantly higher in the non-DMN group than in healthy individuals (p < 0.01). The levels of E-DAF and E-CD59 were significantly lower in the DMN group than non-DMN group (DAF, p < 0.01; CD59, p < 0.0001). Within each genotype of the CR1 gene, the E-CR1 level was significantly lower in the DMN group than in the non-DMN group and healthy individuals (non-DMN, p < 0.01; healthy individuals, p < 0.05). The serum-soluble CR1 level was significantly higher in the DMN group than non-DMN group and control group (p < 0.01 each). However, soluble CR1 did not correlate with E-CR1. Conclusion: Acquired loss of E-CR1 was found among HD patients with DMN. From the viewpoint of host defense, it may be a prognostic factor.

Proanthocyanidin exposure to B6C3F1 mice significantly attenuates dimethylnitrosamine-induced liver tumor induction and mortality by differentially modulating programmed and unprogrammed cell deaths

Proanthocyanidins are of current interest as chemopreventive agents. The potential of the pre-, post- and co-exposure of proanthocyanidin-rich grape seed extract (GSPE) in preventing, reducing and/or delaying dimethylnitrosamine ( N-nitrosodimethylamine, DMN)-induced liver tumorigenesis, carcinogenesis and mortality in male B6C3F1 mice was determined. Animals were divided into six groups: I—control, II—GSPE alone, III—DMN alone, IV—GSPE + DMN, V—DMN exposure (3 months) followed by GSPE diet (9 months) and VI—GSPE diet (3 months) + DMN (3 months) + control diet (6 months). DMN exposure (0–8 weeks: 5 mg/kg; 8–12 weeks: 10 mg/kg, i.p.) was limited to a total period of 3 months. GSPE was incorporated in laboratory chow (ADI: 100 mg/kg b.w.). Animals were sacrificed at 3 month intervals, and serum chemistry, liver histopathology, integrity of hepatic genomic DNA, antioxidant status, and rates of apoptotic and necrotic cell deaths were determined. DMN-induced liver tumor formation (85%) and animal lethality (38%) were powerfully antagonized by co-administration of GSPE + DMN (tumor positive: 45%; death: 11%). More than 75% of the DMN-treated animals had numerous tumors (five or more), which were significantly reduced in the GSPE + DMN group (35%). GSPE also negatively influenced other protocols specifically designed to test initiation and progression phases. Thus, GSPE was instrumental in modulating metabolic cascades and regulated orchestration of cell death processes involved during the multistage tumorigenic process. These results unraveled that long-term exposure to proanthocyanidin-rich grape seed extract may serve as a potent barrier to all three stages of DMN-induced liver carcinogenesis and tumorigenesis by selectively altering oxidative stress, genomic integrity and cell death patterns in vivo.

Right Ventricular Dysfunction in Asymptomatic Diabetic Patients

Diabetes is thoroughly evidenced to increase the risk of development of heart failure even in the absence of frequently coexisting comorbidities such as coronary artery disease and hypertension. Cardiac adverse effects influenced by diabetes have been demonstrated thus far for the left ventricle, particularly systolic and diastolic dysfunction identified at rest (1–9) and/or during exercise (10–11). To date, no information is available regarding the possible involvement of the right ventricle in the pathological process evoked by diabetes. In clinical practice, right ventricular dysfunction is relevant in a variety of disease states affecting both the course and prognosis (12–16), and therefore one may assume that right ventricular performance is also an important issue in diabetic patients. The objective of the present study was to assess right ventricular systolic and diastolic function using conventional echocardiography and tissue Doppler imaging (TDI) in diabetic patients without coronary artery disease and with and without coexisting hypertension. The study group encompassed 27 diabetic and normotensive patients (DMN group) and 36 patients with coexisting diabetes and hypertension (DMHT group) who were prospectively enrolled between October 2002 and May 2003. The control group included 33 healthy volunteers. Coronary artery disease was excluded according to negative dobutamine stress echocardiography, treadmill exercise electrocardiographic test, the absence of myocardial ischemia in 24-h electrocardiographic Holter monitoring, and, in 27 …

Increased leptin concentrations and lack of gender difference in Type 2 diabetic patients with nephropathy

Leptin plays an important role in the regulation of body weight and energy balance. Women have higher circulating leptin level than men. In this study, we examined serum leptin concentrations in Type 2 diabetic men and women with or without nephropathy. Fasting plasma glucose (FPG), lipid profile, and serum leptin concentrations were measured in 34 Type 2 diabetic patients with nephropathy (DMN), 12 normoalbuminuric Type 2 diabetic subjects (DM) and 34 non-diabetic control subjects, all matched for age and body mass index (BMI). Result: Patients with diabetic nephropathy had lower high-density lipoprotein cholesterol and higher triglyceride, FPG, urinary albumin/creatinine ratio (ACR) and serum creatinine than the other two groups. There was a significant trend in serum leptin concentrations ( P<0.001, analysis of variance ANOVA) across the three groups with the main difference being detected between DMN and control subjects (DMN: 17.5±16.8 ng/ml, DM: 14.6±10.5 ng/ml and control: 9.1±7.1 ng/ml). Women had higher serum leptin concentration than men in the control group (12.5±7.3 ng/ml versus 4.2±2.0 ng/ml, P=0.001) and in the DM group (18.9±11 ng/ml versus 8.6±5.9 ng/ml, P=0.07) whereas this gender difference was not observed in the DMN group (18.6±17.0 ng/ml versus 16.8±17.0 ng/ml, P=0.754). On multivariate analysis, ACR ( β=0.411, P<0.001) and BMI ( β=0.240, P=0.002) were independently associated with serum leptin concentrations ( R 2=0.194, F=22.1, P<0.001) in the whole group. In the DMN group, ACR ( β=0.370, P=0.016) was the only independent determinant of serum leptin concentrations ( R 2=0.159, F=11.4, P=0.016). Serum leptin concentrations were higher in Type 2 diabetic patients with nephropathy than normoalbuminuric diabetic patients and controls. Diabetic men with nephropathy had proportionally higher serum leptin such that the gender difference in leptin observed in non-nephropathic individuals was abolished.

Alteration in QT‐RR Relationship in Diabetic Patients with Autonomic Dysfunction

BackgroundImpaired autonomic nervous activity and changes in ventricular repolarization might be related to catastrophic events in diabetic patients.MethodsWe analyzed the QT‐RR relationship and autonomic activity with Holter monitoring in 20 diabetic patients and 10 controls. The diabetic group was divided into two groups: 9 patients with normal coefficient of variance of RR interval (CVRR &gt; 2.5%, DMN group) and 11 with low CVRR (CVRR≤; 2.5%, DML group). Mean hourly QT and RR intervals were determined using the first 10 sinus beats of every hour. Spectral power of heart rate variability was determined with the fast Fourier transform technique.ResultsThe DML group had shorter RR and QT intervals during sleep compared with the DMN group and control group. There was no difference in corrected QT interval among the three groups. The DML group showed decreased circadian variation of high‐frequency (HF) power, and steeper regression line and smaller correlation coefficient of QT‐RR relationship compared with other groups. There was an inverse correlation between InHF and the slope of the regression line of the QT‐RR relationship.ConclusionsThese results indicate that modulation of QT interval with heart rate is impaired along with impairment of cardiac parasympathetic activity in the DML group. Altered modulation of ventricular repolarization might contribute to the pathogenesis of catastrophic cardiac events in diabetic patients with neuropathy.

Synthesis and Surface-Confined Electrochemistry of Dimethoxynaphthalene Fused through Rigid Norbornylogous Spacers to Thiolate, Dithiolate, and Disulfide Groups

A series of compounds (1−8) has been prepared in which a 1,4-dimethoxynaphthalene (DMN) group and either two thiol groups or a disulfide moiety are attached to the respective termini of rigid “norbornylogous” hydrocarbon bridges, 3, 5, 7, 9, and 13 bonds in length. Each of the molecules forms an adsorbate monolayer when its CH2Cl2 solutions are exposed to etched gold. Chemically reversible one-electron oxidation of the DMN group is observed in CH2Cl2 or CH3CN at potentials of 0.54−0.67 V vs ferrocene. Repeated potential cycling demonstates less monolayer degradation (a) in CH2Cl2 compared to CH3CN and (b) at subambient temperatures. Fast cyclic voltammetry experiments establish that the electron-transfer rates between the naphthalene group and the electrode surface are very fast (>103 s-1).

Transcriptional changes in macrophage TNF-α expression following dimethylnitrosamine exposure in vivo

DMN exposure has been shown to increase macrophage cytotoxic activity against tumor targets both in vitro and in vivo. Since the production and expression of the macrophage-derived cytokine tumor necrosis factor-alpha (TNF-α) is associated with such anti-tumor activity, studies were performed to determine whether changes in TNF-α gene transcription and biosynthesis resulted following DMN exposure. Thioglycollate-elicited macrophages obtained from DMN-exposed animals displayed enhanced levels of constitutively expressed TNF-α transcripts compared to vehicle controls. Northern blot analysis of the time course expression of TNF-α following endotoxin (1 μg/ml) stimulation in vitro showed a significantly greater induction of TNF-α transcripts in macrophages from DMN-exposed than control animals, with peak levels detected between 30 and 120 min. Maximum endotoxin-induced TNF-α secretion occurred later than the accumulation of the transcripts, with greater secretion observed between 120 and 360 min. In contrast to endotoxin, stimulation with IFN-γ (100 U/ml) produced no changes in the level of TNF-α transcripts. However, stimulation of macrophages with IFN-γ did greatly enhance the surface expression of membrane-bound TNF-α in cells from the DMN-treated animals. Supernatants from media and endotoxin stimulated macrophage were tested for TNF-α activity against WEHI-164 cells. In media alone, a five-fold increase in TNF-α activity was observed at 6 h in supernatants from macrophage obtained from DMN-exposed animals compared to the vehicle group. Treatment of supernatants with either superoxide dismutase (SOD) or catalase to remove reactive oxygen products did not alter their lytic capacity. However, addition of a neutralizing murine-anti-TNF-α antibody reduced the lytic capacity of the supernatants by 90% in both treatment groups. Accumulation of IL-1β transcripts gradually increased over the 6 h with a concomitant increase in secreted IL-1β that was identical in both DMN and vehicle groups. These results demonstrate that DMN exposure: (1) enhances the expression of TNF-α in peripheral macrophages by transcriptional regulatory mechanism(s) and, (2) does not alter the expression or secretion of IL-1β.

Effect of copper administration on the incorporation of [3H]-thymidine into the liver DNA of rats stimulated by dimethylnitrosamine and diethylnitrosamine.

The incorporation of [3H]thymidine into liver DNA of rats increased 6--8 times 48 h after a single injection of dimethylnitrosamine (DMN, 30 mg/kg) and diethylnitrosamine (DEN, 100 mg/kg). To test the suppressive effect of copper, the incorporation of [3H]thymidine into liver DNA in the DMN groups or DEN groups pretreated with copper was measured 48 h after the administration of DMN or DEN. The incorporation of [3H]thymidine into liver DNA of rats stimulated by the injection of DMN was strikingly suppressed by the injection of cupric acetate (20 mg Cu/kg), but that of rats stimulated by the injection of DEN was not suppressed by the injection of copper. Some other metal salts, silver nitrate (20 mg Ag/kg), nickel acetate (20 mg Ni/kg) and basic lead acetate (20 mg Pb/kg) did not significantly suppress the incorporation of [3H]thymidine stimulated by DMN or DEN. The accumulation of copper was much higher in the liver of copper-administered rats than that of nickel or lead in the liver of nickel-administered rats or lead-administered rats. The accumulation of silver was comparatively high in the liver of silver-administered rats.

The syncarcinogenic effect of methylcholanthrene and dimethylnitrosamine in Swiss mice.

The combined application of MCA (i.g.) and DMN (i.p.) to Swiss mice resulted in an increased incidence of tumors and in a decreased tumor latency, compared with the animals treated with each compound alone. In the MCA+DMN group the rate of lung carcinomas and neoplasms of the kidney was found to be significantly enhanced, while the DMN group showed an increase of vascular tumors. In the MCA group malignant lymphomas were seen more frequently than in the other groups. The findings concerning the lung and kidney were interpreted as a syncarcinogenic effect of the combined application of MCA + DMN.