Central motor conduction studies in motor neurone disease using magnetic brain stimulation

Central motor conduction (CMC) to abductor digiti minimi (ADM) was evaluated in 22 patients with motor neurone disease (MND) using magnetic stimulation of the motor cortex and electrical stimulation at the C7/T1 interspace. CMC was abnormal in 14 patients; prolonged CMC time and absence of response to brain stimulation were more frequent abnormalities than low amplitude responses without prolonged CMC time. The technique can reveal subclinical upper motor neurone involvement and document central motor pathway dysfunction in MND. The patterns of abnormality are not specific to MND; all may occur in other neurological disorders including multiple sclerosis.     

F responses and central motor conduction in multiple sclerosis

Two electrophysiological investigations were used to study 18 patients with multiple sclerosis — F wave characteristics including amplitude, persistence and frequency, which can provide a measure of motoneurone excitability, and magnetic stimulation of the cortex, which enables measurement of conduction along central motor pathways. There was an increase in the mean amplitude and persistence of the F response in patients with abnormal central motor conduction (CMC), although no correlation between the degree of abnormality of CMC and increase in F response amplitude was found. Increase in mean amplitude and persistence of the F response were also found in patients with normal CMC but clinical evidence of a UMN disorder (spasticity and/or weakness); there was no correlation, however, between any single F response characteristic and any particular clinical sign. CMC appears to be the preferred test for detecting subclinical motor lesions in MS: of the patient sides with normal clinical examination, 36% showed abnormal CMC, whereas 23% showed abnormal F responses.     

Cell-mediate cytotoxicity in vitro of human lymphocytes against a tissue culture melanoma cell line (igr3).

Purified peripheral blood lymphocytes from 13 healthy donors, 6 melanoma patients and 1 halo nevus patient were tested for cytotoxic activity against an allogeneic melanoma cell line (IGR3) in, at least, one of the following assays: cell-mediated cytotoxicity (ADCC) and microcytotoxicity assays (ma). The lymphocytes were isolated by Ficoll-Triosil gradient centrifugation (fraction F) followed by removal of iron-phagocytosing and adherent cells (fraction FFF) and by subsequent passage through anti-IgG columns (fraction FFF-C). Leukocytes of each fraction were identified by different methods including morphology, rosette-formation, phagocytic activity, and membrane fluorescence. CMC activity paralled ADCC activity at a log lower level of sensitivity. In both assays lymphocytes of fractions F and FFF had the highest activity, whereas in fraction FFF-C cytotoxicity was strongly reduced. In all three lymphocyte fractions CMC and ADCC activity could be blocked by preincubation of the effector cells in aggregated IgG. Furthermore, depletion of E rosette-forming lymphocytes slightly increased ADCC and CMC activity, whereas depletion of EA and EAC rosette-forming lymphocytes strongly decreased it. Our results therefore indicate that in both CMC and ADCC assays, non-adherent, non-phagocytic Fc receptor-bearing lymphocytes ("K" cells) were the active cytotoxic cells. In MA, on the other hand, mononuclear phagocytes seemed to be the most active cell population. So far no significant difference was observed in CMC, ADCC, and MA between control persons and melanoma patients     

Cell-mediated cytotoxicity in relation to active immunotherapy in acute myeloid leukaemia

Summary Tests comparing lymphocytes from normal controls, remission AML patients not receiving immunotherapy, and AML immunotherapy patients showed that cell-mediated cytotoxicity (CMC) to allogeneic AML cells requires in vivo priming and in vitro stimulation by AML cells. An in vitro lymphoproliferative response to allogeneic AML cells correlated with the development of CMC in immunotherapy-primed lymphocytes, though proliferation did not always lead to CMC. This may be due to differences in the lytic susceptibility of different AML cells. AML-stimulated CMC was cross-reactive on normal allogeneic PHA-transformed lymphoblasts and on lymphoblastoid cell lines (LCL). There was extensive cross-reactivity on allogeneic AML targets not used as in vitro stimulators. However, LCL generally did not induce CMC to allogeneic AML cells. CMC was generally absent, except in a few tests, on autologous PHA-transformed lymphoblasts following in vitro stimulation with allogeneic AML cells. CMC on autologous AML cells was equivocal, with little evidence for cross-reactive tumour-associated antigens in AML. Whilst CMC in this system was correlated with in vivo priming by immunotherapy, it is unlikely that such restimulated lymphocytes are the mediators of host-leukaemic cell cytolysis in vivo.     

Clonal Strain Persistence of Candida albicans Isolates from Chronic Mucocutaneous Candidiasis Patients

Chronic mucocutaneous candidiasis (CMC) is a primary immunodeficiency disorder characterised by susceptibility to chronic Candida and fungal dermatophyte infections of the skin, nails and mucous membranes. Molecular epidemiology studies of CMC infection are limited in number and scope and it is not clear whether single or multiple strains inducing CMC persist stably or are exchanged and replaced. We subjected 42 C. albicans individual single colony isolates from 6 unrelated CMC patients to multilocus sequence typing (MLST). Multiple colonies were typed from swabs taken from multiple body sites across multiple time points over a 17-month period. Among isolates from each individual patient, our data show clonal and persistent diploid sequence types (DSTs) that were stable over time, identical between multiple infection sites and exhibit azole resistant phenotypes. No shared origin or common source of infection was identified among isolates from these patients. Additionally, we performed C. albicans MLST SNP genotype frequency analysis to identify signatures of past loss of heterozygosity (LOH) events among persistent and azole resistant isolates retrieved from patients with autoimmune disorders including CMC.     

黄芪甲甙对实验性肺纤维化大鼠Cathepsin B表达的影响

目的:探讨黄芪甲甙时实验性肺纤维化大鼠肺组织中组织蛋白酶B(Cathepsin B,CB)表达的影响。方法:36只SD大鼠,随机分为对照组、模型组和干预组。模型组和干预组气管内注射博来霉素(BLM,5mg/kg)诱导肺纤维化,对照组在相同条件下给予生理盐水。第二天起干预组大鼠每天经胃管灌服0．1g/L黄芪甲甙2ml,其余两组相同条件下给予助溶剂羧甲基纤维素钠。治疗的第7d和28d,处死动物取出肺组织,进行病理学观察;测定28d肺组织中羟脯氨酸含量;用免疫组化和RT-PCR观察各组鼠肺组织CB蛋白及mRNA表达的水平。结果:病理学观察显示:与模型组比较,干预组肺泡炎和纤维化程度均减轻;模型组羟脯氨酸含量显著增高,而干预组与模型组比较羟脯氨酸含量明显降低;与模型组比较,干预组CB蛋白及mRNA表达显著降低。结论:黄芪甲甙可能通过押制CB的过度表达,对实验性大鼠肺纤维化具有良好的治疗作用。     

Molecular Cloning and Expression of Cellulase Genes from Ruminococcus albus 8 in Escherichia coli Bacteriophage lambda

A genomic library of Ruminococcus albus 8 DNA was constructed by using the Escherichia coli bacteriophage lambdaDASH. Recombinants were screened for cellulolytic activity by plating in soft agar (0.7%) overlays containing either 1% (wt/vol) carboxymethyl cellulose (CMC), 4-methylumbelliferyl-beta-d-cellobioside (MUC, 1 mg/ml), or 1% (wt/vol) Ostazin brilliant red-hydroxyethyl cellulose (OBR-HEC). One hundred and three recombinant phage exhibiting activity against OBR-HEC were found, and these fell into different classes based on the size of the zone of hydrolysis. Twenty-one recombinant phage exhibiting activity against CMC and 19 recombinant phage exhibiting activity against MUC were isolated. Four OBR-HEC, five CMC, and seven MUC clones were further analyzed by restriction endonuclease mapping and cellulase substrate specificity to identify unique clones and to determine their cellulase type. Three different clone types representing endoglucanase activity were identified. Three clones that appeared to encode exoglucanase type activity and four clones that had a mixed specificity, including beta-glucosidase activity, were also identified.     

Plasma protein advanced glycation end products, carboxymethyl cysteine, and carboxyethyl cysteine, are elevated and related to nephropathy in patients with diabetes

In Diabetes Mellitus (DM), glucose and the aldehydes glyoxal and methylglyoxal modify free amino groups of lysine and arginine of proteins forming advanced glycation end products (AGEs). Elevated levels of these AGEs are implicated in diabetic complications including nephropathy. Our objective was to measure carboxymethyl cysteine (CMC) and carboxyethyl cysteine (CEC), AGEs formed by modification of free cysteine sulfhydryl groups of proteins by these aldehydes, in plasma proteins of patients with diabetes, and investigate their association with the albumin creatinine ratio (ACR, urine albumin (mg)/creatinine (mmol)), an indicator of nephropathy. Blood was collected from forty-two patients with type 1 and 2 diabetes (18–36 years) and eighteen individuals without diabetes (17–35 years). A liquid chromatography-mass spectrophotometric method was developed to measure plasma protein CMC and CEC levels. Values for ACR and hemoglobin A1C (HbA1C) were obtained. Mean plasma CMC (μg/l) and CEC (μg/l) were significantly higher in DM (55.73 ± 29.43, 521.47 ± 239.13, respectively) compared to controls (24.25 ± 10.26, 262.85 ± 132.02, respectively). In patients with diabetes CMC and CEC were positively correlated with ACR, as was HbA1C. Further, CMC or CEC in combination with HbA1C were better predictors of nephropathy than any one of these variables alone. These results suggest that glucose, glyoxal, and methylglyoxal may all be involved in the etiology of diabetic nephropathy.     

Ploidy levels and the number of nuclei in cardiomyocytes of the lamprey and fish

It is well known that polyploidization of cardiomyocytes (CMC) is an essential component of heart growth in the warm-blooded vertebrates. Using the Feulgen cytophotometry of alkali-dissociated cells, we determined the ploidy in CMC of the lower vertebrates: lamprey Lampetra fluviatilis (Cyclostomata), skate Bathyraja maculata (Chondrostei), sterlet Acipenser ruthenus, and Russian sturgeon Acipenser güldenst?dti (Ganoids), as well as paradise fish Macropodus opercularis, Amur sleeper Perccottus glehni, and Atlantic salmon Salmo solar (Teleostei). The data obtained have demonstrated a wide variety in CMC ploidy of both cyclostomata and fishes. About 85% of the lamprey CMC contain 2 or more (up to 17) nuclei per cell; with 90 and 10% of the nuclei being, respectively, diploid and tetraploid. Hearts of the skate and sturgeons contain mononucleated diploid CMC. In the perch-like fishes, mononucleated diploid and mononucleated tetraploid CMC make, respectively, 95 and 5%. The salmon heart contains near 50% of mononucleated diploid CMC, 13% of mononucleated tetra- and octaploid CMC, the rest CMC being multinucleated (up to 6 nuclei per cell). In all the examined species, the increased nuclear ploidy is accompanied with a significant increase in the nuclear volume. The number of nucleoli per nucleus does not correlate with the nuclear ploidy level. Evolutionary aspects of CMC polyploidy in chordates are discussed.     

Enhancement of cell-mediated cytotoxicity by recombinant tumor necrosis factor (TNF alpha)

The effects of recombinant tumor necrosis factor (rTNF alpha) on the immune responses were investigated. A single iv injection of rTNF alpha (6 x 10(3) U) caused regression of sarcoma-180 transplanted into BALB/c nu/+ mice, but failed to regress this tumor in nu/nu mice. A higher dose of rTNF alpha (2 x 10(4) U) was necessary to induce antitumor effect in nu/nu mice. A host-related factor seemed to be involved in mediating tumor regression. Therefore, the effects of rTNF alpha on various T-dependent immune responses, including delayed footpad reaction (DFR), cell mediated cytolysis (CMC), and plaque-forming cells (PFC) were examined in BALB/c mice, immunized ip with chicken erythrocytes (CRBC). A single injection of rTNF alpha, at the time of the antigen administration, induced the augmentation of CMC to CRBC in a dose-dependent manner. DFR and PFC were not affected in optimal immunization procedures. The TNF alpha injection, at or after the time of antigen administration, was more effective in inducing augmentation of CMC. The increase in CMC by TNF alpha was mediated by nonadherent, Thy 1.2, Lyt 2.2 positive cells and neutralization of TNF alpha by the anti-TNF alpha monoclonal antibody abolished the effect on CMC. These results indicated that the human recombinant TNF alpha induced changes in the T-cell-mediated responses.     

Atorvastatin inhibits osteoclastogenesis by decreasing the expression of RANKL in the synoviocytes of rheumatoid arthritis

IL-17 and related cytokines are direct and indirect targets of selective immunosuppressive agents for the treatment of autoimmune diseases and other diseases of pathologic inflammation. Insights into the potential adverse effects of IL-17 blockade can be drawn from the experience of patients with deficiencies in the IL-17 pathway. A unifying theme of susceptibility to mucocutaneous candidiasis is seen in both mice and humans with a variety of genetic defects that converge on this pathway. Mucocutaneous candidiasis is a superficial infection of mucosal, nail or skin surfaces usually caused by the fungal pathogen Candida albicans. The morbidity of the disease includes significant pain, weight loss and secondary complications, including carcinoma and aneurysms. This review describes the known human diseases associated with chronic mucocutaneous candidiasis (CMC) as well as the known and proposed connections to IL-17 signaling. The human diseases include defects in IL-17 signaling due to autoantibodies (AIRE deficiency), receptor mutations (IL-17 receptor mutations) or mutations in the cytokine genes (IL17F and IL17A). Hyper-IgE syndrome is characterized by elevated serum IgE, dermatitis and recurrent infections, including CMC due to impaired generation of IL-17-producing Th17 cells. Mutations in STAT1, IL12B and IL12RB1 result in CMC secondary to decreased IL-17 production through different mechanisms. Dectin-1 defects and CARD9 defects result in susceptibility to C. albicans because of impaired host recognition of the pathogen and subsequent impaired generation of IL-17-producing T cells. Thus, recent discoveries of genetic predisposition to CMC have driven the recognition of the role of IL-17 in protection from mucosal fungal infection and should guide counseling and management of patients treated with pharmacologic IL-17 blockade.     

Motilin and a synthetic enkephalin induce colonic motor complexes (CMC) in the conscious dog

In the conscious dog migrating and nonmigrating colonic motor complexes (CMC) were recorded by means of chronically implanted strain gauge force transducers. Intravenous injection of a synthetic enkephalin analogue immediately induced a premature CMC at all three recording sites of the colon. Naloxone inhibited the enkephalin- but not motilin-induced CMC. We therefore exclude that motilin acts by release of endogenous enkephalins. The two peptides stimulate CMC in the canine colon by different pathways. Naloxone alone had no effect on normal colonic motility, suggesting strongly that endogenous enkephalins do not modulate regular CMC in the dog.     

Effects of cellulose, carboxymethylcellulose and inulin fed to rats as single supplements or in combinations on their caecal parameters

We compared the effect of diets containing different nondigestible carbohydrates: cellulose (C), inulin (IN) and carboxymethylcellulose (CMC) as single supplements or in dietary combination on caecal physiology of rats. Sixty male Wistar rats (Rattus norvegicus) were divided into five groups and for 4 weeks were fed a casein diet with the compared carbohydrates (4% of diet) or a combination of IN+C or IN+CMC (both 4+4%). Diet intake and FCR index remained unaffected by the treatments, whereas IN improved the body weight gain of rats compared to CMC. Compared to C group, all diets containing IN and CMC decreased the caecal pH as well as enlarged the caecum, thus increasing the weights of contents and tissue, especially upon CMC treatment. Rats given carboxymethylcellulose (CMC and IN+CMC groups) had watery caecal digesta, and some of them suffered from diarrhoea. In the case of CMC, the caecal enlargement was due to tissue hypertrophy and digesta accumulation mostly in response to an increased bulk of contents. Unlike C+IN, the dietary combination of CMC- and inulin-enhanced fermentation in the caecum of rats, however the proportion of acetate, propionate and butyrate was less beneficial. Compared to CMC, inulin gave a higher concentration of SCFA, especially of butyrate and propionate. The action of inulin in the caecum of rats could be pronounced by dietary treatment combined with CMC.     

Enzyme biotechnology development for treating polymers in hydraulic fracturing operations

Carboxymethyl cellulose (CMC) is a polymer used in many different industrial sectors. In the oil and gas industry, CMC is often used during hydraulic fracturing (fracking) operations as a thickening agent for effective proppant delivery. Accumulations of CMC at fracture faces (known as filter cakes) can impede oil and gas recovery. Although chemical oxidizers are added to disrupt these accumulations, there is industrial interest in developing alternative, enzyme-based treatments. Little is known about CMC biodegradation under fracking conditions. Here, we enriched a methanogenic CMC-degrading culture and demonstrated its ability to enzymatically utilize CMC under the conditions that typify oil fields. Using the extracellular enzyme fraction from the culture, significant CMC viscosity reduction was observed between 50 and 80˚C, at salinities up to 20% (w/v) and at pH 5–8 compared to controls. Similar levels of viscosity reduction by extracellular enzymes were observed under oxic and anoxic conditions. This proof-of-concept study demonstrates that enzyme biotechnology holds great promise as a viable approach to treating CMC filter cakes under oilfield conditions.     

Cell-mediated cytotoxicity in carcinoma of the human urinary bladder

Summary Lymphocytes from patients with tumors of the bladder or other unrelated tissues or with non-malignant genitourinary (GU) conditions and from normal subjects were tested in a microcytotoxicity assay against long- and short-term cultures (T24 and BT) derived from bladder carcinoma and several other target cell types, to determine the validity of the hypothesis that cell-mediated cytotoxicity (CMC) in bladder cancer patients is a specific disease-related phenomenon. At the effector cell level, lymphocytes from bladder cancer patients displayed uniformly greater cytotoxicity for T24 and BT cells than those from normal donors. This proclivity was shared by the lymphocytes of patients with non-GU cancers but not of those with non-malignant GU disorders. At the target cell level, CMC was observed less frequently against non-bladder tumor targets than against T24 and BT cells and the CMC of bladder cancer patients did not differ significantly from that of the other groups. The pattern of CMC observed against the bladder tumor-derived targets was thus one of target cell sensitivity rather than tumor-specificity and disease-related only to the extent that the CMC of patients with cancer was greater overall than of healthy subjects. Abrogation of CMC by passage of lymphocytes through immunoglobulin-coated columns indicated that the effector cells were principally of non-T type, bearing a superficial resemblance to those in normal individuals which induce non-disease related CMC.     

Human influenza viral neuraminidases augment cell-mediated cytotoxicity in vitro

Previously, we reported that influenza virus-induced cell-mediated cytotoxicity (CMC) was largely due to its glycoproteins, hemagglutinin and neuraminidase (NA). These observations were based on the use of a single influenza virus strain, the A/Port Chalmers/3/73 (H3N2), and these were considered insufficient to generalize that all human influenza virus NAs augment CMC. Therefore, antigenically different NAs of human influenza strains were used to study whether (a) all NAs possess the potential to stimulate NK activity and (b) does the enzymatic activity of NA play a role in the CMC stimulation. Biologically active preparations of N1 subtype NA (A/USSR/90/77 (H1N1) and N2 subtype NAs (A/Aichi/2/68 (H3N2) and A/Port Chalmers) were evaluated for NK activity stimulation in an overnight radiolabeled chromium-release assay consisting of human peripheral blood lymphocytes and K562 target cells. The level of CMC stimulation was the same at equivalent protein concentrations with all the NAs tested. The addition of homologous NA-monospecific antibody almost completely reduced the CMC stimulation, while the addition of homosubtypic antibody reduced the CMC by 56-75%. However, in the presence of heterosubtypic monospecific antibody, NA-augmented CMC was reduced by 27-47% in most experiments. The results suggest that the CMC stimulation site is probably the same in all NAs tested. This putative site is thermo-resistant and is independent of the conformational change of the NA molecule. Furthermore, it is distinct from the enzymatic and probably from the antigenic sites.     

Immobilizing Cu,Zn-superoxide dismutase in hydrogels of carboxymethylcellulose improves its stability and wound healing properties

Hydrogels of carboxymethylcellulose (CMC) with 50 and 90% cross-linking degree (CMC50% and CMC90%, respectively) were prepared and loaded with bovine erythrocyte Cu,Zn-superoxide dismutase (SOD) to obtain two drug delivery systems: SOD-CMC50% and SOD-CMC90%. Resistance of native SOD to inactivation by H₂O₂ and the effect of applying SOD-CMC hydrogels to open wounds of rats’ back skin were examined and compared to that of SOD trapped into CMC50% and CMC90% hydrogels. Also, the effect of CMC50% and SOD-CMC90% on human fibroblasts proliferation was evaluated at different times. It was found that SOD in the hydrogel was more resistant to H₂O₂ inactivation than the native enzyme and at the same time it reduced the time necessary for wound healing. Furthermore, the highest cell proliferation value was found for the CMC50% hydrogels, which had a three-dimensional structure suitable for gas and nutrient exchanges and improving cell life conditions. Published in Russian in Biokhimiya, 2006, Vol. 71, No. 12, pp. 1627–1632.     

Immunoassays for carbodiimide modified DNA-detection of unpairing transitions in supercoiled ColE1 DNA.

The water soluble reagent N-cyclohexyl-N'-beta-(4-methylmorpholinium) ethyl carbodiimide-p-toluene sulphonate (CMC) can be used to probe for unpaired and mismatched sites in DNA. Polyclonal antibodies for CMC modified DNA were produced in order to develop immunological assays for the localization and quantitation of CMC adducts. Immunoslot blot analysis of modified DNA exhibited antibody binding proportional to the extent of CMC modification with adduct detection in the femtamole range. Unmodified DNA did not cross react under the conditions of the assay. The distribution of CMC reactivity for supercoiled ColE1 DNA modified at 100, 200 and 300 mM NaCl was determined by immunoanalysis of EcoRI-Hae2-NruI restriction fragments Southern transferred to nylon membranes. Reactivity above random expectation occurred in the A2-II fragment which can be accounted for by its high A-T content of 71.3%. Reactivity below random expectation occurred in the C fragment which can be accounted for by its low AT content of 43%. CMC modification for the other restriction fragments appeared random.     

Differences in thymus dependency among the alloreactive T-cell subpopulations in their development

Effects of thymectomy at various times after birth (Tx-1, Tx-3, Tx-7, Tx-14) on mixed lymphocyte reaction (MLR), cell-mediated cytotoxicity (CMC), and graft-versus-host reaction (GVHR) were examined by experiments with spleen cells obtained at 8 weeks of age. All of MLR, CMC, and GVHR were detected in spleen cells of Tx-14 mice. However, the ability of spleen cells to induce GVHR was abolished by thymectomy at 7 days after birth. On the other hand, MLR and CMC were not affected in such mice. In Tx-1 or Tx-3 mice, only MLR was detected in spleen cells. These results suggest that thymus dependency of T cells responsible for MLR are lower in their maturation than those for CMC and that thymus dependency of T cells responsible for GVHR are the highest among these T-cell subpopulations.     

Endothelins specifically recognize lysophosphatidylcholine micelles

Lysophosphatidylcholine (LPC), a major phospholipid component of oxidized low‐density lipoprotein (ox‐LDL), is implicated in numerous inflammatory diseases, including atherosclerosis. Here, to clarify the relationship between bioactive endothelins (ETs) (which are considered to be potent proinflammatory mediators) and LPC/ox‐LDL, we investigated the interaction between ETs and LPC/ox‐LDL by fluorescence spectroscopy and western blotting. Tryptophan fluorescence measurements revealed ETs specifically interacted with LPC at concentrations that exceeded the critical micelle concentration (CMC). The tryptophan residue in ETs was not likely to be involved directly in the interaction between ETs and LPC micelles. Tryptophan fluorescence quenching revealed tryptophan residue in ETs where LPC concentrations were below the CMC may be buried deeply in the peptide or may interact with other amino acid residues, whereas tryptophan residue in ETs in the presence of LPC at concentrations exceeding the CMC was exposed outside of the peptide. Furthermore, ETs bind to ox‐LDL in a concentration‐dependent manner. These results strongly suggest that ox‐LDL contains micelle‐rich LPCs and that ETs specifically interact with the bioactive LPC micelles. Further study of the interaction between ETs and LPC micelles contained in ox‐LDL will provide important information on the development and progression of many inflammatory diseases, including atherosclerosis. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.