Concentration Of 2μM Research Articles

Differential dose-response effect of cyclosporine A in regulating apoptosis and autophagy markers in MCF-7 cells.

Cyclosporine A (CsA) is an immunosuppressant primarily used at a higher dosage in transplant medicine and autoimmune diseases with a higher success rate. At lower doses, CsA exhibits immunomodulatory properties. CsA has also been reported to inhibit breast cancer cell growth by downregulating the expression of pyruvate kinase. However, differential dose-response effects of CsA in cell growth, colonization, apoptosis, and autophagy remain largely unidentified in breast cancer cells. Herein, we showed the cell growth-inhibiting effects of CsA by preventing cell colonization and enhancing DNA damage and apoptotic index at a relatively lower concentration of 2µM in MCF-7 breast cancer cells. However, at a higher concentration of 20µM, CsA leads to differential expression of autophagy-related genes ATG1, ATG8, and ATG9 and apoptosis-associated markers, such as Bcl-2, Bcl-XL, Bad, and Bax, indicating a dose-response effect on differential cell death mechanisms in MCF-7 cells. This was confirmed in the protein-protein interaction network of COX-2 (PTGS2), a prime target of CsA, which had close interactions with Bcl-2, p53, EGFR, and STAT3. Furthermore, we investigated the combined effect of CsA with SHP2/PI3K-AKT inhibitors showing significant MCF-7 cell growth reduction, suggesting its potential to use as an adjuvant during breast cancer therapy.

New silver complexes with bioactive glycine and nicotinamide molecules – Characterization, DNA binding, antimicrobial and anticancer evaluation

This study introduces a pair of newly synthesized silver complexes, [Ag2(HGly)2]n(NO3)2n (1) and [Ag(Nam)2]NO3·H2O (2) (Gly – glycine, Nam – nicotinamide), that were prepared and characterized by relevant methods in solid state (elemental, spectral, thermal and structural analysis) and their stability in solution was verified by 1H NMR measurements. Moreover, suitable reaction conditions were observed by potentiometry depending on pH in case of binary system Ag-Gly. X-ray analysis confirmed argentophilic interactions in complex 1 with an Ag1-Ag2 distance of 2.8018(6) Å. Antimicrobial testing indicates higher growth inhibition effect of complex 1 than complex 2. Moreover the effectivity of both complexes against bacteria (Staphylococcus aureus and Escherichia coli) is superior (or similar) to that of the commercially available Ag(I) sulfadiazine, AgSD (used, for example, in Dermazine cream). The binding of the Ag(I) complexes to calf thymus DNA was investigated using electronic absorption, fluorescence and circular dichroism spectrophotometry. The Stern–Volmer quenching constants obtained from the linear quenching plot were estimated in the range from 2.01×103 to 20.34×103M−1. The results of topoisomerase I and topoisomerase II (Topo I and Topo II) inhibition assay suggested that complex 2 inhibits the enzyme activity of both enzymes at a concentration of 2μM. The cytotoxicity of both complexes on L1210 leukemia cells was revealed to be approximately three times higher than that of cisplatin. Moreover, the new Ag(I) complexes also induced apoptosis of the leukemia cells. The high DNA binding activity of these complexes is considered to be responsible for their cytotoxic effects.

Low-Dose Deguelin, Selective Targeting of Mt NPM1, Induces Differentiation in NPMc+ AML Cells By Regulation the Axis of SIRT1/CEBPβ/G-CSFR, Instead of HDAC1/3

AML with NPM1 mutation (Mt NPM1) showing aberrant cytoplasmic dislocation of NPM1 exhibit relative good response to induction therapy, however, still a proportion of NPMc+ AML can not be cured by conventional treatments. Therefore, the aim of the present study was to determine the therapeutic efficacy of deguelin, a natural rotenoid isolated from several plant species, which has been demonstrated extensive biological activity in various tumors with low lethal effect. In the 54th ASH meeting in 2012, we reported deguelin selectively reduced Mt NPM1, induced differentiation and potentiated apoptosis in acute myeloid leukemia cells carrying NPM1 mutation.In this paper, we will focus on the investigation of the effect and molecular of mechanisms of deguelin treatment in nontoxic concentration with long time duration on AML cells carrying Mt NPM1. Deguelin treatment at the differentiation-induced concentration of 2µM for 5 days significantly induced granulocytic/monocytic differentiation such as a lower nucleocytoplasmic ratio, peripherally located nucleus, and chromatin condensation in 66.7% of OCI-AML3 cells carrying Mt NPM1. However, these phenomena were not presented in the OCIM2 cells harboring Wt NPM1. The NBT reduction assays that used as another marker of differentiation of myeloid leukemia exhibited increased NBT reduction activity, and Fluorescence-activated cell sorting (FACS) analysis demonstrated significantly increased expression levels of CD11b/CD14/CSF-3R in deguelin-treated OCI-AML3 cells (CD11b: from 2.8% to 62%; CD14: from 2.6% to 32.3%, CSF-3R: from 0.7% to 21.6%, P£¼0.01). Furthermore, immunoblot analysis revealed deguelin significantly reduced the levels of Mt NPM1 and HDACs (HDAC1/HDAC3/SIRT1), but not Wt NPM1, accompanied by increasing the expression of proteins and mRNAs of CEBP¦Â and G-CSFR. In addition, immunoprecipitation and immunoblot analysis confirmed deguelin significantly down-regulated Mt NPM1 via the ubiquitin-proteasome pathway. In consistent with above results, sh-Mt NPM1 and sh-SIRT1 treatment induced differentiation of OCI-AML3 (sh-Mt NPM1: CD11b/CD14/CSF-3R respectively: from 15.5% to 62.3%; from 13.9% to 47.1%; from 0.6% to 18.4%; sh-SIRT1: CD11b/CD14/CSF-3R respectively: from 18.2% to 47.1%; from 19.2% to 43.7%; from 0.4% to 13.4%, P£¼0.01), which confirmed by morphological examination and NBT reduction assays, and also upregulated the levels of proteins and mRNAs of CEBP¦Â and G-CSFR. sh-Mt NPM1 treatment reduced the expression of SIRT1, which did not change the levels of HDAC1/3. In other sides, sh-SIRT1 treatment did not alter the levels of Mt NPM1, suggesting that the decline of SIRT1 was partially accountable for granulocytic/monocytic differentiation of AML cells with Mt NPM1. Finally, overexpression of Mt NPM1 was performed to determine whether the depletion of Mt NPM1 directly induces granulocytic/monocytic cell differentiation. Compared to GFP-expressing cells, the proportion of the differentiation antigens (CD11b/CD14) and differentiation gene CSF-3R declined in Mt NPM1-expressing cells after deguelin treatment (Mt NPM1-expressing-treated deguelin-induced CD11b/CD14/CSF-3R-positive cells respectively: from 47.5% to 23.1%; from 29% to 18.8%; from 23.2% to 6.5%, P£¼0.01). Importantly, the upregulation of CEBP¦Â and G-CSFR by deguelin was appreciably hampered in Mt NPM1 expressing cells, compared with GFP-expressing cells.Taken together, our studies suggest deguelin, selective silencing of Mt NPM1, which induces differentiation via the axis of SIRT1/CEBP¦Â/G-CSFR (Fig1) may represent a novel active differentiating agent like ATRA for NPMc+ AML.Footnotes * Asterisk with author names denotes non-ASH members [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Novel pyrazolopyrimidine derivatives targeting COXs and iNOS enzymes; design, synthesis and biological evaluation as potential anti-inflammatory agents

A novel set of 4-substituted-1-phenyl-pyrazolo[3,4-d]pyrimidine and 5-substituted-1-phenyl-pyrazolo[3,4-d]pyrimidin-4-one derivatives were synthesized and evaluated as potential anti-inflammatory agents. The newly prepared compounds were assessed through the examination of their in vitro inhibition of four targets; cyclooxygenases subtypes (COX-1 and COX-2), inducible nitric oxide synthase (iNOS) and nuclear factor kappa B (NF-κB). Compounds 8a, 10c and 13c were the most potent and selective ligands against COX-2 with inhibition percentages of 79.6%, 78.7% and 78.9% at a concentration of 2μM respectively, while compound 13c significantly inhibited both COX subtypes. On the other hand, fourteen compounds showed high iNOS inhibitory activities with IC50 values in the range of 0.22–8.5μM where the urea derivative 11 was the most active compound with IC50 value of 0.22μM. Most of the tested compounds were found to be devoid of inhibitory activity against NF-kB. Moreover, almost all compounds were not cytotoxic, (up to 25μg/ml), against a panel of normal and cancer cell lines. The in silico docking results were in agreement with the in vitro inhibitory activities against COXs and iNOS enzymes. The results of in vivo anti-inflammatory and antinociceptive studies were consistent with that of in vitro studies which confirmed that compounds 8a, 10c and 13c have significant anti-inflammatory and analgesic activities comparable to that of the control, ketorolac. Taken together, dual inhibition of COXs and iNOS with novel pyrazolopyrimidine derivatives is a valid strategy for the development of anti-inflammatory/analgesic agents with the probability of fewer side effects.

Zooplankton in the eastern tropical north Pacific: Boundary effects of oxygen minimum zone expansion

Oxygen minimum zones (OMZs) may be expanding in their worldwide spatial and vertical extent as a result of global climate change. Here, we highlight structural and functional features of pelagic OMZ zooplankton communities, especially the little known mesopelagic lower oxycline zooplankton assemblage in the eastern tropical north Pacific (ETNP). Day and night vertically-stratified zooplankton samples were collected with a MOCNESS plankton net system to depths of 1000m during two cruises in 2007 and 2008 as part of the Eastern Tropical Pacific Project. Size-fractionated biomass and selected species distributions were analyzed. A spatial comparison between two stations, Tehuantepec Bowl (TB) and Costa Rica Dome (CRD), with different OMZ thicknesses especially in 2008, was used as a proxy for future potential climate change to show how variability in oxycline depth at the upper and lower OMZ boundaries may have broad consequences for biological distributions and ecosystem function.The same zooplankton biomass features were present at both locations but responded differently to changes in OMZ thickness and boundary depths. The impact of habitat compression was quantified by the change in proportion of zooplankton biomass in the upper water column. The thermocline was the location of peak zooplankton biomass regardless of OMZ extent. At the lower oxycline, a unique zooplankton assemblage and secondary biomass peak (sharp order of magnitude jump from OMZ core biomass levels) occurred that was strongly associated with a specific oxygen concentration of 2μM. In 2008, the lower oxycline biomass peak deepened by over 200m at TB with its expanded OMZ compared to CRD, despite a temperature difference of ∼2°C, while remaining at the same oxygen level. In contrast, another secondary biomass peak corresponding to the daytime depth of diel vertical migration, and occurring within the upper oxycline or OMZ core, was present at the same depth and temperature at both locations, despite different oxygen concentrations. A dense monospecific layer of the copepod Eucalanus inermis was sometimes present just above the distinctive more persistent lower oxycline assemblage. While the general ontogenetic migration of this species was previously known, its precise positioning relative to oxygen concentration and the lower oxycline community is a new finding. The vertical re-positioning of biomass layers at the thermocline and lower OMZ boundaries, and the increased depth range of low oxygen water that diel vertical migrators and sinking particles must transit in an expanded OMZ, could have widespread effects on species distributions, the biological pump, and benthic–pelagic coupling.

Amphiphilic porphyrin-cardanol derivatives in Langmuir and Langmuir–Blodgett films applied for sensing

This study presents the preparation of a nanostructured films with an amphiphilic meso-porphyrin whose side chains are derived from cardanol, a byproduct of the cashew industry. The Langmuir films were prepared on acidified water to reducing the self-aggregation of the complex and the mean molecular area for the meso-porphyrin measured by surface pressure isotherms was 177Å2 and confirmed by compressibility modulus. The 700nm band observed in the Langmuir films supports the coexistence of monomers and dimers similar to the spectra of the solution (CHCl3/CH3COOH). The formation of bright block domains with different sizes and shapes for the monolayer disappears after surface pressure starts to increase (π≥2mNm−1), as observed by Brewster angle microscopy measurements. Results of UV–vis spectra of the Langmuir films and FTIR reflection of the Langmuir–Blodgett (LB) films of meso-porphyrin indicated that the molecules exhibit preferential orientation with the planes perpendicular to the water (Langmuir) and parallel to the substrate (LB) surface. The LB films were tested as a promethazine sensor, which detected a concentration of 2μM, a value greater than those obtained by other nanostructured systems.

Comparison of the binding characteristics of [18F]THK-523 and other amyloid imaging tracers to Alzheimer’s disease pathology

Extensive deposition of senile plaques and neurofibrillary tangles in the brain is a pathological hallmark of Alzheimer's disease (AD). Although several PET imaging agents have been developed for in vivo detection of senile plaques, no PET probe is currently available for selective detection of neurofibrillary tangles in the living human brain. Recently, [(18)F]THK-523 was developed as a potential in vivo imaging probe for tau pathology. The purpose of this study was to compare the binding properties of [(18)F]THK-523 and other amyloid imaging agents, including PiB, BF-227 and FDDNP, to synthetic protein fibrils and human brain tissue. In vitro radioligand binding assays were conducted using synthetic amyloid β(42) and K18ΔK280-tau fibrils. Nonspecific binding was determined by the addition of unlabelled compounds at a concentration of 2μM. To examine radioligand binding to neuropathological lesions, in vitro autoradiography was conducted using sections of AD brain. [(18)F]THK-523 showed higher affinity for tau fibrils than for Aβ fibrils, whereas the other probes showed a higher affinity for Aβ fibrils. The autoradiographic analysis indicated that [(18)F]THK-523 accumulated in the regions containing a high density of tau protein deposits. Conversely, PiB and BF-227 accumulated in the regions containing a high density of Aβ plaques. These findings suggest that the unique binding profile of [(18)F]THK-523 can be used to identify tau deposits in AD brain.

Cytotoxic and apoptotic effects of constituents from Millettia pachycarpa Benth

The aim of this study is to investigate the cytotoxic and apoptotic effects of constituents from the seeds of Millettia pachycarpa Benth. Fourteen compounds (1–14) including one novel chalcone (10) were isolated as active principles from Chinese herbal medicine M. pachycarpa Benth. Their structures were identified by using spectroscopic methods. All isolates were then evaluated for their cytotoxic effects against several cancer cell lines (HepG2, C26, LL2 and B16) with cisplatin as a positive control. And their apoptosis-inducing effects were tested against HeLa-C3 cells with taxol as a positive control. Both studies showed that compounds 1, 2, 7 and 10 demonstrated significant cytotoxic and apoptotic effects against cancer cells. Moreover, in the apoptosis assay the novel chalcone (10) showed strong apoptosis inducing effects at a concentration of 2μM within 36h. It was found to be the most potent apoptotic inducer of the compounds isolated from M. pachycarpa Benth.

Activation of TRPV1 by a Recombinant Double-Knot Toxin from a Chinese Bird Spider

TRPV1 is a tetrameric voltage-sensitive cation channel that is activated by heat and vanilloids. The architecture of TRP channels are thought to be related to Kv channels, with each subunit containing six transmembrane segments. In addition to being activated by capsaicin, TRPV1 is activated by Double-Knot Toxin (DkTx), a protein toxin purified from the Chinese Bird Spider, Selenocosmia huwena (Bohlen et al. 2010, Cell 141, 834-35). DkTx is unique in that it contains two Inhibitor Cysteine Knots (ICK) motifs connected by a peptide linker. Although these ICK motifs are related to those found in tarantula toxins that target voltage sensors in Kv channels, DkTx does not appear to interact with classical voltage-activated cation channels. We set out to explore the mechanism of DkTx activation of TRPV1, and began by producing the toxin in E. Coli and testing for activity against TRPV1. DkTx was expressed as a fusion with bacterial Ketosteroid Isomerase (KSI), cleaved from KSI using hydroxylamine, and purified using reverse phase HPLC. Reduced DkTx was folded in vitro in a solution containing (GSH/GSSG) and guanidine HCl, and the folding reaction monitored by HPLC. Using this procedure we obtained a predominant species of the toxin that was further purified by reverse phase HPLC. When tested for activity on TRPV1 expressed in Xenopus laevis oocytes, DkTx produced robust and slowly reversible activation of the channel when voltage clamped at −60 mV. At a concentration of 2μM, DkTx produced comparable activation to 2 μM capsaicin, suggesting that the apparent affinity of the recombinant toxin is similar to that reported for the native toxin. We are currently working to solve the structure of DkTx using NMR and further investigating it's mechanism of activation.

Histidine-rich glycoprotein binds factor XIIa with high affinity and inhibits contact-initiated coagulation

AbstractHistidine-rich glycoprotein (HRG) circulates in plasma at a concentration of 2μM and binds plasminogen, fibrinogen, and thrombospondin. Despite these interactions, the physiologic role of HRG is unknown. Previous studies have shown that mice and humans deficient in HRG have shortened plasma clotting times. To better understand this phenomenon, we examined the effect of HRG on clotting tests. HRG prolongs the activated partial thromboplastin time in a concentration-dependent fashion but has no effect on tissue factor–induced clotting, localizing its effect to the contact pathway. Plasma immunodepleted of HRG exhibits a shortened activated partial thromboplastin time that is restored to baseline with HRG replenishment. To explore how HRG affects the contact pathway, we examined its binding to factors XII, XIIa, XI, and XIa. HRG binds factor XIIa with high affinity, an interaction that is enhanced in the presence of Zn2+, but does not bind factors XII, XI, or XIa. In addition, HRG inhibits autoactivation of factor XII and factor XIIa–mediated activation of factor XI. These results suggest that, by binding to factor XIIa, HRG modulates the intrinsic pathway of coagulation, particularly in the vicinity of a thrombus where platelet release of HRG and Zn2+ will promote this interaction.

Modeling of the structure and interactions of the B. anthracis antitoxin, MoxX: deletion mutant studies highlight its modular structure and repressor function

Our previous report on Bacillus anthracis toxin-antitoxin module (MoxXT) identified it to be a two component system wherein, PemK-like toxin (MoxT) functions as a ribonuclease (Agarwal S et al. JBC 285:7254-7270, 2010). The labile antitoxin (MoxX) can bind to/neutralize the action of the toxin and is also a DNA-binding protein mediating autoregulation. In this study, molecular modeling of MoxX in its biologically active dimeric form was done. It was found that it contains a conserved Ribbon-Helix-Helix (RHH) motif, consistent with its DNA-binding function. The modeled MoxX monomers dimerize to form a two-stranded antiparallel ribbon, while the C-terminal region adopts an extended conformation. Knowledge guided protein-protein docking, molecular dynamics simulation, and energy minimization was performed to obtain the structure of the MoxXT complex, which was exploited for the de novo design of a peptide capable of binding to MoxT. It was found that the designed peptide caused a decrease in MoxX binding to MoxT by 42% at a concentration of 2μM in vitro. We also show that MoxX mediates negative transcriptional autoregulation by binding to its own upstream DNA. The interacting regions of both MoxX and DNA were identified in order to model their complex. The repressor activity of MoxX was found to be mediated by the 16N-terminal residues that contains the ribbon of the RHH motif. Based on homology with other RHH proteins and deletion mutant studies, we propose a model of the MoxX-DNA interaction, with the antiparallel β-sheet of the MoxX dimer inserted into the major groove of its cognate DNA. The structure of the complex of MoxX with MoxT and its own upstream regulatory region will facilitate design of molecules that can disrupt these interactions, a strategy for development of novel antibacterials.

Polychromatic Fingerprinting of Excitation Emission Matrices

The ability of a ubiquitous polychromatic excitation to support the fingerprinting of the Excitation Emission Matrices (EEM) of fluorescent indicators was reported. Solutions of fluorescent molecules, with concentration of 2µM for the absorption and 10µM for the emission measurements were prepared. Spectroscopic grade THF and distilled water passed through Milli-Q purification system were used as solvents. EEM measurements were carried out on a spectrofluorimeter, operating in the 390-700 nm detection range, with excitations at 20 nm intervals within the same range. During the measurement, the screen displayed a sequence of 50 colors and for every color, the transmission and emission spectrum of both the reference and the sample were recorded. Illuminating colors in the red region are not able to excite fluorescence and this can be seen in the spectra for red light.

AT 101, an Inhibitor of Bcl-2 Family Members Is Cytotoxic to a Heterogeneous Group of CLL Samples and Synergistic with Rituximab.

Bcl-2 family proteins include both pro-survival and pro-apoptotic factors. The balance of these regulatory proteins determines a cell's threshold for death. Bcl-2, Bcl-XL, and Mcl-1 are structurally related anti-apoptotic proteins of the Bcl-2 family. Constitutive expression of Bcl-2 is characteristic of CLL and has been implicated in the leukemia's pro-survival tendency. In addition, high level expression of Mcl-1 and Bcl-2 have been shown to correlate with important clinical outcomes in CLL such as poor response to chemotherapy and decreased overall survival. Bcl-2 prevents mitochondrial cytochrome c release and inhibits apoptosis through direct regulation of caspases. The BH3-only proteins dimerize and neutralize their pro-survival counterparts via interaction with the BH3 binding grove on the anti-apoptotic proteins. This interaction results in the release of cytochrome c, subsequent caspase activation, and resultant cell death. Gossypol is a naturally occurring product found in cottonseed oil. AT 101, an orally bio-available derivative of gossypol is currently being evaluated in cancer clinical trials. AT 101 is a small molecule that mimics the inhibitory BH3 domain of endogenous antagonists of Bcl-2 negating its cytoprotective role. AT 101 induces apoptosis via caspase activation in cancer cells that over-express Bcl-2 or Bcl-XL, but had no significant cytotoxic effect on normal blood mononuclear cells (Mohammad, 2005). Fluorescence polarization assays demonstrate that AT 101 binds to Bcl-2, BcL-XL, and Mcl-1 with greater affinity than gossypol (Prada,ASH 2005). We hypothesized that inhibiting Bcl-2 and Mcl-1 by AT 101 may allow the CLL cells to enter apoptosis more readily and render them more sensitive to standard CLL therapeutics. We tested 10 different primary CLL patient samples in vitro and found gossypol to be cytotoxic in a dose and time dependent manner. We observed 50% reduction in CLL cell viability at a concentration of 2μM after 48 hours of treatment. Significant cytotoxicity was observed irrespective of ZAP-70 expression or IgVH mutational status. Increased ratios of Bcl-2/Bax and Mcl-1/Bax have been demonstrated in CLL patients who do not respond to rituximab (Bannerji 2003). To examine the ability of gossypol to enhance the cytotoxic effects of rituximab in CLL we evaluated samples from 6 different patients affected with the leukemia in vitro. CLL cells treated with rituximab at 10μg/ml showed no significant decrease in viability compared to untreated cells. When AT 101 was added at 5μM concentrations in combination with rituximab 10μg/ml for twelve hours the average CLL viability was decreased by 67.2% over that of cells treated with rituximab alone (P<0.001 by Bonferroni multiple comparison test) and by 45.3% over cells treated with AT 101 alone (P<0.001). The combined effect of AT 101 and rituximab appears synergistic and is displayed throughout CLL subtypes. In our studies we were unable to reach the IC50 of rituximab on CLL using doses up to 100μg/ml. However, the addition of AT 101 to rituximab 10μg/ml surpassed the IC50 and demonstrated similar cytotoxicity to that of high dose rituximab100μg/ml when used in conjunction with AT 101. Together our results suggest that AT 101 may have therapeutic potential in CLL as a single agent or in concert with other known CLL therapeutics. In particular, AT 101 was found to have synergistic cytotoxicity when combined with rituximab. Currently a phase I clinical trial is underway to evaluate the activity of AT 101 in previously untreated patients with CLL.

Antisense Oligodeoxynucleotide Complementary to CXCR4 mRNA Block Replication of HIV-1 in COS Cells

CXCR4 is both a chemokine receptor and an entry co-receptor for the T-cell line-adapted human immunodeficiency virus type 1 (HIV-1). To find a more efficacious therapeutic treatement of acquied immunodeficiency syndrome, we exmined the effects of antisense oligonucleotides on CXCR4 production. COS cells, stably expressing CXCR4 and CD4, were incubated with several kinds of oligonucleotides. Total human p24 antigen production was determined using an enzyme-linked immunosorbent assay system. An antisense phosphorothioate-modified oligonucleotide, complementary to the translation region of the CXCR4 mRNA, showed minimal inhibition of p24 antigen production at the high concentration of 2μM. On the other hand, the antisense phosphorothioate oligonucleotide, when used with transfection reagents, showed high efficiency at low concentrations, and confirmed the sequence-specific action. Interestingly, the oligonucleotide with the natual phosphodiester backbone, when used with the transfection reagents, also had high functional effects, comparable to the modified oligonucleotide. This defines the prerequisite criteria necessary for the design and the application of antisense oligonucleotides against HIV-1 in vivo.

ADP-induced Platelet Aggregation in Diabetes mellitus

ADP-induced platelet aggregation at final concentration of 2μM (ADP-Agg.) was performed according to the method by Born using a Bryston aggregometer model 350. The sensitivity of the platelets was expressed in terms of the maximum percentage of light transmittance to that of platelet-poor-plasma within 5 minutes after the addition of ADP. Peripheral neuropathy was evaluated by motor nerve conduction velocity (M. C. V.). Retinopathy was evaluated by usual ophthalmoscopy.The subjects studied were 20 age-matched healthy subjects and 47 diabetic patients. These 47 diabetic patients were divided into following four groups according to their M. C. V. and with or without retinopathy. N (-) indicates the patient whose M. C. V. was not less than the mean M. C. V. of age-matched normal subjects. R (-) indicates the patient whose fundoscopic findings revealed nothing remarkable as to diabetic retinopathy, e. g. microaneurysm, retinal hemorrhage etc. The results of two-sample t test of ADP-Agg. were significant between the groups listed as follows. The figures in parenthesis indicate the number of subjects.P<0.001…Control (20)<Diabetics (47)P<0.01…R(-) N(-) (16)<R(+) N(+) (13)R(+) N(+), R(-) N(+), R(+) N(-) (31)>R(-) N(-) (16)P<0.05…R(-) (22)<R(+) (25)N(-) (28)<N(+) (19)R(+) N(-) (12)<R(+) N(+) (13)R(-) N(-) (16)<R(-) N(+) (6)These results indicate that platelets of N (+) patients are more sensitive to ADP than those of N (-) patients regardless of retinopathy.In the diabetic patients FBS and ∑BS (50gOGTT up to 2hrs.) correlated with ADP-Agg. but the serum cholesterol, triglycerides β-lipoprotein and ΔIRI30/ΔBS30 did not.From the foregoing, it appears that in the diabetics hyperglycemia per ce might play an important role in the observed platelets abnormalities of patients with peripheral neuropathy.